frühjahrstagung der deutschen pharmakologischen gesellschaft

13. FrOhjahrstagung der Deutschen Pharmakologischen Gesellschaft in Mainz vom 19. bis 22. M&rz 1972

EROFFNUNGSANSPRACHE

Introductory Address

SchUmann, H.-J.

In diesem Jahr treffen wir uns zum 13. Mal zu einer FrGh-

jahrs-Arbeitstagung in Mainz. Als Vorsitzender unserer Ge-

sellschaft m~chte ich Sie alle, G~ste wie Mitglieder, auch im

Namen des Vcrstandes herzlich willkommen heiBen.

Dem Herrn Rektor der Universit~t Mainz, Herrn Prof. Dr. P.

SCHNEIDER mGchte ich fGr die freundlichen Worte der BegrGBung

sehr herzlich danken. Ftthlen wir uns doch der Unlverslt~t

Malnz durch die Jahrelang gew~hrte Gastfreundschaft besonders

verbunden. Ich glaube, die meisten unserer Mitglieder kG~uen

sich den Ablauf unseres J~hrlichen Tagungsprogrammes ohne die

Mainzer FrUhjahrstagung nlcht mehr vorstellen, so sehr geh6rt

diese Veranstaltung in das Programm unserer Gesellschaft.

1960 land die 1. Arbeitstagung in Mainz start, dank der Inl-

tiative yon Herrn Kollegen KUSCHINSEY und der seiner Mitar-

beiter.

Die Deutsche Pharmakologische Gesellschaft hat durch ihre

Sprecher immer wieder unseren Mainzer Kollegen fGr ihre be-

reitwillige Mitarbeit bei der Gestaltung dieser Tagung ge-

dankt, und sie weiB die Bedeutung dieser Arbeitstage, die

ausschlieBlich den wissenschaftlichen Problemen unseres

Faches gewidmet sind, wohl zu wGrdigen. In diesem Jahr m6chte

ich jedoch Ihnen, verehrter Herr Kollege KUSCHINSKY, der Sie

mit Ihrer Emeritierung Ihre Funktionen der jtLugeren Genera-

tion Gbergeben, unseren ganz besonderen Dank aussprechen fGr

die Schaffung der Mainzer Tagung, der Pr~gung ihres Stiles

und der F~rderung ihres wissenschaftlichen Niveaus. - Wir

m~chten in diesem Zusammenhang auch den Beitrag Ihrer SchGler

besonders anerkennen und der Hoffnung Ausdruck geben, dab sie

R2

die schon zur Tradition gewordene Ausrichtung der Mainzer

Tage welter fGr uns alle Gbernehmen.

In diesemJahr haben wir ein sehr umfangreiches Progr~mm zu

bew~ltigen. Der frGhe Tagungsbeginn wurde erforderlich, um

trotz der stattfindenden je 3 Parallelsitzungen die angemel-

deten 241 Vortr~ge im Programm bis Mittwochabend unterzubrin-

geno 0bgleich der sprunghafte Anstieg der Vortragszahl sicher

mitbedingt ist durch die vielen gemeinsamen Tagungen mit be-

freundeten Gesellschaften, so meine ich doch, dab wir uns in

unserer Mitgliederversammlung und im Vorstand erneut mit die-

sem Problem befassen sollten. Wenn auch eine gewisse kriti-

sche Selbstbeschr~nkung bei der Anmeldung sicherlich nicht

von Schaden w~re, so wird sie doch zur LSsung unserer Proble-

me nur einen begrenzten Beitrag liefern kSnnen.

Die im vorigen Jahr vonder Mitgliedervers~mmlung beschlosse-

ne Bildung der Sektionen T0XIKOLOGIE und KLINISCHE P~AK0-

LOGIE soll durch die Verabschiedung je einer Gesch~ftsordnung

in der morgigen Mitgliederverssmmlung zu einem formellen Ab-

schluB gebracht werden. Da welter eine Erg~nzungswahl des

Vorstandes sowie die Anderung des Gesch~ftsordnungspunktes

Uber die Mitgliedsbeitr~ge und die freie Abgabe der Verhand-

lungsberichte an die Mitglieder zur Diskussion stehen wird,

bittet der Vorstand um das Erscheinen mSglichst vieler Mit-

glieder.

Meine Damen und Herren, da wir auch in diesem Jahr, wie Sie

wissen, keine eigenst~ndige Herbsttagung veranstalten, fGhle

ic~ mic~ aufgerufen, der im vergangenen Jahr verstorbenen

Mitglieder unserer Gesellschaft zu gedenken.

Wit beklagen den Tod von Prof. Dr. Arthur STOLL, Prof. Dr.

Leopold THER, Prof. Dr Heinrich HOFNANN, Dr. Dr. Max

SLA2ENGER, Prof. Dr. Richard LABES und Prof. Dr. Ludwig

SCHMIDT.

Am 19o Januar 1971 verstarb Prof. Dr. Arthur STOLL im geseg-

neten Alter von 84 Jahren. Am 8. Januar 1887 in Schinznach-

Doff als Sohn des Bezirksschulrektors geboren, studierte er

Chemie und promovierte 1911 in ZUrich bei seinem Lehrer

Prof. Richard WILST~TTER, der 1915 den Chemie-Nobelpreis fur

R3

seine Studien Gber das Chlorophyll erhielt. STOLL hatte schon

fGr das Diplom, dann fur die Promotion und schlieBlich als

Assistent im Privatlabor seines verehrten Lehrers auf dem

Chlorophyllgebiet mitgearbeitet. Er folgte WILST~TTER von

ZUrich nach Berlin-Dahlem ins Kaiser-Wilhelm-Institut und

1916 nach ~nchen, wo ihm schon 1917 der Professorentitel

verliehen wurde. Die mit WILSTATTER gemeinsam durchgefGhrten

Arbeiten fanden ihren Niederschlag in zwei Monographien:

t'Untersuchungen Gber Chlorophyll, Methoden und Ergebnisse"

und "Untersuchungen Gber die Assimilation der Kohlens~ure".

Die Verschlechterung der Lebensbedingungen w~hrend des 3.

Kriegsjahres veranlaBten STOLL,zum 1. Oktober 1917 seine Po-

sition in I~nchen aus und als p~armazeutischer Chemi-

ker zur Fa. S~DOZ nach Basel zu gehen. Seine Aufgabe als

Chemiker in der pharmazeutischen Industrie sah er besonders

darin, die sc~onenden Extraktionsmethoden WILSTATTERS, die im

Umgang mit dem empfindlichen Chlorophyll erarbeitet wurden,

fGr die Gewinnung und Reindarstellung anderer pflanzlicher

Wirkstoffe nutzbar zu machen, und diese damit der Medizin als

definierte Stoffe fGr eine rationelle Therapie zur VerfGgung

zu stellen~ 0bjekte dieser Untersuchungen waren Heilpflanzen,

die schon in der Volksmedizin angewendet wurden, wie Secale

cornutum, Digitalis, Scilla, Strophantus und Rauwolfia.

Bereits im ersten Anlauf gelang nach wenigen Monaten Arbeit

die Reindarstellung des Alkaloids Ergotamin aus dem Mutter-

kornpilz. Diese Leistung leitete eine neue Epoche ein, in der

die Gesamtdroge mit ihrer mSglichen Vielzahl von Wirkkompo-

nenten verdr~ngt wurde vom spezifisch wirksamen Einzelstoff.

Es ist das Verdienst Arthur STOLLS und seines Mitarbeiter-

teams, in den folgenden Jahren neben der Reindarstellung der

gesamten Mutterkornalkaloide auch die der genuinen Herzgly-

koside aus Digitalis lanata und purpurea sowie der Scilla

maritima erarbeitet zu haben. Auf dieser Basis entstanden in

Zus~mmenarbeit mit den Pharmakologen ins0esondere ROTMLI~ und

seiner Gruppe sowie der Klinik wichtige, exakt dosierbare

Heilmittel. Mit der weiteren Verbesserung der chemischen Me-

thodik wurde mit Beginn der 40er Jahre auch die Strukturana-

lyse der verwendeteten Wirkstoffe und schlieBlich ihre S~-

R4

these mSglicn. • diese Zeit f~llt die Strukturaufkl~rung der

Mutterkornalkaloide als Lysergs~urederivate, die Darstellung

von Lysergs~uredi~thylamid sowie der hydrierten Mutterkorn-

alkaloide, die sich als weniger toxisch erwiesen als die ge-

nuinen Alkaloide, wie z.B. das Dihydroergotamin. Das Ver-

zeichnis seiner Publikationen umfaBt 298 Titel.

Schon aus dieser kurzen Darstellung der wissenschaftlichen

Leistung von Arthur STOLL l~Bt sich sein Format als pharma-

zeutischer Chemiker ablesen. Er gehSrt zu den GroBen seines

Faches. Zahlreiche Ehrungen und Anerkennungen sind ihm zuteil

geworden. Insgesamt 19mal wurde ihm die Ehrendoktor~rde fGr

die Fachgebiete Naturwissenschaften, Pharmazie und Medizin

verliehen. Er wurde u.a. zum Mitglied der Deutschen Akademie

der Naturforscher (Leopoldina) zu Halle, der KSniglich Schwe-

dischen Akademie der Wissenschaften zu Stockholm und zum

Foreign Member of the Royal Society of London gew~hhlt. Die

Ehrenmitgliedschaft von mehr als 20 wissenschaftlichen Ge-

sellschaften und Akademlen wurde ihm angetragen u.a. der

Schwelzerlschen Akademie der Med. Wissenschaften, der

Schweiz. Gesellschaft fGr Analytische und Angewandte Chemie,

der Gesellschaft Deutscher Chemiker, der Deutschen Pharma-

zeutischen Gesellschaft und der Pharmaceutical Society of

Japan.

Wenlge Tage nach seinem 62. Geburtstag starb am 25. April

1971 nach schwerer Erankheit Prof. Leopold THER. 1909 in

Wien als Sohn eines Apothekers geboren, studierte er zu-

n~chst Pharmazie und nach seinem AbschluB sp~ter auch noch

Medizin an der deutschen Universit~t in Prag. Seine pharma-

kologische Ausbildung erhielt THER bei Prof. Richard LABES

in Jena von 1939 - 1945 und nach dem Kriege bei Prof. Otto

RIESSER in Frankfurt am Main. Von ibm empfing er entschei-

deride Impulse. Die Arbeiten Gber die pharmakologische Beein-

flussung des Kohlehydratstoffwechsels lassen seinen EinfluB

deutlich erkennen. ~ach der Habilitation im Jahre 1949 und

dem frGhen Tode RIESSERS trat THER als Pharmakologe in die

Farbwerke HOECPLST AG ein und Gbern~m 1953 die Leitung der

Pharmakologischen Forschungslaboratorien. Seine weitangeleg-

te Ausbildung sowie seine guten Sachkenntnlsse, insbesondere

R5

auf dem Gebiete der pharmakologischen Methodik, kamen ihm bei

der Bew~ltigung der Vielfalt von Problemen, die mit dem Aus-

bau der pharmakologischen Forschung verbunden waren, sehr zu-

gute. Seine Vorliebe fGr die pharmakologische Methodologie

wird durch die beiden von ihm verfaBten BGcher "Pharmakolo-

gische Methoden zur Auffindung der Arzneimittel und Gifte"

sowie "Grundlagen der experimentellen Arzneimittelforschung"

offenbar. Leider ist der 2. Teil des letztgenannten Buches,

der die speziellen pharmakologischen Methoden behandeln soll-

te, nicht mehr vollendet worden. 1957 wurde THER zum apl.

Professor fur Pharmakologie und Toxikologie an der Universi-

t~t Frankfurt ernannt. Er war ein stets hilfsbereiter, viel-

seitig interessierter Kollege, dessen Rat und Kritik Gewicht

batten, dessen aufgeschlossenes Wesen gepaart mit menschli-

cher W~rme ihm viele Freunde geschaffen habeno

Am 30. April 1971 starb Prof. Dr. Heinrich HOFMANN, Leiter

der Ern~hrungstoxikologischen Abteilung am Institut fGr Er-

n~hrungswissenschaft der Universit~t GieBen. 1909 in Aue

(Sachsen) geboren, studierte er Medizin an den Universit~ten

Rostock, Innsbruck und Leipzig. Seine pharmakologische Aus-

bildung erhielt HOFMANN im Leipziger Institut bei Prof. GROS.

Seiner Habilitation im Jahre 1941 folgte aus politischen

Grthaden die Ernennung zum Dozenten erst 2 Jahre sp~ter. 1949

wurde er auf den Lehrstuhl seines Faches nach Jena berufen,

den er schon seit 1945 kommissarisch leitete. Aus dieser Zeit

entst~mmt eine groBe Zahl wissenschaftlicher Publikationen,

die sich u.a. mit der Pharmakologie von Lokalanesthetika,

Analgetika, Muskelrelaxantien und insbesondere eines neuen

Barbiturs~urederivates, n~mlich dem Schlafmittel Kalypnon,

befassen. - 1961 wurde Heinrich HOFMAi~ aus politischen GrGn-

den aus seinem Amt entlassen und zu 13 Jahren 2ucnthaus ver-

urteilt. Durch Intervention von Freunden wurde 1967 seine

vorzeitige Entlassung mGglich. Seit dieser 2eit war Prof.

HOFMA~ in GieBen t~tig, wo er sich vornehmlich mit Toxizi-

t~tsproblemen yon SGBstos befaSte. Sein tragisches Schick-

sal weist uns erneut auf ~ie FroDleme im zweigeteilten

Deutschland bin.

R6

Im Alter yon 68 Janren verstaro Dr.rer.pol. et med. Max

SLAZE~GER. 1903 in Berlin geboren, war er w~hrend seiner

Assistentenzeit in den Jahren 1929 - 1931 Assistent am DGssel-

dorfer Pharmakologischen Institut bei HEUBNER un~ JARISCH

und bis zum Juni 1933 an der 2. s Abtellung aes ~ranken-

hauses ~erlin-~eukSlln. Danach ginger in die Emigration und

~nderte seinen Namen. Aus SCHLESINGER wurde SLAZENGER. Nach

langj~hriger T~tigkeit als Internist in Burma und Indien kehr-

te Max SLAZENGER 1959 nach Europa zurUck, wo er tells in der

Schweiz, tells in Deutschland t~tig war. Sein Schicksal wurde

wesentlich bestimmt durch den Nationalsozialismus, der ihn aus

seiner Heimat verdr~ngte und seine Entwicklung als wissen-

schaftlich t~tiger Arzt j~h unterbrach, wie mehrere Arbeiten

vor seiner Emigration,zusammen mit SCHLOSSMANN und RANDERATH

verfaBt, beweisen.

Am 21. Juni 1971 verschied Prof. Dr. med. et phil. Richard

LABES im Alter von 81 Jahren. Der in Dirschau geborene West-

preuBe studierte von 1909 - 1914 Medizin in Freiburg, Berlin

und ~thachen, nahm dann als Unterarzt am 1. Weltkrieg in RuB-

land tell und promovierte 1920 zum Dr. med. in Berlin. Nach

abgeschlossenem Chemiestudium promovierte er 1924 zum Dr.phil.

in Leipzig. 1925 ging Richard LABES als Assistent zu FUHNER

nach Bonn. Hier erhielt er seine Ausbildung in der Pharmakolo-

gie und Toxikologie und habilitierte 1928. Diese Jahre in Bonn

waren fur LABES wissenschaftlich besonders ertragreich. Seine

Arbeiten besch~ftigen sich, entsprechend seiner vielseitigen

Ausbildung in Medizin und Chemie, u.a. mit Problemen der Ner-

venerregung und insbesondere mit chemisch toxikologischen Pro-

blemen wie z.B. den Beziehungen zwischen der Wirkung von

Krampfgiftenund StSrungen der Gewebsatmung. 1935 wurde er auf

den Lehrstuhl fGr Pharmakologie an der Universit~t Jena beru-

fen, den er his zu seiner Zwangsevakuierung dutch die Amerika-

net im Jahre 1945 nach Heidenheim a.d.Brenz innehatte. Von

1950 his zur Emeritierung 1956 war er Gastprofessor an der

Universit~tBonn.

FUr uns alle unerwartet kam in den Tagen vor dem Weihnachts-

fest die Nachricht vom Tode unseres Kollegen Prof. Ludwig

SCHMIDT. Er wurde am 15. Dezember 1971 im Alter von 50 Jahren

R7

mitten aus seiner erfolgreichen T~tigkeit herausgerissen.

1921 im rum~nischen Kronstadt geboren, studierte er Medizin

in Danzig und Freiburg. Nach einer insgesamt 4-j~hrigen Aus-

bildung in der Anatomie, ~iochemie und Physiologie wurde er

1950 Assistent bei Prof. JANSEN im Freiburger Pharmakologi-

schen Institut. Schon 1953 erfolgte die Habilitation und

1959 die Ernennung zum apl. Professor. Nach seinem Eintritt

in die Bundeswehr wurde Prof. SCHMIDT 1964 zum Leiter der

Wissenschaftlichen Abteilung am Schiffahrtmedizinischen In-

stitut der Marine in Kiel ernannt. Diese Position hat er his

zuletzt innegehabt. Sein Schriftenverzeichnis enth~lt 95

Titel. Spezielle Interessengebiete waren die pharmakologische

Beeinflussung der Coronardurchblutung, Wirkungen und Neben-

wirkungen von Laxantien, die Pharmakologie und Toxikologie

von L8sungsvermittlern und LGsungsmitteln sowie Probleme der

UnterkUhlung und Wiedererw~rmung. Wir verlieren mit Prof.

Ludwig SCHMIDT einen vielseitig interessierten Kollegen, der

sich durch seine verst~ndnisvolle und tolerante Grundein-

stellung auch seinen Mitarbeitern gegenGber auszeichnete und

der neben seiner Arbeit eine kultivierte Geselligkeit

sch~tzte.

Ich bitte Sie, sich zum Andenken und zur Ehre der verstorbe-

nen Kollegen zu erheben.

Ich wGnsche, dab trotz des groBen Progrsmmes die wissen-

schaftlichen Diskussionen nach den Vortr~gen und auBerhalb

der H8rs~le, aber auch der menschliche Kontakt nicht zu kurz

kommen mGgen und erkl~re hiermit die Tagung fGr erSffnet.

Prof. Dr. med. Hans-Joachim Sc~Gmann, Klinikum Essen der

Ruhr-Universit~t, Pharmakologisches Institut, 43 Essen,

Hufelandstr. 55

R8

VERLEIHUNG DES FRITZ-KULZ-PREISES 1971

Presentation of the Fritz-KUiz-Award for 1971

Wir haben tuns heute zur Verleihung des Fritz-KUlz-Preises

hier versammelt, der zum 2. Mal vergeben werden soll. Ich

begrUBe Sie im Namen des Vorstandes sehr herzlich. Mein be-

sonderer Grub gilt jedoch Frau Ida KULZ, der Stifterin des

Preises, der zur Erinnerung an das Werk von Fritz KULZ alle

2 Jahre verliehen wird und der F~rderung des wissenschaft-

lichen Nachwuchses in der Pharmakologie dienen soll. Es ist

uns eine besondere Ehre, dab Sie, sehr verehrte gn~dige Frau,

auch in diesem Jahr unserer Einladung gefolgt sind. Ich freue

mich besonders, Ihnen im Namen des Vorstandes der Pharmakolo-

gischen Gesellschaft fur die ErhGhung des Stiftungskapitals

um 20.000.- DM auf nunmehr insgesamt 70.000.- DM unseren be-

sten Dank auch bei dieser Gelegenheit aussprechen zu kSnnen.

Wir sind damit in die Lage versetzt, einen grSBeren Betrag

als bisher zur Preisverteilung bereitzustellen. Der Vorstand

hat sich deshalb im Einvernehmen mit Frau KULZ auf neue Moda-

lit~ten der Preisverleihung geeinigt, die im neugefaBten w 4

der Stiftungsurkunde zum Ausdruck kommen. Es heiBt da jetzt:

"Aus dem Ertrag dieser Verwaltung sollen alle 2 Jahre 2 ex-

perimentell pharmakologische Arbeiten aus deutschsprachigen

pharmakologischen Instituten, die in dieser Periode zur Pu-

blikation eingereicht wurden, ausgezeichnet werden. Die Ver-

fasser dieser Arbeiten dGrfen das 30. Lebensjahr nicht Uber-

schritten haben". Welter heiBt es: "Der Preis kann sowohl s

Dissertationen als auch fur 0riginalarbeiten verliehen werden.

Habilitationsschriften werden nicht berUcksichtigt". Die we-

sentliche ~nderung des w 4 besteht einmal darin, dab es 2

Preistr~ger geben wird und dab nicht nur Dissertationen, son-

dern auch andere Originalarbeiten eingereicht werden kSnnen

und schlieBlich, dab Habilitationsschriften nicht mehr be-

rUcksichtigt werden sollen.

Den Mitgliedern des Preisrichter-Kollegiums mGchte ich fur

die mtthevolle Arbeit der Auswahl herzlich danken und gleich-

zeitig darauf hinweisen, dab der n~chste Bewerbungstermin der

1. April 1973 ist.

R9

Das Preisrichter-Kollegium hat den Fritz-KGlz-Preis 1971

Herrn Dr. Ulrich ABSHAGEN, geboren am 18. Juli 1943, aus dem

Institut fur Klinische Pharmakclogie der Freien Universit~t

Berlin, fur seine Arbeit "Eliminationskinetik und Stoffwech-

sel von 2-Propanol im Vergleich mit anderen aliphatischen

Alkoholen von C 1 bis C5" zuerkannt. In der BegrGndung des

Preisrichter-Kollegiums heiBt es u.a.: "Alle untersuchten

Alkohole werden einfach exponentiell aus dem Plasma elimi-

niert, mit Ausnahme von Methanol und ~thanol. Methanol folgt

einer Exponentialfunktion 2. Grades. Die ~thanol-Elimination

l~uft bei allen Versuchsanordnungen und selbst bei niedrigen

Konzentrationsbereichen rein linear ab.

Die gegenseitige Beeinflussung der verschiedenen Alkchole

erlaubt SchlUsse auf gemeinsame Abbauwege. So werden pharma-

kokinetische Hinweise dafGr beigebracht, dab nicht nur ~tha-

nol und 1-Propanol, sondern auch 2-Propanol durch Alkohclde-

hydrogenase abgebaut werden, nicht dagegen tert. Butanol oder

Methanol.

Die Arbeit bringt eine FGlle von pharmakokinetischen und

Stoffwechseldaten, welche sauber erarbeitet und mathematisch

dargestellt sind. Sie liefert ein abgerundetes Bild der

Pharmakokinetik, einer wichtigen Stoffgruppe, und stellt da-

her nicht nur eine gute experimentelle, sondern auch eine

besondere geistige Leistung dar. Diese selbst~ndige Synthese

zahlreicher Befunde zu einer aus biochemischen und physika-

lischen Gedankeng~ngen entwickelten relativ einfachen Theorie

machen die Arbeit in besonderem MaBe preiswUrdig".

Im Namen der Stifterin und der Pharmakologischen Gesellschaft

mSchte ich Ihnen, Herr Dr. ABSHAGEN, den Fritz-KGlz-Preis

1971 und die Urkunde Gberreichen, verbunden mit den besten

GIUckw~nschen und ~Gnschen fur eine weiterhin erfolgreiche

wissenschaftliche T~tigkeit.

Prof. Dr. med. Hans-Joachim Scht[mann, Klinikum Essen der

Ruhr-Universit~t, Pharmakologisches Institut, 43 Essen,

Hufelandstr. 55

R 11

PHARMACOKINETICS OF DIGOXIGENIN-BIS AND MON0-DIGITOXOSIDE IN RATS WITH RESPECT TO ENTEROHEPATIC CIRCULATION (e.c.) (Pharmakokinetik yon Digoxigenin-bis und mono-digitoxosid bei der Ratte unter be- sonderer BerGcksichtigung des enterohepatischen Kreislaufes) U. Absha6en t K. v. Ber~mann I N. Rietbrock After i. d. admin, of DH-Digoxin (D) to biliary fistula (b. f.) rats total radioactivity in blood and bile is eliminated with an half time of 7 h in both fluids. In rats without b. f. an half time of 13.5 h was ob~rvedin blood and of 22 h inbile. To explain the longer half time in bile than in blood, pharmacokineties were studied on all substances, which could participate in e. c. after D admin. 0nly small e. c. of the CHCl3-insoluble fraction was found. Digoxigenin-bis-(B) and monodigitoxoside (M) showed ap- proximatively the same absorption kinetics as D. After i. d. admin, of these metabolites in b. f. rats the blood levels of radioactivity were 5 - 6 times lower than those of D because of higher biliary excretion. 90 - 95 ~ of the absorbed amounts of B and M were excreted in bile within 11 h compared w~h61% of D. After i. d. admin, of D to rats without b. ~ the by far longer half time of elimination of radioactivily of B and M in bile than in blood seemed to be due to a circulation of B and M between intestine and liver. TLC gives evidence of metabolic pattern in bile of these animals, which show that significantly more B is present in bile in rats without b. f. than in rats with b. f. No differences were found in the case of M, which is a priori formed to a lesser extent and further converted rapidly to CHCI 3 insoluble metabolites, which cannot be reabsorbed. Dr. U. Abshagen, Institut fGr Klinische Pharmakologie der Freien Universit~t Berlin, D-IO00 45, Hindenburgdamm 30

ACID-BASE BALANCE OF CATS DURING STIMULATION OF GASTRIC ACID SECRETION. Der S ~ u r e - B a s e n - H a u s h a l t der Katze w~hrend der s t i m u - l i e r t e n M a g e n s e k r e t i o n . M a r g i t t a A l b i n u s and K . - F r . Sewing

In the most d e t a i l e d s tudy on ac id~base ba lance (ABB) produced by loss of H § d u r i n g g a s t r i c ac id s e c r e t i o n h i s t a l o g was used a s . s t i - mu lan t (D.G. McQuar r ie and J.Ao Vennes, J .Lab . C l i n . M e d . 7 5 , 6 1 - 7 3 , 1970) . The marked s ide e f f e c t s o f h i s t a m i n e (H) or h i s t a m ~ n l i k e compounds made i t d e s i r a b l e to compare the e f f e c t s o f H on ABB w i t h those of p e n t a g a s t r i n (PG). In cats a n a e s t h e t i z e d w i t h c h l o r a l o s e and equ ipped w i t h a g a s t r i c cannu la g a s t r i c ac id sec re - t i o n was c o l l e c t e d in 15 min p e r i o d s d u r i n g s t i m u l a t i o n w i t h 0.2 u g . k g - l . m i n - I PG or 5 ~ g . k g - l . m i n ~I H i . v . f o r 2 h r , In b lood samples from c a r o t i d a r t e r y and g a s t r o s p l e n i c ve in c lose to the stomach pH, pC02, HCO~ and p02 were measured in 30 m i n - i n t e r v a l s . Both s t i m u l a n t s produced a g a s t r i c s e c r e t o r y p l a t e a u of about 0.65 resp . 0.87 mEq H+/15 min. PG caused a marked r i s e of venous pH. The venous ac tua l pC02 was c o n s i d e r a b l y reduced but r e t u r n e d imme- d i a t l e y a f t e r P G - i n f u s i o n above p r e s t i m u l a t o r y v a l u e s , The HCO~ c o n t e n t o f a r t e r i a l and venous b lood rose g r a d u a l l y . The venous p02 was reduced. Dur ing H s t i m u l a t i o n a r t e r i a l and venous pH rose s l i g h t l y a f t e r an i n i t i a l sharp drop. Venous pC02 d id no t f a l l d u r i n g s t i m u l a t i o n , however , the p o s t s t i m u l a t o r y r i s e was the same as a f t e r PG. The changes of HCO~ were s i m i l a r as d u r i n g PG, A r t e - r i a l p02 was reduced and the re m igh t be a s l i g h t r i s e in venous p02. I t i s conc luded t h a t the changes in ABB d u r i n g PG are m a i n l y due to a loss of H + and t h a t those observed d u r i n g H i n f u s i o n are ove r l apped by e x t r a g a s t r i c e f f e c t s o f H.

Dr. M. A l b i n u s , Pharmakol . I n s t i t u t , D-74 TUb ingen, W i l h e l m s t r . 56

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DIFFERENCES IN THE [Ca] -DEPENDENCE OF THE CARDENOLIDE ACTION IN ISOLA- TED HEART MUSCLE PREPARATIONS. (~ber Wirkungsunterschiede einiger Carde- nolide an isolierteu Herzmuskelpr~paraten bei ~nderung der extracellu- l~ren Calciumkonzentration). R. Alken, U. Fricka und W. Klaus The dependence of the toxic and therapeutic effects of digitoxin (DTO), digoxin (DO), acetylstrophanthidin (AS) and strophanthidin-3-bromoace- tare (SBA) on the extracellular [Ca3 (o.45 - 7.2 mM) was tested in experiments on isolated papillary muscles (guinea pig). Both parameters showed a significant difference between (a) DTO resp. AS and (b) DO resp. SBA: with increasing [Ca] the well known Ca-digitalis-synergism on the positive inotropic (ED 5o) and toxic effects (TDM = minimal dosis producing irregularities, contracture) of DTO and AS was shown, whereas the ED 5o as well as the TDM of DO and SBA was increased by raising the [Ca] in the range from o.45 to 3.6 mM. In order to test wether this discre- pancies could be explained by a different influence of increased [Ca3 on the glycoside uptake by the heart, the accumulation of 3H-digitoxin and 3H-digoxin in isolated guinea pig hearts perfused for 3o min at a constant rate was determined. The perfusion medium contained o.45, o, 9 and 1.8 mM [Ca] resp. In these experiments a significant decrease both of the DT0 and the DO content in the heart was obtained with increasing [Ca] in the perfusion medium. On the basis of above observations the cardenolides might be classified into two groups corresponding to the [Ca]-dependence of their pharmacological actions but this behaviour cannot be explained by a difference in the glycoside uptake into the heart.

Inst. f. Pharmakologie der }{HE, D-3ooo Hannover, Roderbruchstr. 1oi

A pH-STAT METHOD FOR THE INVESTIGATION OF THE KINETICS OF CARBONIC ANHYDRASE (E.C.4.2.1.I.) (Enzymkinetische Untersuchungen an der Carboanhydrase (E.C.4.2.1.1.)) C.Alsen und F.K.Ohnesorge

The common methods for the estimation of CAH-activity are either not applicable for exact enzymkinetic studies or too many experimental difficulties are encountered (Review: R.P.Davies in Methods of Biochem. Analysis ii, 1963). In the present paper a micromodification of the technique described by K.J.Leibman (J. Pharmacol. Exp. Ther. 131, 271, 1961)is presented. The modified metho~ is hydration reaction of CO9, a Km-Value of 3.45 x IO-~M was determined. For acetazDlamide (Ac), we obtained a Ki-value of 3.9 x IO-9M. The method proofed to be useful for investigating enzymkinetics in environmentaltoxicological problems, such as disorders of calcium metabolism.

Dr.C. Alsen, Institut fur Pharmakologie der Universit~t Kiel D-23OO Kiel, Hospitalstr. 4-6

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EFFECT OF PYRIDINENUCLEOTIDES (Pns) AND NICOTINAMIDE (N) ON GLU- COSE OXIDATION AND INSULIN RELEASE IN ISOLATED PANCREATIC ISLETS 0FRATS (Einflu~ von Pyridinnucleotiden und Nicotinamid auf die Glucoseoxydation und die Sekretion von Insulin bei isolisrten Pankreasinseln der Ratte~ H.P.T.Ammon and J.Steinke

Methods:Collagenase prepared islets were obtained from either normal rats or rats pretreated with 6-aminonicotinamide (6-AN), an antimetabolite of Pn synthesis,(35 mg/kg i.p.,6 h previously). The effect of NAD(H),NADP(H) or N (I or Io mM) was studied on I) insulin (IRI) released into the medium and 2) on 14-C02 production from glucose labeled either in the I or 6 position (S.A. I~C /mg glucose),in the presence of 3 mg/ml glucose.Results: I) Pns~ but not N, added to islets of either normal or 6-AN-treated rats increased significantly the oxidation of 14-C-l-glucose whereas little change took place in oxidation of 1r Islets from normal rats in the absence of glucose showed major response only to~ NADPH,however,with 3 mg/ml glucose both NAD and NADH ~x- hibited marked,N only small insulin release.3)Islets of 6-AN- treated rats exhibited decreased pentosephosphate shunt (PPS) activity and decreased glucose induced insulin release.Addition of any Pns and to a small extent also N restored insulin secretion. Conclusions:Pns added to the incubation medium of isolated islets exert an intracellular effect as evidenced by increased PPS activity.This in turn is associated with increased insulin release.As all Pns tested exhibit a similar effect it is postulated that they act by being converted to intracellular NADP.

Dr,H.P.T.Ammon and Dr.J.Steinke,E,P.Jos!in Res.Lab.,Harvard ~edi- cai School,17o Pilgrim Rd.,Boston,Nass.o2215

ON THE NORADRENALINE DEPLETING ACTION OF OXYFEDRINE IN RATS (Uber die noradrenalinverarmende Wirkung yon Oxyfedrin an Ratten) E._u~- pelTG.planz,D.Palm,H.Grobecker According to several authors (cf.~.K~ovetz and G.P6ch,Arzneimit- tel-Fcrsch. 19,1562-1566,1969) Oxyfedrine (0) acts as a direct ~- sympathomimetic agent in isolated heart preparations,-O which is an N-aralkyl-substituted norephedrine derivative, can be assumed at least after its metabolic degradation in rive-to act as an indirect sympathomimetic amine.Oral administration of 2Omg/kg 0 caused a pronounced and long lasting depletion of noradrenaline in rat iris,heart and brain.Also,l;3 and lOmg/kg/day for 7 days signi-

c ficantly diminished the noradrenaline ontent of the heart ~26~ - 76%~-After oral administration of 2Omg/kg O,the occurrence of norephedrine and p-hydroxy-norephedrine in urine could be demonstrated by means of column and thin layer chromatography. Quantitati- ve gaschromatographic measurements revealed an excretion of 5mg/ kg norephedrine within 48 hrs. This dose of norephedrine decreased the noradrenaline content of rat heart to nearly the same extent as 20 mg/kg O.-It is concluded that because of an extensive metabolic degradation, 0 may also act as an indirect sympathomimetic a- gent in vivo.This assumption is supported by the finding,that in rats pretreated with S]s 525A, the noradrenaline depleting action of 0 is significantly diminished.

H.Grobecker, Pharmakelogisches Institut der Universit~t D-6000, Frankfurt/H., Theodor-Stern-Kai 7

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EFFECT OF O{-TOCOPHEROL TREATMENT ON CERIUM INDUCED CHANGES IN DRUG METABOLISM IN RATS (Die Wirkung yon ~-Tocopherol auf die yon Cerium verursachten ~nderungen des Arzneimi t teIs tof fwechsels in der Rattenleber) P. Arvela and N.T. K~rki Cerium (Ce) is a h ighly hepatotoxic agent when administered i.v., causing fa t t y l i v e r (F.Snyder et a l . , Fed.Proc. 18~ 327,1959) and an impairment of drug metabolism in rats (P.Arvela et a-r., Experient la 27, 1189-1190, 1971). Ant iox idants have been reported to prevent the fat E i n f i l t r a t i o n caused by hepatotoxic agents such as CC14 or ethanol (N.R. D$ Luzio et a l . , Exp.Molec. Path. 11 t 38-52, 1969). In the present study we have tested the e f fec t of a--n ant iox idant , ~-Tocopherol acetate (o(-TP), on Cerium induced l iver in)ury.

~ w~ntyfive male rats (180-200 g) were div ided in$o.4 groups: ( l ) Controls, 2) {eceiv ing one i .vo i n i ec t i on of 2 mg Ce/kg, (3) treated with 10 mg

~-TP/IO0 g for 5 days and (4) treated with Ce and~(-TP as in groups 2 and 3. The test-enzymes for the metabolic a c t i v i t y were: 3,4-benzpyrene hydroxylase, giucose-6-phosphatase, ur id ine diphospho glucuronyl t ransferase (UDPGT), Cyt P-450 and NADPH-Cyt C reductase and these a c t i v i t i e s were determined from the microsomai f rac t ion of the l i v e r . In addi t ion the t r i g i y c e r i d e content of plasma and l i v e r and the pIasma FFA leve i were measured. Cerium induced alterations in enzyme activities (decrease with the ex- ception of UDPGT) were improved after the treatment with O(-TP. The leve l of Cyt P-450 which decreased in group 2 to 34 % was 73 % in group 4 of corresponding cont ro ls . In addi t ion the elevat ion of plasma FFA (156 %) and i i v e r t r i g I yce r i des (296 %) caused by Ce were returned aImost to the cont ro l leve ls . P. Arvela, Department of Pharmacology, SF-90 100 Oulu 10, Finiand.

HISTAMINE METHYLTRANSFERASE IN MAN, DOG AND PIG: DISTRIBUTION, PURIFICATION AND PROPERTIES (Vorkommen, Reinigung und Eigenschaften der l t istaminmethyi t ransferase (HMT) in Mensch, Hund und Schwein) H" Barth, W. Lorenz, K. Goecke, K" Herrmann, W. Seidel , H. T ro id l

In human t issues the fo l lowing a c t i v i t i e s of HMT were determined by the method of Lorenz et al.,Arch.Pharmak.267,421,1970,(pmoles/min and mg pro te in ;n=2-8) : antrum 299,duodenum 550, ieiunum 707, ileum 270,coecum 600,coion 691,sigma 754,rectum 346,ga l l -b iadder 127,pancreas 96,kidney 341,lung 258,spleen 264, thyro id gland 127, skin 243. HMT-act iv i ty and histamine (HA) content in the gas t r ic mucosa did not show any co r re la t i on . Therefore HMT very probably is not loca l ized in mast c e l i s . HMT-act iv i ty in gas t r ic musculature i s d i s t r i bu ted un i - formely over the whole stomach and about 3 - fo ld lower than in the mucosa. HMT was pu r i f i ed from pig an t ra l mucosa 126-fold by u l t r a c e n t r i f u g a t i o n , ammonlumsulphate praecipi tat ion,chromatography on DEAE-Cellulose and Sephadex G 100. The proper t ies at t h i s preparat ion were:K m for HA 2.3xlO-5M; Km for S-adenosylmethionine 4.3x10- M,pH-optlmum 7.4;complete i n h i b i t i o n by the an t ima la r ia l drugs amodiaqulne,chloroquine,quinacr ine (10-4M) and none b~ the f lavonoids (+ ) - ca tech in , t r i - and te t rahydroxy- e thy l ru tos ide (lO-~M). In the dog's stomach amodiaquine i nh ib i t ed HMT both in v i t r o and in v ivo. A dose of 2 mg/kg i .m. increased the maximum HA response of a Heidenhain pouch by 100 %.

Dipl.Chem.H.Barth, Abtei lung fur exp.Chirurg ie und pathol.Biochemle, Chirurgische U n i v . - K l i n i k , D-355 Marburg, Robert-Koch-StraBe 8

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THE EFFECT OF PRE-LOAD ON THE ACTION OF DBA ON ISOLATED _~T ~TRIA (EinfluB der Vorspannung auf die Wirkung von cyclischem N ,0 - Dibutyryladenosin-3',5',monophosphat (DBA) am isolierten Ratten- vorhof) W. Bartsch u. P. Schaub

An isolierten RattenvorhSfen wurden die inotropen und chronotropen DBA-Effekte unter variablen Vorspannungen (200 - 1500 mg) mit an- n~hernd isometrisch arbeitenden DehnungsmeBstreifenaufnehmern untersucht. DBA bewirkt mit 400 ug/ml Badl~sung bei 200 mg Vorspannung eine geringe, bei 1500 mg Vorspannung eine deutlich st~rkere positiv inotrope Wirkung. Die gleiche Dosis verursacht, unabh~ngig yon der Vorspannung, initial einen maximal 6 min dauernden negativ chronotropen Effekt. Bei 200 mg Vorspannung h~it diese Frequenz- minderung l~nger als 15 min an, mit zunehmender Belastung kommt es dagegen vonder 6. min ab zu einer positiv chronotropen Wir- kung. Am Beispiel von DBA wurde gezeigt, dab unterschiedliche Vorspan- nung nicht nur zu quantitativ, sondern auch zu qualitativ diffe- renten Ergebnissen fGhren kann. FGr Versuche dieser Art am isolierten Rattenvorhof wird deshalb empfohlen, eine Vorspannung zwischen 500 und i000 mg zu w~hlen und entsprechende Angaben zu machen. Bei Vorspannungen unter 200 mg ist mit einem mangelhaften Ankopplungsgrad und bei Vorspannun- gen von ~ber 1500 mg mit Insuffizienzerscheinungen des Pr~parates zu rechnen.

Dr. W. Bartsch und Dr. P. Schaub, Boehringer Mannheim GmbH, 6800 Mannheim 31, Sandhofer Str. 112-132

REPRODUCTION AND MUTAGENICITY TRIALS OF 2-METHYL-4-NITRO-I- (4-NITROPHENYL)IMIDAZOLE (Reproduktions- und Mutagsnit~tspr~fun- gen mit 2-Methyl-4-nitro-l-(4-nitrophenyl)imidazol) A.Bauer, H.Frohber~ G. Jochmann~ B.v. Sehillin 6

In Lan~zeitversuchen an Ratten erzeugte das Nitroimidazolderivat in hohen Dosen Hodenatrophie und Schwund des lymphatischen Gewe- bes. Derartige Ver~nderungen sind z.B. auch yon teratogen und mutagen wirkenden alkylierenden Cytostatika bekannt. Deshalb wurden Fertilit~ts-, Teratogen- und Mutagenversuche durchgef~hrt. 2-monatige t~gliche orale Gabe yon je 200 mg/kg fGhrte zur Steri- lit~t der m~nnlichen Ratten, die sich 9-18 Wochen nach Behand- lungsende zurGckbildete. FGr Wistar-Ratten (bis I0 x 200 mg/kg), NMRI-M~use (bis i0 x i00 mg/kg) und fGr weiBe Neuseel~nder Kanin- chen (bis 13 x 50 mg/kg), vom 6.-15. bzw. 18.Tag p.c. oral appliziert, wirkte das Nitroimidazolderivat trotz Adulttoxizit~t in den hohen Dosen nicht teratogen. Nach einmaliger i.p.-Behandlung m~nnlicher M~use mit i00, 200 und 400 mg/kg wurden w~hrend der gesamten Dauer der Spermatogenese (8 WQchen) keine dominanten Letalmutationen festgestellt. Trenimon[ R) (0,125 mg/kg i.p.) erzeugte in den ersten 3 Paarungswochen 40 % dominante Letalmutati- onen. An Knochenmarkzellen yon Ratten und chinesischen Hamstern verursachte das Nitroimidazolderivat (400 mg/kg), 48, 24 und 4 Stunden vor der TSSung oral appliziert, keine Chromosomenaberra- tionen. Trenimon[ R) ( 0, 25 mg/kg i.p.) erzeugte dagegen bei beiden Spezies etwa 70 % aberrante Metaphasen.

Dr.A.Bauer, Institut fGr Toxikologie, E.Merck, 61 Darmstadt, Postfach 4119.

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PERMEABILITY OF CHOLEPHILIC SUBSTANCES WITH ISOLATED LIVER CELT~ (Permeabilit~t cholephiler Substanzen bei isolierten Leberzellen) H. Baur I and E. Pfaff

In liver there is a directed transport system which makes a net transport of large organic anions from the blood to the bile. An example for a physiological substance of this class is bilirubin, others are numerous xenobiotics, such as bromsulphthalein (BSP). In the overall transpor~ from blood to bile some knowledge has been collected on binding proteins in blood and cytoplasm (of. Fleischner and Arias, Amer. J. Mad. 89, 576, 1970), on microsomal enzymes rendering these substances more hydrophilic, but little is known on the mechanism of uptake and excretion. For this reason, we isolated liver par- enchymal cells. To exclude permeability changes introduced by the isolation, we modified the procedure of Berry and Friend (J. Cell Biol. 83, 506, 1969). Thus, intact cells could be separated from damaged cells and could be con- trasted to cell preparations with defined permeability changes. The kinetics of BSP uptake under various conditions were measured by centrifugal filtration. Whereas damaged cells show a fast, but limited uptake, intact cells may put up a concentration ratio inside/outside of up to i00. Since the uptake is in part osmotically active, the cells may burst before a saturation point can be reached. The uptake of BSP into intact cells increases unproportionally with higher outside concentrations (above 200 ~M). The uptake is pH dependent and accompanied by proton uptake.

Dr. E. Pfaff, Institut fdr Toxikologie der UniversitKt, D-7~O0 TUbingen, WilhelmstraBe 56

EFFECT OF THEOPHYLLINE ON 45CA UPTAKE IN ISOLATED PERFUSED GUINEA- PIG HEARTS (45Ca-Aufnahme am isollert perfundlerten Meerschweinchenherzen unter dem Einflul3 von Theophyllln) P. Bellemann, H. Scholz

The effect of theophylllne (T) on mechanlcal performance, tissue Ca and Ca exchange (exchange of extracellular 45Ca against cellular Ca) was studied in isolated, electrically stlmulated (frequency 18o beats/mln) Langendorff perfused

45 a i guinea-pig hearts. Following a 3o-mlnute exposure to C , t ssue Co and 45Ca activity were measured in right ventrlcular samples. Right ventrlcular tension was recorded Tsometrlcally as described by Beckett (J.Pharm.Pharmac. 22, 818, ]970). The investigations were performed at 35~ in Tyrode solutlon containing o.45 mM CaCI 2 . -"Therapeutic" T concentrations (5 x lo -5 - 1o -3 g,/ml) which increased contractile force without producing arrhythmlas or contractures had no effect on tissue Co and did not alter the exchangeable Ca fraction. "Toxic" T concentrations (2 x 1o -3 g,/'ml) induced contractures, increased tissue Ca and enhanced the amount of exchangeable cellular Ca. - It is concluded that T contractures may be due to an increment in the exchangeable cellular Ca fraction. The positive inotropic effect of T, however, is thought to be independent of this mechanism. (Supported by a grant from the Deutsche Forschungsgemelnschaft).

P. Bellemann, Pharmakologlsches Instltut der Unlverslt~t Malnz D-65oo Malnz, Obere Zahlbacher Str. 67

R17

INTRAVENOUS AND ORAL ACTIVITY OF OLEANDRIN DERIVATES (Intrave- n~se und enterale Wirksamkeit yon Oleandrinderivaten) H.F.Benthe, GoHaberland

Durch geeignete Veresterung und Ver~therung yon herzwirksamen Glykosiden l~t sich ihre enterale Wirkmamkeit erh8hen.Dieser Zusammenhang wurde an einer gro~en Zahl yon Acylverbindungen des Oleandrins (~) untersucht.Die Bestimmung der enteralen Wirksam- keit erfolgte an Katzen dutch Vergleich der intravenSs und oral tGdlichen Dosen.FGr die mGglichen Veresterungen bieten sich im Aglucon C16 und im Zucker C4'an.Unabh~ngig yon der Substitution am C4'ist Verschlu~ des CI6-OH Voraussetzung fGr eine Resorption, dabei f~llt die enterale Wirksamkeit mit steigender Kettenl~nge des Substituenten:Diformyl bis Dibutyryl und hSher.Auch bei al- leiniger Variation des Esters am C4'im O.erweist sich C4'-buty~l als oral wenig wirksam,ebenso C1-Acetyl.Enteral fast wirkungslos sind C4'-Alkyl-Kohlens~ureester des O.,demgegenGber zeigt der Phenyl-Kohlens~ureester eine gute enterale WirksamkeitoV0n den Alkyl-substituierten Aminokohlens~ureestern des O.sind die ge- prGften Methyl-Athyl- und i-propyl Verbindungen oral wirksam, die Wirkung erlischt bei Butylsubstitution. In Stoffwechselunter- suchungen mit 3H-Acytyl-Oleandrin am Menschen wurden im Harn mindestens 3 CHCl3-15sliche Metaboliten nachgewiesen:0o,4'Ac-16- Desacetyl-O. und in zeitlich zunehmend gr~erer Menge die entsprechenden Verbindungen vom C3'-Demethyl-Oleandrin.

H.F.Benthe, Pharmakologisches Institut der Universit~t Hamburg, 2000 Hamburg 20, Martinistr. 52

INVESTIGATIONS ON THE ROLE OF CATECHOLAMINES IN ANAPHYLACTIC SHOCK (Untersuchungen Gber die Rolle der Catecholamine im anaphylaktischen Schock) W. Bernauer and P. Filipowski

In previous investigations we found a considerable increase of serum catecholamines (especially adrenaline) in ~uaphylactic shock of guinea pigs (W. Bernauer et al., Naunyn-Schmiedeberg's Arch. Pharmak. 270, 326-334, 1971). Now the consequence of this mechanism for t e~survival in the shock was examined. Acute anaphylactic bronchospastic death was prevented by mepyramine, but about 60% of the animals died later in the protracted shock. Pro- pranolol, or adrenalectomy abolished the effect of mepyramine showing that ~-adrenergic action of adrenaline is necessary for the broncholytic effect of antihistamines. Mepyramine treated animals did not die in orotracted shock when pretreatment with reserpine (I mg/kg; 24h) was performed. Also in these animals propranolol led to acute anaphylactic death. In reserpinized animals a significant increase of serum adrenaline after antigen injection was measured which was much smaller than in non-reserpinized animals. When reserpine was given immediately before antigen, protracted shock was not prevented but intensified. Animals of this experimental group which died in protracted shock had very high serum noradrenaline levels. Dibenamine inhibited the lethality increasing effect of acute reserpine injection. - In- fusion of adrenaline or noradrenaline in mepyramine treated, anaphylactic animals enhanced the death rate in protracted shock, dibenamine reversed the effect to a beneficial one. Isoproterenol infusions had a life-saving effect in these experiments. Priv.-Doz. Dr.w. Bernauer, Pharmakologisches Institut der Uni- versit~t, D-7800 Preiburg i.Br. (Germany), Katharinenstr. 29.

R18

EFFECTS OF NEUROLEPTICS AND THYMOLEPTICS ON PHOSPHODIESTERASE ACTIVITY AND ON CYCLIC 3', 5'-AMP LEVELS IN RAT BRAIN (Wirkungen yon Neuroleptika und Thymoleptika aus die Phosphodiesterase und den 3', 5'-AMP-Gehalt im Rattenhirn) S. Berndt und U. Schwabe

Recently, several psychotropic drugs have been shown to change cyclic 3', 5'-AMP (cAMP) metabolism in brain. The effects of neuroleptics and thymoleptics on phosphodiesterase (PDE) activity in vitro and on cAMP content of rat brain in vivo was studied. Chlor- promazine, triflupromazine and chlorprothixene inhibited PDE in a non-competitive manner, whereas imipramine, desipramine and ami- tryptiline proved to be competitive inhibitors. Haloperidol had virtually no effect on PDE activity. The content of cAMP was determined in tissue zylinders of rat frontal brain taken by needle punction and immersed in liquid nitrogen within 3 sec. Following intravenous injection of chlorpromaz~ne (5 mg/kg) the cAMP content was elevated 2,5 fold within I minute end had declined to control values after 3-5 minutes. Pretreatment with theophylline (10 mg/kg i.p.) completely prevented the chlorpromazine induced increase os cAMP in brain but did not change the drug induced effects on behaviour. Desipramine (10 mg/kg) and haloperidol (0.25 mg/kg) also elevated the cAMP content in rat brain, but this effect was only partly antagonized by theophylline (20 mg/kg). The inhibition of PDE by psychotropic drugs is obviously o9 minor importance for the observed accumulation o9 cAMP and is not directly related to the psychopharmacological effects of th%se agents.

Dr. S. Berndt, Institut f~r Pharmakologie, Medizinisehe Hochschule Hannover, 3000 Hannover-[leefeld, Roderbruchstr. I01

THE INFLUENCE OF PENTOBARBITAL ON CALCIUM DEPENDENT CHANGES OF CONTRACTILE FORCE (Pentobarbital und die Calcium-bedingte Anderung der Kontraktionskraft) V.S. Bishop, A.H. Brigqs, G.O. carrier und T. Peters

The influence of Na-Pentobarbital (P) (I0-4-IO-3M) on the rates of tension decline and tension recovery were determined in stimulated rabbit left atria when the Ca concentration in Ringer's solution was changed. The rate of recovery was significantly prolonged in the presence of P. The time course of tension decline is not only governed by the size of ionized Ca fraction but is also influenced by intracellular bound Ca. In order to determine P's action on intracellular Ca stores, the rate of Ca uptake by cardiac sarcoplasmic reticulum in the presence of this drug was investigated. P enhanced the rate of Ca uptake by this system. Since the rate of decline is proportional to the size of the intracellular ionized Ca fraction, a three compartmental model was designed which represents extracellular, intracellular ionized, and intracellular stored Ca. A computer was programmed to describe the rate of tension decline based on this model.

The experimental data for the influence of P on tension decline agreed with the theoretical curves under the condition that the relative size of thestored Ca is increased and it's dissociation rate is diminished. These data indicate that P can shift the equilibrium for Ca towards intracellular storage sites which may explain the negative inotropic effect.

Dr. T. Peters, Institut ftir Pharmakologie der Universit~t Kiel D-23OO Kiel, Hospitalstr. 4-6

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DRUG METABOLISM ANDPROTEIN SYNTHESIS IN ISOLATED PERFUSED RAT LIVER (Arzneimittelmetabolismus und Proteinsynthese in der isoliert-perfundierten Rattenleber) K. W. Bock, S. Matern I and W. FrShling

Livers from male Sprague-Dawley rats (200-250 g) were perfused with Eagle minimal essential medium and bovine erythrocytes. When added at a concentration of 0.5 mM, hexobarbital disappeared from the perfusion fluid at an initial rate of 0.22mmoles/min/g liver. Hexobarbital metabolism was identical in perfusions with rat blood. Albumin (determined with antibodies against purified rat albumin) was synthesized at a constant rate (0.4 mg/h/g liver) within 4 h. Hexobarbital oxidation and albumin synthesis were reduced in perfused livers from rats which had been starved for 20 h and in perfUsions lacking erythrocytes. Adaptive changes of protein synthesis after the addition of drugs to the perfusion medium were assayed by following the induction of tyrosineaminotransferase with dexamethasone and of C-amino- le~linic-acid-synthetase with the porphyria-produeing Sedormid congener allylisopropylacetamide and dexamethaso~e. When dexamethasone was omitted from the medium ~-aminolevulinie-acid-synthetase decreased rapidly and could not be induced by allylisopropylacetamide indicating that corticoids or their metabolites are necessary for heme and consequently hemoprotein synthesis in the isolated liver. The results demonstrate the functional integrity of the isolated organ over a 4 h period.

Dr. K. W. Bock, Institut fHr Toxikologie der Universit~t, D-7400 T~bingen, WilhelmstraSe 56

EXPERIMENTAL STUDIES ON THE POSITIVE INOTROPIC ACTION AND THERA- PEUTIC RANGE OF METABOLITES OF DIGITOXIN (Untersuchungen Hber die positiv inotrope wirkung und die therapeutische Breite von Digi- toxin-Metaboliten) H. B~ttcher and D. Proppe

The positive inotropic action and therapeutic range of monodigitoxoside and bisdigitoxoside of digoxigenin, metabolites of digitoxin, were investigated by means of cat heart-lung preparations. Ouabain was used as reference compound. The serum-calcium level was lowered to about 55 % by hemodilution with i0 %-dextran solution (mol.wt. 40000) to Dbtain a moderate cardiac failure. The perfusion medium was kept at a temperature of 37 ~ C.

Ouabain and the metabolites of digitoxin were applied cumulatively at intervals of ten minutes in equimolar amounts. Each dose led to an increase of the previous glycoside concentration of the blood- dextran medium by 2 x iO -~ M. The influence of the glycosides on the following parameters was measured: ECG, heart rate, dp/dt, stroke volume, and left ventricular stroke work. Results: The maximum positive inotropic effect obtained by monodigitoxoside and by bisdigitoxoside of digoxigenin were 1.35 and 2.0 times larger than that evoked by ouabain. The toxic doses, characterized by occurrence of arrhythmia, were determined to be 3.6, 6 and 9 times the threshold doses of bisdigitoxoside, ouabain and monodigitoxoside, respectively.

D. Proppe, Institut f~r Pharmakologie der Universit~t Kiel D-2300 Kiel, Hospitalstr. 4-6

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EXCRETION, METABOLIC FATE AND RETENTION OF MESTRANOL IN RATS (Ausscheidung, Stoffwechsel und Retention von Mestranol bei Ratten) H. M. Bolt and H. Kap~us

After i.v. application of #-14C-mestranolto female r~ts the total fecal and urinary excretion within 3 days amounted to 48,2 ~ 8.9%, whereas the excretion of • was 74.7 ~ 9.5 % - An accumulation of mestranol radioactivity occured in fat, brain and adrenals. This radioactive material has been identified as unaltered mestranol by isotope dil~tion. The whole body content of radioactivity after a dose of i*C-mestranol was assayed in mice. The CH2C12/EtOH-extraetable radioactivity fell within 48hrs to i.# %. The radioactivity not extractable with these solvents amounted 15.4 % after 48 hrs and remained constant after 72 hrs. Most of this remaining radioactivity was not extractable with water and also not extractable with ether after hot acid hydrolysis. Some metabolites of mestranol could be separated from feces extracts by column chrg~atography on alumina. After application of doubly labelled mestranol (#-I*C and (3)-methoxy-)H) it is demonstrated, that some of the metabolites have an unaltered 3-methoxy group, while others had undergone a demethylation and thus derive from ethynylestradiol. Details of this work will be presented elsewhere (XENOBIOTICA2, 1972, in press).

,Dr. H. M. Bolt, Institut fur Toxikologie der Unlversit~t, D-7400 Th~Ingen, WilhelmstraBe 56

ON THE POISONING OF THE ISOLATED FROG RETINA BY CU-IONS (Uber die Intoxikation der isolierten Froschretina mit Cu-Ionen) U. Borchard, K.U. Schneider Cu-lonen sind im Gewebe als Bestandteil yon Enzymsystemen in geringer Men- ge vorhanden. HShere Konzentrationen, wie sie beirn Morbus Wilson oder nach Eindringen Cu-haltiger FremdkSrper ins Auge auftreten, zeigen eine gewebs- schfidigende Wirkung. Um quantitative Angaben ~iber den toxischen Konzentra- tionsbereich ffir das Nervengewebe machen zu kSnnen, wurde die isolierte Re- tina mit Badlbsungen umstrSmt, deren Cu-Gehalt yon 3 bis 60 pMol/l variierte. Die Dauer der Cu++-Intoxikation betrug zwischen 30 und 90 min. Unter den Ver- suchsbedingungen erwies sich der VergiftungsprozeB, meBbar an der GrS/~e des Belichtungspotentials (ERG), fiir Konzentrationen unterhalb yon 6 pMol/1 als geringfiigig und war nach UmstrSrnung mit Cu-freier TyrodelSsung riickg~ngig zu machen. Sowohl die Steigerung der Cu-Dosis wie auch die Verl~ingerung der Einwirkungsdauer fiihrten zu einer erhShten Netzhautsch~idigung, die auch nach l~ngerer Einwirkung yon TyrodelSsung bestehen blieb. Komplexbildner wie D- Penicillarnin, Dimercaprol, Na-Thiosulfat und EDTA zeigten gegeniiber der Ty- rodeiOsung einen deutlich giinstigeren Einflu6 auf die Wiederherstellbarkeit des urspriinglichen Funktionszustandes der Retina. Die beste Wirkung ergab sich in Ubereinst immung mit der klinischen Erfahrung fiir D-Penici l lamin. Der mit der Methode der Atomabsorpt ionsspektrometr ie best immte Cu-Gehalt der Netzh~iute macht den EinfluB yon Dosis und Einwirkungsdauer der BadlSsung auf die Cu-Speicherung des Gewebes deutlich, die paral lel zur Abnahme des ERG verEiuft. Dr. U. Borchard, Pharmakologisches Institut der Universit~it zu KSln, 5 KSln 41, Gleueler Str. 24

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THE BIOSYNTHESIS OF THE CO-NEUROTRANSMITTER OCTOP.~INE IN COMPA- RISON WITH THE NEUROTRANSMITTER NORADRENALINE (Die Biosynthese des Co-Neurotransmitters Octopamin im Vergleich zum Neurotrans- mitter Noradrenalin) K. Brandau and J. Axelrod

Octopamine (p-hydroxyphenylethanolamine) is subcellularly distributed, taken up, stored, and released in the same way as noradrenaline, its effect at the receptor is quantitatively weaker than that of noradrenaline. Because of its normal occurance in sympathetic nerves it is suggested that it serves as a co-neurotransmitter (Kopin et al., Proc. Natl. Acad. Sci. U.S.A. 52, 716,1964; Molinoff and Axelrod, Science 164, 428, 1969, J. Neurochem. 19, 157, 1972). In this paper by measuring endogenous concentrations of octopamine in heart and brain of rats it is shown that octopamine is synthetized by the same enzymes as noradrenaline is and using the same biosynthetic precursors except tyramine. Dopamine and DOPA can be ring-dehydroxylated in the brain of rat~ Together with former results from this laboratory which showed ring-hydroxylation of octopamine forming noradrenaline an interesting orbit is pointed out. The pharmacological relevancy of these findings concerning the treatment of Parkinsonism and hepatic coma with L-DOPA is interpreted.

Dr. K. Brandau, Farbenfabriken Bayer AG, Institut fur Pharmakologie, 56 Wuppertal I, Friedrich-Ebert-Str. 217

ON THE MODE OF ACTION OF 0XYFEDRINE (Zur Wirkungsweise yon 0xyfe- drin) W. Brandt~ M. Reiter

Die positiv inotrope Wirkung yon 0xyfedrin (OF) auf den Meer- schweinchen-Papillarmuskel (Krebs-Henseleit-Lsg., 35~ 3,2 mM Ca, i/see) ist gegen mehrmaliges Auswaschen relativ resistent. Sie wird durch Reserpin-Vorbehandlung (5 mg/kg, 24 h) nicht beeinflusst und durch Propranolol in nicht kompetitiver Weise gehemmt. Dies deutet in Ubereinstimmung mit den yon anderen Autoren an anderen Versuchsanordnungen erhobenen Befunden auf eine direkte Er- regung der Herz-~-Rezeptoren hin. - Die isometrische Spannungs- kurve zeigt naeh OF die fHr Noradrenalin (NA) typische Verk~rzung der Erschlaffungszeit bei erh~hter Anstiegssteilheit. Die An- stiegszeit wird verk~rzt. Die Wirkungsst~rke yon OF betr~gt ca. 40 % und die Wirkungsgeschwindigkeit ca. 20 % der yon NA. OF hemmt die NA-Wirkung in kompetitiver Weise in Konzentrationen ab 10-SMo

Die positiv inotrope Wirkung yon OF ist in Konzentrationen bis ca. 10-6M mit der yon NA vergleichbar. In h~heren Konzentrationen nimmt sie dagegen wieder ab. Dies beruht m~glicherweise auf Auto- inhibition am ~-Rezeptor oder auf einem funktionellen Antagonis- mus.

Dr. W. Brandt, Institut fHr Pharmakologie u. Toxikologie der Tech- nisehen Universit~t M~nchen, 8 M~nchen 23, Biedersteiner Str. 29

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SUSTAINED URiNARYEXCRETIONOF PERAZINEMETABOLITES IN MANFOLLOWINGTERMI- NATION OF TREATMENT (Langdauernde Harnausscheidung yon Perazin-Metaboliten beim Menschen naeh Beendigung elner Behandlung) U. Breyer

In 3 male patients the excretion of various perazine (Taxilan R) metabolizes was measured during ingestion of the drug and up to 31 days following termination of treatment. The quantities of perazine sulfoxide and its dioxo derivative 4-[3-(phenothiazlnyl-lO)-propyl]-l-methyl-2,5-dioxo-piperazine sulfoxide declined within a few days to_trace amounts. Desmethyl perazine sulfoxide and its monooxo derivative 4-L3-(phenothiazinyl-10)-Propyl]-2- oxo-piperazine sulfoxlde were detected in decreasing concentrations for 2-3 weeks. No clear-cut decrease during the time interval studied was observed for the concentrations of products resulting from piperazine ring degradation. These are N-/3-(phenothiazinyl-lO)-propyl]-ethylenediamine sulfoxide, its Nt-methyl analogue and 3-(phenothiazlnyl-lO)-propylamine sulfoxide (PPA-SO). These findings indicate that desmethyl perazine persists in the body longer than perazine and that there is probably an aeewm/lation of phenothiazlnyl- propyl-ethylenediamine and its N~methyl derivative in the organs of humans analogous to that demonstrated in the tissues of rats and other species (U. Breyer e% al., Experientia, in press; U. Breyer, Biochem. Pharmacol., in press). Following termination of perazine administration, rats excreted PPA-SO in urine for 5 days, whereas the sulfoxides of perazine and desmethyl perazine were detected only for 1 or 2 days, respectively.

Dr. Dr. U. Breyer, Institut fur Toxikologie der Universit~t, D-7400Th%ingen, WilhelmstraSe 56

INFLUENCE OF AESCIN ON SURFACE PROTEINS OF YOSHIDA ASCITES TU- MOUR CELLS (Einflu2 yon Aescin auf Oberfl~chenproteine yon Yoshida-Asci- testumorzellen). F.v. Bruchhausen u. K. Gruber.

Smaller concentrations of the triterpene glycoside aescin stimulate glucose uptake into fat pads, whereas higher concentrations inhibit glucose transport also into Yoshida ascites tumour cells (GRUBER, HEINj v. BRUCHHAUSEN, Naunyn-Schmiedebergs Arch. Pharmk. 271, 361-373j 1971). Incubation of these tumour cells with 3 �9 10 -5 M aescin diminishes the surface coat (glyco- lemm, glycocalix). This was revealed electronmicroscopically by ruthenium red staining without any detectable alterations of the cell structure. In the supernatant the glycoprotein-like material is enriched. This could particularly be demonstrated by means of reaction with l-anilino-8-naphthalene sulfonate and pretreatment with (14C)-fucose, a particular precursor of cell surface glycoproteins. On fractionation of the released material by isoelectric focussing, it could be seen that several fractions and especially fragments of minor molecular weight could be separated. (3H)-Dihydroaescin added to the incubation vessel is not bound to these materials to a greater extent. The delayed release of surfaee material by aescin does not seem to correlate with the immediate alterations in transport processes for glucose amino acids and other substrates.

Prof. Dr. F. v. Bruchhausen, Pharmakologisches Institut der Freien Universi- t~t Berlin, D-1000 Berlin 33~ Thielallee 69/73

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ANTIBACTERIAL ACTION OF THE SURFACTANT OCTYL PHENOXY POLYETHANOL (TRITON X-IOO) R IN VITRO (Antibakterielle Wirkungen von Octyl- phenoxypolyaethanol (Triton X-IOO) R in vitro)K.Brune

In 1951 Cornforth et al. (J.W.Cornforth et al.,Nature (London) 168,150,1951) described the tuberculostatic action of the non- ionic detergent octyl phenoxy polyethanol (Triton X-IOO) (Rohm and Haas Company) inn viv__~o. However this detergent failed to demonstrate measurable effects on the growth of Mycobacteria in cul- ture media (R.J.W. Rees.,Proc. Roy. Soc.Med.46,581,1958). Since that time the mechanism of the antibacterial action of Triton X-IOO remains a matter of speculation (P.D.A.Hart.,Science 162,686,1968; C.De Duve et al.,Amer. Rev. Physiol.28,635,1966). Using a modification of the method described by Muschel et al. (L. H.Muschel et al.,J.Intmun.7__66,1,1956) we found measurable effects of Triton X-IOO on the growth of Escherichia coli in vitro,especially if it was allowed to act upon the bacteria together with components of mammalian or avian polymorphonuclear leukocytes, serum,or even purified serum albumin. This synergistic effect suggests a new interpretation of the antimicrobial effect of Triton X-IOO in vivo and in vitro.

Dr.K.Brune,Pharmakologische Anstalt der Universit~t Basel, CH-4056 Basel,Klingelbergstrasse 70.

INHIBITION OF ACTIVE INTESTINAL TRANSPORT OF SUGARS AND AMINO A- CIDS BY PRENYLAMINE (SEGONTIN R) (Hemmung des aktiven intestinalen Transportes yon Zuckern und Aminosiuren dutch Prenylamin (Segon- tinR). W. F. Caspary and W. Creutzs

SuccessFul treatment os the dumping syndrome with prenylamine and an improvement of glucbse tolerance after an oral glucose load has been reported recently. We examined the eFFect oF prenylamine on active intestinal transport oF sugars and amino acids in rat small intestine in vitro. Prenylamine inhibited active transport oF substrates oF t e~testinal hexose transport system (D- glucose, D-galactose, 3-O-methyl-D-glucose) as well as active transport os amino acids (L-leucine, L-methionine, L-lysine, L- proline, glycine andS- amino-isobutyric acid). The lowest inhibitory concentration oF prenylamine on active transport oF D-galactose was 0.05 mg/ml. The inhibitory eFFect increased on increasing incubation time and was not reversible. Kinetic analysis showed a non-competitive type oF inhibition. Prenylamine markedly decreased transmural potential diFFerences (PD) induced by D-glucose, indicating that Nat-dependent translocation oF glucose a- cross the brush border membrane is inhibited.

The potent direct inhibitory eFFect oF prenylamine on active intestinal transport processes suggests that the above clinical observations after pretreatment with prenylamine may be induced by an inhibition or delay os intestinal nutrient absorption.

Dr. W. F. Caspary, Medizinische Universititsklinik, D-3400 G~t- tingen, Humboldtallee I.

Supported by the Deutsche ForschungsgemeinschaFt (Ca 71/I).

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EFFECT OF ANAPHYLACTiC REACTIONS ON ZINC, MAGNESIUM, AND CALCIUM IN PLASMA AND LUNG OF GUINEA PIG (EinfluB anaphylaktischer Reak- tionen auf Zink, Magnesium und Calcium in Plasma und Lunge des Meerschweinchens) J. Christians, K. D. Friedber$

High amounts of heparin, histamine, and Zn in rat peritoneal mast cell have suggested a histamine-binding complex of these 3 components (L. Kerp, Int. Arch. Allergy, 22, 112-123, 1963). Accord- ingly, in anaphylactie shock of guinea pigs, release of i atom Zn/2 molecules of mast cell histamine could have raised plasma Zn by 7o-loo %. We measured Zn in plasma and lung and, additionally, Mg and Ca, by atomic absorption spectrophotometry. 1,5 min after induction of acute and protracted anaphylactic shock, anaphylatoxin shodk, and histamine shook, plasma Zn (4o,5 +o,6/uval/l, n=56) failed to change significantly. However, normal ~lasma Mg (1,85+o,o3 mval/l, n=56) rose by 20 %. Plasma Ca (5,o6+o,o3 mval/l, n=56) was elevated io %. In protracted shock, there was a further increase of Mg (6o %) and Ca (15%) shortly before death. Lung Zn (~ 63• mval/kg TG, n=8) did not change. Provided that histamine release in anaphylactic shock is mainly from mast cells, our results indicate that Zn liberated simultaneously may be either retained in the tissue or turned over extre- nely fast. Histamine binding without participation of Zn (B. Uvn~s et al., Acta physiol, scand., Suppl. 336, 1-26, 197o) seems more probable. Rise of plasma Mg is thought to be due to transitory hypoxia, while the cause of Ca elevation is as yet unknown.

Dr. J. Christians, Institut fur Pharmakologie und Toxikologie, D-34 G~ttingen, GeiststraBe 9

EXPERIMENTAL ELEVATION OF THE PULMONARY VASCULAR RESISTANCE IN CATS BY THE INTRAVENOUS ADMINISTRATION OF DECYL SODIUM SULFATE AND DODECYL SODIUM SULFATE (Experimentelle ErhShung des pulmonalen Ge- f~widerstandes bei Katzen dureh intravenSse Zufuhr yon Na-Decylsulfat und Na-Dodecylsulfat) H.G. Classen, K.A. Schumacher, P. Marquardt, M. Sp~th

n/100 or n/10 aqueous solutions of the following sodium alkylsulfates were injected

or infused intravenously into cats: octyl sodium sulfate (1), decyl sodium sulfate (II), dodecyl sodium sulfate (III), tetradecyl sodium sulfate (IV) and hexadecyl sodium sulfate (V). The following phenomena were measured: Pulmonary vascular resistance (PVR), systematic vascular resistance (SVR) and pulmonary ventilation volume/min (PVV). - (1) proved inactive in amounts up to 115 mg

infused during 25 rain. - I.v. injections of 0.5 to 5 mg/kg of (II) and (Ill) were followed by a strong increase of PVR, a less pronounced increase of SVR and a reduction of PW. Infusions during about 12 rain of 5.2 rag/rain (II) or 5.7 mg/min (Ill) raised the PVR by the factor 4. - (IV) and (V) were active only in 3 of 6 experiments, (V) being less active than (IV), and both being less active than (III) or (II). - Decapitation and pretreatment with reserpine or phentolamine did not influence the effects of (If) and (III). Since (If) and (Ill) increased the perfusion pressure of isolated lungs or hind - limbs as well, a direct effect on smooth muscles is supposed. - (If) and (Ill) proved active only on i.v. administration but neither on i.p. (500 mg/kg) nor on i.m. (250 mg/kg) injections nor per os (i g/kg).

Univ. -Doz. Dr. H.G. Classen, Institut f~ir Experimentelle Therapie der Universitiit D-78 Freiburg/Br., Hugstetter StraI3e 55.

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A RESPIRATOR FOR SI~[ULTANEOUS ARTIFICIAL VENTILATION OF FOUR S}{ALL AN!}{ALS (Ein Respirator zur gleiehzeitigen ]dinstlichen Beatmung yon viem Kleintieren) G.CunitztK.H.~,eis und H.Lidl

Um Arbeiten mit kleinen Labortieren,die kqnstliche Beatmung er- fordern,zu beschleunigen,wurde ein Respirator gebaut,der es erlaubt,gleiehzeitig un4 v~llig unabhingig voneinander 4 Tiere in der ~Tarkose zu boatmen. Seine Arbeitsweise beruht auf der Funk- tion eines T-StHekes(nach Ayre,P.:Brit. J.Anaesth. 28,52o-523 1956) :Den Respirator besteht im wesentliehen aus 3 Teilen.Einem Druckbehilter,aus Drosselventilen und aus einem Unterbrecher. Der Druekbehilter,der 6 eingeschwei~te Stutzen ha%,nimmt die aus einem ~arkoseappana% zuflie1$enden Gasgemisehe auf. In ihm wird /bet eine Schraubenklemme ein Druek yon 40o em Wasser ge- halter.4 Stutzen f/hren ~ber Schlauchverbindungen zu Prizisions- nadelventilen,mi% welchen die AtemvolUmina der Tiere reguliert werden k~nnen.Die Nadelventile leiten welter zu TrachealkanGlen. Deter Expirationsschenkel ist mit dem Unterbrecher verbunden. Letzterer besteht aus einer motorgetriebenen metallenen Welle (Durchmesser 11 mm) und aus einem umgebenden Stahlzylinden.Wel- !e und Zylinder weisen in aer Zahl der Tieranschl~sse Bohrungen ~on 5 mm auf.Die vorderen Bohrungen des Zylinders tragen Stut- zen zur Aufnahme der Verbindungsschliuche,die hinteren BohrlS- chen endigen frei. Je nach Position der Welle wird der Casstr0m und damit das ganze System unterbroehen oder freigegeben.Wie anhand yon Blutgasanalysen gezeigt %~rde,kSnnen Ratter auch ~ber l~nge~e Zeit mit diesem zeitgesteuerten Druekgenerator beatmet werden. Dr. G. Cunitz,Abteilung fur Anaesthesiologie der Universitit, 87 W~rzburg, Josef-Schneider-Str. 2

MORPHOLOGICAL CHANGES OF RED CELLS BY DIRECT LYTIC FACTOR AND PHOSPHOLIPASE A, STUDIED BY SCANNING ELECTRON MICROSCOPY (Mor- phologische Ver~nderungen der Erythrozytenmembran dutch den Direkt-Lytischen-Faktor und Phospholipase A. Untersuohungen mit dem Rasterelektronenmikroskop) B. Damerau, P.G. Lankisch, H. Alberti, H. Scholz

Morphological changes by the direct lytic factor of cobra venom (DLF) and phospholipase A of bee venom (Ph-ase A) on washed guinea-pig red cells were investigated with the scanning electron microscope. Ph-ase A induced slight swelling and changing of the discocyte forms to echinocytes with distinct intermediate forms. After prolonged incubation spherocytes with small protrusions appeared. In controls echinocytes occurred much later, whereas in hypotonic medium the cells changed from discocyte to spheroid forms without echinocyte intermediates. With DLF swelling of cells was less extensive but haemolysis was much stronger. The surface of the but slightly swollen and still discoid cells were covered with granular or fiber-like apposi- tions, probably extruding haemoglobin.

Dr. B. Damerau, Max-Planck-Institut f~r experimentelle Medizin, Abt. Biochemiscbe Pharmakologie, D-34OO G~ttingen, Hermann-Rein- StraSe 3.

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EFFECTS OF PROBENECID ON THE DISTRIBUTION AND BILIARY EXCRETION OF 3H-DIGITOXIN AND ITS METABOLITES (Uber den EinfluR yon Pro- benecid auf Verteilung und bili~re Ausscheidung yon 3H-Digitoxin und seine Stoffwechselprodukte) K.HoDamm,HoVorbringer,H.Heckert und W.Braun

Probenecid effects on the distribution and biliary excretion of 3H-Digitoxin and its metabolites were studied in the rat and mouse. Measurements of radioactivity in either ligated jejunal loups (rat and mouse) or after direct catheterization of the biliary duct (rat) showed a decreased biliary excretion of Digitoxin metabolites in both species after Probenecid. This effect is most probably due to a competitive inhibition of Probenecid on an active transport of Digitoxin metabolites during their biliary secretion~ The distribution of 3H-Digitoxin between plasma and liver,heart- and skeletal muscle, kidney, gut, and erythrocytes was not changed in rats through Probenecidwhere the plasma/organ quo~ent was found to be 0,2. In contrary to this the same quotient was found in the mouse between 2,0 and 4,0 and was reduced after Probenecid to values between 0,5 and 1,0o This effect may be explained through an interaction between Digitoxin and Probenecid on binding sites of plasma proteins~

K~176 Pharmakologisches Institut der Universit~t Hamburg, D-2000 Hamburg 20, Martinistro 52~

BILIARY EXCRETION OF BSP IN RATS PRETREATED WITH POLYCHLORINATED BIPHENYLS(PCB)o (Bili~re Ausscheidung yon BSP bet Rattan nach Vorbehandlung mit polychlorierten Biphenylen(PCB)o) H.G.Dammann und G.Czok

Biliary excretion was studied in rats(200g b.w.)pretreated with PCB(500mg/kg iop.)after 1,3,14,28,56 days.Similtaneously with an increased liver weight there was a closel7 related increase of bile flow and BSP-Tm. At the same time I~P content of the liver and BSP plasma concentration were decreased.These effects correspond to the increased BSP plasma clearance and were most pronounced at the 3rdday,and reached control level after 8 weeks. The rats with an average bow.of 220 g had already 3 days after TCB the same liver weights as those tested after 8 weeks (boW. 400g).This was the same in the case of BSP-Tm.Bile flow and BSP- Tm related to the liver weight were not influenced by TCB. In the bile electrolyte concentration remained unchanged,whereas bile salt concentration was below control levels.This indicates that the increased bile flow corresponds to an increased excretion of electrolytes and not of bile salts.BSP metabolism is accelerated by TCB.Only the excretion of BSP-conjugated was increased,BSP- unconjugated remained unchanged.The increased BSP-Tm is caused by the following 3 effects of TCB: accelerated uptake of BSP resulting in lower BSP plasma concentrations, enhanced metabolic transformation of BSP and accelerated BSP excretion into the bile corresponding to the increased bile flow.

Dr.HoG.Dammann, Pharmakologisches Institut der Universit~t Hamburg, 2000 Hamburg 20, Martinistr. 52

R27

INFLUENCE OF ASCORBIC ACID ON WARFARIN METABOLISM (Der EinfluB yon Ascorbin- s~ure auf den Warfarin-Metabolismus) F.W. Deckert and K.P. Link

An increased sensitivity of vitamln-C_-deficient guinea pigs to single doses of coumarin anticoagulants was manifested by marked increases in clot time elevation, followed by greatly prolonged recovery periods, and by higher mortality rates after either single or multiple warfarin (W) doses. Since the biotransformation of W (F.W. Deckert et al., Acta Pharmacol. et Toxicol. 29 (Suppl. 4), 9, 1971) primarily involves hydroxylation and since increased half-lives of some drugs due to their decreased hydroxylation in C-deficient guinea pigs has been demonstrated by others (e.g.J. Axelrod et al., J. Pharmacol. Exptl. Therap. ill, 176, 1954; A.H. Conney et al., Ann. N.Y. Acad. Sci. 92, 115, 1961), impaired W metabolism might account for the observed prolongation of the hemorrhagic tendencies in scorbutic animals. Hence, the effect of dietary l-ascorbic acid on the rate of metabolite excretion, as well as on the nature of the metabolites excreted (as a function of time), was studied over the range 1-100 mg/kg W. No significant and consistent differences in the rate of C-I~-W metabolism (as measured by the gross urinary excretion of labeled compounds) could be detected at any dose level for control vs deficient guinea pigs. Smallchanges in metabolitepatterns could be detected at high dose levels, but these changes cannot be readily interpreted. Thus, the prolonged hypoprothromblnemic response to W in scorbutic guinea pigs probably cannot be accounted for solely by alterations in W metabolism due to C-deficiency. Alternate explanations include the following: i) Vitamin K metabolism, 2) clotting factor or other protein synthesis, and 3) capillary fragility; the latter seems to be indirectly supported by these studies.

Dr. F. Deckert, Department of Biochemistry, University of Wisconsin, Madison, Wisc. 53706 (USA) (Presently: Institut fur Toxikelogie der Unlversit~t, D-7400 TGblngen, WilhelmstraBe 56)

DISTribUTION OF 125j-TOXIN IN GENERAL TETANUS OF RATS (Verteilung von ---J-Toxin beim qeneralisierten Tetanus der Ratte) W,Dimpfel und E, Habermann General tetanus was produced by i.v, administration of al�89 dose ( 2Ong/kg ) of highly purified toxin labelled with J. Radioactivity was examined in plasma, liver, muscle and different parts of the CNS at different times. No radioactivity except that transported by the blood stream was found in cerebrum and cerebellum, whereas high amounts were measured in the brain stem, cervic~ cord and the lumbar cord. The main binding site of tetanus toxin applied i,v, is located in the grey matter.- Antitoxin (500 U/kg) injected i.m, together with toxin or iO hours later prevents the entry of toxin into the CNS as well as the symptoms of tetanus. When administered 48 hours after toxin, it fails to diminish the ~ ioactivity of the spinal cord or the severity of symptoms. ---J labelled toxoid ( 1 pq/k~2 ~ i.v, entered the spinal cord to

a lesser extent than toxin. - -J labelled antitoxin given i.v. in comparable doses could not be detected in the CNS. The correlation between our findings with ~eneral tetanus to those of Habermann (Naunyn-SchmiedebergJs Arch, Pharmacol. 272p 75-88 (1972) with local tetanus is so close that we postulate the same mechanism of entering the CNS, i,e. by nerve roots. E, Habermann, Pharmakologisches Institut der Universit~t Giessen, D-6300 Giessen, s I,

R28

EFFECT OF PARAOXON ON THE LIPOLYTIC SYSTEM OF FAT CELLS (Einfluss yon

Paraoxon anf das tipolytische System der Fettzelle) V. Dinnendahl, H.-D. Peters , K.U. Helm und P.S. SchSnhSfer

The inhibitory effect of organophosphorous compounds has often been investigated with lipase from several t issues. In our experiments with isolated epididymal fat cel ls paraoxon caused a dose-dependent inhibition of lipolysis in the LD50 dose-range (10 -7 - 10 -5 M). The inhibition occured in unstimulated as well as stimulated lipolysis elicited by 10-6M norepinephrine or 3 x 10-3M dibutyryl-A-3 ' , 5' -MP. Since the hormone-sensi t ive lipolytic system of fat cells is activated by adenyl cyclase the influence of paraoxon on enzymes and precursors of this system was tes ted. Paraoxon did not affect adenyl cyclase activity, phosphodiesterase activity, ATP-levels , A-3 ' , 5 ' -MP accumulation in intact cells nor the binding of A-3' , 5 ' -MP to the receptor protein of muscle protein kinase. Obidoxime, HS 6 and atropine (up to 10-3M) did not abolish the 50% inhibition of maximum tipolysis elicited by 10-6M paraoxon. The results indicate that paraoxon affects only the target enzyme tr iglyceride lipase of the adenyl cyclase system. Furthermore, this inhibition is not revers ible by antidots tested. A direct interaction between paraoxon and oximes was not observed in this test system.

Dr. V. Dinnendahl, Pharmakologisches Iustitut der Rheinischen Fr iedr ich-Wilhelms- Universit~it, D-53 Bonn, Reuters t rasse 2 b.

CHARACTERISATION OF PULMONARY-STRETCH-RECEPTORS (PSR) IN GUINEA PIGS BY BRADYKININ AND HISTAMINE (Zur Charakterisierung yon Lungendehnungsrezeptoren (LDR) des Meerschweinchens durch Bradykinin und Histamin) E. Ch. Dittmann, U. Kilian When recording action potentials of the whole N.vagus in guinea pigs, MAR- QUARDT (Naunyn-Schmiedebergs Arch. Pharmacol., 253,207-220, 1966) found that bradykinin applied i.v. had a pure myotropic effect, whereas ETSCHEN- BERG (Diss., KGln 197o) stated a combined myotropic/neurotropic effect after applying histamine i.v.. On the basis of these results we examined bradykinin and histamine in functionally isolated single fibres of the N.vagus. In lo3 experiments under intermittent positive pressure breathing 2o% of psr showed a low, 50% a middle and 30% a high threshold. All types of psr, especially those with the high threshold, were sensitized by the myotropic/neurotropic acting histamine. When bradykinin was applied, only the psr with the high threshold were sensitized, whereas the discharges of the other types were reduced. It is probable that a decrease in alveolar space is the result of an increase in capillary space after bradykinin is applied (ROCHA E SILVA et al., J. Pharma- col. Exp. Ther., 128,217-226,196o). Therefore we suppose that the psr with the low and middle threshold can be localized in the region of the pulmonary alve- ols and the terminal bronchioles (tissue with only a few smooth muscles and a low resistance against the intermittent positive pressure breathing). In case of psr with a high threshold we suppose a localisation in the region of the bron- ches (tissue with a abundant smooth muscles and high resistance).

Dr. Dr. E. Ch. Dittmann, Pharmakologisehes Institut der Unlversit~it zu KSln, 5 KSln 41, G1eueler Str. 24

R 29

POLYAMINES IN RAT LIVER DURING STARVATION AND REFEEDING

(Polyamine in Rattenleber bet Hunger und WiederfGtterung) S Domschke,

H D. S61ing

Polyamines (spermidine, spermine) and their precursor putreseine are sug-

gested to be growth factors and to be involved in regenerative processes via

regulatory influences on RNA metabolism (Raina A., J~nne J., Fed. Proc.

29:1568, 1970). The present study was designed to elucidate the role of polyamines in livers from starved and refed rats. Polyamines were determined fluoromeirically after they had been purified in an isotope dilution procedure employing dansylation and separation by TLC. Activities of ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase (SAN~D) which are essentially engaged in polyamine formation were measured by use.of C-14-substrates. Following 12 hrs of starvation, ODC activity was reduced markedly (13 per cent of normal) and was decreased to values near zero after 3 days of fasting, whereas SAMD activity declined not so drastically (50 per cent of normal). Putrescine and spermidine contents of livers from starved rats fell compatible with decreased enzyme activities. However, spermine contents remained almost unchanged. On refeeding, within 3 hrs ODC activity increased about 7- fold as compared to controls, whereas the rise in SAMD activity was less pronounced. Attributable to increased enzyme activities in the refeeding period, the hepatic polyamine pattern showed strong tendency towards normalization. It is of considerable interest that hepatic polyamine and RNA contents fluctua- ted concomitantly during starvation and refeeding thus providing another aspect of possible interactions between polyamine and nucleic acid metabolism.

Dr~S. Domschke, Biochem. Abtlg., Med. Univ.-Klinik, D-3400 GOttingen

THE SIGNIFICANCE OF THE BL00D-BRAIN-BARRIER FOR CHOLINE PENE- TRATION (Die Bedeutung der Blut-Hirn-Schranke fGr Cholin) K. Dross and H. Kewitz

The influx of choline into the brain can not be demonstrated if this compound itself is administered even i. v. since choline disappears from the blood within a minute or two to the normal level again. But injecting phosphorylcholine (PCh) in a dose of 160 mg/kg as calcium salt its concentration in the blood went up to almost 100 times of the initial value of 9 x 10-9 moles/ml. This was followed by a rise of the choline concentration in the blood from 1.1 x 10 -8 to 9 x 10 -8 moles/ml within two minutes declining to 3.4 x 10 -8 moles/ml within 20 minutes, Three minutes after the injection of PCh the choline in brain was elevated 3 fold from 3 to 9 x 10 -8 molee/g whereas POh was increased by 15 ~ only that means from 2.3 to 2.7 x 10 -7 moles/g which is not a significant difference. From these data it can be concluded that choline penetrates easily into the brain. The hindrance towards an equilibrium of labelled choline between blood and brain is due to the concentration gradient which is directed from brain t o blood and which is derived by the formation of choline within the brain.

Dr. K. Dross, Institut fGr Klinische Pharmakologie der Freien Universit~t Berlin, D-IO00 Berlin 45, Hindenburgdamm 30

R 30

METABOLISM OF ANISOLES IN LIVER MICROSOMES (Stoffwechsel von Anisolen in Lebermikrosomen) W. DUnges, H. Heinemann und K.J. Netter

A micro technique of incubation, extraction, extract-concentration and gas chromatography has been developed ~or the study o~ mlcrosomal metabolism, p-Methyl- anisole was used as a model substrate. After incubation with microsomes from rabbits stimulated with phenobarbitone, the O-demethylation product (p-cresole), the ring-hydroxylated compounds o-hydroxy-p-methyl-anisole and m-hydroxy-p- methyl--anisole could be identified through comparative gas chromatography. In addition to these metabolites, first described by J. Daly (Biochem. Pharmacol. 19., 2979-2993, 197o), p-methoxy-benzyl-alcohole and probably p-hydroxy-m- methyl-anisole could be found. As only small losses of the volatile substrate occur with this method, we could show the decrease of the p-methyl--anisole peak simultaneously with the increasing occurance of peaks from metabolites with time.

Dr. W. D~Jngesr Pharmakologlsches lnstitut der Univers~t~t Mainzr D-65oo Mainz, Obere Zahlbacher Str. 67

ZINC, MAGNESIUM, AND POTASSIUM IN SERUM AND TISSUES FOLLOWING SUBLETHAL CYANIDE POISONING (Der Zink-, Magnesium- und Kalium- gehalt in Serum und Gewebe nach subletaler Cyanidvergiftung) H. G. Cure, K. D. Friedber~

Typical symptoms of sublethal cyanide ~oisoning (s.c.p.) are quickly reversible. However, high complex-affinity of the cyanide ion to Zn and other metals might possibly cause late effects. Consequently, the influence of s.c.p. (75/uMol NaCN/kg s.c.) on Zn levels in serum and several tissues of S.C. rats was studied by atomic absorption spectrophotometry. Additionally, we investigated involvement of Mg, Ca, K, and Na in serum.

Two hours after s.e.p., Zn in liver and kidneys was significantly elevated. A maximal increase (35-50 %) in liver, lungs and kidneys was measured 1-2 days later. After 8 days, Zn content of liver and kidneys still remained higher than normal. Repeated induction of s.c.p, failed to enhance these effects. 3o minutes following a single s.c.p, there was a rise in serum levels of Zn (30 %), Mg (6o %), and ~ (7o %). The origin of such amounts of Zn mobilized is unknown, since none of the tissues tested (liver, lungs, kidneys, spleen and heart) showed a reduction of Zn. Elevation of K parallels results of Tauberger and Karzel in cat-plasma (Naunyn-Schmiedebergs Arch. exp. Path. Pharmak. 2~9, 218-219, 1968). Rise of serum-Mg in s.e.p, has not been described so far. Both effects are thought to result from transitory hypoxia.

H. G. Duve, Institut fHr Pharmakologie und Toxikologie der Universit~t, D 34oo GGttingen, GeiststraBe 9.

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INFLUENCE OF TOLBUTAMIDE AND GLIBENCLAMIDE ON LIPOLYSIS, PHOS- PHODIESTERASE AND CYCLIC 3',5'-AMP LEVEL IN ISOLATED FAT CELLS (EinPluB yon Tolbutamid und Glibenclamid auf Lipolyse, Phospho- diesterase und den 3',5'-AMP-Gehalt in isolierten Fettzellen) R. Ebert, O. Hillebrandt, U. Schwabe

Tolbutamide and glibenclamide inhibited isoproterenol-induced lipolysis in isolated Fat cells os rats in a dose dependent manner, whereas carboxytolbutamide slightly enhanced glycerol production. Lipolysis induced by dibutyryl cyclic AMP was reduced by very low concentrations of glibenclamide (o,I ~M) and tolbutamide (20 ~M) by about 40 %, but markedly increased by higher concentrations oF glibenclamide. Phosphodiesterase activity in adipose tissue was inhibited by high concentrations of tolbutamide and glibenclamide. In addition, tolbutamide further increased the isoproterenol-induced @levation os cAMP levels in isolated fat cells, whereas glibenclamide shoved no significant effect on cAMP. The antilipolytic potency of glibenclamide and tolbutamide was highly dependent on the Ca ++ concentration in the medium: When the fat cells were prepared in the previous collagenase step without Ca ++ , the inhibition os isoproterenol-induced lipolysis was largely enhanced in the presence os Ca ++ up to 1,2 raM, but completely blocked by further elevation of the Ca ++ concentration to 2,4 mM. Our results suggest, that the antilipolytic actiQn of sulEonylureas is independent from the level of cAMP, but is possibly related to changes in calcium metabolism.

Dr.. R. Ebert, Institut s Pharmakologie, Medizinische Hochschule Hannover, 3000 Hannover-Kleefeld, Roderbruchstr. 101

TRE PHYSIOLOGICAL DISPOSITION OF FENFLURAMINE IN THE RAT (Vertei- lung und Ausscheidung von Fenfluramin bei der Ratte). M. Eichelbaum and I.G. Sipes

Plasma and tissue concentrations and urinary excretion of fenfluramine (F) and norfenfluramine (NF) were determined in the rat after the i.p. administration of 30 mg/kg F. The concentration of F and NF was highest in lung, salivary glands, brain, spleen, and adrenal medulla. Lower concentrations in decreasing order were found in the liver, kidney, heart, intestine, stomach, muscle, fat, and plasma. The plasma and tissue levels of F and NF decline monoexponentially suggesting an open one- or two-compartment model. The half life time for F ranges from 3.9 to 6,2 h and for NF from 8.7 to 15.3 h. 24% of the dose are excreted in the urine as F and 13% as NF. The markedly lower tissue concentrations of F and NF as found after chronic F treatment imply that F induces its own metabolism. In consequence only 12% of the dose is excreted as F and 8.5% as NF. As the N-deethylation in vitro is increased 2-fold after chronic F treatment the lower tissue concentrations and urinary excretion of NF can be explained only by assuming an additional indu- cible pathway of F degradation. Since appreciable amounts of NF are found in all tissues, the possibility exists that part of the pharmacological actions may be caused by NF.

Dr. M. Eichelbaum, Medizinische Poliklinik, D-63 Giessen, Friedrichstrasse 27.

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NEW PHENYL-AMINOETHANOLS WITH EFFECTS ON THE ADRENERGIC ~- AND B-RECEPTORS (Neue Phenyl-amino-aethanole mit Wirkung auf die adrenergen ~- und B-Rezeptoren) G. En6elhardt

The efficacy of a series of new 1-(4-amino-3,5-dichloro-phenyl)- 2-amino-ethanols was tested in respect to the arterial pressure and heart-rate in anaesthetized cats; antagonis~ ~gainst tachy- cardia and lowering of the diastolic pressure caused by Isopro- terenol after i.v. application; furthermore broncholytic effects in guinea-pigs after i.v.-application (KONZETT-ROSSLER) and oral administration (KALLOS-PAGEL), in dependence of the substitution of the side chain nitrogen atom. Small alkyl groups up to 2 C atoms as substituents of the aliphatic nitrogen show an ~- mimetic and 8~-blocking effect as well as a slighter 89-blocking efficacy. Unbganched alkyl groups with 3 and 4 C atoms~are responsible for the blockade of the 8~-receptors. Branched alkyl groups at the nitrogen atom of the side chain cause a distinct B,-mimetic activity accompanied by a blocking effect and an only slight intrinsic activity on the 81-receptors. The relative efficacy decreases from t-butyl, i~opropyl, t-pentyl to sec butyl. These Bg-mimetics posses an intensive and long lasting broncholytic e~fect also after oral administration.

Dr. G. Engelhardt, Abteilung Biologische Forschung, Pharma- kologie D der Fa. Dr. Karl Thomae GmbH., Biberach/Riss

INHIBITORY EFFECTS OF SPIROLACTONES ON THE SYNTHESIS OF ALDOSTERONE IN VITRO (Wirkung von Spirolaktonen auf die Aldosteronsynthese in vitro) H. Erbler

Adrenal quarters os sodium deficient rats were incubated in a modified KRB-bufs with increasing amounts, of ACTH, dibutyryl-cAMP (DBA), potassium chloride and Angiotensim-II respectively. Aldo- sterone was determined by gas chromatography with electron-capture detection, corticosterone was assayed fluorometrically.

Addition of Spironolactone (SC 9420) or Aldadiene (SC 9376) to the incubation medium (I0-5M - I0-4M) resulted in a dose dependent inhibition os aldosterone production. The ACTH- and DBA-stimulated synthesis of corticosterone was not inhibited by Spironolactone indicating that the synthesis os aldosterone is inhibited at a step following corticosterone. This assumption is supported by a highly significant inhibition of the conversion of 1~C-corticoste- tone to 14C-aldosterone by Aldadiene.

In addition, the basal production rate of aldosterone in adrenals os normal fed as well as os sodium deficient rats was inhibited by Aldadiene.

Dr. H. Erbler, Institut s Pharmak01ogie , Medizinische Hochschule Hannover, 3000 Hannover-Klees Roderbruchstr. 101

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KINETICS OF OXIDATIVE TRICHLOROETHYLENE METABOLISM IN MAN (Kinetik des oxidativen Trichlor~thylen-Umsatzes im Menschen) T.Ertle, D.Henschler, G.MGller, M.Spassowski

Groups of 5 volunteers were exposed to analytically controlled concentrations of trichloroethylene (Tri) vapor: 6 hrs/day for 5 days at (I) 50 ppm, (II) iOO ppm, or (III) 250 ppm - 12 min/hr 6 times/day (equivalent to a mean of 50 ppm). Blood trichloroethanol (TCE) concentrations, as determined in 0.2 ml samples by a gas chromatographic (e.c.d.) technique, rose to (I)1.6, (II) 3.2, (III) 1.7 ~g/ml at the end of the first 6 hr-exposure, accumulating at the end of the 5 day period to (I) 2.0, (II) 5.0 and (III) 2.4 ~g/ml, respectively. Urinary excretion of TCE and trichloroacetic acid (TCA), single and combined, increased from day to day in linear progression with factor 1.3, approaching - at day 5 -24-hr values of 375 mg (I+III) and 520 mg (II); thus, evaluation of Tri exposure from TCE+TCA excretion is only possible when taking into account the previous body load. - TCE blood levels of two individuals were determined following ingestion of 15 mg/kg chloral hydrate (therapeutic dose). Extra- polation to TCE levels after Tri inhalation reveals that 8 hrs Tri exposure to 50 ppm corresponds to an interval of 7-8 hrs, lO0 ppm to only 2 hrs after chloral hydrate intake. Accumulation of TCE is believed to account for the "psycho-organlc syndrome" in persons with chronic exposure to Tri.

Prof.Dr.D.Henschler, Institut f~r Pharmakologie und Toxikologie der UniversltMt, D-87 WGrzburg, KoellikerstraSe 2.

THE EFFECTS OF SOME DRUGS ON THE PROTEIN BINDING OF PHENOL RED AND DIGITOXINo(Uber den Einflu~ verschiedener Pharmaka auf die Eiwei~bindung yon Phenolrot und Digitoxin~ R,Erttmann,R,Beeck, K,H,Damm und W,Braun

In der vorliegenden Arbeit werden die Konstanten K Iund n fGr die Albuminbindung yon Phenolrot und 3H-Digitoxin bestimmtoDie ~nd~ung dieser GrG~en durch den Einflu~ yon Phenylbutazon,Sali- cyls~ure,Prohenecid und Benzbromaron wurde in Dialyseversuchen untersuchtoDer verdr~ngende Effekt yon Phenylbutazon und Pro- benecid auf die Phenolrotbindungsrate wird durch die ver~nder- ten K Iund n-Werte quantitativ erfa~t.FGr Phenylbutazon, Sali- cyls~ure,Probenecid und Benzbromaron konnten nur bei hoher Do- sierung und niedrigem Albumingehalt Verdr~ngungseffekte gegen- dher Digitoxin nachgewiesen werden.Mit einer klinischen Bedeu- tung der dargestellten Kompetitionsph~nomeae ist demnach nicht zu rechnen.

RoErttmann, Pharmakologisches Institut der Universit~t Hamburg, 2000 Hamburg 20, Martinistro 52

R 34

INVESTIGATIONS ON CARBON TETRACHLORIDE-INDUCED FATTY INFILTRATION OF THE KIDNEYS (Untersuchungen zur Fetteinlagerung in die Nieren nach Tetrachlorkohlenstoff) C,-J.Estler~ H.-J.Pesch, B.Schmidt

In male mice treated with 3 ~l/g carbon tetrachloride i.p. the kidney weight had increased from 6.4 to 8~Img/g body weight.This was mainly due to a higher water content but the dry weight of the kidneys was also increased. The microscopic examination revealed albuminous swelling, increased protein permeability and a few scattered necroses. An increased deposit of fat in the epithelium of the tubules could not be evaluated under the light microscope. Quantitative biochemical analysis,h0wever,showed that 12 hours after the administration of CC14 the triglyceride content of the kidneys had significantly increased from 6.9 to 8.8 mg/g.Parallel to the increase of the renal fat content the level of non-esterified fatty acids in the serum rose by 18~% within 12 hours and then declined gradually.After 24 hours the non-esterified fatty acids in the serum and the triglyceride content of the kidneys had reached normal values again.B-Pyridylcarbinol(50~g/g i.m.every 6 hours) prevented both the rise of the non-esterified fatty acid level in the blood and the increase of the triglyceride content of the kidneys. This suggests that these changes are causally interrelated and that the deposition of fat in the kidneys of mice poisoned with CCla is the result of an enhanced uptake of fatty acids by the kidneys with subsequent esterification to triglycerides.

Prof. Dr. C.-J.Estler, Pharmakologisches Institut der Universit~t Erlangen-NGrnberg, D-852 Erlangen, Universit~tsstr. 22

THE INHIBITION OF THE PHOSPHORYLATION OF CHOLINE BY CPD 48,/80 AND ITS RELATIONSHIP TO THE HERPES-INDUCED GIANT CELL FORMATION. (DIE HEMM- WlRKUNG VON CPD 48/80 AUF DIE PHOSPHORYLIERUNG VON CHOLIN UND IHRE BEZIEH UNG ZUR H ERPESI NDUZI ERTEN RI ESENZELLBI LDUNG.) D. FALKE, I. JUST, S. DUNDAROFF.

Besldes its hlstamin releaslng propertles Cpd 48/80 inhibits the penetration Of Herpesvlrus and the Herpes-induced glant cell formation~ The DNA-, RNA- and protein synthesls of both uninfected and herpes-infected cells is not affected by Cpd 48,/80. In contrast, the incorporation of radioactive choline or ethanolamine is reduced considerably. Tritium- labelled 48/80 is strongly bound by the cells (Falke, Kahl: J. gen. Virol. , 10, 273 (1971) Falke, Kahl, Netter: Naunyn-Schmledebergs Arch. Pharmak., Suppl. Vol. 270, R 30 (1971).

The inhibition of the blosynthesls of leclthine- or of colamlne-cephaline could be due to a blockade of the penetration of choline or an inhlbltion of its phosphorylation. - The experiments demonstrated an inhibition of the choline-kinase and furthermore a reduction of the uptake of choline into cells. The dimer of CI0d 48/80 only weakly affects the choline- kinase, whereas the inhibition of the biosynthesis of leclthine is as strong as that exerted by Cpd 48,/80. The choline-kinase does not seem to be a membrane bound enzyme. Its type of inhibition is of the mixed type. There is no indication of a causal relationship between the inhibition of the lecithine-blosynthesis and the inhibition of the giant cell formation. Both activities rather seem to be effected by reactions of Cpd 48/80 with different "receptor" sites.

Institut fiJr Med. Mikrobiologie der Universit~f, D 65 Mainz, Augustusplatz 1 - Germany.

R 35

EFFECT OF BUPHENIN ON CAPILLARY FLUID EXCHANGE IN THE CAT HINDLIIVfB (t~ber den Einflu~ von Buphenin auf den kapill~ren Flfissigkeits- austausch im Hinterlauf der Katze) W. Felix, J. Remien

Hindlimbs of cats (chloralose anesthesia) in situ were perfused at constant arterial flow and constant venous outflow pressure (W. Felix, J. Remien, K. H~llfritzsch, Pflfig. Arch. 329, 352-359, 1972). Buphenin i. a. and i.v. ( 0, I - 1,0 mg/kg) effected a net fluid movement into the blood vessels. Probably this effect was elicited by a decrease of capillary filtration pressure due to an opening of precapillary sphincters: at constant flow the enlarged capillary cross-section area leads to a decrease of resistance and mean capillary pressure, respectively. The increased capillary filtration coefficient indicated that capillary surface area was enlarged. It cannot be excluded that Buphenin influenced capillary fluid movement by mechanisms other than hemodynamic, as net fluid absorption did not always parallel the dilation of the resistance vessels. The response of the resistance vessels, however, does not indicate a response of precapillary sphincters. During repeated applications of the drug, the effect on capillary fluid exchange was diminished or abolished.

Prof. Dr. med. W. Felix, Pharmakologisches Institut der Universit~t, D-8000 Miinchen 2, Nu~baumstr. 26

EXOCRINE PANCREATIC SECRETION IN THE RAT AS AFFECTED BY ANTIBIOTICS. Die exocrine Pankreasfunction der PLatte unter der Behandlung nit Antibiotica. K. FLEISCHER Exocrine pancreatic secretion was studied in rats being treated subchronically with orgtetraeycline or chloramphenicol, respectively. Volume, total protein and amylase activity excreted were determined in urethan anesthesized, cannu- lated rats ( 400 g b.w.). Excretion was followed up to two hours after operation in two sampling periods of one hours duration. Maximal stimulation of exocrine pancreatic secretion was achieved by infusion of bo~h secretin(25U/kg/hr) and pancreocymin (25 U/kg/hr). 0xytetracycline (VENDARCIN ~) was given subcu- taneously ~n a three days course ( 2oo and 4oo mg/kg/d), chloramphenicol (CHL0- ROMYCETIN ~L) for three days by mouth (6oo and 12oo mg/kg/d)o Volume of pancreatic secretion was not consistently altered by both regimens as compared to controls. In contrast, total protein (mg/ hr) and protein concentration (mg/g/ hr) of pancreatic juice was found to be significantly decreased in both groups. Amylase secretion (U/hr and U/g/hr, respectively) was also significantly diminished following both regimens. The decrease in protein excreted, ~s well as in amylase activity produced, were most prominent in the second sampling period (2.h~), both effects showing a clear dose-response relationship.

Blood levels of oxy~etracycline within 15 hrs. after dosage fell from 29 to 1ojug/ml (for 2o0 mg/kg/d) and from 56 to 34jug/ml (4oo mg/kg/d), chlorampheni- collevels ranged at about 8~g/ml at the same time for both doses given. The results obtained are interpreted to show an impairment of pancreati~ protein biosynthesis an~ secretion by certain broad spectrum antibiotics. Clini- cal implications will be discussed in relation to the pathogenesis of side - effects of these antibiotics on the G.-I.-tract.

Authors adress: Dr.K.FLEISCHER, D-87-WURZ~3URG, Medizin. Uni~.-Klinik

R 36

EXPERIMENTS WITH RATS ON THE MODE OF ACTION OF CYSTEA- MINE ON THE PITUITARY ADRENOCORTICAL SYSTEM (Untersuchungen an Ratten fiber die Wirkungsweise von Cysteamin auf das Hypophysen- Nebennierenr inden-System) K. Flemming, B. Geierhaas , V. Seydewitz

Prev ious invest igat ions w~h ra t s had shown that the co r t i cos te rone in- c r e a s e in the adrena ls and in the blood following whole body X - i r r a d i a - tion was inhibited by cys teamine . Now it was tes ted if cys teamin were a lso able to influence the co r t i cos t e rone i nc rea se caused by drugs and which effects the substance exer t s on the pi tui tary adrenocor t i ca l sys tem. For this r ea son the content of the adrena ls of co r t i cos te rone and DCC was de te rmined as well as the act ivi ty of definite enzymes at var ious t imes a f te r the injection of the drugs. It was found that cys teamine inhibits the inc rease in the cor t i cos te rone content of the adrena ls that is caused by a r e l e a s e of ACTH f rom the pi tui tary due to var ious drugs. The mechanism of this effect of cys te - amine is an inhibition of the l l - ~ h y d r o x y l a s e sys tem which is followed by a shift f rom cor t i cos te rone to DOC in the adrenals and in the blood. In addition to this specif ic effect of cys teamine on the adrenal cor tex itself, the substance has a non-speci f ic effect on the pi tui tary causing a r e l e a s e of ACTH like many other drugs.

Prof . Dr. K. Flemming, Institut fiir Biophysik und Strahlenbiologie der Universi t~t F re iburg i. Br . , D-7800 Freiburg, Alber t s t r . 23

Resorption und Stoffwechsel yon Tri- und Tetrahydroxy~thylrutosid bei Ratten. H. FSrster

Die Resorption yon Tri- und Tetrahydroxy~thylrut6sid wurde dutch Messung der Substanzkonzentration im Pfortaderblut und im arteriellen Blur bei narkotisierten Ratten w~hrend Durch- strSmung des D~nndarms mit rutosidhaltiger LSsung Eemessen, Beide Substanzen wurden im Pfortaderblut in hSherer Konzentration gefunden als im peripheren Blur. Die %eilweise hohen arterio- portalen Konzentrationsdifferenzen sprechen f~r eine relativ rasche Resorption aus dem Darmlumen. ~r der Resorption yon Trihydroxy~thylrutosid wurde ein Metabolit im Blur nachgewiesen, dessen Bildung in der Leber bei Leberperfusionen best~tig% ~rde. In der isoliert perfundier%en Rattenleber wird Trihydroxy~thylrutosid rasch aufgenommen (biolo~ische Halbwer%szei~ ca. 90 Min) und teilweise unver~ndert in der Galle aus~eschieden oder zu dem Metaboliten umEesetzt, der sowohl in der Galle als auch im Perfusionsmedium gefunden w11rde. TetrahydroxyHthylrutosid wird dagegen yon der Rat%enleber nur sehr lanEsam aufgenommen, wahrscheinlich nicht umgesetz%, jedoch ebenfalls in der Galle ausgeschieden.

H. FSrster, Ins%i%ut f~r vegetative Physiologie der Universi%~t Frankfurt, D-6000 Franks 70, Ludwig-Rehn-StraBe 14.

R 37

ISOLATION OF RIBONUCLEIC ACID FROM CALFS' HEARTS AND THE BINDING OF CARDIAC GLYCOSIDES TO THIS RNA (0her die Isollerung von Ribonukleins~ure aus Kalbsherzen und die Bindung von Herzglykosiden an diese kardlale RNA) J. Franke, U. Wollert

Cardiac glycosides and cardiac glycoside-like acting bisguanylhydrazones (BGs) can be accumulated in the mlcrosomal fraction of guinea pigs' heart muscles. BGs are highly bound to RNA. It had to be testedt whether the binding of cardiac glycosides to RNA is of importance for the accumulation of these substances in the microsomal fraction of the heart. RNA was extracted from the ribosomal fraction of calfs' hearts. The yield was 23.5 mg RNA/loo g wet weight of the heart muscle. The interaction of RNA and cardiac glycosides was determined by means of ultracentrifugatlon using tubes with a capacity of 2 ml. Binding measurements were made in the presence of o.2 M tris buffer, pH 7.4o, using a RNA concentration of o.5 o~ and 3 �9 lo -7 g/ml and 3 �9 lo -6 g,/ml of 3H-digitoxin (DT), 3H-digoxin (DG) and 3H-ouabain (OU). The percentage cx of free substance was found as shown below:

3 �9 lo -7 g/mh DT 85.2 + 4.7; DG 91.5 + o.8; OU 92.2 + o.3 3 �9 lo -6 g/mh DT 86.o + 3.1; DG 93.8 + 1.9; OU 91.8 + o.2

It is suggested that the binding of cardiac glycosides to RNA is of no importance for the accumulation of these substances in the mlcrosomes of the heart muscle.

Dr. J. Franke, Pharmakologlsches Institut der Universlt~t Mainz D-65oo Mainz, Obere Zahlbacher Str. 67

CIS- AND TRANS-I,2-DICHLOROETHYLENE:TWO NEW INHIBITORS OF MICRO- SOMAL ENZYMES (Cis- und trans-l,2-Dichlor~thylen: Zwei neue Hemm- stoffe mikrosomaler Enzyme) K.J.Freundt, J.Macholz

Aliphatic halogenated hydrocarbons include several compounds (CC14, C2HC13, CHCI~, CH3CI , CgHFqBrCI) which influence microsomal enzymes. We hav~ found thas two further substances from this group, cis- and trans-l,2-dichloroethylene (DCE), inhibit hepatic mixed-function oxidases already at low doses. 8-hr-exposures of female rats (strain Wistar, weight abt. 220 g) to staggered concentrations (between 200 and i000 pp~.) of cis- and trans-l,2-DCE, respectively prolong the hexobarbitone sleeping time and zoxazolamine paralysis time in a dose-dependent and significant manner. The hexobarbitone effect is prolonged to a higher degree than the zoxazolamine effect. In rats equal doses of the two agents inhibit the formation/excretion of total 4-amino-antipyrine (4-AAP) in a dose-dependent and pronounced fashion following oral uptake of amidopyrine (20 mg/kg). This (reversible) effect persists for approx. 6 hrs after exposure. The excretion deficit is fully compensated for during the later excretion phase. Five 8-hr-exposures to iOOO ppm trans-l,2-DCE on consecutive days do not inten- sify the inhibitory effect. However,N-acetylation of 4-AAP and O- glucuronidation of 4-OH-phenazone remain unaffected even after exposure to iOOO ppm trans-l,2-DCE/8 hr. The hemcytochrome-P-450 level in isolated rat liver microsomes decreases significantly by 17% after 8-hr-exposure to iOO0 ppm trans-l,2-DCE. The same dose causes no marked changes in conventional liver function tests.

Klaus J.Freundt, Institut fHr Pharmakologie und Toxikologie der Universit~t W~rzburg, D-87 W~rzburg, KoellikerstraBe 2

R 38

EFFECTS OF TEMPORARY FASTING ON FETAL DEVELOPMENT IN SMALL RODENTS (Beeinflussung der Fetalentwicklung kleiner Nagetiere durch vorUbergehenden Nahrungsentzug) H.Frohber~, A.Bauer

Bei Wistar-Ratten ffihrte Nahrungsentzug (N) vom 6.-lO.Tag p.o. zu Aborten, vom ll.-15.Tag p.o. beeinfluSte N die Fetalentwieklung jedoeh kaum. FHr NMRI-M~use wirkte 5-t~giger N vom 6.-10. oder ll.-15.Tag p.e. zumeist letal. N vom 6.-8. oder 9.-ll.Tag p.c. fHhrte zu Aborten oder Resorptionen (R), vom 12.-14.Tag p.o. zu R und Gaumenspalten (GS). GS wurden bei den Feten yon NMRI-M~usen aueh nach einmaliger oraler Applikation yon Dexamethason (D) zur Zeit des Schlusses der Processi palatini erzeugt: am 12.Tag p.e. 11,3 %, am 13.Tag 54,5 % und am 14.Tag 4,4 %. Wistar-Ratten erwiesen sich auch gegen Corticoide weniger empfindlich: i x 50 mg/kg p.o. D am 13.Tag p.c. erzeugte 17,3 %, bei NMRI-M~usen 81,5 % GS. Die bei M~usen durch 3-t~gigen N verursachte Embryotoxizit~t dGrfte teilweise Folge einer vermehrten endogenen Corticosteronproduktion sein. Massenhaltung auf engem Raum vom 12.-14.Tag p.c. erzeugte bei NMRI-M~usen GS und wirkte adulttoxisch. Bezogen auf die GS- Frequenz w~re Massenhaltung mit 3 x 25 mg/kg, N vom 12.-14.Tag p.c. mit 3 x i00 mg/kg Cortison oral vom 12.-14.Tag p.c. vergleichbar. Im Gegensatz zu diesen Cortisondosen erzeugte 3-t~giger N auch vermehrt R. Ursache dieses st~rkeren embryotoxischen Effektes dGrfte die dutch den N ausgelGste Hypoglyk~mie sein, da bei M~usen die hungerbedingte Embryotoxizit~t durch Glucose ver- mindert wird und Insulin allein auch teratogen wirkt.

Dr.H.Frohberg, Institut fur Toxikologie, E.Merck, 61 Darmstadt, Postfach 4119

EFFECTS OF INHIBITORS OF RENAL ELECTROLYTE TRANSPORT ON GLUCONE0- GENESIS IN VITRO. ( Die Wirkung yon Hemmstoffen des renalen Elek- trolyttransports auf die Glueoneogenese in vitro) G. FNlgraff, H. NNnemann

The effects of ethacrynic acid, chlorothiazide and ouabain on the rate of glucose formation from several substrates by rat renal cortical tissue slices were investigated in vitro. Slices of 2-6 mg dry weight were incubated for 60 min in Krebs Ringer bicarbo- nate buffer gassed with 5% C02 at 38 ~ C and pH 7.4.

Ethacrynic acid and ouabain increased the rate of glucose synthesis concentration dependent with fructose, pyruvate or lactate as substrates. The lowest effective concentrations were 10 -7 M ethacrynic acid and I0 ~5 M ouabain. Ten times higher concentrations exerted maximal effects. Both, ethacrynic acid and ouabain did not influence gluconeogenesis when intermediates of the Krebs cycle were added as substrates. Chlorothiazide in the wide concentration range of I0 -Io - 10 -3 M, however, did not increase the rate of glucose formation whatever the substfate was.

It has to be further investigated, which regulating enzymes of glycolysis and/or gluconeogenesis are activated or inhibited to explain these effects and how they are related to the effects on electrolyte transport.

Prof. Dr. G. FNlgraff, Abt. Pharmakologie der Med. Fak. der RWTH, D-5100 Aachen, MTI II.

R 39

PIPERAZINERINGDEGRADATION IN RING-SUBSTITUTED PHENOTHIAZINES AND ANTIHISTA- MINIC DRUGS RESULTING IN CUMULATING METABOLITES (Abbau des Piperazinrlngs bei kernsubstituierten Phenothiazinen und Antihlstaminika zu kumulierenden Metaboliten) H. J. Gaertner~ and U. Breyer

In analogy to findings on perazine (U. Breyer et al., ~xperientia, in press) we could demonstrate piperazine ring degradatio~to ethylenediamine in # additional piperazine-substituted phenothiazines. N-/~-(2-chloro-phenothiazinyl-lO)-propylJ-ethylenediamine was found in the tissues of male rats following chronic oral administration of prochlorperazine or perphenazine. Its concentrations in peripheral parenchymatous organs measured 12 hours after the last dosage of a 14 days' treatment with 2x25 mg/kg prochlorperazine were between 50 and 100~g/g tissue. An analogous 2-trifluoromethyl metabolite occur~edafter treatment of male rats with trifluoperazine or fluphenazine. It was also detected in a dog treated for 8 days with increasing doses of fluphenazine (up to 40 mg/kg). Twenty-four hours after the last dosage its concentration in the liver amounted to 20~g/g. Daily oral administration of 50 mg/kg chlorcyclizine to female rats resulted in the accumulation of N-(p-chloro-benzhydryl)-ethylenediamine besides chlorcyclizine and norchloreyclizine which has been observed by Kuntzman et al. (J. Pharmacol. exp. Ther. 149, 29-35, 1965). Twenty-four hours after the last dosage of a 14 days t treatment the concentrations in peripheral tissues were around lO00~g/g for norchlorcyclizine and 100 ~g/g for the ethylenediamine derivative. Investigations after longer intervals showed that this metabolite disappeared at a still slower rate than the demethylated compound.

Dr. H. J. Gaertner, Institut fur Toxikologie der Universit~t, D-7400 THbingen, Wilhelmstra2e 56

NEW MITODEPRESSIVE SUBSTANCES, COMPARATIVELY STUDIES WITH A SERIES OF ANT ICAN CEROU S DRU GS. VII. 2-HALOACETYLAM!N O --5-R- 1,3, 4-TH lAD IAZOLES. (Neue mitodepressive Verbindungen, vergleichend mlt einer Reihe yon antikarzinoma- tSsen Arzneimitteln studiert. VII. 2-Haloazetylamino-5-R-1,3,4-thiadlazole). F.Gag~u, AI.Cacoveanu, C. Nistor, U. Binder, E.Bebesel

AIs Fortsetzung ihrer Studien Liber neue 2-Amino-I, 3,4,-thiadiazolverbindungen pr~Jften die Verfasser dfe mitodepresslve Wirkung von 43 neuen 2-Haloazetylamino-5-R-thiadia- zolen. Die PrLifung erfolgte durch einen vergleichenden Biotest auF Radik~Jle von Lepi- dium satTvum L, die mit versch~edenen Konzentratlonen tier zu testenden Substanzen in Methylzelluloseemulsion (so k~nnen auch wasserunlSsliche Verbindungen geprUft werden) versetzt wurden. Der Test wurde vergleichend mit 20 klinisch bew~ihrten Antikrebsmitteln durchgefUhrt. Die Autoren fanden beachtliche mitodepressive Wirkung (Hemmungskoeffi- zient 50%, vergleichend mit SinalostR, 3 mg/ml, Mustin R, 0,3 mg/ml, Natulan R, 3,3 mg/ml) be1 folgenden Verblndungen : 2-Dichlorazetylamino-l,3,4-thiadiazol, 2-Di- chlorazetylamino-5-n-propyl- 1,3, 4-thladiazol, 2-Trichlorazetylamino-5-n-propyl- 1,3, 4- thlodlazol, 2-Dichlorazetylamino-5-n-butyl-1,3, 4-thiadiazol, 2-Trichlorazetylamino-5- benzy I- 1,3, 4- th lad iazo I, 2- Bromazety lamino-5- (3')-pyridi I- 1,3, 4- thiadiazol, 2- Bromazetyl- amlno-5-(4' )-pyridi I-1,3, 4- thiadiazol.

Dr. F. Gagiu, Onkologisches Institut- Clu b S.R.RumUnlen, Str.Republicii nr.34-36.

R 40

NEW MITODEPRESSIVE SUBSTANCES, COMPARATIVELY STUDIED WITH A SERIES OF ANTICANCEROUS DRUGS. VIII. N4-HALOACETYLAMINO-N1-R-BENZENESULFONA- MIDE-DERIVATIVES. (Neue mTtodepressTve Verbindungen~ verglelchend mlt ether ReThe von antikarzinomat~sen Arznelmitteln studiert. VIII. N4-Haloazetylamlno-N1-R-benzol- sulfonamTd-Derivate). F. Gaglu~ C. Nistor~ A. Cacoveanu~ E. Bebeseb U.Binder

Ausgehend von Angaben der Literatur, dab Sulfonamide AntTmetaboliten der Fols~ure sind und potentiell cytostatische Aktivitdt aufweisen~ synthetisierten die Autoren 53 neue N4-Haloazetylamino-N1-R-benzolsulfonamide und priJften ihre mitodepressTve Wirkung an Hand eines vergleichenden Biotestes auf RadikUle von kepidium sativum k. Bel 27 der untersuchten Verbindungen fanden die Verfasser verglelchend mit Sinalost R (3 mg/ml), Must~nR (0,3 mg,/ml) und Natulan R (3,3 mg/ml) eTnen Hemmungskoeffizien- ten von uber 50 ~ .

Dr. F. Gagiu, Onkologisches Inst i tut- Cluj, S.R. Rumdnien, Str. Republlci| nr.34-36

NEW MITODEPRESSIVE SUBSTANCES, COMPARATIVELY STUDIED WITH A SERIES OF ANTICANCEROUS DRUGS. IX. DERIVATIVES OF. AMINO ACIDS AND PEPTIDES. (Neue mitodepresslve Verbindungen~ vergleichend mit einer Re|he von antTkarzinomat~- sen Arzne|mitteln studiert ~ IX. Der|vate von Amlnosduren und Peptiden). F. Gaglu, C. Daicoviciu, A. Cacoveanu, C. NTstor.

Die Verfasser synthet|sTerten 10 neue Derivate von Aminosduren und Peptiden : AC-Asp (NH2)-OK, I ; BA-Asp(NH2)-OK, II ; AC-Glu (NH2)-OH.DCHA, I11 ; BA-Glu (NH2)- OH.DCHA, IV ; CP-Glu (NH2)-OH. DCHA, V ; AE-Ser-Ala-OPAC, VI ; AE-Ala-Phe- Ileu-Gly-OMe~ V I I ; Z-Pro-Leu-Glu(OBzl)-Phe-OMe, V I I I ; HCb H-Ala-Phe-Ileu- OPAc, IX ; AE-Ala-Glu(OBzl)-Glu(Obzl)-OPAc, X. Sie prLIften Thre mitodepresslve Wirkung an Hand eTnes vergleichenden Biotestes auF RadikUle von Lepidium sativum L. Ir l ib IV~V, IX zeigten einen Hemmungsfaktor von 53-91%.

Dr. F. Gagiu, OnkologTsches Inst[fut - Cluj, S.R.Rumunien, Str. Republicli 34-36.

R 41

Effect of Heparin and Protamine on Triglycerid-Lipase in Guinea- Pig Liver. (Einfluss yon Heparin und Protamin auf Triglycerid-Lipa- sen in der Meerschweinchenleber.) Ch. Glanzmann, I.Glanzmann Polyaniones and Polycationes liberate histaminase from guinea-pig liver. (Hahn F. et al. Biochem. Pharmacol.15,155-160). For further investigation of this effect, we investigated the influence of He- parin and Protamine on Triglycerid-lipase after in vivo administr~ tion in homogenized guinea-pig liver. Estimation of enzyme activity was performed in accordance with W.Guder et al. (Biochem. Bio- phys.Acta, 187,173-185,1969). At pH 4,5 Triglycerid-lipase showed an initial increase followed by decrease, whereas at pH 7,O and 8,5 only decrease could be registered. Protamine 5 mg/kg results in increase of enzyme activity at 4,5 pH, whereas at 7,0 pH and 8,5 the same decrease as after Heparin results. These results are compatible with release of Triglycerid-lipase from the guinea-pig liver by Heparin and Protamine. A causal relationship between this effect and histaminase liberation by Heparin and Protamin is ruled out by the investigation about the inhibitory effect of~-Butyro- lacton on the histaminase liberation by Heparin, but not by Prota- mine (Ch. Glanzmann et al. Arch. Int.Pharmacodyn.in Press). Dr. Ch.Glanzmann, Pharmakolog. Institut d.Universitit Freiburg, D-78-Freiburg, Katharinenstr.29, z. Zt. Univ. Klinik fdr Nuklearmedizin und Radiotherapie Kantonsspital ZGrich, CH-8OO6-Zdrich, Rimistr.lO0

MECHANISM OF HYPERGLYCEMIA DURING ACUTE CARBON MONOXIDE POISON~G (Zum Entstehungsmechanismus der Hyperglyk~mie wahrend der akuten CO.-Vergiftung) M.GGthert, P.HGck, G.Malorny~ H~

In rabbits breathing 2400 ppm CO for 1 h we measured COHb concentrations of 45-50%(mean values)in several experimental series: If rabbits are not pretreated with drugs, the glucose concentration in blood increases from Ii0 to 264 mg%,and the adrenalin~A) concentration increases from 0,8 ~g/l to 5,4 ~g/l.lf rabbits are pretreated with propranolol(8 mg/kg),reserpine(0,33 mg/kg each day in 3 successive days),tetraethylammonium(40 mg/kg),and aza- methonium(20 mg/kg)~yperglycemia is inhibited either completely (as in the case of propranolol and reserpine) or at least for more than 50%.After pretreatment with cyclazenin(5 mg/kg 24hbe- fore experiment)there is no inhibition of hyperglycemia.This drug only exerts a depleting action on peripheral sympathetic NA stores and does not inhibit the stimulating action of aeetyl- cholin on A secretion from adrenal medulla (result of perfusion experiments in isolated bovine adrenals)~ Normal and adrenal- ectomized rats breathed 2400 ppm CO for 30 min: In normal rats glucose concentration in blood increases from 109 to 283 mg~ (70~ COHb),whereas there is no hyperglycemia in adrenaleetomized rats(71~ COHb)o- During acute CO poisoning central nervous stimulation of adrenal A secretion is observed which induces hyperglycemia by increased glycogenolysis.

Priv.Doz.Dr.M.G6thert, Pharmakologisches Institut der Universi~t Hamburg, 2000 Hamburg 20, Martinistr. 52

R 42

STEREOSELELCTIVITY OF THE NEURONAL UPTAKE OF NORADRENALINE (NA) IN THE ISOLATED NICTITATING MEMBRANE (Stereospezifit~t der neuronalen Noradrenalin- Aufnahme in der isolierten Nickhaut) K.-H. Graefe and E. Eckert

Paires of muscles isolated from nictitating membranes of reserpine-pretreated cats (i mg/kg s.c. 24 h prior to experiment) were incubated for 7.5 min with 1.2 ml of I0 ng/ml of (Z)- H-NA (plus 18 ~g/ml U-0521 to block COM~ and ascorbic acid + EDTA). At the end of the incubatio~ the removal of the H-amine from the bath and the formation of ~he deaminated H-catechols were measured (with correction for the deaminated H-catechols remaining in the muscle).3Denerva- tion of the muscle and/or cocaine (iO ~g/ml) reduced the removal of H-NA by 40% while deamination was reduced by 75%. Deamination of the -H-amine was linear over a period of I0 min, removal was not 3 Apparently, under our experimental conditions the production of deaminated M-catechols is a reliable measure of neuronal uptake of H-NA. In order to test for stereoselectivity of neuronal ~ptake, the inhibitory effect of either isomer of NA on the deamina tion of (Z)- H-NA was studied. Muscles were ~ncubated four times for 7.5 min each (at i~tervals of 1 h) with iO ng/ml of H-NA in the absence and in the presence of i0 to I000 ng/ml of unlabelled (-)- or (+)-NA. (-)-NA was tested on the muscles of one side, (+)-NA on those of the other. The IC50's (concentrations producing 50% inhibition) of (-)- and (+)-NA did not differ significantly from each other. The pooled data gave an IC50 of 0.98 ~M, if 75% inhibition is regarded as maximum inhibition. If rates of deamination are calculated for total NA, the relation between bath concentration and rate of deamination was linear for denervated and showed saturation kinetics for innervated muscles. It is proposed that neuronal uptake is the rate limiting step. K.-H. Graefe, Institut fHr Pharmakologie und Toxikologie, 87 WUrzburg, Koellikerstr. 2

EFFECT OF DIPHTHERIA TOXIN ON CONTRACTILITY AND ULTRASTRUC - TURE HEART MUSCLE (Die Wirkung yon Diphtherietoxin auf Kontraktilit~it und Ultrastruktur des Herzmuskels). U.Greilich, D. Preiss and J.A. Ro~ner Papillary muscles from the right cardiac ventricle of 18 guinea pigs (9 controls),

which had been given diphtheria toxin (dlm) on three consecutive days, showed no differences in their maximal isometric contractility when stressed with 400 mg. Another group of 18 animals served as controls for the dlrn. Reduc- tion of the stress from 2500 mg to 400 mg failed to affect the contractility. The papillary muscles injured by toxin revealed a delay in the relaxation period of the first contraction after a five-minute pause. The papillary muscle in 2 x 3 groups with a stress of 60, 400 and 2500 mg were fixed and three electronmicroscopic preparations of each made. The lengths of the random'y selected sarcomeres showed no differences between treated and untreated animals (er- ror of probability ~,. = 0, 05). The contractility of the toxin-injured muscles corresponded to a frequency of stimulus between 0,25 and 6/sec. of thai of the controls. Changes noted were a uniform hyposarcolemmal and interfibrillary fatty dege- neration of the myocardial cells and a regular dilatation of the cisternae of the T-system. We did not find displaced or disrupted fibrils but did detect large and sr~all vacuoles containing osmiophilic cloudy and discrete electrondense material at the level of the Z-bands. We believe the first changes are membrane associated. Alysosomal reaction probably develops later. Dr. J. A. Ro~ner, Pathologisches Institut der Universit~t Heidelberg,

D-6900 Heidelberg, Berliner Strafe 5

R 43

THE EFFECT OF DIPHENYLTHIOHYDANTOIN ON SPINAL MOTOR ACTIVITY (Die Wirkung yon Diphenylthiohydantoin auf spinalmotorische Aktivitit)* W. Grossmann and I. Jurna

The amplitude and the rate of rise and fall of motoneurone action potentials are increased by diphenylthiohydantoin (DPTH) (GROSSMANN et al., Arch. Pharmak. Suppl. 270, R 47, 1971). In further experiments, it was found that EPSPs and IPSPs are enhanced by DPTH (40 mg/kg i.v.). The rheobase remained unchanged while the membrane resistance was increased. Not all motoneurones were influenced in this way. In reflex experiments it was found that alpha motoneurone discharge increased whereas gamma discharge remained unchanged. The effect on neuronal membranes was analyzed with the double sucrose gap method on rat spinal roots. DPTH did not influence depolarization produced by increasing extracellular potassium, but inhibited the effect of sodium or calcium deficiency on resting and action potentials. The results suggest that DPTH counteracts a mechanism of sodium-calcium exchange which might be similar to the one described by BAKER et al. [J. Physiol., Lond. 200, ~31, 1969) for the squid axon.

Dr. med. W. Grossmann, Institut fdr Pharmakologie und Toxikolo- gie der Universitit des Saarlandes, D-665 Homburg/Saar * Supported by the Sonderforschungsbereich 38 Membrane Research at the University of the Saarland

DER EINFLUSS DES PH AUF DEN CA++-ABH~NGIGEN TONUS DER CORONAR- GEFASS-MUSKULATUR. G. GrOn, M. Bayer u. A. Fleckenstein.

Der kontraktile Tonus der Coronargef~sse kann nut in Anwesenheit yon Ca++-Ionen aufreehterhalten werden (I). Kalium-depolarisierte Streifenpraparate yon Rinder- und Schweine-Coronarien in einer TyrodelSsung mit 43 mM K+/I ersehlaffen daher bei einfachem Ca ++- Entzug oder bei Zusatz yon EDTA infolge elektromechaniseher Ent- koppelung vollst~ndig. Minimale Konzentrationen yon Ca++-antago - nistischen Inhibitoren der elektromechanischen Koppelung (Nitro- glycerin und Bay a 1040 (jeweils 25 - 50 ~g/l); Verapamil, D GO0 und Prenylamin (0,5 - 2 mg/l); Papaverin (20 - 40 mg/l)) oder hohe

-- . " �9 " .

~ n ~ , ( ~ c ~ d ~ ) r ~ e ~ n d ~ e ~ W ~ d~k~~.Auch an isolierten Myofibrillen (2) verlaufen bei gleichem Ca++-Gehalt der L6sung ATP-Spaltung und Spannungsproduktion im alkalischen Be- reich um ein Vielfaches intensiver als bei Sauerung, die zu einer vSlligen Inaktivierung der Ca++-Effekte fGhren kann. 02-Mangel , Cyanid und DNP scheinen auf diesem Wege d.h. dutch intracellul~re S~uerung den Coronar-Tonus zu beseiZigen. Extra-Calcium oder Alka- lisierung wirkt daher nicht nut den relaxierenden Einfldssen der Ca++-antagonistischen Coronardilatatoren entgegen, sondern kann auch die durch 02-Mangel , Cyanid oder DNP bewirkte Erschlaffung wieder aufheben.

I) grQn, G. u. A. Fleckenstein: Naunyn-Schmiedebergs Areh.Pharma- kol. Suppl. to Vol. 270, R 48 (1971).

2) Schadler, M: PflGgers Arch. ges. Physiol. 296, 70-90 (1967). Ansohrift der Verfasser: Physiologisches Inst~-ut der Universitat, D-78 Freiburg i. Br., Hermann-Herder-Str. 7

R 44

ON THE EFFECT OF CHLORAL HYDRATE AND TRICHLOROETHANOL ON THE EEG OF THE ISOLATED PERFUSED RAT BRAIN (0ber die Wirkung yon Chloral- hydrat und Trichlor~thanol auf das EEG des isoliert perfundierten Rottenhirns) J. GrcJner, J. Krieglstein, H. Rieger

The effect of chloral hydrate on the EEG of the isolated perfused rat brain could be studied over prolonged periods of time, since only part of the substance is metabolized by the isolated organ. This enabled us to compare the effects of chloral hydrate and its main metabolite trichloroethanol. The concentrations of either chloral hydrate or trichloroethanol added to the perfusion fluid ranged between 1.5 to 5.5 raM. The EEG recordings taken 3o min after the beginning of the perfusion were compared for both substances at equimolar concentrations. Chloral hydrate and trichloroethanol changed the general pattern of the EEG in almost the same way, the higher dosages causing a continuous decline in activity leading finally to a picture with silent periods interrupted by spikes, as can be found Tn severe intoxications. An approach to quantification of the EEG was made by count- ing all waves over 50 microvolts. A linear relation between the number of such waves per sec and the concentration of chloral hydrate or trichloroethanol could be established. There was no statistical difference between the regression lines for both substances. From these results we concluded that both agents seem to be very similar in anesthetic potency.

Dr. J. GrcJner, Pharmakologisches Institut der Universitdt Mainz D-65oo Mainz, Obere Zahlbocher Str. 67

Smooth muscle contracting substances in the lung washings in rats and guinea-pigs. (Glattmuskul~r kontrahierende Stoffe in der SpUlflUsslgkeit des broncho-alveolaren Raumes bel Versuchs- tieren.) M. GrUnspan und S. Pallade~ In order to study the pharmacological properties and the partial composition of the pulmonary surfactant, rats anaesthetized with hexcbarbital sodium were exsanguinated and the lungs washed using a ca~nula inserted into the trachea.-3 ml of Tyrode solution at 37 C were introduced into the lungs and 2 ml were recovered. The samples from 12 animals were pooled and lyophilised. Af- ter extraction with chloroform-methanol 2:1 the phosphollpids were determined by thin layer chromatography. In the lyophilisate we found o,68/uMol phcsphatidylcholin per 2 ml lung washing.-The lyophilisate ~as redissolved in Tyrcde solution and tested subsequently on the guinea-pig~ileum. The solution produced a rapid, dose-related contraction. It corresponded to an activity of about o.o25-o.o5/ug histamine.2 HC1/ml lung washing which could not easily be ~ashed out. It exhibited a self-sensitizing action upon repeated administration of the same dose. It also induced sensitization of the ileum to histamine and bradykinin, but not to serotonln.-Pretreatment of the ileum with atropine did not abolish the contraction, mepyramine, however, had an inhibitory action. It appears therefore that there are at least 2 components: a rapid one, probably histamine and a slow-contracting component which is resistent to the antihistamines.-It is possible that the histamine component is accompanied by a still unknown substance of the pulmonary surfactant which may be a phospholipid.

M.GrGnspan, Institut f~r Lufthygiene und Silikoseforschung, Universit~t DGsseldorf, 4 D~sseldorf q, Postfach 56~

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INHIBITION OF THE CATECHOL-O-METHYLTRANSFERASE BY FLAVONOIDS (Hemmung der Cateohol-O-Methyltransferase durch Flavonoide) R. Gu$1er and N.J. Den$1er

Since Wilson and DeEds described a protection of epinephrine by flavonoids in 1949, various authors reported on the inhibition of the catechol-O-methyltransferase by flavonoids. The question

arises whether and to what extent different flavonoids inhibit the catechol-O-methyltransferase of human liver. Moreover, the in vivo effects of administration of flavonoids to the human on the oate- cholamine metabolism were investigated. Incubations were carried out with a partially purified enzyme preparation from human liver using norepinephrine as substrate and ]4C-S-adenosylmethionin as cofactor. Flavonoids were added in three different concentrations and 15o (concentration of flavonoid causing 50% inhibition) was determined graphically. All of the ]9 flavonoids of various types showed marked inhibitory effects, compounds with catechol structure of the side ring being most effective. The inhibition by quercetin and rutin was competitive, by isorhamnetin mixed type. The k~ value for norepinephrine was 3.3 x 10 -6 M, the inhibitor constantS'for quercetin was 5.32 x 10 -6 M and for turin 2].5 x I0 -6 M resp. In consequence to these studies, investigations in vivo were started infusing quercetin over a period of 4 h and injecting H3-norepinephrine after ! h. Preliminary results showed a marked decrease in the elimination of the O-methylated metabolites whereas norepinephrine excretion remained unchanged.

Dr. R. Gugler, Medizinische Poliklinlk, D-63 Giessen, Friedrich- strasse 27.

RELATION BETWEEN TOXICITY OF COMMERCIAL IRON DRUGS AND Fe(II)- CONTENT (Abh~ngigkeit der Toxizitfit handelsfiblicher Eisenpr~parate vorn Eisen-II-Gehalt) K. Gutschow, A. Schmid

Bei frfiheren Untersuchungen (K. Outschow und A. Sehrnid, Arch. Pharrnakol. Suppl. zu Vol. 270 1971 Ref 50) der LD handelsiiblicher Eisen-III-Pr~i-

' ' ' 50 parale wurden zurn Tell Vergiftungssyrnptome beobachtet, die der akuten Ei- sen-II-Vergiftung entsprechen. Deshalb wurden diese Prfiparate rnit 2, 2'-Bi- pyridin bei 522 nrn rnit folgendern Ergebnis auf ihren Eisen-II-Gehalt untersucht (Fe(II) in % des deklarierten Gesarnteisens; x + s-): Ferkornin 0, 20

- x + 0,009, Parkefer 0,20 + 0,005, Dorefer 0,24 + 0,008, Ferrophor 0,25 T 0,012, Eisendextrin CYANAMID 0,30 + 0,015, Centoferrurn B 0 34

, , + 12 ' 0,014, Ferrocid 0,54 + 0 065 Eisen'7-50" plus 0,55 _0,012, Centoferrum

~, 60 + 0, 023, Ferrlecit O, 78 + 0,022, Myofer 0, 88 _+ 0,015, Eisendextran- glyzeringlykosidkomplex 0, 95-$ 0,019, Eisenarnylosekornplex 5% I, 28 +0, 109 Ferrosanol 1,78 + 0,008, PecuferolBl2 1,98+0,015, Percuferol 2, 90-$0,015 Ferrovet 3, 77 +-0,015, Ferrodextran 100 4, 33 + 0,010, Pygrnia 5,08Z0,018 Jeetofer 5, 37 + 0, 113. Zwischen den Eisen-II-Gehalten und der akuten Toxizitfit der Pr~parate besteht eine positive nichl lineare Korrelation rnit einer Irrturnswahrscheinlich- keit yon weniger als i%. Daraus ergibt sich die Konsequenz, daf3 derartige imr~iparate keine reduzierenden Liganden oder Zus~tze (z. B. Vitamin C) ent- halten oder rnit reduzierenden LSsungsrnitteln (z. B. GlueoselSsung) verd~innt werden sollten.

Dr. K. Gutschow, Institut fiir Pharrnakologie, Toxikologie und Pharrnazie der Tier~irztlichen Fakult~t, 8000 Milnchen 22, Veterin~rstr. 13.

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EFFECTS OF 9~-FLUORHYDROCORTISONE, DOCA, AND CORTISONE ACETATE ON 24-HOUR AND 12-HOUR URINARY SODIUM, POTASSIUM, AND WATER EXCRETIONS IN RATS. (Einflu~ yon 9~-Fluorhydrocortison, DOCA, und Cortisonace: tat auf die 24-Stunden- und 12-Stunden-Urinausscheidungen von Na= trium, Kalium und Wasser.)

D. Haack~ E. Homsy~ B. M~hrin~ and J. M~hring.

In male Wistar rats, the effects of DOCA (2 x 5 mg/kg/day s.c.), 9~-FF (2 x 2 mg/kg/day p.o.), and cortisone acetate (2 x Io mglkg/ day i.m.) on the 24-hour and 12-hour urinary excretions of electrolytes and water have been studied comparatively. During the first 24 hours after the beginning of DOCA treatment, a positive Na and a negative K balance were found. 9~-FF induced negative K and water balances, and a moderate Na loss. Cortisone caused water and Na loss, without affecting K balance. Urinary Na concentration was decreased after DOCA, as well as after 9~-FF, whereas it was increased within the first 24 hours after cortisone. K concentration in the urine was increased after DOCA and 9x-FF administration, but did not change under cortisone treatment. During the 12-hour period at night, DOCA, as well as 9x-FF, caused Na and water retention and K loss, while cortisone induced a decrease in urine volume only. Dur- ing the day, DOCA also induced Na and water retention and K loss, but 9~-FF caused loss of K, Na, and water. Cortisone induced Na and water loss only. These findings indicate a DOCA-like effect of 9~- FF on urinary Na and K concentrations. During the night, 9~-FF seems to act mainly DOCA-like, however, during the day, mainly cortisone-like. Accordingly, changes in Na and K balances after the administration of 9~-FF reflect the sum of these effects. Dr. D. Haack, Department of Pharmacology, University of Heidelberg, 69 - Heidelberg, Hauptstra~e 47-51, Germany.

INFLUENCE OF AMANTADIN (A) ON MONO- AND POLYSYNAPTIC REFLEXES AND RENSHAW CELLS IN THE CAT (Die Wirkung des Amantadins auf mono- und polysynaptische Reflexe und Renshaw-Zellen der Katze) J. Haase und K.-H. Sonta@

In intercol!icular decerebrate or low spinal cats without anesthesia 1,4-28 mg/kg (A) i.v. regularly raises the threshold of antidromically excited Renshaw cells. An increase in the height of mono- and polysynaptic reflexes is frequently observed, but in other cases they are smaller. In decerebrate cats the antidromic (recurrent) inhibition of single tonic ~-motoneurones is diminished after 14 mg/kg (A) i.v. if the strength of the antidromic stimulus train is about the threshold for recurrent inhibition.

Conclusions: i. (A) can act at the level of the spinal cord. 2. To some extent it removes recurrent inhibition from extensor motoneurones. 3. The correlate in the monosynaptic pathways may be an increase of reflex height. 4. The net result could point to an improvement of motor activity. However, the results on reflexes are as yet inconclusive and require further investigation.

Prof. Dr. J. Haase, Dr. K.-H. Sontag, Max-Planck-Institut f~r experimentelle Medizin, Pharmakologische Abteilung, D-34OO G~ttingen, Hermann-Rein-Str. 3.

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IST DER CA++-KANAL DER MEMBRAN V0N WARMBL[~fERMYOKARD-FASERN F[~R CA§ STRENG SPEZIFISCH? H.-P. Haastert, M. Kohlhardt u. H. Krause

In Voltage-Clamp-Experime~en a~.isoliert~n WarmblGter-Trabekeln wurde der Einfluss yon Sr , Ba -~ und Mg ~ auf den langsamen Ein- w~rtss~om untersucht, der physiologischerweise in erster Linie vow+Ca - getragen wird. Dementsprechend nimmt dieser Strom in Ca -armer I~s~g in bekannter We~e ab. Bei Zusatz yon Sr ++ (5~ raM/l), Ba ~ (2 raM/l) oder Mg ~ (5.5 bzw. ll.O raM/l) in die Ca -arme L~sung kommt es jedoch nach unseren Befunden zu einer RGckkehr des langsam~ Einw~rtsstromes, da diese Katlonen offen- ba~+ebenfalls den Ca -Kanal benutzen kSnnen. Wird der langsame Ca T -Einw~rtsstrom dutch D @~0 (0.5 m~l) blockiert, so l~sst sich dieser Effekt dutch Sr oder Ba trotz weitere~+Anwesen- heit yon D 600 - in gleicher Weise wie dutch Extra-Ca - neutra- lisieren. Mg-Ionen sind dagegen nicht in der Lage, die D 600-in- duzlerte Hemmung des langsamen Einw~rtss~omes zu durchbrechen. Die Befunde ~igen, dass der langsame Ca -Kanal der Membran nicht fGr Ca -spezifisch ist, sondern auch yon anderen zweiwerti- gen Kati~en mitbenutzt werden kann. Inhibitoren des transmembra- n~ren Ca --Fluxes kSnnen offenbar auch die transmembran~ren Be- wegungen anderer zweiwertiger Kationen in unterschiedlichem Masse hemmen.

Anschrift der Verfasser: Physlologisches Institut der Universit~t D-78 Freiburg i. Br., Hermann-Herder-Str. 7

THE INHIBITION OF THE FORMATION OF A CENTRAL STIN~JLATING THEOBRO- MINE-N[ETABOLITE BY ALLOPURINOL IN WHITE MICE (Die Hemmun E der Bildung eines zentralerregenden Metaboliten aus Theobromin durch Allopurinol) BoHach und P. Mitznegg

Dr. B. Hach und Dr. P. Mitznegg, Pharmakologisches Institut der Universitit Erlangen-NGrnberg, 8520 Erlangen, Universititsstr.22

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CALCIDMANTAGONISM AND INHIBITION OF PHOSPHODIESTERASE BY VERAPAMIL (Calcium- antagonismus und Phosphodiesterasehemmung durch Verapamil) G. Haeusler, R. Eigenmann, W.P. Burkard. The relaxation of uterine smooth muscle produced by verapamil (VE) seems to be due to a specific Ca++-antagonistic effect of the drug (Fleckenstein et al., Klin.Wschr. 49,32,1971). In the current experiments VE and its stereoisomers competitively antagonized Ca++-induced contractions in depolarized renal arte- ries of the rat and in rabbit main pulmonary artery (RNPA); the (-)isomer was 5-6 times more potent than the (+)isomer with pal0 values of 7.4 and 6.6, respectively. VE (10-6M) had no influence on the degree of depolarization by K + as measured with intracellular microelectrodes, but prevented K+-induced contraction. In anaesthetized cats VE lowered BP and cardiac contractile force (CF) in a dose-dependent manner, the (-)isomer being more potent. Papaverine, whose muscle relaxation was claimed to be due to phosphodiesterase (PDE) inhibition, (Kukovetz and P~ch, N.-S. Arch.Pharmak. 267,189,1970), lowered BP and increased CF. High concentrations of VE (10-3M - 3xl0-3M) were necessary for a 50% inhibition of PDE in the homogenate of rat brain and heart and rabbit heart and main pulmonary artery. The inhibition of PDE was equal for both isomers. It is concluded: a) VE (in low concentrations) eompetetively antagonizes the contractile effect of Ca ++ in depolarized vascular smooth muscle without influencing K+-induced depolarization and dependent on its configuration, b) inhibition of PDE occurs only in high concentrations of VE and without stereospeci- ficity. These results exclude a causal relationship between vascular smooth muscle relaxation and inhibition of PDE by VE, A decrease in Ca ++ permeability by VE of the cell membrane may best explain the experimental findings.

Dr. med. G. Haeusler, Department of Experimental Medicine of F. Hoffmann-La Roche & Co. Ltd., OH 4000 Basel, Switzerland.

FINE STRUCTURAL FINDINGS IN THE GASTROINTESTINAL MUCOSA FOL- LOWING THE ADMINISTRATION OF PHENYLBUTAZONE AND ACETYLO-SALI- CYLIC ACID (Ultrastrukturelle Befunde an Magen- und Darmschleim- haut nach Gabe von Phenylbutazon und Acetylsalicyls~ure) K.-J. Hahn, E. Morgenstern, D. Krischkofski and E. Weber

Mitochondrial damage in human intestinal mucosa under therapy with tetracyclines (TC) has been regarded as morphologic correlate for irritative diarrheas occuring in this therapy (K.-J. Hahn et al., Arch.Pharmak., 266, 347,1970). On account of the frequency of gastrointestinal--adverse reactions, the effect of phenylbutazone (PBZ) and acetylosalicylic acid (ASA) on the ul- trastructure of the gastrointestinal mucosa was investigated in patients undergoing abdominal surgery and in guinea pigs. A marked increase of secondary lysosomes in the intestinal epithelial cells of both species was noted. ASA induced lysosomes derive from multivesicular bodies, retaining vesicular elements. Lysosomes induced by PBZ contain myelin figures and their origin is uncertain. Very similar lysosomes were also found in the parietal cells of the gastric mucosa in the treated patients. This finding was not reproducable in the animal. In both species both drugs cause desquamation of the mucous cells of the gastric surface with the known ultrastructural signs of cyto- lysis. The increase of lysosomes indicates cellular injury. Since no specific alterations of any of the organelles could be detected the drug induced damage is assumed to occur on a molecular level.

Dr.K.-J. Hahn, Medizinische Universit~tsklinik, D-69 Heidelberg, PrQf.Dr.E.Morgenstern, Universit~t D-665 Homburg (Saar)

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INVESTIGATIONS ON THE EFFECT OF PHALLOIDIN ON ISOLATED RAT LIVER MEMBRANES (Untersuchungen Gber die Wirkung v0n Phalloidin an isolierten Plasmamembranen der LeberzeIle)D.Hegner

In the perfused isolated rat liver phalloidin induces release of potassium (FRIMMER et al., Arch. Pharmak. 258, 197, 1967). To prove wether the membrane permeability might be caused by change of membrane structures the effect of phalloidin on isolated plasma mem- branesl was tested. Trealtment of spin-labeled plasma membranes (3- (2-Bromacetamido)methyl -2,2,5,5-tetramethyl-E-pyrrolidinyloxyl)

according the technique of McCONNEL (Proc.N.A.S.5__55,8,1966) with 30 ~g phalloidin/mg protein showed a decrease of the intensity of the ESR signal due to immobilised spin-label, whereas the intensity of mobile spin labels increased. Using infrared absorption spec- troscopy dried films of phalloidin trmated plasma membranes showed changes in ionized carboxyls absorbing at 1590 cm -1 and in the Ami- de II region. The isoelectric point of plasma membranes shifted from h. 9 • 0.26 to ~.67 • 0.31 after phalloidin treatment. Experi- ments involving the fluorescence of l-anilino-8-naphthalene-sulfo- nic acid reflects temperature dependent decrease of the hydrophobic structures of membranes in the presence of phalloidin. After treatment of membranes with phalloidin release of feline positive material with trypsin was much greater than in controls. The data suggest that phalloidin could promote conformational changes in membrane protein. This work was supported by the Deutsche Forschungsgemeinschaft.

Prof,Dr.D.Hegner, Institut fur Pharmakologie und Toxikologie im Fachbereich Veterin~rmedizin der Justus Liebig-Universit~t Giegen, 63 Gie~en,{S~hubertstraBe 1

CHANGE OF TRANSEPITHELIAL FLUID TRANSPORT OF THE GALL BLADDER BY DIURETICS (Beeinflussung des transepithelialen FlHssigkeitstrans- portes der Gallenblase durch Diuretika) K.Heintze, J. Weinert, 0. Heidenreich

The isolated in Ringer solution suspended gall bladder of guinea pigs was found to be a particular suitable model for the study of the action of diuretics for following reasons: it is in contrast to the amphibian skins an organ from a warm-blooded animal which posseses one layer of epithelial cells only and reabsorbs NaCI and water isotonically without creating a potential difference. Therefore it shares some properties with the proximal tubule. Amiloride, given to the bath solution at the serosal side fails to inhibit the fluid transport. Given to the mucosal side it inhibits dos~-dependently the reabsorption at concentrations higher than 3x10 -~ M. In lower concentrations however it stimulates fluid transport unexpectedly. Mersalyl given to the mucosal side evoked similar effects. 3xlO -5 M stimulated the transport. Given to the serosal side, the inhibition was similar to the effect on the mucosal side. A sti- mulatory action however was absent at all concentrations used. HgCI 2 showed no differences compared with Mersalyl except that it was effective at 10-50 times lower concentrations. The results show that the fluid transport of the gall bladder is stimulated by low concentrations of amiloride, mersalyl and HgCI 2 and inhibited dose-dependently by high concentrations.

Dr. K. Heintze, Pharmakologisches Institut der RWTH Aachen, D-5100 Aachen, Alter Maastrichter Weg I

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CENTRAL ALPHA-REGEPTORS AND THE BLOOD PRESSURE DEPRESSING EFFEOT OF ALPHA-METHYLDOPA (Zentrale ~-Receptoren und die blutdruckde- pressorische Wirkung von ~-Methyldopa) A. Heise, G. Kroneber~

Since we have shown in previous investigations, that the stimulation of ~-receptors in the brain can mediate a blood pressure decrease (A. Heise, G. Kroneberg, K. Schlo~mann, Naunyn-Schmiede- berg's Arch.Pharmak. 268, 348-360, 1971), and since a central component in the mode of the hypotensive action of ~-methyldopa has been proven (M. Henning, P.A. van Zwieten, J.Pharm.Pharmac. 20, 409-417, 1968), we have studied the possible relationship between the existence of central nervous ~-adrenergic receptors and the mode of action of ~-methyldopa. The posterior part of the third and the whole fourth brain ventricle of the cat were perfused with ~-methyldopa, ~-methyldopamine, ~-methylnoradrenaline, tyramine, noradreualine and dopamine. All substances induce a marked decrease in blood pressure. If the ventricles were in addition perfused with yohimbine or phentolamine, the blood pressure lowering effect of all substances except dopamine was significantly inhibited. This showed the specificity of the blocking effect of phentolamine and yohimbine. If cocaine was used instead of phentolamine or yohimbine the blood pressure depressing effect of tyramine was inhibited, whereas that of ~-methylnoradrenaline was potentiated. It is concluded, that ~-methyldopa exerts its central blood pressure lowering effect by mediation of central nervous ~-adrenergic receptors. A. Heise, G. Kroueberg, Farbenfabriken Bayer AG., Institut f~r Pharmakologle 56 Wuppertal I, Friedrich-Ebert-Str. 217, W.-Germany

INFLUENCE OF ~-ADRENERGIC BLOCKING AGENTS ON THE EXCITIBILITY OF FROG NERVE AND ~OCARDIUM (Der EinfluB yon ~-Sympatholytica auf die Erregbarkeit yon Nerv- und Herzmuskel-Pr~paraten am Frcsch) D.H, 11enbrecht~ G.Wiethold, H.Grobecker

In order to evaluate the correlations between nonspecifie pharmacological effects of O-adrenergic blocking agents and physicochemical properties the following parameters were studied: lowering of surface tension, octanol-water partition coefficient, local anesthesia (isolated frog nerve), prolongation of myocardial conduction velocity (frog heart muscle strips). The results obtained from dose response curves are summarized in the table:

surface activ, partition local an- myocardial -iO ~ynes/cm coefficient esthesia_ cond.vel~c. xlO-~M pH 7.0 ICSOxlO-3M ICSOxlO--M

Alprenolol 2.8 3.270 4.5 2.6 Propranolol 4.0 5.390 6.5 3.3 KL 255 2.2 3.750 20 5.7 K~ 592 8.O 0.530 45 9 Oxprenolol 8.5 0.430 35 12 INFEA 70 0.270 70 15 Pindolol 180 0.120 130 25 Sotalol N 600 O.Oll ~lO00 ~350 Practolol ~2000 0.009 N 3000 ~ 900 From the highly significant statistical correlations between all parameters studied it is concluded that nonspeeific pharmacological effects of ~-sympatholytics can be predicted quantitatively from their physicochemical properties. D.IIellenbrecht, Pharmakol. Inst.der Univ.,D-6000 Frankfurt/Main, Theodor-Stern-Kai 7

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ENTERAL ABSORPTION AND METABOLISM OF DL-I-(m-HYDROXYPHENYL)-2- ETHYLAMINOETHANOL IN THE HUMAN (Enterale Resorption und Metabolis- mus von dl-l-(m-Hydroxyphenyl)-2-~thylamino~thanol (Etilefrin) beim Menschen). J.H. Hengstmann, U. Weyand, H.J. Dengler.

The fate of tritiated Etilefrin in man following intravenous and oral dosage was investigated. After oral administration 83.13 3.23% of the radioactivity are excreted in the urine (Nffi8) compared with 78.21 + 1.23% following intravenous injection (N=4). But in contrast t~ 28.31 + 2.14% of the i.v. dose, of the oral dose only 6.]8 + 0.75% were identified as the free amine after column- and thin-Tayer chromatographic separation. The major part of the labelled material consisted of the conjugated amine (p.o. 67.57 + 3.64%, i.v. 40.]2 + 3.47% of the dose). Less than 2% only were d~aminated to the corresponding m-hydroxymandelic acid, less than 2% were excreted as HTO. Following oral application there was a rapid increase in the serum H3-activity with peak concentration between 30 and 60 minutes, followed by an exponential decline with a half-life time of ]67 minutes. The serum curve for the free amine had after both routes of administration nearly the same half-life time (p.o. 141 + 10, i.v. 126 + ]3 minutes). ComparatiVe experiments with similar hydroxyphenylalkylamines

support our hypothesis that a 3-hydroxy-group, independent of add~ tional ring hydroxylation, favours conjugation as the major metabolic step pathway after oral administration.

Dr. J.H. Hangstmann, Medizinische Poliklinik, D-63 Giessen, Friedrichstrasse 27

CHOLINESTERASE INACTIVATION BY DIFFERENT CONCENTRATIONS OF PARA- OXON AND TABUN IN THE COD (GADUS CALLARIAS) (Inaktivierung der Cholinesterase von Dorsch (Gadus callarias) in vivo durch im Meer- wasser gel~stes Paraoxon und Tabun) A. Herrlingerund F.K.Ohnesorqe

The purpose of the present investigation was to detect small concentrations of Tabun (T) in sea water by means of measuring the ChE-inactivation in cod tissues. As a reference compound paraoxon (P) was used. - Brain and skeletal muscle contain AChE (E.C.3.1. 1.7) exclusively. Red cells and plasma are free of any ChE-activity. - The inactivation of AChE (E.C.3.1.I.7) in brain and muscle of living cods exposed to P and T in sea water was found to be time and concentration dependend. The inactivation is characterized by kg-values of 1.15 x 104 for P and about 8 x IO3M -I x min-l'for T. In vitro as well as in vivo, the brain AChE of cods is much slower inactivated than that of mammals. In addition, the unusual high resistance of cods against an intoxication by P and T results from the fact, that they survive an inactivation of brain AChE up to 95%. In contrast, mammals die at inactivation levels of about 80%. - The determination limit for P and T is 1 x 10 -7 - 3 x 10-SM or 0.015-0.005 ppm. The method is, therewith, sufficiently sensitive for detecting concentrations endangering mammals.

Prof.Dr~med.F.K.Ohnesorge, Institut fttr Pharmakologie der Universit~t Kiel, D-23OO Kiel, Hospitalstr. 4-6

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MODIFICATION OF POSITIV INOTROPIC AND METABOLIC ACTIONS OF IN- CREASED [CA]e , OUABAIN AND NOREPINI~HR!NE BY PO 2 ((Jber den Einflu8 des O2-Partialdruckes auf die Kontraktionskraft und den Stoffwechsel isolierter Meer- schweinchenherzen be| Einwirkung erh~hter [Ca]e , Strophanthln und Noradrenalin. k. Hertle, F. Kersfing, R. Krebs

The action of ouabain (Ou), norepinephrine (NE) and increased extracellular Ca concentrations (ECa]e) at different pO 2 (670, 350, 100 mm Hg) on Tntraventrlcular pressure (IP) and O2-consumption (Qo2) was investigated on Tsolated gdnea pig hearts. The electrically driven (2,5 Hz, 4 msec, 40 %0 hearts were perfused with an constant volume (10 ml/mln) at 32 ~ C. Under control conditions contractile force was decreased by about 3o~ if pO 2 was lowered to 100 mm Hg. This was accompanied by an increase of QO2 from 50 to about 150 ~,l/g dry weight/min and an rise of lactate-output from 0,25 to 4,43 pM/g dry welght/mln. At all levels of pO 2 ouabain lead to an equivalent increase in IP. In ~:ontrast the action of NE and [Ca]e on IP were significant deminlshed parallel to induced changes in pO 2. In spite of increasing IP at 100 mm Hg pO 2 Ou, NE and [Ca]e did not further increase QO2, but Ou and NE lead to an elevation of lactate-output. Ou increases QO2 at 670 and 350 mm Hg pO 2 by identl- col amounts. The increase of QO2 by NE and [Ca'le at 350 mm Hg pO2 was reduced. The results show that the good correlation between heart performance and QO2 seen at high pO 2 is lacking under conditions of reduced pO 2.

k. Hertle, Pharmakologisches InsHtut der Universlt~t, D-6500 /V~inz, Obere Zahlbacher Strasse 67

PRODUCTION OF H202 DURING N-DEMETHYLATION OF ETHYLMORPHINE IN RAT LIVER MICR0SOMES (Die Entstehung yon H202 w~hrend der N-Deme- thylierung yon Athylmorphin in Rattenlebermikrosomen) A. G. Hildebrandt t I. Roots and W. Kohlen

Andreae(Nature 17~, 859, 1955) and Perschke et al. (Nature I~0, 257, 1961) have devised a method to determin@ small amounts of H202 in biological material by its reaction with the fluorescent dye scopoletin (7-hydroxy-6-methoxycoumarin) in the presence of horse radish peroxidase. This method has been modified to determine H202 in rat liver microsomes during oxidation of NADPH. Azide was employed to inhibit degradation of exogenously added or endogenously formed H202 by catalase. Azide neither influences the rate of NADPH oxidation nor the rate or extent of HCH0 production during N-demethylation of ethylmorphine. Thus, about 70 per cent of added H202 have been recovered from microsomes by this method. During oxidation of NADPH three to five nanomoles of H202 have been formed per mg of protein and minute. This number increases about twofold applying microsomes from phenobarbital treated rats. Although NADPH dependent N-demethylation of ethylmorphine had no influence on the rate of H202 production it decreased the extent of its formation. This suggests that H202 is rather a substrate of cytochrome P-450 dependent peroxidase activity (Hrycay et al., Arch. Bioch. Biophys. 147, 14, 1971) than an intermediate during N-demethylation of ethylmorphine. (Supported in part by Deutsche Forschungsgemeinschaft) Dr. A. G. Hildebrandt, Institut f~r Klinische Pharmakologie der Freien UniversitMt Berlin, D-1000 Berlin 45, Hindenburgdamm 30

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HYDROPHOBIC INTERACTION EFFECTIVE IN BINDING OF MORPHINE-LIKEANAIGESICS TO ALBUMIN, PLASMA PROTEINS AND TISSUE HOMOGENATES. (Hydrophobe Wechsel- wirkung bei der Bindung morphinartiger Analgetika an Albumin, Plasmapro- teine und Gewebshomcgenate) V. ESllt~and H~. Teschemacher.

The bindin~ of labelled morphine-like analgesies(14C-morphine, 3H~dihydro morphine, ~H-etorphine, 3H-fentanyl) to h t~msn albumin and plasma proteins and to tissue homogenates and plasma proteins of rabbits was determined using equilibrium d~alysis and ultrafiltration. In all media, hydrophobic interaction was found to be effective: at pH 7.4 and 37~ the lipophilic compounds fentanyl and etorphine showed B-values (fraction bound) between 60 and 90 %; the hydrophilic compounds morphine and dihydromorphine had B-values between 10 and 50 %. Lowering temperature from 37~ to3~ decreased B-values as expected for hydrophobic interactions. Raising the pH from 5.0 to 9.0 increased the partition coefficient heptane/water and binding to albumin and plasma. Curves similar to titration curves resulted but points of inflexiou differed from the pK measured in water suggesting a pK-shift of the compounds in protein solutions. Parameters for binding of the compounds to human serum albumin were obtained by a SCATCHARD plot. Different n-values and nearly equal k-v~lues rssulted. Almost constant binding was found for ~-vslues from 10-*to 10-'. A linear relationship for fentanyl was obtained between the quotient cg/cf and low concentrations of brain and liver homogenates. Extrapolation to undiluted tissue homogenates revealed 6-values of about 90 %. No evidence was found for saturable unspecific or specific binding in brain homo- gen~tes (g/v = 0.25) with morphine concentrations ranging from 10 -3 to 10-" mol.

Dr. V. HSllt, Msx-Planck-Institut fdr Psychiatrie, D-8000 Mfinchen 23 KraepelinstraBe 2, Germany.

DOPAMINE B-HYDROXYLASE: A MAJOR PROTEIN COMPONENT OF THE MEM- BRANES OF CHROMAFFIN GRANULES (Dopamin B-hydroxylase: eine Haupt- komponente der MembraneiweiBe chromaffiner Granula) Heide H6rtnag_!l , H.W~inkler und H.Lochs

In chromaffin granules the enzyme dopamine B-hydroxylase (DBH) is distributed equally between the soluble content and the membranes. Both types of DBH have now been isolated and characterized.

For solubilizing the membrane-bound DBH N-cetylpyridinum chloride (N-CPCI) was used. When membranes were treated with this detergent (2,5mg/mg protein) 8o% of the protein and 90 % of the DBH activity were solubilized. The solubilized proteins were fractionated by Sephadex chromatography in the presence of N-CPCI (O,O1% w/v). The DBH-activity was eluted together with the major component of the membrane proteins which has been named Chromo- membrin A (see Winkler, Phil.Trans.B 261, 293, 1971)

When N-CPCI was added to the soluble lysate of chromaffin granules (0,55 mg N-CPCI/mg protein), about 95 % of the proteins but only 25 % of the DBH-activity were precipitated; the only protein remaining in the supernatant was the protein which precedes Chromogranin A in electrophoresis. This protein has been identified previously as DBH.

The identity of the membrane-bound and soluble DBH could be demonstrated by polyacrylamide gel electrophoresis, by immuno- logical methods and by amino acid analyses.

Dr.Heide H6rtnagl, Pharmakologisches Institut der Universit~t Innsbruck, Peter MayrstraBe i, 602o Innsbruck, Austria

R 54

EFFECTS OF RENIN SUBSTRATE INFUSIONS ON VASCULAR RESISTANCE IN THE ISOLATED RAT KIDNEY (Wirkung yon Reninsubstrat Infusionen auf den Gef~sswiderstand der isolierten Rattenniere) K.G. Hofbauer, H. Zschie- drich, H. Orth, and F. Gross

Isolated rat kidneys were perfused in an "open system" with an electrolyte solution containing 3.5 g% Haemaccel. Renal "plasma" flow, glomerular filtration rate and urinary sodium excretion remained constant during the second hour of perfusion. Renin content of the perfused kidneys was equal to or lower than the amount present in the contra- lateral kidneys removed at the beginning of the experiment. Renin release was in the range of 1300 to 2300 ng A I �9 rain-1. Since the perfusion fluid was free of renin substrate, no angiotensin formation took place within the kidney.

When renin substrate in doses of 20 to 220 ng�9 m1-1 �9 rain -I was added to the perfusate for 1 or 2 min., renal vascular resistance (RVR) increased immediately and remained elevated during the infusion. Afterwards, RVR returned to control values within several minutes. Infusions of angiotensin I and II caused similar increases in RVR. From these experiments it is concluded that angiotensin I is released from renin substrate and converted to angiotensin II within the kidney in amounts sufficient to influence renal hemodynamics. This indicates that the two enzymatic steps of formation and conversion of angiotensin I occur within seconds.

Dr. Karl G. Hofbauer, Pharmakologisches Institut der Universit~t Heidelber~

6900 Heidelberg, Hauptstrasse 47-51.

STIMULATION OF PROTEIN KINASE PREPARATIONS FROM RAT CEREBELLUM BY GUANOSiNE-3':5'-MONOPHOSPHATE AND ADENOSINE-3':5'-MONOPHOSPHATE! (Stimulierung von Proteinkinase Preparationen aus dem Kleinhirn durch Guanosin-3':5'-monophosphat und Adenosin-3':5'-monophosphat) F. Hofmann~ G. Sold The influence of cyclic nucleotides on histone phosphorylation has been studied in the 50,000 x g supernatant of rat cerebellum in the presence of 50 mM sodium glyceroD~osphate/P ~ buffer, pH 7.5. When ~ -> P-ATP was replaced by ~-> P-GTP no ~hosph2~ylation took place~ 0nly in the presence of Mg ~T (i-20mM) or of Co ~T (2.5mM) did both cyclic nucleotides stimulate the phosphorylation of his- to~, whereas t~ reaction was not stimulated in the presence of Zn , Mn or Ca . When Guanosine-3':5'-monophosphate (cGMP;3xI0-TM was added,protein kinase activity increased about 85%.The apparent K for cGMP was 3.6x10-SM.Adenosine-3':5'-monophosphate (cAMP) had aasimilar effect but higher concentrations (10-SM) were necessary, the apparent K being 9x10-7M. This difference was not due to a phosphodiester~se, since enzymatic degradation of both nucleotides was minimal. In a phosphate free buffer,cGMP had only a slight sti- mulatory effect on the rate of phosphorylation,whereas cAMP stimulated the reaction by 100%.Addition of P~ resulted in a 230% increase of the cGMP stimulated reaction a~d a 60% decrease of the cAMP stimulated reaction. It is concluded that in rat cerebellum, the effects of cGMP are meadiated by a cGMP-dependent protein kinase which has been partially separated from a cAMP-dependent protein kinase. Dr. F. Hofmann, Pharmakologisches Institut, D-6900 Heidelberg, Hauptstr. 47 - 51

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EFFECT OF CLONIDINE ON SYSTEMIC AND CORONARY HEMODYNAMICS DURING EXERCISE IN DOGS WITH a) INTACT CONDUCTION SYSTEM b) AV-BLOCK (Wirkung yon Clonidin auf H~modynamik und Ceronardurchblutung bei arbeitenden Hunden mit und ohne atrio-ventrikular Block) J. Holtz, E. Bassen~e and W. yon Restorff

In dogs with chronically implanted pressure tubes (aorta) and flow transducers (aorta asc., left circumflex coronary artery) exercising on a treadmill at various speeds, 20 ~g clenidine i.v. resulted in a decrease in mean arterial pressure (MAP) of: a) 25 % in dogs with intact conduction system exercising at 9 km/hr, b) 16 % in dogs with AV-block and ventricular automatici- ty exercising at 3 km/hr. In a) and b) the fall in MAP was mainly caused by a fall in cardiac output (CO) and less by a fall in peripheral resistance. The fall in CO was brought about by a fall in heart rate,whilest decrease in stroke volume was minimal Coronary flow during exercise under clonidine remained unchanged compared to control exercise in a) and b) resulting in a relati- velmcrease in coronary flow per CO under clonidine. The extent of the clonidine induced decline in heart rate and cardiac output varied with the strength of exercise and was substantially modified by atropine (60 ~g/kg i.v.). This indicates an augmentation of the parasympathetic activity in addition to the central inhibition of sympathetic activity ( W. HSffke and W. Ko- binger, Drug Res. 1_~6, 1038, 1966) brought about by clonidine in the unanesthetized dog. Dr. J. Holtz, PD Dr. E. Bassenge and Dr. W. v. Restorff, Physio- logisches Insitut der Universit~t M thachen, D 8000 MGnchen 2, Pettenkoferstr. 12

EFFECTS OF 9~-FLUORHYDROCORTISONE ON SODIUM, POTASSIUM, AND WATER BALANCES IN THE RAT. (Einflu~ yon 9~-Fluorhydrocortison auf Natri= um-, Kalium- und Wasser-Bilanzen bei der Ratte.)

E. Homsy~ D. Haack~ B. MShrin~ and J. M6hring.

In male Wistar rats, the effects of four different dosages of 9g- FF (2 x o.oo5, 2 x o.o25, 2 x o.I, and 2 x o.5 mg p.o. per animal and per day) on electrolyte and water balances were studied. Dur- ing the first 24 hours after drug administration, a dose-dependent negative potassium and water balance was found. Sodium balance was affected only by the highest dose, which induced a slight but highly significant natriuresis. During a 5-day period o# 9m-FF administration (2 x o.5 mg), potassium balance was markedly negative, whereas sodium balance varied from slightly negative to slightly positive values. Food intake decreased on the third to fifth day after the beginning of 9m-FF treatment. On the first day of drug administration, water intake decreased, while urine volume increased. Thereafter, water intake further decreased with a concomitant decrease in urine volume. During the first day of 9~-FF treatment, the decrease in body weight was due mainly to water loss, whereas subsequently also due to decreased food intake. After cessation of 9~-FF treatment, sodium, potassium, and water balances returned to control values within four days. In contrast to man and dogs, in the rat 9m-FF has no typical socalled mineralocorti- cold effects on electrolyte and water balances.

Dr. D. Haack, Department of Pharmacology, University of Heidelberg, 69 - Heidelberg, Hauptstra~e 47-51, Germany.

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IRON ABSORPTION AND CELLULAR TRANSFER PROTEIN IN THE INTESTINA~ MUCOSA OF MICE WITH HEREDITARY ANEMIA ( E i s e n r e s o r p t i o n u n d z e l l u l i r e s T r a n s f e r p r o t e i n b e i M ~ u s e n m i t h e r e d i t i r e r A n ~ m i e ) H. H f i b e r s , E. Hf ibe r s~ W. F o r t h a n d W. Rummel

I n t h e p a r t i c l e f r e e s u p e r n a t a n t o f m u e o s a l c e l l s f r o m d u o d e n u m a n d j e j u n u m two p r o t e i n f r a c t i o n s ( I a n d I I ) o c c u r e w h i c h b i n d i r o n . I I i s i n v o l v e d i n t h e t r a n s f e r o f i r o n a c r o s s m u e o s a l cells during absorption (H~bers et al., Life Sci. I0, part I, ii~i, 1 9 7 1 ) . I r o n a b s o r p t i o n i s i n s u f f i c i e n t i n m i c e w i t h s e x l i n k e d a n e m i a ( s l a ) . The q u e s t i o n s h o u l d b e a n s w e r e d , w h e t h e r a c o n n e c t i o n e x i s t s b e t w e e n t h e i n s u f f i c i e n c y o f i r o n a b s o r p t i o n a n d t h e b e - h a v i o u r o f I I i n d u o d e n a l m u e o s a o f s l a i n c o m p a r i s o n t o n o r m a l m i c e . The 5 9 - F e c o n t e n t o f I I f r o m m u e o s a l c e l l s o f s l a m i c e a m o u n t s t o o n l y 25 % o f t h a t f r o m n o r m a l a n i m a l s . A f t e r h a v i n g f e d a low i r o n d i e t f o r 6 w e e k s , t h e 5 9 - F e c o n t e n t o f I I i n c r e a s e s 5 - f o l d i n n o r m a l m i c e a n d o n l y 2 - f o l d i n s l a m i c e . H o w e v e r , t h e t o t a l a m o u n t o f 5 9 - F e b o u n d by I I o f s l a m i e e f e d w i t h a low i r o n d i e t i s e v e n s m a l l e r t h a n t h a t d e t e r - m i n e d i n n o r m a l a n i m a l s f e d w i t h n o r m a l d i e t . The 5 9 - F e c o n t e n t o f m u c o s a l t i s s u e a s w e l l a s t h e 5 9 - F e a b s o r p t i o n ( d e t e r m i n e d i n a b o d y c o u n t e r ) c o r r e l a t e w i t h t h e 5 9 - F e c o n t e n t o f I I i n n o r m a l a n d i n s l a m i c e . T h e r e f o r e , i t was c o n c l u d e d t h a t t h e d e f e c t o f i r o n a b s o r p t i o n o f s l a m i c e may be due t o e i t h e r a d e f e c t o f s y n t h e s i s o r t o a d e f e c t o f i r o n b i n d i n g p r o p e r t i e s o f I I w h i c h i s i n v o l v e d i n t h e i r o n t r a n s f e r a c r o s s t h e m u e o s a l c e l l s d u r i n g a b s o r p t i o n . H. H U b e r s , I n s t . f . P h a r m a k o l . u n d T o x i k o l d e r U n i v . d . S a a r l . D-665 H o m b u r g / S a a r ( S u p p o r t e d by a g r a n t i f SFB 38 m e m b r a n e r e s . )

ON THE ACTION OF CHOLINOMIMETIC DRUGS UPON THE CHRONICALLY DENERVATED RAT DIAPHRAGM (Uber eine dualistische Wirkung von Cholinomimetika am denervierten Rattenzwerchfell) W. HHbner: H. LHllmann and A. Zie~ler

Cholinomimetic drugs evoke a transient contracture and decrease the contraction amplitude in chronically denervated, electrically stimulated isolated hemidiaphragms as long as the compounds are present. The extent of the two effects differ according to the drugs applied, as shown by comparison of ACh, carbachol, tertiary and quarternary arecaidine-esters and pyrrolidine-derivatives. The sizes and the time courses of the effects depend on the tim~ elapsed after denervation, the activity of the cholinesterase present in the muscle and on the mode of adding the drug (single or fractionated). Based upon the results a kinetic receptor model is proposed indicating that the contracture is proportional to the changes of the drug receptor complex (dAR/dt), whereas the decrease of the contraction amplitude is proportional to the concentration of the drug receptor complex (AR).

W. H~bner, Institut f~r Pharmakologie der Universit~t Kiel D-2300 Kiel, Hospitalstr. 4-6

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INHIBITION OF MICROSOMAL DRUG METABOLISM BY COUMARIN COMPOUNDS (Hemmung des mikrosomalen Arzneimittelstoffwechsels durch Cumarinderivate) B. HUthwohl, E. J~ihnchen, G.F. Kahl, H. S.tamm, W. Weber

Coumarin compounds which are substrates of the hepatic microsomal drug metabolizing system may inhibit the biotransformation of other drugs. Therefore, the influence of a number of 4-hydroxycoumarin anticoagulants on demethylation and aromatic hydroxy- latlon was studied in phenobarbital stimulated rat liver microsomes. Formaldehyde production from aminopyrine (1 raM) resp. p-aminophenol production from aniline (1 raM) were 7.92+o.65/o.27o+o.o13 nmoles/mg protein x mln in control microsomes and were decreased to 3.17+o.3o/o.128+o.oo6 by phenprocoumon, to 4.78+o.43/o.18o+ o.o17 by warfarin-3.33+o.32/o: 17o+o.o16 by acenocoumarol, to i . 7o+o , lo/o.o61 +o.ool by dlcoumarol, and to 2.32+o.16/o.115+o.oo4 by ethylbiscoumacetate (inhibitor concentrations o.5 raM/1 mM).-lnhibitor constants K / (IJM) for inhibition of formaldehyde production from ethylmorphine were 25 for phenprocoumon, 24o for warfarin, 73 for acenocoumarol, 9 for dicoumarol, and 58 for ethylbiscoumacetate. Binding of coumarin compounds to oxidized cytochrome P-45o was determined by recording difference spectra. Monocoumarins produced type II, dicoumarlns type I spectra. Spectral dissociation constants K S (MM) were 1.5+o.1 for phenprocoumon, 4-5.5+6.2 for warfarin, 3.4+o.3 for acenocoumarol, 4.9+o-~8 for dicoumaro, and 2.8+o.4 for ethylblscoumacetaTe. Binding of coumarins to b~vine serum albumin was measured by sephadex gel Filtration. A correlation was found between the binding constant K 1 and the logarithm of the octanol/water partition coefficient of the substances.

(Supported by the Bundesministerium fur Jugend, Familie und Gesundheit, Bonn) Pharmakologisches Institut der Universit~t, D 65 Mainz, Obere Zahlbacher Str. 67

INFLUENCE OF RIFAMPICIN IN VITRO ON THE PROTEIN SYNTHESIS IN MICROSOMAL, MITO- CHONDRIAL AND NUCLEAR FRACTIONS FROM RAT LIVER AND EMBRYOS (EinfluB yon Rlfam- picin auf die Proteinsynthese in Microsomen, Mitochondrien und Kernen in Ratten- leber und -embryonen) E. J~ger~ R. Bass~ D. Neubert

Toxic effects of rifampicin (Rif) cannot be explained by inhibition of a DNA- dependent RNA polymerase since this drug inhibits this enzyme in procaryotes only. In order to test protein synthesis as a possible target for the toxic action of Rif we have studied 14C-phenylalanine-incorporation into microsomal, mitochondrial and nuclear fractions from rat liver and -embryos in vitro. Rif was found to inhibit mitochondrial protein synthesis at a concentration about one order of magnitude lower than that necessary to affect protein synthesis in microsomes and nuclei. Cell fractions from embryonic tissues are found to be somewhat less sensitive to Rif.

_Liver~ ~ poly-U Microsomes Nuclei Mitochondria

Emetine 50 ~g/ml 0 - 5 % 0 - 5 % 80 - 95 % Chloramphenicol 30 ~g/ml 90 % 80 % 5 - 20 % ONase 2oo ~g/ml 9o % 5o % 8o - 9o % Puromycin 1oo ~g/ml 0 - 5 % 0 - 5 % 0 - 5 %

Rifampicin Io ~g/ml Ioo % Ioo % 7o - 8o % 2oo pg/ml 8o % 60 - 7o % 3o - 4o %

(Values as % of controls).

This work was supported by the Deutsche Forschungsgemeinschaft (SFB 29). Rif was a generous gift from Dr. Helm (Chemie-Gr~nenthal).

Dr. med. Elke J~ger, Pharmakologisches Institut der Freien Unlversit~t Berlin, Abt. "Embryonal-Pharmakologie", D-1ooo Berlin 33, Thielallee 69/73

R58

ON THE TOXICOLOGY OF 2-METHYL-4-NITR0-1-(4-NITROPHENYL)-IMIDAZOLE (Beitrag zur Toxikologie von 2-Methyl-4-nitro-l-(4-nitrophenyl)- imidazol) G. Jochmann, M.v. Eberstein, H.Frohberg, W.Kovac, A.Metallinos, P.Rogulja

EMD 15700, ein in 1-Stellung substituiertes Methylnitroimidazol, wurde fGr die Behandlung bestimmter protozo~rer Erkrankungen ent- wickelt. Bei allen geprGften Tierarten lag die LD~o naeh einmaliger oraler Applikation und einer Nachbeobachtungs~eit yon 14 Ta- gen zwischen 3-6 g/kg und nach i.p.-Injektion zwischen 1,1 - 1,5 g/kg. Weiterhin wurde die Substanz 3-4 Monate an Wistar-Ratten und Beagle-Hunde oral verabreicht. EMD 15700 wurde in einer t~g- lichen Dosis yon 25 mg/kg yon Hunden und Ratten toleriert. 4/6 Hunde der 50 mg/kg-Gruppe starben nach 9, alle Hunde der 100 mg/ kg-Gruppe nach 7 1/2 und alle Tiere der 200 mg/kg-Gruppe nach 2-3 Versuchswochen unter zentralnervSsen Symptomen. Pathohistolo- gisch wurden Ver~nderungen im GroS-, Stamm- und Kleinhirn nachgewiesen. Alle Ratten der 200 mg/kg-Gruppe Gberlebten, 26/30 Tiere der 400 mg/kg-Gruppe verendeten im Durchschnitt nach 7-wSchiger Behandlung. Proportional zur verabreichten Dosis und nach t~glicher Applikation yon 50 mg/kg und darGber wurden bet Ratten dege- nerative Hodenver~nderungen und ab 200 mg/kg auch Schwund des l~n- phatischen Gewebes festgestellt. EMD 15700 wirkte somit ffir Hunde kumulativ-toxischer als fGr Ratten. Dies ist dureh die am Vergldch zu Ratten verzSgerte Ausscheidung bei Hunden bedingt, wie die yon H.Wenzl u.a.(1971) mlt l~c_markierter Substanz durchgef~hrten pharmakokinetisehen Untersuchungen zeigten.

Dr.G.Jochmann, Institut fGr Toxikologie, E.Merck, 61 Darmstadt, Postfach 4119

BINDING OF ALDADIENE WITH THE CORTICOSTEROID ANTIBODIES (Bindung von Aldadien an Corticosteroid Antik6rper A. Joumaah und N. I. Salloum)

In rabbits and sheep, antibodies against aldosterone, desoxycorticosterone, and cortisol were produced by injection of bovine serum albumin complexes of 21-hemisuccinates of the corresponding corticoids. Aldadiene (3-oxo- 17~-hydroxy-4.6-androstadien-17~-yl propionic acid), a metabolite of the aldosterone antagonists Saldacton and Aldacton, showed substantial cross reactions with the various antibodies. The percentage of cross reaction was in the range of 0.1% - 0.85% for aldosterone, of 0.4% - 0.7% for cortisol, of i. 8% - 2.9% for corticosterone, and of 0.91% - 2.6% for desoxycorticosterone. No correlation between the antibody titre and the percentage of cross reactions was found. The possibility of such a cross reaction has to be considered when aldosterone, cortisol, corticosterone or desoxycorticosterone are determined by radioimmunological methods in patients who are under treatment with Aldactone.

A. Joumaah, Pharmakologisches Institut der Universit~t Heidelberg, 69 Heidelberg, Hauptstrasse 47-51.

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Interferenz der Wirkung yon Metrifudil, Adenoein, Dipyridmmol, Hexobendin und Lidoflmzin mit natOrlich vorkommenden Nucleins~urederivmten (INTER- ACTION OF METRIFUDIL, ADENOSINE, DIPYRIDAMOLE, HEXOBENDINE and LIOOFLA- ZINE WITH NATURALLY OCCURING NUCLEIC ACID DERIVATIVES) W. 3uhran~ K. Dietmenn

Am ooronaren Gef~ewideretand wacher Hunde wurden die Triphosphors~urs- ester dee Guanosin (GTP), Cytidin (CTP), Uridin (UTP) und Adenosin (ATP) geprQft. Unter physiologiechen Bedingungen bewirkten die Nuoleotide stets sine dosisabh~ngige Widerstandeabnahme. Bei intrecoronarer Infusion nshm die Wirkung in der Reihenfolge ATP >UTP >CTP >GTP ab. Intraven0ee Injek- tionen dieser Stoffe verminderten ebenfalls den intravaekul~ren Coronar- wideretend. Die durch Adenosin, Metrifudil, Hexobendin, Lidoflazin und Dipyridamol dilatierten Coronargef~Be reagierten auf intreven~se und in- tracoronare Gaben yon GTP, CTP und UTP mit einer Widerstandezunahme. Die konstriktorieche Wirkung war am et~rketen ausgepr~gt naoh GTP, dem Nucle- otid mit der schw~cheten Gef~Beigenwirkung. Oagegen bewirkte ATP nach Adenoein und Metrifudil eine additive, nach Hexobendin, Lidoflazin und Dipyridamol eine Obermdditive weitere Widermtandeebnahme. Die konetrikto- rische Wirkung yon GTP lie8 sich nach Carbochromen, Nitroglycerin oder hypoxischer Coronardilatation nicht nachweieen. Nach anderen 8efunden k6nnen die Nucleosidtriphoephate wsgen ihrer star- ken Polarisierung nicht die Zellgrenzfl~chen peesieren (RICHRAN and WYBORNY, Amer. 3. Physiol. 207, 113g, 1964). Die antagonistische Wirkung yon GTP, CTP und UTP l~et sich danach am einfachsten Ober sins Hemmung der d i r e k t an de r g l a t t e n Ge f~Bmueku l s tu r oder an den Ne rvenend igungen angreifenden Coronardilatatoren erkl~ren.

Drs. W. 3uhren, K. Dietmann, Pharmakologisohe Lsboretorisn Boeh~Ingsr Mennheim GmbH, D-6B00 Mannhsim 31, Sandhofer StreSe 112 - 132.

FORMATION OF GUANOSINE-3~5&MONOPHOSPHATE INHUMAN PIATELETS (Bildung yon Guanosin-3~Simonophosphat in Thrombozyten des Mensohen) R. Jun~, I. Mechler, S. Mocikat, E. Behme Guanylate Cyolase (GC), which catalyzes the formation of guanosine-3~Simono- phosphate (Guo-3~SIP), has been found in human platelets (0.5 - 2.5 nMol/min/ mg Prot.), leukocytes (80 pMol/min/mg Prot.) and erythrocytes (i pMol/min/ mg Prot.). The activity of GC was determined as described previously (Neumann et el. this journal). More than 95 % of GC activity is found in the I00,000 g supernatant. The activity was reduced after dialysis. The formation of Guo- 3~5iP in the presence of Mn ++ was higher than with Ca ++ or Mg ++. No activity was detectable when Ba ++, Co ++ , Cu ++, Hg ++ or Zn ++ (0. i - I0 mM) were added. In the presence of i mM Mn ++ the formation of Guo-3~SIP was reduced by addition of Mg ++, Ba ++, Co ++, Zn ++ (0.i -i0 raM), Ca ++ (0. i or 1 mM), and was higher with i0 mM Ca ++. The Mn++ dependent activity is completely inhibited by Cu ++ or Hg ++ (0.I - i0 mM). I', Br-, Cl-, P-, and Na +, K +, Li + had no effect. ATP, ITP, UTP, CTP, ADP, IDP, GDP, and AMP (0. i - i mM) diminished the formation of Guo-3~SLP; cyclic nucleotides have no effect. Formation of Guo-3~5~P is inhibited by pyro- phosphate (i mM), dithioerythrite (0.i mM) and reduced gluthatione (5 mM) by 97 %, 48 % and ~5 % respectively. Triton X-IO0 reduced the activity of GO. The formation of Guo-3~5LP was inhibited by i mM epinephrine and 0. i or 1 mM norepinephrine, but was not altered by histamine and acetylcholine (0.01 - i mM). In the presence of 1 mM Ca ++ , however, the activity was increased by acetylcholine. Mn ++ dependent activity was decreased by serotonine (0.01 - 1 mM). This effect was inhibited by the addition of 1 mM Ca ++ .

Dr. E. Behme, Pha~makologisches Institut, D-6900 Heidelberg, Hauptstr. 47-51.

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DEPRESSION BY MORPHINE OF POSTSYNAPTIC INHIBITION AND TEMPORAL SUMMATION IN SPINAL MOTONEURONES (Dimpfung postsynaptischer Hemmung und zeitlicher Summation an spinalen Motoneuronen durch Morphin)* I. Jurna and W. Grossmann

Experiments with intracellular recording from gastroenemius- soleus (GS) motoneurones were performed in intercollicularly decerebrated cats spinalized at the lower thoracic level. Inhibitory postsynaptic potentials (IPSPs) were produced by electrical stimulation of the sural nerve. Temporal summation of excitatory postsynaptie potentials (EPSPs) was elicited by repetitive stimulation of the GS-nerves with a strength sufficient to activate Ia-afferents (monosynaptic activation). Morphine (2 mg/kg intravenously) reduced the IPSPs only to a small extent. Temporal summation of EPSPs was depressed. The effects of morphine on IPSPs and on temporal summation of EPSPs were antagonized by levallorphan (0.2 mg/kg intravenously).

Institut fdr Pharmakologie und Toxikologie der Universitit des Saarlandes~ Homburg/Saar D-665

w Supported by the Sonderforschungsbereich 38 Membrane Research at the University of the Saarland and a grant of the Deutsche Forsehungsgemeinschaft

A QUANTITATIVE DETERMINATION IN NANOMOLE-RANGE OF ~I_ and ~i(@_ TETRAHYDROCANNABINOL IN BLOOD,URINE AND SALIVA (~p~ quantitative Bestimmung im Nanomol-Bereich fOr~ l- und~•176 - cannabinol in Blut,Harn und Speiehel)W.W. Just~G.Werner und M.

Wiechmann. AI - und~l(6)_Tetra - Das Bestimmungsverfahren zum Naehweis yon hydrocannabinol naeh Extraktion aus biologischem Material beruht auf der Reaktion der phenolisehen Hydroxylgruppe der beiden Isomeren mit Dimethylamino-naphthalin-sulfonsiurechlorid (DANS-Cl)unter Bildung des Sulfonsiureesters(N~ u.M.Wiech- mann,Progress in Thin-Layer-Chromatography and Related Methods, Vol.l~Ann Arbor-Humphry Science Publishers 1970~London).Die quantitative Bestimmung dieser fluoreszierenden Verbindung erfolgt dureh direkte Messung der Fluoreszenz auf der DUnnschieht- platte;die Erfassungsgrenze liegt bei O,OOl Nanomol. Komplettierend zu diesem Nachweis kann die massenspektroskopi- sche Identifizierung der DANS-THC-Verbindung treten, die neben dem MolekUlpeak eine charakteristische Fragmentierung zeigt. Die Nachweisgrenze liegt bei 0,I Nanomol. Der Cannabis-Abusus beim Menschen ist mit dieser Methode nachweisbar.

P r o f . D r . G . W e r n e r , M a x - P l a n c k - I n s t i t u t f ~ r H i r n f o r s c h u n g , A r b e i t s - g ruppe N e u r o c h e m i e , 6 0 0 0 - F r a n k f u r t / M a i n , D e u t s c h o r d e n s t r a s s e 46

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BINDING OF PYRIDINE DERIVATIVES TO REDUCED CYTOCHROME P-450 (Bindung vonPyridinderlvaten an reduziertes Cytochrom P-450) R. Kahl The experiments are concerned with the alteration of metyrapone (MP) binding to reduced cytochrome P-450 after pretreatment with either phenobarbital (PB)or 3-methylcholanthrene (3-MC). The spectral dissociation constant KS obtained from ~O.D. 446-490 in dithlonlte reduced rat liver mlcrosomes was 2.4 + 0.3 pM for MP in PB stimulated mlcrosomesr but 390 + 80 pM in 3-MC stimulated microsomes. No comparable differences were observed for the bindlng of 3-(3-pyrldyl)-propanol and pyridlne. Additional pretreatment with cyclohexi- mlde (CH) inhibited the induction of cytochrome P-450 by PB and 3-MCr but did not influence the different substrate dependency of MP binding after PB and 3-MC pretreatment. Isooctane extraction of P B stlmulated microsomes, which did not alter cytochrome P-450 content, considerably decreased MP binding to reduced cytochrome P-450.

nmales P-450/mg protein A O. D. 446_490/nmole P-450 (MP 0.04 raM) (MP 2.5 raM)

Controls 1.08+0.18 0.0058 + 0.0009 0.0061 § 0.0010 PB 2.10u 0.0172 + 0.0012 Q.0170 ~ 0.0016 PB + CH 1.41 u 0.18 0.0148 u 0.0020 - - - - - - - PBextr. 1.87u 0.0055 u 0.0007 0.0125 + 0.0010 3-MC 1.91 70.18 0.0026 u 0.0004 0.0091 ~ 0.0010 3-MC + CH 1.03 ~ 0.14 0.0020 u 0.0005 0.0075 u 0.0017 The results suggest that the alteration of MP binding to reduced cytochrome P-450 is rather due to changes in the phospholipid environment of the heine moiety of the cytochrome than to the synthesis of specific hemoproteins induced by PB and 3-MC. (Supported by a grant from the Deutsche Forschungsgemeinschaft) Dr. Regine Kahl, Pharmakologisches Institut der Unlversitat, D 6500 Mainz, Germany

THE INFLUENCE OF NEPHRECTOMY DETERMINED AT THE TOXIC AND LETAL KOMBINED EFFECTS OF CARDIOTONIC STEROIDS (0her den Ein- fluB der Nephrektomie auf das toxische und letale Kombinationsverhalten yon Herzglykosiden) G. Kalversiep, J. Hamacher Fiir das polare g-Strophanthin und ConvaUatoxin sowie fiir das apolare Digoxin und Digitoxin und deren bin~ire Kombinationen wurden in drei Mischungsver- h~iltnissen die Dosis bis zum Beginn der Herzsch~idigung sowie die Infusions- toxizit~it bei kurzer, mittlerer und langer Uberlebenszeit nach KNAFFL- LENZ bestimmt, um den EinfluI~ der r e n a I e n A u s s c h e i d u n g als eine der mSglichen Ursachen - neben Serum-Gewebe-Verteilung, PlasmaeiweiB- bindung, Metabolisierung in der Leber und spezifischer Wirkungsweise - fiir dasn i c ht a d d i t i v e Kombinationsverhalten herzwirksamer Glykoside zu ermitteln. Optimaltiter ergaben sich dabei fiir g-Strophanthin, Convallatoxin, Digoxin und Digitoxin nach I a n g e r Uberlebenszeit. Der Kurzzeittiter blieb fiir g-Strophanthin, Digoxin und Digitoxin unbeeinflu~t. Die Isobolographie nach LOEWE l~t bei Kurzzeit-Untersuchungen eine Wandlung yon ursprfing- lich additivem bzw. unteradditivem zu [iberadditivem, bei Langzeit-Untersu- chungen dagegen yon iiberadditivem zu unteradditivem Kornbinationsverhalten erkennen. Mittelzeit-Untersuchungen zeigten mit a b n e h rn e n d e r Polarit~it der Kombinationspartner bei urspriinglich additivem bzw. [iberadditivem ein unter- bis iiberadditives bzw. ein geringer iiberadditives bis unteradditives Kombinationsverhalten.

Prof. Dr. rned. J. Hamacher, Pharmakologisches Institut der Universit~it zu KSln, 5 KSln 41, Gleueler Str. 24

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METABOLISM OF MESTRANOLDUE TO THE MICROSOMALMIXEDFUNCTION OXIDASE IN VITRO (Die Umwandlung yon Mestranol in vitro mit Hilfe der mlschfunktionellen mikrosomalen Hydroxylase) H. KapDusa H. M. Bolt t and H. Remmer

This report deals with the O-demethylation of mestranol (ethynylestradiol-3- methyl ether), which results in the formation of the hormonally active ethynylestradiol. Incubations of radiolabelled mestranol were performed with rat liver microsomes and a NADPH-regenerating system. The kinetics of the reaction were examined and resulted in a KM-value of 3.1 x 10-5 M. The enzymic reaction was completed after 20 min. The products have been separated by means of column chromatography on alumina. After incubation of doubly labelled mestranol (4-I#C and methoxy 3}{) the main metabolite has been identified as ethynylestradiol by crystallization to constant specific activity. In addition, a minor metabollte was found, which possessed the unaltered methoxy group. Pretreatment with phenobarbital caused a threefold increase of ethynylestradiol production. The demethylatlon of mestranol could also be demonstrated in the isolated perfused rat liver. Further details will be given elsewhere (Acta endocrin., 1972, in press). Our results might be of considerable interest regarding the usual sequential medication of the contraceptives. If mestranol is used as estrogen component, the effective dose of estrogen depends on the demethylation rate. The individual differences of microsomal oxidations are well known (H. Ren~mer, Amer. J. Med. 49, 617, 1970).

Dipl.-Biochem. H. Kappus, Institut fur Toxikologie der Unlversit~t, D-7400 T~bingen, Wilhelmstra~e 56

COMPARATIVE STUDIES ON PINEALECTOMY-INDUCED, RENAL AND SPONTANEOUS HYPER- TENSION IN RATS (Vergleichende Untersuchungen Uber die Epiphysektomie- induz ier te t renale und spontane Hypertonie bet Ratten) H. Karppanen I H. Vapoatalo t S. Lahovaara and P. Mdnnist~ Pinealectomy (PE) produces moderate hypertension in rats (A. Zanoboni et at. t L i fe Sci. 6t2327-2331 , 1967). To induce th is type of hypertensidn we developed a new"stereotaxic method fo r PE. The skul l is perforated in two points (1 mm apart) above the pineal body in the midl ine. The electrocoa- gulat ion needle is brought through these holes to a depth of 1.7-2.3 mm depending on the weight of the rats (160-260 g). This method with a low mor ta l i t y rate is rapid and successful in 95 % of the cases. Renal hypertension (RH) was produced by removing the right kidney;and tying a f igure eight l i ga tu re around the l e f t kidney. 81ood pressure (BP) of conscious rats was measured with a t a i l cuf f method. By the f i f t h week of obser- vat ion the 8P values were: contro ls ; 125• PE rats; 140• RH rats; 160• and SH rats: 182• mm Hg. The plasma renin a c t i v i t y (PRA) of the controt rats was 28• and that of PE rats 39• ng angiotensin/ml x 3 h. PRA of the RH and SH rats did not d i f f e r from the contro l leve l . The plasma potassium was sl igh$1y lowered in RH and SH rats (4.6~0.1 and 4.5• mEq/1; contro l 5.1-0.1) . In a p i l o t in v i t r o experiment adrenal quarters from contro l , PE, RH and SH rats were incubated with ACTH, d i - butyry l cyclic-AMP and 5HT. No c lear -cu t d i f ferences in the aldosterone production were observed concerning various types of hypertension. PE-hypertension is less marked than RH or SH. Increased PRA plays a part in PE- but not in other types of hypertension that have been tested. Grants: The Finnish Heart Associat ion, The Y.Jahnsson Foundation. We are gra te fu l to Dr. H.Erbler, Hannover for the aldosterone measurements.

Dr. H. Vapaatalo, Dept. Pharmacol., Univers i ty of Oulu, SF-90 100 Oulu 10.

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SYMPATHICOLYTIC ACTIVITY AND CARDIODEPRESSIVE EFFECTS OF D- AND L- FORMS OF 6-RECEPTOR BLOCKING AGENTS ON PERFORMANCE, 02-CONSUMPTION AND SUBSTRATE OXIDATION IN ANTAGONISM TO g-STROPHANTHIN. E" Kastner , H.J. Mensing and M. Siess (TUbingen/Marburg)

02-consumption, ox ida t i on ra te of 14C-glucose or 1"C-hexanoate and performance of spontaneously beat ing a t r i a of guinea pigs were measured in Ty rode -so lu t i on at 30~ (M.Siess et a l . , J . M o l e c . Ce l l . C a r d i o l . l , 2 6 1 (1970). Concent ra t ion- response-curves obtained a f t e r enhancement of the performance wi th 2xlO -6 g/ml o r c i p r e n a l i n e show d i f f e r e n t s y m p a t h i c o l y t i c a c t i v i t y of 6 - recep to r b lock ing agents (6-RBA). The concent ra t ions of ~-RBA which re tu rn the increased frequency to the i n i t i a l value show a wide range from the high ac- t i v e L -oxpreno lo l (SxlO -8 g/ml) > DL-p indolo l > L - a l p r e n o l o l > L- p roprano lo l to D-oxprenolo l > D-propranolo l > DL-p rac to lo l > D- a l p r e n o l o l (8xlO -6 g/ml) wi th low a c t i v i t y (1 :160) . L-forms are to 10-32 f o l d more e f f e c t i v e than the D-forms. In unt reated a t r i a a l l ~-RBA show between 5x lO-~-5x lO -~ g/ml d i r e c t negat ive i n o t r o p i c e f f e c t s . Independend of the catecholamine content of the a t r i a the t o x i c increase of f requency and ar rhy thmia caused by g-s t rophan- th in (g-S) can be antagonized only by high ca rd iodepress ive con- c e n t r a t i o n s of D- or L-forms of 6-RBA. D-forms are t he re fo re of i n t e r e s t of d i g i t a l i s i n t o x i c a t i o n because of t h e i r low sympathi- c o l y t i c a c t i v i t y . Using high concent ra t ions of 6-RBA the t o l e r a t e d concen t ra t ion of g-S can be increased 3 x and the p o s i t i v e ino- t r o p i c maximum enhanced 2 x. g-S reverses the decreased e f f i c i e n c y caused by 8-RBA. The s e n s i t i v i t y to 6-RBA is more enhanced in a t r i a incubated wi th hexanoate than wi th glucose.

Prof . Dr. M.Siess, Pharmakol. I n s t i t u t , 74 TUbingen, Wi lhe lms t r . 56

NEUROGENIC AND MYOGENI0 REACTIVITY OF THE CORONARY ARTERY (Unter- suchun~en Gber die neurogene und myogene Reaktivit~t der Coronar- gef~Be) S. Kazda and G. Kroneberg

The influence of intraarterially injected sympathomimetic and myogenic vasoactive substances on the peripheral resistance of the coronary and femoral artery was investigated in anaesthetized open-chest dog. The ~-mimetic isoproterenol as well as the myogenic vasodilator BAY a 1040 produced a dose dependent decrease of the peripheral resistance of both arterial beds. There was a marked difference in the vasoconstrictor response: whereas angiotensin increased the peripheral resistance of both arterial beds, the ~-sympathomimetic BAY 1470 increased only that of the femoral artery. The coronary arterial resistance was not essentially influenced by BAY 1470 even after pretreatment with propranolol. Only in those experlments, in which the initial blood flow values were extremely high, BAY 1470 had a slight coronary constrictor effect. In contrast to the intravenous injection intraarterially injected Phentolamin does not alter the coronary resistance. The results give evidence for the suggestion that there are far less s-receptors in the coronary than in the femoral vessels and that the constrictor tone of the coronary vessels is mainly due to myogenic mechanisms.

Dr. S. Kazda, Prof. Dr. G. Kroneberg Farbenfabriken Bayer AG, Institut fGr Pharmakologie, 56 Wuppertal S, Postfach 130105

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THE EFFECT OF DECREASED EXTRACELLULAR NA-K-CONCENTRATIONS ON MYOCARDIAL OXYGEN CONSUMPTION AND MECHANICAL ACTIVITY OF ISOLATED GUINEA PIG HEARTS (Zur Wirkung vermlnderter extrazellulBrer Na-K- Konzentrationen auf den Sauerstoffverbrauch und die Kontraktilit~it isolierter Meer- schwelnchenherzen) F. Kerstlng, R. Krebs

Reduction of extracellular potassium ~<+]e and sodium [_Na+]e concentration leacl to an increase of contractility of cardiac muscle. There is probably involved a change in the activity of a special membrane-bound carrier system influencing cellular Ca- metabolism (Reuter, H., N. Seitz: J.Physlol. 195, 451-47Ol 1968)_. If the changes in calcium-metabolism induced by decreasing co--~centrations of J-K~e and rNa+]e are dependent on aerob metabolism, changes of contractility should correlate with oxygen-consumption. The changes in heart performance and oxygen-consumption were studied on isolated isovolumetric (lo ml/min) perfused guinea pig hearts beating with constant frequency (2.5 Hz). After reduction of [Na+]e from 145 mM to 26.9 mM and of [K+] e from 5.4 mM to 1.o mM heart performance increased. The increasing heart performance was accompanied under ~+]e-reduction byFan enhanced QO2, whereas under comparable increases of heart performance [1~ ~e-reduction induqes Qo2-reduction, i .e. the efficiacy (relation of heart performance/Qo2 ) has been increased. Simultaneously reducted [Na+'l e and [K +] e concentrations with constant Na/K-relatlon increases contractility without changin~ efficiacy. The determination of the positiv inotropic effect by the relation Ca/Na z was not found under condition of decreased J-K +]e-concentration.

Dr. F. Kersting, Pharmakologisches Institut der Unlversit~t Mainz D-65oo Mainz, Obere Zahlbacher Str. 67

GAS CHROMATOGRAPHIC ESTIMATION OF ACETYLCHOLIN5 IN RABBIT HEART (Gaschromatographische Bestimmung von Acetylcholin im Kaninchenherzen) H. Kilbinger

Rabbit hearts were homogenized with an "Ultraturrax" in 5o ml o.4 n perchloric acid. After 45 mln the extract was centrifuged. An internal standard of 5oo ng propionylchollne (PCh) was added to 5 ml of the supernatant. The choline esters were precipitated as reineckates, treated with Biorex 9 and demethylated according to the method of I. Hanin and D.J. Jenden (Biochem.Pharmac. 1_88, 837-845, 1969). The tertiary analogs of acetylcholine (ACh) and PCh were extracted with diethyl ether and the ether extract reduced to a volume of 3-4 pl by micro distillation (K. Beyermann et a~., Z.AnaI.Chem. 25___11, 289-293, 197o). Of the final extract 1 Isl was injected into a HP 762o gas chromatograph equipped with a nitrogen detector (N-FID). Gas chromatography was carried out at 11o~ on Gas Chrom Q 1OO/o12O mesh coated with 7 ~ Carbowax 2o M; the temperature of the N-FID was 42o C. The ACh concentration of the whole rabbit heart was o.33 + o.ol pg/g (mean weight of base +_ S.E.; n=13). Concentrations (pg/'g) in different parts of the heart were as follows: Both atria, o.98 + o.o7; n=4; right atrluml 1.2o + o.12, n=4; left atrium, o.86 + o.11, n=4; vent-"rlcles, o.16 + o.ol , n---4. Endog-enous PCh was not detected. The ~,Ch concentrations of the rlght-and left rabbit atria estimated by bioassay (J. VIk, Naunyn-Schmiedebergs Arch.exp.Path.Pharmak. 23___55, 19-22, 1958) are in agreement with the above data.

Dr. H. Kilbinger, Pharmakologisches Institut der Universitdt Mainz D-65oo Mainz, Obere Zahlbacher Str. 67

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SIGNIFICANCE OF THE CENTRAL BIOGENIC AMINES FOR THE THRESHOLD IN ELECTRO AND PENTETRAZOLE SEIZURES (Bedeutung der zentralen biogenen Amine fur die Krampfschwelle im Elektro- und PentetrazoI-Kramp0 Marion KILIAN and H.- H. FREY

Seizure threshold for maximal electroconvulsions was 113 V in mice and 325 V in rats. In both species the threshold was significantly elevated by pretreatment with 5-HTP (100 mg/kg i .p . , 30 rain) or L-dopa (200 mg/kg ~.p., 30 rain) and lowered after p-CI- phenylalanlne (300 mg/kg p.oo on 3 days before the test in mice, or once 48 h before the test in rats), a-methyltyrosine (100 mg/kg i .p. , 2 h), dlsulfiram (400 mg/kg p.o., 6 h) or FI.A-63 (40 mg/kg i .p. , 2 h). The threshold was also lowered by cyproheptadine (0.2 mg/kg i .p. , 30 mln) and phentolamlne (1 m~/kg i .p. , 20 rain) but not by haloperidol. Pentetrazole threshold for clonic convulsions (49 m~"kg i .v. in m~cer 38 mg/kg in rats) was altered in the same direction as that for electroconvulslons by 5-HTP~ a-methyltyroslne I disulfiram and FLA-631 but p-CI-phenylalanlne, cyproheptadine and a-adrenolytics were without effect. Contrary~ the threshold for the tonic component of the chemoseizure (98 m~/'kg i .v. in mice) was predominantly altered by manipulations on the tryptaminergic side. The results point to a role of 5-HT and noradrenallne metabolism in both seizure models~ whereas dopamine seems to be of minor importance. Results with amphetamine and p-CI- amphetamine, the former stimulating predominantly adrenergic, the latter tryptaminerglc sitest flt into this picture~ since both provoked elevations of the seizure thresholds in both tests. Marion Killan, Inst. f. Vet.- Pharmakol. u. Toxikol~., Freie Univ., D-1000 Berlin 33~ Koserstr. 20

THE CORRELATION BETWEEN SOME PARAMETERS OF THE MECHANICAL FUNCTION AND THE HEAT PRODUCTION OF ISOLATED HEARTS UNDER SOME PHARMACOLOGICAL AND PHYSIOLOGICAL INFLUENCES. (Uber die Korrelation einige~ Parameter der Herzfunktion mit der myokardialen W~rmeproduktion unter verschiedenen physiologischen und pharmakologischen Einfl~ssen.) W. Klaus, K. G~ttler, M. Theisohn und I. Theisohn-Schwedhelm Durch Messung der myo~ardialen W~rmeproduktio~ unter dem Einflu~ yon Adrenalin (1o-9-3xlo-SM), Strophanthin (5xlo-8-2.5xlo-7M) und bei Va- riation des intraventrikul~ren Volumens (o.5 - 1.4 ml) am isolierten Kaninchenherzen (Tyrodel~sung, 37 ~ C~ 15o Reize/min sollte gepr~ft wet- den, wie sich die Relation des Energieverlustes durch W~rmeabgabe zu verschiedenen cardialen Funktionsparametern (LVP, EDP, ~P, dP/dt, T ~P~ TPP) unter diesen Bedingungen verh~lt. Die beste Korrelation fand sich f~r den Vergleich mit den w~hrend der Kontraktion auftretenden Druck- ~nderungen. Die funktionsunabh~ngige W~rmeproduktion betrug bei Volumen- variation 168 • 19~ unter Strophanthin 11o i 17 und unter Adrenalin 1o5 • 13 mcal/g FG/min, die funktionsabh~ngige W~rmeproduktion unter diesen Bedingungen 1.65 • o.53, 3.87 • bzw. 4.25 • o.32 meal/g FG/ min bezogen auf I mm Hg Druckentwicklung. Die Unterschiede zu dem durch Volumenvariation bedlngten Verhalten k~nnen wegen der Komplexlzit~t dieser Gr~e nicht eindeutig interpretiert werden, der fehlende Unter- schied zwischen der Adrenalin- und Strophanthlnwirkung steht in Uber- einstimmung mit dem Verhalten des 02-Verbrauches unter entsprechenden Bedingungen.

Inst. f. Pharmakologie der MHH~ D-3ooa Hannover, Roderbruchstr. loi

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DIFFERENT MODES OF ACTION OF METRAZOL AND STRYCHNINE AND SOME AN- TAGONISTS ON APLYSIA NEURONS (Unterschiedliche Wirkungsmechanismen yon Metrazol und Strychnin und einigen Antagonisten an Aplysia Nervenzellen) M.R. Klee and D.S. Faber

Metrazol (5-70 mM) and strychnine (0.1-I mM) both change the fir- ing pattern os identified neurons in the abdominal ganglion os Aplysia Californica in that they induce doublets or multiple discharges characterized by abnormally short inter-spike intervals. High calcium or low sodium concentrations in the bathing medium counteracts these strychnine effects. Treating these doublets as a model of "convulsive activity" we have used voltage clamp analysis to determine their mode os genesis. We have found that strychnine changes the I-V-characteristic os these neurons in a specific way by shifting the threshold for the inward current to lower and that of the outward current to higher voltages. At the same time the inward and outward currents are equally reduced and their time courses prolonged. Metrazol only reduces the inward current but it enhances potassium inactivation and causes pronounced anomalous rectification, resulting in oscillations of the membrane potential os even normally silent neurons. Both drugs reduce the early fast Rotassium current which exists in bursting pacemaker cells of Aply- sia similarely to the reduction o9 the delayed rectification potassium current. Mephenesin (5-25 mM), an antidote to strychnine, blocks both components of the inward current (Na + & Ca++), while trimethadione, generally considered to be a metrazol antagonist, does not seem to have that property in this preparation.

Dr. M.R. Klee, Max-Planck-Institut fur Hirnforschung, Neurobiolo- gische Abteilung, D-6000 Frankfurt-Niederrad, Deutschordenstr. 46

STUDIES ON THE EFFECT OF CYCLOPHOSPHAMIDE, RIFAMPICIN AND SPIRONOLACTONE ON LI - VER-REGENERATION (Untersuchungen Uber den EinfluB yon C., R. und S. auf die Le- ber-Regeneration). H.P. Kley t D. Neubert In order to elucidate toxicological ef fec ts of the 3 drugs on p r o l i f e r a t i n g t i s - sues, DNA-, RNA- and N-content per l i ve r as well as the a c t i v i t y of RNA-polyme- rases In isolated nuclei were studied in rats during a 4 days period fo l lowing 2/3 hepatectomy. No c lear -cu t e f fec t on these parameters was found with spironolactone ( loo mg/kg d a i l y ) . Both r i fampic in (R.) and endoxan (E.) effected some parameters connected with the regeneration process. Surpr is ing ly , both drugs did not e f fec t regeneration wi th in 48 h a f t e r hepatectomy, but only in ter fered revers ib ly wi th the second phase of the regeneration process. While the e f fec t of R. was confined to RNA- and prote in-synthes is , E. also delayed DNA-synthesis. The s ign i f icance of the react ions proceeding in the second phase of the regene- ra t ion process is discussed. R. was k indly donated by Dr. HELM (Chemie GrUnenthal) and E. by Prof. Dr. BROCK, (ASTA-Werke)~and S.by Dr.TIMMLER(Boehringer,Mannheim).

DNA-content RNA-content days a f te r hepatectomy days a f t e r hepatectomy

2 3 4 2 3 4

Rifampicin 91% 95 % - 82 % 86 % 30o mg/kg (da i l y )

Cyclophosphamide 5 mg/kg (once) loo % 79 % lo5 % loo % 67 % 134 %

Data: % of cont ro ls . _ _ = p < 0.0027

Prof. Dr. D. Neubert, Pharmakologisches I n s t i t u t der Freien Unlvers i tBt Ber l in , Abt. "EmbryonaI-Pharmakologle" , D-looo Ber l in 33, Th le la l l ee 69/73

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SUBCELLULAR DISTRIBUTION OF CYCLIC NUCLEOT!DE PHOSPHODIESTERASES (PDE) IN ISOLATED FAT CELLS OF RATS (Die subzellul~re Verteilun@ yon zyklischen Nukleotid-Phosphodiesterasen in isolierten Fett- zellen der Ratte) U. Klotz and K. Stock

Isolated Fat cells were prepared after Rodbell (1964) and s nated by diFFerential and gradient centriFugation after McKeel and Jarett (1970). The isolated subcellular Fractions were identified by marker enzymes. In the homogenate, nuclear, microsomal, mitochondrial and cytoplasmatic Fraction PDE-activities were measured in the mM-substrate range after Butcher and Sutherland (1962) and in the ~M-substrate range after Thompson and Appleman (1971). In all Fraction tested c-AMP, c-GMP and C-UMP were hydrolyzed, but to a markedly diFFerent degree. The highest specific activity towards c-AMP and c-GMP was Found in the cytoplasma while c-UMP was best hydrolyzed in the microsomes. The cytoplasma contained 75 % oP the total PDE-activity, being more specific For c-AMP (Km 0.7 ~M) than For c-GMP (Km 9.3 ~M), but c-GMP had a ten Fold higher Vmax-value. The microsomes contained only 4 % oP the total c-AMP resp. c-GMP activity but about 30 % oP the total c-UMP activity; the Km values were: c-UMP 0.67 mM, c-AMP 0.17 mM, cGMP 0.1 mM. The highest Vmax value (nmoles Pi/mg protein x min) was Pound with c-UMP (15.0), while c-AMP (2.0) and c-GMP (1.1) were hydrolyzed much slower. Papaverine, a competitive inhibitor in all Fractions tested, was most potent in the microsomes (Ki 1.8 ~M) and less potent in cytoplasma (Ki 8.3 ~M).

U. Klotz, Pharmakologisches Institut der Medizinischen Hochschule, D-3000 Hannover-KleePeld, RoderbruchstraBe 101, Germany.

INVOLVEMENT OF ADRENERGIC NEURONES WITHIN THE CNS IN VAGALLY MEDI- ATED CARDIODEPRESSOR REFLEXES (Beteiligung von adrenergen Neuro- nen an der ~bertragung vagaler Herzreflexe im ZNS) W.Kobin~er and A.Walland

In a previous publication (W.Kobinger and A.Walland, Europ.J. Pharmacol. 16, 120; 1971) it was shown that intracisternal (i.ci.) injection of 1 ~g/kg Clonidine (CLON, hypotensive, direct a-adrenergic agent) facilitated vagal reflex bradycardia, and this facilitation was antagonized by Phentolamine (PHENT) 0,5 mg/kg,i.ci. I.v. injections of both substances were ineffective (to be pub- lished). These experiments were continued in dogs, pentobarbital (25 mg/kg), adrenergic B-blockade with Doberol (K~ 592, 5 mg/kg). Blood pressure and heart rate were recorded. In approx. 50% of the dogs Angiotensin (ANG) elicited a reflex bradycardia which was diminished by PHENT 0,5 mg/kg i.ci. but not i.v. In all dogs pretreated with Reserpine (2 mg/kg, 18 hrs) a distinct reflex bradycardia was elicited by ANG; this reflex was not influenced by PHENT 0,5 ~g/kg, i.ci. In Reserpine pretreated dogs CLON (i ~g/kg,i.ci.) facilitated bradycardia, and this facilitation was antagonized by the following injection of PHENT 0,2 mg/kg, i.ci. It is concluded that the central transmission of the vagus reflex is not entirely dependent of adrenergic neurones(not influenced by Reserpine and s-receptor block), but the reflex can be amplified by adrenergic mechanisms (facilitation by CLON, inhibition by s-receptor blockade).

Doz.Dr.W.Kobinger and Dr.A.Walland, Arzneimittelforschung GmbH, Pharmakologisches Labor, A-1121 Wien, Laskegasse 5-11.

R 68

INFLUENCE OF CARDIOTROPIC AGENTS ON dp/dtmax AND VCEmax OF THE LEFT VENTRICLE (Der Einfluss kreislaufwirksamer Pharmaka auI dp/dtmax und VCEmax des linken Ventrikels) Eo KGbler und K. Greeff

Dp/dtmax wird nicht nur yon Anderungen der Kontraktilit~[t, sondern auch von Anderungen des enddiastolischen Druckes und des Blutdruckes beeinflusst. Die Verldirzungsgeschwindigkeit der kontraldilen Elemente (VCE) ist ein direktes Mass der Kontraktilit~t. An narkotisierten Katzen wurde der Druck im linken Ventrikel gemessen und dp/dt sowie die Quotienten dp/dt/P und dp/dt/(P-EDP) analog (Analogreehner Fa. ifd Meseher) er- rechneto Nut bei konstantem enddiastolischem Druck ist dp/dt/P proportional zu VCE, anderenfalls muss der Quotient dp/dt/(P-EDP) gebildet werden. Aeetyleholin senkt dp/dtmax signifikant; VCE bleibt unbeeinilusst. Die Abnahme von dp/dtmax wird also dutch die B1utdrucksenkung und nicht durch eine Kontraktilit~tsminderung verursachto Histamin bewirkt zum Zeitpunkt der maximalen Blutdrucksenkung keine signifikante ~tuderung von dp/dtmax~ VCE steigt an. Die am isolierten Herz bekannte positiv inotrope Wirkung des Histamins Igsst sich in situ wegen der Blutdruckwir- kung mit dp/dtmax nicht nachweisen. Druckgeschwindigkeitskurven des Ventrikels haben im xy-Oszillographen die Form einer Hyperbel, wie sie vom isolierten Papillarmuskel bekannt ist (E. H~ SONNENBLICK~ Fed. Proco 21, 975 (1962)). Die Interpolation der Kurven auf die Wandspannung Null is--t- unsicher.

Dr. E. KGhler, Pharmakol. Institut der Universit~t, 4 IMisseldorl, Moorenstr. 5

DIFFERENZIERUNG ZWISCHEN CA ++- UND NA+-ANTAGONISTISCHEN INHIBI- TOREN DER TRANSMEMBRAN~REN KATIONENFLUXE DURCH VOLTAGE-CLAMP-EX- PERIMENTE AM WARMBLUTERMYOKARD.

M. Kohlhardt, B. Bauer, H. Krause u. A. Fleckenstein

In den+~etzten Jahren wurde in unserem Laboratorium eine Reihe yon Ca -antagonistischen Hemmstoffen (Verapamil, D 600, BAY a i040) analysiert, die den elektromechanischen Koppelungsprozess am WarmblGtermyokard spezifisch blockieren k~nnen. Wie Voltage- Clamp-Experimente an isolierten Ventrikel-Trabekeln der Katze zeigten, reduzieren diese Substanzen selektiv den transmembra- n~ren Ca+§ ohne nennenswerte Ver~nderung des schnellen Na ~ Influxes. Umgekehrt kann durch Lok~lan~sthetika wie z.B. Novo- cain oder Xylocain der s~nelle Na -Strom gehemmt werden, ohne dass gleichzeitig die Ca -Konduktivit~t der Membran beeinflusst wird. Andere cardio-inhibitorische Pharmaka wie z.B. Prenylamin nehmen eine Mittelstellung ein, indem sie beide Einw~rtsstr~me vermindern. Die Befunde zeigen- dass die Membran der WarmblGter- Myokardfaser Gber getrennte Ca d+- und Na+-Kan~le verfGgt, die dutch Pharmaka unabh~ngig voneinander blockierbar sind.

Anschrift der Verfasser: Physiologisches Institut d. Universit~t D 78 Frelburg i. Br., Hermann-Herder-Str. 7

R 69

KINETICS OF THE ACCUMULATION AND RELEASE OF OUABAIN AND DIHYDR0- 0UABAIN BY THE ISOLATED GUINEA PIG HEART (Kinetik der Aufnahme und Abgabe yon Ouabain und Dihydroouabain am isolierten Neer- schweinchenherzen) N. Kolassa and F. Lauterbach

The time course of the accumulation and release of ouabain and dihydroouabain by the isolated perfused guinea pig heart was examined by measuring the concentration of H3-ouabain and H3- dihydroouabain, respectively, in the effluent perfusion fluid during a 20 min influx and 20 min efflux period. For the amount of glycosides in the extracellular space two com- partments with rate constants of 4.3 and 0.46 minT7 were compu- ted by means of the simultaneous measurement of C1~-inulin. Cellular ouabain was described by a third compartment. The size of this compartment was used for the calculation of a tissue- medium-ratio (T/N) which amounted to 1.3 at 10 -8 M ouabain after 20 min. The T/M value decreased with increasing ouabain concentrations (0.5 at 10 -6 N ouabain); it was dependent on the K +- concentration in the perfusion medium (T/M = 2.9 at 2.2 mM K+; T/N = 0.9 at 10 mMAK+); it remained unchanged when the heart was preloaded with 10 -~ or 10-7 ~ non-radioactive ouabain. The rate constant of 0.085 min -~ for this third compartment showed no significant changes at the tested experimental conditions. 10.-8 M dihydroouabain yielded a T/N value of only O.q2. Thus, uptake in this third compartment parallels the positive inotropic efficacy and not the very similar physieochemical properties of both glycosides.

Dr. N. Kolassa, Institut fur Pharmakologie und Toxikologie der Ruhr-Universit~t Bochum, 463 Bochum, im Lottental.

THE EFFECTS OF CORONARY DILATATORS AT DIFFERENT CA++-CONCENTRA - TIONS OF THE PERFUSION FLUID (Die Wirkunge~+von Coronardilata- toren bei verschiedenen extracellul~ren Ca -Konzentrationen) F.KOlSChe ~ W.K. Raff and W. Lochner

The effects of coronary dilatators were studied in isolated~++ isovolumetrically beating guinea pig hearts at different Ca - concentrations of the perfusion fluid. In all experiments hyp- oxic dilatation of the vessels was absent because the coronary venous partial pressure of oxygen was higher than 50 mmHg. The coronary dilatation caused by six of the investigated substances - Nifedipine, Prenylamin, Verapamil, Lidoflazin, Dipyridamol and Hexobendin - could be suppressed by increasing of the Ca ++- concentration of the perfusion fluid. Therefore the mechanism of action on t~e smooth muscle of the coronary vessels has to be explained by a calcium antagonistic effect of these substances. The contractility of the myocardium is influenced in different ways. Nifedipine, Verapamil and Prenylamin show negative inotropic effects, which are also compensated by increasing the Ca ++- concentration. In contrast Hexobendin, Lidoflazin and Dipyrid- amol show no negative or even a small positive inotropic effect. The effects of nitroglycerine and carbochromen are not affected by changes of the extracellular Ca++-concentration.

Prof.Dr.W.Lochner, Physiologisches Institut der Universit~t DGsseldorf, D-4000 DGsseldorf, MoorenstraBe 5.

R 7 0

INFLUENCE OF VARIOUS DRUGS O N THE BINDING OF CHLORPROMAZINE TO ERYTHROCYTES AND ALBUMIN (Der Einflu6 verschledener Pharmaka auf die Bindung von Chlorpromazln an Erythrocyten und Albumin) G. Krieglstein, I. Hahn t J. KrlegJsteln, K. Tschentscher

The distribution of the highly l ipoph;l lc chlorpromazlne (CPZ) was studied under the influence of various drugs in a simplified blood (31.5 + o.25 ~ well-washed bovine erythrocytes and 4 g o~ bovine serum albumin in o.o2 7~4 phosphate buffer solution containing o.15 M NaCI). CPZ was added to the simplified blood to give a total concentration of loo /JM and after an incubation period of 3 hrs at 22~ the CPZ concentration was measured in the albumin solution after centrifugation at 3ooo x g and in the buffer solution after ultracentrifugation at 15o ooo x g. 68.1 + o.8 ~ of CPZ were bound to erythrocytes~ 28.5 + o.9 ~ were bound to albumin and 3.5 + o.2 ~ were unbound (x + s- ; n = lo).-Several substances ( i .e. chlorimipramine,-

- - X , . . . .

diphenylhydantoin, indometkiacm, chloroth~azide, chlort~tracyclme, suramm, olelc acid, ioglycamlc acid) added in a concentration of lo -v M to the simplified blood were able to change the distribution equilibrium of CPZ. In most cases considerable changes were found between the erythrocyte bound CPZ and the albumin bound CPZ whereas the free CPZ in the aqueous phase was not markedly changed. Evidence is presented that salicylic acid and imipramine did not displace CPZ from albumin binding, as it is reported in the literature, in the simplified blood used. It is concluded that binding studies merely with albumin solutions or plasma may be mis- leading when pharmacokinetic implications are argued.

Dr. G. Krieglstein, Pharrnakologisches Institut der Universitat Mainz D-65oo Malnz, Obere Zahlbacher Str. 67

E F F E C T S OF ISOPRENALINE~ H I S T A M I N E AND O X Y F E D R I N E ON T H E C Y C L I C AMP C O N T E N T OF T H E H E A R T (Wirkung yon Isoprenal in~ H i s t a - min und O x y f e d r i n auf den Z y k l o - A M P - G e h a l t des Herzens) W.R. Kukovetz T G. PBch I A. Wurm

In o r d e r to f u r t h e r e luc ida te the poss ib le r o l e of cyc l i c AMP (cAMP) as cel lu- : lap med ia to r of the pos i t i ve i no t rop i c e f fec ts of drugs which s t imula te a d e n y l - cyc lase , the t ime course of the e f fec ts of O. 5 IJ, g i sop rena l i ne (IS), 10 #g h i - stamine (HIST) and 10 IJ, g o x y f e d r i n e (OF) on the cAMP-con ten t (estima,ted accord ing to A .G . Gi lman, Proc . Nat. Acad. Sc i . 6?, 305-312,11970) and on the ampl i tude of i so ton ic con t rac t i ons (ram i . c . ) was s tud ied in the g u i n e a - p i g - hear t (Langendor f f ) . With a l l 3 substances the magni tude on the i no t rop i c r e s - ponse was c l o s e l y p a r a l l e l e d , however p receded, by s i m i l a r changes in cAMP. D o s e - r e s p o n s e - c u r v e s were es tab l i shed wi th O. 5 - 5 0 0 n9 IS and wi th O. 2 - 10 #,g H IST . The r i ses in mm i . c . we re c l o s e l y re f l ec ted by s i m i l a r r i ses in cAMP. In pa r t i cu la r~ a f t e r the lowest doses of both drugs which inc reased mm i . c . by-1/4-% above c o n t r o l ~ t h e r e w e r e a l r e a d y s ign i f i can t r i ses in cAMP by '~'43%. The a d r e n e r g i c B -b locke r K6 59:z (50 P,9) which by i t se l f d id not af fect mm i . c . and cAMP, comp le te l y abo l i shed the r i se in mm i . c . and in cAMP~ as produced by 1OO ng IS whereas s i m i l a r e f fects of 2, IJ,9 H I S T remained unchanged. The resu l t s add c i r cums tan t i a l ev idence to the v iew that cAMP me- d ia tes the pos i t i ve i no t rop i c response of drugs which inc rease its c o n c e n t r a - t ion in the hear t .

P ro f , D r . W,R. Kukovetz , Pha rmako log i sches lns t i tu t der U n i v e r s i t ~ t A - 8 0 1 0 Graz~ U n i v . - P l a t z 4.

R 71

INFLUENCE OF PHENYLBUTAZO~,TE-BINDING TO PROTEINS ON THE ACTION OF TOLBUTAIVIID ON THE GLUCOSE BLOOD LEVEL OF RABBITS (EinfluS der Bindung yon Phenylbutazon an Proteine auf die blutzucker- senkende Wirkung yon Tolbutamid beirn Kaninehen) H. Kurz

Phenylbutazone and oxiphenbutazone liberate tolbutamid from its bindirig to plasma proteins. However, this did not prod~uce the expected effect on the action of tolbutamid on glucose level in rabbits. Although oxiphenbutazone increased the effect of tolbutamid on glucose level, this happened to a far higher degree than could be explained by liberating tolbutamid from the plasma proteins. On the other hand, phenylbutazone even decreased the effect of tolbufamid. There is evidence that the effects of phenylbutazone and oxiphenbutazone are rather due to the specific pharmacological action of these substances on glucose level. The missing effectiveness of the liberation of tolbutamid from plasma could be explained by calculating the changes of drug concentration in the entire body water. This indicates that the amount of the drug liberated from the plasma produces only a slight increase of the effective concentration if equally distributed within the entire body water. According to that, liberation from the plasma proteins should be of no considerable effect on phar- macologieal action. Therefore, it seems that the infhenee of liberation on drug action is rather caused by the liberation of the drug from its binding to tissue proteins, espeeially muscle proteins, than by liberation from the quantitatively small amount of the plasma proteins.

Prof. Dr. H. Kurz, Pharmakologisches Ins t i tu t der Universit~t Miinehen, D-8000 Miinehen 2, Nussbaumstrasse 26

SERUM DOPAMINE-BETA-HYDROXYLASE (DBH) DURING DEVELOPMENT OF IMMOBILIZATION-INDUCED AND GENETIC HYPERTENSION IN RATS (Serum Dopamin-Beta-Hydroxylase (DBH) w~hrend der Enfwicklung von Immo- bilisation induziertem und genetisehem Hochdruck in Raften) F. Lamprecht, R.F. Williams and I.J. Kopin

Serum DBH activity, which reflects peripheral sympathetic nerve function, was measured in rats subjected to immobilization for two hours daily over a period of four weeks. Both serum DBH and blood pressure (BP) increased significantly after two weeks of repeated immobilization (p (DBH) ~ 0. 001; p (BP) 0.02). The BP difference between control and experimental animals remained the same under ether anesthesia. After four weeks of immobilization, there was a further increase in both serum DBH and BP. Upon cessation of stress intervals, serum DBH returned to normal within five days; BP remained elevated for two additional weeks before declining to normal. In contrast, DOCA- salt-induced BP elevation was accompanied by a decrease in serum DBH (p 0.01). DBH levels in both genetic strains tested (spontaneously hypertensive rats and salt-sensitive rats) were lower than their corresponding controls (p < 0. 001). It appears likely that the decreased DBH levels reflect diminished sympathetic nerve activity in non-stress-induced hypertension. Presumably the BP elevation is a consequence of other factors. Similarly, the persistence of BP elevation after repeated immobilization at a time when serum DBH levels have returned to normal also suggests that the hypertension is not a consequence of peripheral sympathetic nerve activity.

Dr. F. Lamprecht, NIMH, Building 10, 2D46, Bethesda, Maryland 20014, USA.

R72

COMPARATIVE STUDIES ON THE EFFECT OF PROCAINAMIDE, LIDOCAINE AND ~-DIETHYLAM!NO-n-BUTYRYLANILIDE ON THE ELECTRIC FIBRILLATION THRESHOLD OF THE GUINEA PIG HEART IN HYPOTHERMIA (Vergleichende Untersuchungen zur Wirkung yon Procainamid, ~idocain und ~-Di~thylamino-n-butters~ureanilid aufdi 9 elektrische Flimmer- schwelle des Meerschweinchenherzens in Hypothermie) H. Lange-Asschenfe!dt

Hypothermia augments the susceptibility to cardiac arrhythmias and esp. to cardiac fibrillation. The aetiology of these disturb- ances has not yet been clarified. Since hypothermia induces a complex abnormal condition in intact organisms, we believe that experiments on whole animals are required for the investigation of antifibrillatory drugs with this respect. In this study rectangular impulses (duration IO ms, frequency 50 ips) were applicated by an intracardial electrode in anaesthetized female guinea pigs at a body temperature of 30~ Stimul- ation intensity was increased gradually until cardiac fibrillation occurred. In comparison to the control group procainamide, lidocaine and ~-diethylamino-n-butyrylanilide (ABA) elevated the fibrillation threshold in a dosage of 7.5 micromol/kg each. Lidocaine and ABA are equally effective but more potent than procainamide. Since ABA is reported to be less toxic than lidocaine, this compound should be subjected to further investigations.

Dr. H. Lange-Asschenfeldt, Schiffahrtmedizinisches Institut der Marine, D-2300 Kiel-Kronshagen, Kopperpahler Allee 120

CHANGES IN NA/K CONTENT OF ERYTHROCYTES INDUCED BY THE DIRECT LYTIC FACTOR OF COBRA VENOM AND PHOSPHOLIPASE A (Ver~nderungen im Na/K-Gebalt yon Erytbrocyten durch den direkt lytischen Fak~r aus Cobragift und Phospholipase A) P.G. Laukisch, C. Jacobi, W. Vo~t~ K. Sohoner

The effects of the direct lyric factor (DLF) of cobra venom, bee venom phospholipase A and of combinations of these two lysins on guinea-pig red cells have been studied. DLF caused an increased permeability to Na ions, swel~ing of the cells and moderate haemolysis. No prelytic loss of potassium was seen. Pbospholipase A produced loss of potassium, considerable gain of sodium, cell swelling, but no remarkable lysis. Combinations of the two venom constituents, more strongly baemolytic, mimicked the effect of the predominant component of the mixture. Thus prelytic loss of potassium was observed when higher concentrations of phospholipase were combined with low concentrations of DLF, but was absent when DLF predominated. The development of spherocytosis at rather low critical volumes of baemolyzing DLF treated red cells, and the effect of DLF on osmotic stability suggest that DLF has other effects on the cell membrane in addition to those of an osmotic haemolysin.

Dr. P.G. Lankisch, Max-Planck-Institut fur experimentelle M~izin, Abt. Biochemische Pharmakologie, D-3400 GSttingen, Hermann-Rein- Strafe 3.

R73

INVESTIGATIONS ON THE ABSORPTION MECHANISM OF 3-O-METHYL-)-GLU- COSE IN THE ISOLATED MUCOSA OF GUINEA-PIG SNALL INTESTINE. (Untersuchungen zum Nechanismus der Resorption yon 3-@-Methyl@- Glucose an der isolierten Nucosa des MeerschweinchendGnndarmes). F. Lauterbach Cardiac glycosides and quaternary ammonium salts are taken up by the intestinal epithelial cell by means of a transport mechanism situated in its basal membrane. In this connection, trans- elNthelial fluxes and intracellular uptake of 5-0-methyl-glucose (5-O-mg) and galactose (gal) were studied on an isolated preparation of guinea-pig small intestinal mucosa. Transepithelial fluxes of inulin-H-5 and 5-0-mg-C-14 are significantly correlated, but 5-0-mg flux at zero inulin flux is four times higher in the direction from the lumen to the blood than vice versa. Intracellular concentrations of 5-0-mg are 0.2~ 0.25, and 0.55 mN after incubation with 1 mN on the lumen side, the blood side, and both sides respectively. When 1 mM 5-O-mg or 28 mN gal is added to both sides of the tissue, sugars are transported uphill from lumen to blood side, but the tissue concentration remains below that of the medium. After preloading the tissue with sugar and incubating in a sugar-containing medium, sugar is released against a concentration gradient, an effect that is not influenced by iodoacetate nor anaerobiosis. Uptakes of both sugars from the blood side are similar linear functions of concentration. The present experiments provide no evidence for a basal transport system comparable to that of cardiac glycosides.

F. Lauterbach, !nstitut fur Pharmakologie und Toxikologie der Ruhr-Universit~t Bochum, 465 Bochum~ Im Lottental.

ABOUT THE DIFFERENCE BETWEEN DIRECT AND INDIRECT (HEMODYNAMIC) INFLU- ENCE ON CAPILLARY PERMEABILITY (Zur Differenzierung yon direkter und indirekter (himodynamischer) Beeinflussung der GefiBpermeabiIit~t) H. D. Lehmann

The influence on capillary permeability was studied in the hindlimbs of anesthetized dogs by measurement of lymph flow, lymph/plasma-qmetients of protein and dextran (mw 60000), arterial flow, arterial and venous pressure. Papaverin (0,6 mg/kg x min i.a.) enhanced lymph flow and arterial flew, the other parameters remained unchanged. Partial occlusion of Vena ilica ext. enhanced lymph flow, 1/p-dextran and arterial pressure. These are examples of indirect (hemodynamic) influence on capillary permeability. Very similarly bradykinin (0,1 ug/kg x min i.a.) increased lymph flow, 1/p-dextran, arterial flow and venous pressure. For distinction between direct and indirect influence on capillary permeability it is necessary to determine the effect of antagonizing substances on capillary permeability and hemodynamics. The increase of lymph flow induced by bradykinin was inhibited by phenylbutazone, but the hemodyaamic effects of bradykinin were not inhibited (Lehmann, Arch. Pharmak. 270 (Suppl.): R 83, 1971). The increase of lymph flow induced by papaverin was not inhibited by phenylbutazone. It is concluded that phenylbutazone acts as a specific antagonist against bradykinin and the influence of bradykinin on capillary permeability is a direct one.

Dr. H. D. Lehmann, Biologisches Institut Madaus, 5 KSln 91, 0stmerhei- mer StraBe 198

R74

INFLUENCE OF HEAVY WATER ON THE MEMORY OF FISHES (Der EinfluB yon Sehwerem Wasser auf das Ged~ehtnis bei Fischen) E. Lehr, M. Wenzel u. G. Werner

2-j~hrige Rotfedern (Scardinius erythroph%halmus,Teleostei) er- lernten in mit Futter belohnten Dressuren rote und griine Signal- liehter zu unterscheiden. Schon w~hrend der Dressur oder unmit- telbar naeh dereu AbschluB wurdeu solehe dressierte Fisehe in Wasser mit 12,5 oder 25% D20-Gehalt eingesetzt. Bereits 2 Std. sp~ter zeigte auch das KSrperwasser dieser Tiere die gleiche D20-Konzentration. GegenHber Kontrolltieren in normalem Aqtk~rie~ wasser behielten in vier voneinander unabh~ngigen Versuchsreihen (insgesamt 12570 Testwahlen) die Fische in Sehwerem Wasser ihre erlernte Aufgabe signifikaut (p > 0701 ) besser, gleichgUltig,ob beim Ged~chtnistest alle Wahlen belohnt waren oder ob dabei die unbelohnte Extinetionsmethode angewandt wurde(E. Lehr, M. Wenzel und G. Werner, Naturwissenschaften 57, 521-24, 1970). Das stabilere Ged~ehtnis kSnnte durch die gegenfiber normalen WasserstoffbrHeken st~rkeren Deuterium-Bindungen verursacht sein, indem diese die Ged~ehtnismolekfile selbst festigen und die Akti- vit~t jener Enzyme herabsetzen, die normalerweise durch den Abbau yon Ged~chtnisstrukturen zum Vergessen ffihren.

Dr. E r i c h Lehr u. P r o f . Dr. G. W e r n e r , M a x - P l a n c k - I n s t i t u t f u r H i r n f o r s c h u n g , A r b e i t s g r u p p e N e u r o e h e m i e , D-6000 F r a n k f u r t / M . - N i e d e r r a d , D e u t s c h o r d e n s t r . 46 P r o f . Dr. M. W e n z e l , B i o l . - C h e m . A b t . de s P h a r m a z e u t i s e h e n I n s t i - t u t e s d e r F r e i e n U n i v e r s i t ~ t , 1 B e r l i n 3 3 , K S n i g i n - L u i s e - S t r . 2 - ~

NEURONAL EFFLUX OF EXOGENOUS CATECHOLAMINES FROM THE ISOLATED RABBIT HEART (Der neuronale Efflux von exogenen Catecholaminen aus dem isollerten Kanlnchenherzen) K. L~ffelholz Neuronal efflux of noradrenaline (NA) was found previously (L~ffelholz et al . , Naunyn- Schmiedebergs Arch.Pharmak. 27._~o, R87, 1971) and in the present experiments on isolated rabbit hearts after block of the Tntraneuronal inactivation of NA, i .e. after pretreatment wlth pargyllne and reserpine. NA (2oo and 3o ng/ml) or adrenaline (A, 2oo ng/ml) were infused for various periods (1-18o rain). The concentration of the amine in the venous effluate during and after infusion was measured fluorimetrically. After the end of the infusion the neuronal efflux declined exponentially. The rate of the decline (t I/2) and the intercept (Yo) with the y--axis at t o (end of' the infusion) were calculated. Both t_ and Yo increased with higher total amounts of NA infused; a steady state (t 1/2 u= 43 min, Yo = 26 ng/g �9 rain) was reached after 4o min with NA infusions of 2oo ng,/ml and after 12o rain with NA infusions of 3o ng/ml. Similar values were obtained after infusions of A (2oo ng,/ml). After the start of the infusion the NA removal was marked, but then it declined gradually and disappeared after 2o- 6o rain (2oo ng/'m] NA) or 12o-18o rain (30 ng/ml NA). Inhibition of the re-uptake of NA by DMI (5 �9 lo -6 g,"ml) during the postlnfusion period elevated Yo to about 6o ng/g �9 m|n and decreased t 1/2 to about 3o rain. It is concluded that the neuronal efflux is partly responsible for the decline of the NA removal during NA infusion. Further, the rate of distribution of exogenous NA in the adrenergic axon seems to be slow and, therefore, is most probably the rate-limiting step for an equilibrium betweer the extraneuronal and axoplasmlc NA and for the neuronal efflux of exogenous NA.

Dr. K. ksffelholz, Pharmakologisches Institut der Universitdt Mainz D-65oo Mainz, Obere Zahlbacher Str. 67

R75

HISTAMINE RELEASE DURING ORTHOTOPIC HOMOLOGOUS LIVER TRANSPLANTATION IN PIG (Histaminfreisetzung bei orthotoper homoioger Lebertrans- pIantat ion am Schwein) W. Lorenz, E. Hel i , O. Boeckl, H.J. Reimann, G. Zimmermann

During l i v e r t ransplantat ion in man, dog and pig severe hypotension in the anhepatic phase (AHP) and fol iowing revascular izat ion (REV) were observed. A vasoactive substance, histamine (HA), was therefore determined f luorometr ica l ly (W.Lorenz er at.,Europ.J.Pharmacol.14,155-175, 1971) in the plasma of r igh t atrium of 14 young pigs during l i v e r t rans- p lantat ion (E.Hell et al.,Wien.kl in.Wschr.83,82-85,1971). The surviv ing time of the 7 recip ients was 4-16 days, the average time of cold ischaemia 35 min. Basal HA levels observed at various times during and also a f ter operation were 21~ng/mi both in the donors and rec ip ients. An increase of plasma HA concentration was measured (1) about 1 h a f ter beginning the anaesthesia with pentobarbitone, galIamine and N20 both in the donors (54~10 ng/ml) and recip ients (44~12 ng/ml), (2) in the eariy.AHP of the donors (40• ng/mi), (3) immediateiy af ter REV ~86~6 ng/mi) and, (4) a f ter opening the hepatic ar tery anastomosis (43-8 ng/ml). Severe hypotension (50-100 mm Hg) occured during the la te AHP and immediately af ter REV, but was related r HA release only in the i a te r phase. Thus, HA release is probably an important factor in the development of hypotension fol iowing REV,but not in the AHP of I i v e r t rans- p iantat ion. Prof.Dr.W.Lorenz, Abteilung fur exp.Chirurgie und pathol.Biochemie, Chirurgische Un iv . -K l in ik , D-355 Marburg, Robert-Koch-StraBe 8

HISTAMINE RELEASE FROM TYPICAL AND ATYPICAL MAST CELL STORES BY CREMOPHOR EL (Histaminfreisetzung aus typischen und a• Mastzei1- speichern dutch Cremophor El) W. Lorenz r H.J. Reimonn r A. Schmal, R. Tauber r R. UhIi9

Cremophor E1 (CEL), a tensid, which was shown %0 be a speci f ic histamine (HA) releaser in dogs (W.Lorenz e• al.,Arch.Pharmak.269,417,1971) in - creased the HA concentration in whole blood by more than 600 % and caused a severe hypo• and copious gastr ic secretion in 5 dogs with a acute gastr ic f i s t u l a . Tachyphylaxis was induced by repeated in ject ions of 0.2 - 1.0 ml/kg CEL. The tensid Lensodel NP 40 (0.05 ml/kg i . v . ) , a condensation product of octylphenole and ethylenoxide caused hypotension, but no HA release. In order to invest igate from which t issues HA was released by CEL, 12 mongrel dogs (pairs of the same i i t t e r ) were used. Six of them received i . v . 0.2 ml/Eg CEL at the f i r s t day, 0.4 ml/kg at the second and 0.6 ml/kg at the th i rd day whereas the other s ix dogs (as controls) received the same volumes of sal ine. HA was determined f luorometr ica l ly according r W.Lorenz et a l . (Europ.J.Pharmacol.14, 155-175,1971). A s igni f lcan~ HA release was found in skin (83%), pa.creas (75~), antrol musculature (54%), ieiu.um (39~), colon (37%), ontral mucoso (36%) and ileum ( 3 ~ ) . No HA release could be shown in lung, l i v e r , spleen and diaphragm. Thus CEL is the f i r s t HA releaser which acts both on typ ica l and on atypical mast ce l l stores.

Prof.Dr.W.Lorenz, Abteilung fur exp.Chirurgie und pathol.Biochemie, Chirurgische Un iv . -K l in ik , D-355 Harburg, Rober• 8

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COMPARATIVE INVESTIGATIONS ON THE INFLUENCE OF VARIOUS SUBSTRATES ON THE RESPIRATION OF A SPONTANEOUSLY CONTRACTILE VASCULAR SMOOTH MUSCLE AND A NON-CONTRACTILE CONTROL VESSEL (PORTAL VEIN AND VENA CAVA INFERIOR OF THE RABBIT) IN VITRO (Vergleichende Untersuchun- gen ~ber die Substratoxydation dutch einen autorhythmischen und einen nicht spontan aktiven, glatten Gef~amuskel (Pfortader bzw. Vena cava des Kaninchens) in vitro) H. L~bcke und D. Voth

Electrons microscopic studies of vessels with active contractility show for this type of vascular smooth muscle certain features of fine structure which are related to the special activity (R. Schipp et al., Z.Anat. 134, 81-100~ 1971). There are also differences of the intracsllL'i~r cationic concentrations and of the proportion of energy rich phosphates (ATP, ADP, CP) between the both types of vascular smooth muscle. We studied the oxygen consumption of both tissues by measuring the respiration with the conventional Warburg manometric apparatus. The following substratss are used: glucose; fructose; sorbitol; ribose; the sodium salts of succinate, pyruvate, lactate; valine. The respiratory rats of the spontaneously contractile muscle exceeds that by the control vessel significantly and reaches the threefold value. The are not only quantitative but also qualita- tive differences of the utilisation of the substrates used for the experiments between the two types of vascular smooth muscle. The autonomously contractile smooth muscles differ definitely from those without this functional speciality not only in regard to the fine structural characteristics but also to the metabolic behaviour and activity. Prof.Dr.D.Voth, Neurochirurgische Universit~tsklinik mainz, D-6500 M a i n z , LangenbeckstraSe I

CATECHOLAMINE TURNOVER IN THE RAT BRAIN:DIFFERENT RESULTS OBTAINED IN DIFFERENT EXPERIMENTAL CONDITIONS (Catecholaminturnover im Rat- tengehirn: unterschiedliche Ergebnisse aus verschiedenen Methoden) L. Maitre, P. Baumann and M. Staehelin

The effects of chlorpromazine (CPZ) and benzoctamine (B) were compared in several systems used for studying catecholamine(CA) turnover in the rat brain: l)accumulation and release of 3H-CA formed from intravenously injected ~H-tyrosine 2)accumulation of 3H-CA in brain slices incubated with ~H-tyrosine 3)depletion rate of the endogenous CA stores after blockade of CA biosynthesis with ~-methyltyrosine. Accumulation and release of 3H-dopamine(DA) formed from i.v. injected ~H-tyrosine were increased by both CPZ and B, whereas accumulation and release of 3H-noradrenaline(NA) were increased only by B. 3H-CA accumulation was also enhanced in striatal and hypothalamic slices obtained from rats treated with these drugs. A particularly strong effect on NA accumulation was seen in the hypothalamus of B-treated rats. After blockade of CA biosynthesis, NA depletion was accelerated by CPZ and B, but DA depletion remained unaltered.

The results show a good correlation between the methods involving newly synthesized CA and a profound discrepancy between these models and that using blockade of CA biosynthesis. It seems therefore that different CA pools are measured in the two sets of experimental conditions.

Dr. L. Maitre, Biologische Forschungslaboratorien,Division Pharma- zeutika, CIBA-GEIGY AG, CH 4002 Basel (Schweiz)

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INTERACTION BETWEEN USVC 6524 AND VARIOUS ADRENERGIC BETA-STIMU- LANT AGENTS ON THE HISTAMINE-INDUCED BRONCHOSPASM IN GUINEA-PIGS (Zwischen~irkungen zwischen USVC 6524 und verschiedenen beta-sti= mulierendenAgenten auF den Bronchospasmus dutch Histamin in Meerschweinchen) E. Marmo, R. Saini and A.P.Caputi

The present study was undertaken to investigate the antagonistic properties os a new beta-receptor blocking agent, namely USVC 6524 with respect to various beta-stimulants,on the histamine- induced bronchospasm in guinea-pigs.A 0.4% histamine aerosol was used. The animals were kept For a maximum period os 4 minutes,For the induction oF bronchospasm. EDSO (anti-bronchospastic activity) os the beta-stimulant drugs, expressed in mcg/kg was: isoprenaline (22.97), salbutamol (28.86), terbutaline (33.14), soterenol (37. 90), Th 1165a (37.90), nylidrine (65.68), metaproterenol (70.16), isoxsuprine (71.49), orthoxine (I03.13), chlorprenaline (119.59) and quinterenol (432.74). By pretreatment with USVC 6524 (250 mcg/kg, i.v., given 2 minutes before the beta-stimulant drug),the EDso was:isoprenaline (49,613), Th 1165a (108.41), soterenol (110. 39), terbutaline (161.01), salbutamol (206.64),chlorprenaline (287.48), orthoxine (450.71), metaproterenol (481.93), nylidrine (>500), isoxsuprine (~750) and quinterenol (~I000). Our results clearely indicate that USVC 6524 has a high blocking activity on ~-l and ~-2 receptors.

ProF. E. Marmo, Pharmakologisches Institut der Unlversltat Neapel, Via Cimarosa 37, 80127 Napoli (Italia)

METABOLIC ACTIVATION OF CARCINOGENIC POLYCYCLIC HYDROCARBONS? THE EFFECT OF MICROSOMAL ENZYMES ON HYDROCARBON-INDUCED CARCINOGENESIS IN VITRO.(Metaboli - sche Aktivierung polyzyklischer Kohlenwasserstoffe? Der Einfluss mikrosomaler Enzyme auf die Kohlenwasserstoff-induzierte Carcinogenese in vitro). Hans Marquardt (Sloan-Kettering Institute for Qancer Research, New York/USA).

Es ist eine -bisher nicht gesicherte- Vermutung, dass carcinogene polyzyk- lische Kohlenwasserstoffe (PK) vor Entfaltung ihrer hiologischen Aktivitaet metabolischer Aktivierung (durch -induzierbare- mikrosomale Enzyme)beduerfen. Diese Frage wurde untersucht an einem Carcinogenese-Modellsystem, der malignen Transformation von Zellen (stammend vonder C3H-Maus Prostata) in Zellkultur. Addition von "Feeder"-Zellen zu oder Induzierung mikrosomaler Enzymaktivitaet in diesen Zellen erhoehte die Metabolisierungsrate yon und die Transformations- rate nach Benz(a)anthrazen (BA) und Methylcholanthren. Hemmung der mikrosomalen Enzymaktivitaet mittels @-Naphthoflavon oder Roentgenstrahlen verhinderte Transformation. Diese Ergebnisse bestaetigen die Vermutung, dass -zumindest diese- PK metabolischer Aktivierung beduerfen. Vorhergehende Untersuehungen (H.Marquardt et al~ Cancer Res., im Druck : April 1972) hatten gezeigt, dass Epoxyde als "proximate carcinogens" dieser PK angesehen werden koennen.

Nicht im Einklang mit diesem Schluss sind unsere Ergebnisse mit Dimethyl- benz(a)anthrazen (DMBA). Induzierung mikrosomaler Enzymaktivitaet fuehrte - trotz Steigerung der Metabolisierungsrate- nicht zu einer Erhoehung der Trans- formationsrate nach DMBA. Ausserdem war die transformierende Aktivitaet sowohl des K-Region-Epoxydes von 7-Methyl-BA als auch die der beiden Brom-substituierten PK (7-Brommethyl-BA und 7-Bron~nethyl-12-methyl-BA), die als Modellsub- stanzen fuer eine moegliche metabolische Aktivierung an einer Methylgruppe synthetisiert worden waren, geringer als die yon 7-Methyl-BA oder DMBA. Moeg- licherweise sind solche PK mit chemisch reaktiven Methylgruppen carcinogen ohne vorhergehende metabolische Aktivierung. Dr.H.Marquardt, Sloan-Kettering Institute, New York/New York IOO21/USA .

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PERITUBULAR AND INTRA~UMINAL EFFECTIVE CONCENtRAtIONS OF DIURE~ TICS. (Die peritubul~r und intraluminal wirksamen Diuretika- Konzentrationen). K.Meng

The isotonic fluid reabsorption (Jv) from cortical tubules of the rat kidney was measured with the split oil droplet technique of Gertz. In the same tubule the peritubular blood capillaries were perfused with artificial solutions using glass capillaries of 8 ~ outside diameter. The intraluminal solution for splitting the oil contained diuretics and (in mEq/1) 145 Na, 6 Ca, 141C1, 10 acetate (pH 6,9). ~he blood capillary perfusate contained diuretics, 145 Na, 6 Ca, 111C1, 35 HC03, 5 acetate and was equilibrated with 7/93% 002/02 mixture prior to use (pH 7,4).

JV was markedley reduced when diuretics were present in the capillary perfusate. Diuretics caused also inhibition of JV when applied intratubularly but only after higher concentrations. The diuretic concentrations (Mol/1) for 50% or 15% inhibition of JV are s~mmarized in the table:

Acetazolamide (N=141 Ohlorothiazide (N= 97 Mefruside(Lactone, N=127 Furosemide (N=207 Ethacrynic Acid (N=I05

peritubular intraluminal 50% 15% 15% inhibition

3.10 -5 5.10-~ 1,5.10 -5 2 . 1 0 - ~ 1 . 1 0 - / 3 . 1 0 - 5 4-.10 - ~ 2 . 1 0 - ' _ ( 6 10 -5 5 . 1 0 - 4 5 . 1 0 -./. 2 . 1 0 - 4

-- I �9 10-5

Priv.Doz.Dr. Karl Meng, Institut fur Pharmakologie, BAYER AG., 56 Wuppertal I, Postfach 130105

EFFECT OF 6-MERCAPTOPURINE ON NUCLEIC ACID METABOLISM OF RAT EMBRYOS IN VIVO (Wirkung yon 6-Mercaptopurin auf den NucleinsSure-Stoffwechsel in Rattenembry- onen in vivo) Peter MEWES, Gerd BOCHERT The incorporation of i~C-u-glucose fragments and 32p-orthophosphate into DNA and RNA of 12-days-old rat embryos is found to be drastically altered after injection of 1oo mg/k9 6-mercaptopurine (6-MP). The specific activity is decreased about the same degree in ribose moieties and phosphate groups of DNA purine nucleotides. The specific activity in the base part of G is reduced to an even higher degree. The de-novo-synthesis of pyrimldine nucleosides is scarcely affected, but the incorporation of 32p-orthophosphate is inhibited. As 6-MP is known to be a strong inhibitor of the de-novo-synthesis of the purine skeletons a deficiency in purine bases may be compensated by a higher rate of the salvage pathway. Our results suggest that purine nucleotides - possibly derived from nucleic acids - are preferably broken down to bases and re-synthesised to poly- nucleotides. As expected the de-novo-synthesis of pyrimidine bases is not affected by 6-MP, but a decreased incorporation of 32p in the s-position is found.

DNA Bases (lhC) Nucleotides (I~c) Nucleotides (32p)

G 7o + 5 78 -+ 3 8o -+ 3

A 82 _+ 6 7o +_ 2 68 _+ 2

C 94 _+ 7 loo + 4 84 • 3

T lo7 • 8 117 + 4 73 • 2

Data are expressed in % of c o n t r o l s ; 1oo mg/kg 6-MP; 3 h.

Dr. Peter Mewes, Pharmakologisches i n s t i t u t der Freien Un i ve rs i tE t B e r l i n , Abt . "Embryonal-Pharmakologie", D-looo Ber l i n 33, T h i e l a l l e e 69/73

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INFLUENCE OF AMINOREX (MENOCIL R) ON PULMONARY PRESSURE AND ON THE CONTENT OF BIOGENIC AMINES IN THE LUNGS OF RATS (Der Einfluss von Aminorex auf den Blutdruck im kleinen Kreislauf und den Gehalt an biogenen Aminen in der Rattenlunge). H. Mielke, K.-U. Seller Ursula Stumpf and O. Wassermann

Pulmonary hypertension (H.LHllmann et al., Drug Res,in press) and a remarkable degranulation of mast cells (unpublished observation) result from treatment of rats with aminorex (A). Therefore we looked for a correlation between these two phenomena.- Pulmonary pressure ((PP), right ventricle, closed chest) and the content of serotonin (5-HT), norepinephrine (ICE), and dopamine (D) in lungs- according to Sadavongvivad (Br.J.Pharmac. 38,353,1970) - were measured in the same animal (male Sprague-Dawley rats, chloralose (80 mg/kg) - urethane (iO0 mg/kg) anaesthesia) I0 minutes after a single injection or 24 hrs after chronic treatment (4 weeks) of A (I0 mg/kg/day).- After a single dose of iO mg A/kg or 5 mg 5-HT/kg i-P. both PP and 5-HT content are increased, whereas NE and D remain unchanged. Pretreatment with phentolamine (lO mg/kg i.p.) prevented both effects. Propranolol (0.75 mg/kg i.p.) caused a liberation of 5-HT and enhanced the effects of A.- The pulmonary hypertension due to chronic treatment with A is completely prevented by simultaneous application of methysergid (5 mg/kg/day~ Under these conditions NE and D contents are also increased, which may indicate an additional indirect sympathomimetic action of A without any importance for the pulmonary circulation, since NE infusions show only a transient effect on PP. The A-induced increase of 5-HT in the lungs can be directly correlated to the PP.

Dr.K.-U.Seiler, Institut f. Pharmakologie, D-23 Kiel, Hospitalstr.

INFLUENCE OF METYRAPONE AND HYPOXIA ON NEOPRONTOSIL AND ACET- ANILIDE METABOLISM IN ISOLATED RAT LIVERS (Metabolismus von Neoprontosil und Acetanilid an der isolierten Rattenleber unter dem Einflu8 von Metyrapon und Hypoxie) K. Minck, G.F. Kahl, R. Kern, A. Lier, G. Nissen The interaction of metyrapone(MP)with oxidative and reductive drug metabolism and the oxygen sensitivity of azo reduction were studied in vitro and in isolated rat livers which were perfused with a recirculating hemoglobin free medium under continuous re- gistration of arterial and venous oxygen pressure. In vitro reduction of neoprontosil (NP)was measured continuously at 546 nm in phenobarbital stimulated rat liver microsomes under anaerobic conditions. Activity of azo reductase was decreased by carbon monoxide from 5.93 +0.14 to 1.45 + 0.05 nmoles/mg prot. x min. 1 mM MP did not alter NP reduction, the-activity amounting to 5.78+ 0.67 nmoles/mg prot. x rain. Addition of MP (lmM) to CO inhibited microsomes reT:cti- vated NP reduction up to 2.50+0.13 nmoles/mg prot. x min (P(0.001). In isolated rat livers 0.38+0.05 nmoles of-free sulfanilamide/ml medium x g liver were formed within 35 min under oxygen, the activity was not altered by MP (1 raM). Decreasing arterial oxygen saturation enhanced sulfanilamide production, the concentration in the medium was increased to 1.90+0.12 nmoles/ml x g after 30 rain of anoxia. NP reduction was completely blockecT by CO and was reactivated by 1 mM MP. The activation of acetanilide hydroxylation by MP described in vitro (Leibman, MoI.Pharmacol. 5_z_~ 1, 1969) was confirmed on the level of the surviving organ.- The difference of CO and MP action may possibly elucidate some aspects of the mechanism of the reactions involved.

(Supported by a grant from the Deutsche Forschungsgemeinschaft) Pharmakologisches Institut der Universlt~t, D 6500 Malnz, Obere Zahlbacher Str. 67

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THE INFLUENCE OF VARIOUS CONCENTRATIONS OF EXTRACELLULAR HYDRO- GEN AND CALCIUM IONS ON THE BLOOD PRESSURE EFFECT OF CATECHOLA- MINS (Der EinfluB unterschiedlicher extracellulgrer Wasserstoff- und Calciumionen-Konzentrationen auf die Blutdruckwirkung von Katecholaminen). K. aus der MHhlen~ O. Heidenreich

In anaesthetized rats the blood pressure effects of 2 ~ug/kg noradrenaline and IO 7ug/kg orciprenaline were tested ~urlng metabolic acidosis (pN ~7,05), hypercalcemia (plasma calcium levels 9,5 + 0,2 mequ/l) and metabolic acidosis with simultaneous hypercalcemia. Compared with preceding control periods the systolic an d diastolic blood pressure increase by noradrenaline was diminished by 50 % during the acidotic state. The blood pressure decrease by orciprenaline was also reduced by 73 and 86 % resp. During simultaneous acidosis and hypercalcemia the systolic pressure effect of noradrenaline was only reduced by 26 % whereas the diastolic pressure reached control values. The blood pressure decrease by orciprenaline, on the contrary, was completely

�9 +

abolished in this metabolic state. The antagonistlc effect of H and Ca ions with respect to noradrenaline and the synergistic one with respect to orciprenaline reveal definite differences in the mechanism of action by which these ions influence the circu- latory effects of ~- and ~-sympathomimetics.

Prof. Dr. O. Heidenreich, Abt. Pharmakologie der Med. Fakult~t der RWTH Aachen~ Alter Maastrichterweg 1

INFLUENCE OF VARIOUS SUBSTRATES ON THE RESPIRATION OF THE CHOROID PLEXUS AND BRAIN CORTEX SLICES OF THE RABBIT IN VITRO (Uber den EinfluO einiger Substrata auf den Sausrstoffverbrauch des Plexus chorioideus und von Hirnrindenschnittsn des Kaninchens in vitro) N. Nakayama und D. Voth

The oxygen consumption of both tissueswas comparatively studied by measuring the respiration with the conventional Warburg manometric apparatus. The following substrates are used: D-glucose; D-ribose; D-sorbitol; the sodium salts of succinats~ pyruvate, lactate, glutamate; ~-aminobutyric acid; malic acid; valine. The respiratory rat~ of the plexus exceeds that by brain cortex slices significantly with all substr~tes. The rates wit h glucose and fructose are identical. Pyruvate and lactate yield a much higher rate of the plexus, whereas the unstimulatad respiration of cortex slices was less than that found in the presence of glucose. Succinate increases the respiration of the plexus to a very surprising extent, the rate of oxygen consumption reaches the high value of -48,6 + 2,6 p10~/mg dry weight x hr. The oxygen uptake by cortex sli~es was no~ at all affected in the presence of succinste. The significance of these variations of respiratory rate with succinata as substrata is quite unknown. Possibly succinate takas part in the control of the metabolic activity of the choroid plexus (D. Voth st al., Brain Research (1972) - in press -, D. Voth et el., 3. Hirnforsch. (1972) - in press - ).

Prof. Dr. D. Voth~ Neurochirurgische Universit@tsklinik mainz, D-6500 M a i n z , LanganbeckstraBe I

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DIBUTYRYL-3',5'-AMP -AND 45CA UPTAKE IN ISOLATED RAT ATRIA (Dibutyryl-3', 5'-AMP und 45Ca-Aufnahme an isolierten Rattenvorh~fen) H. Nawrath, T. Meinertz, H. Scholz

6 , The effects of cyclic N -2 -O-dibutyryl adenosine 3',5'-monophosphate (DB-AMP; lo -3 M) on mechanical performance, 45Ca uptake and total tissue Ca concentration were investigated in electrically stimulated (12o beats/min) left atria isolated from female Sprague-Dawley rats (weight 18o-22o g). For comparison, the effects of adrenaline (A; 5.5 x lo -6 M) and theophylline (T; 1.7 x lo -3 M) were studied as well. The experiments were performed at 35~ in Tyrode solution containing o.9 mM CaC~2.~. -Each drug increased contractile force. But neither DB-AMP (exposure to "+3Ca 1-6o mln) nor A and T (incubation time 5 mln) enhanced 45Ca uptake or altered total cellular Ca concentration. - Since A and T, which both increase 45Ca uptake in guinea-pig atria under similar conditions (H. Reuter, Arch. exp.Path.Pharmak. 251, 4ol, 1965; H. Scholz, Arch.Pharmak. 271, 396, 1971), were ineffective in rat atria, it cannot be excluded that in this preparation membrane Ca fluxes possibly may be masked by so far undefined mechanisms. Thus1 the nature of the positive inotropic action of DB-AMP remains to be elucidated. (Supported by a grant from the Deutsche Forschungsgemeinschaft).

Dr. H. Nawrath, Pharmakologisches Institut der Universitdt Mainz D-65oo Mainz, Obere Zahlbacher Str. 67

ABOLITION OF THE APPARENT N~-IMPERMEABILITY OF THE COLON MUCOSA BY OX~fPHENISATINE (Aufhebung der scheinbaren Natriumimpermeabi- liter der Mucosa des Colons dutch Oxyphenisatin) G. Nell~ W. Rummel~ W. Forth and R. Wanitschke

In spite of the steep concentration gradient, there is nearly no nettransfer of Na+ from blood to lumen in tied off segments of rat colon filled with isotonic choline chloride solution. i mg ~ oxyphenisatine (0.) causes a significant increase of the nettransfer of Na + from blood to lumen. Na+ may pass from blood to lumen either on the transcellular or the intercellular route. In order to distinguish between these two possibilities the intracellular concentrations of Na + and K + were calculated from the Na + and K + content in mucosal tissue and the volume of ECS determined by 5iCr-EDTA. In control rats the intracellular concentrations are: 13 • 1.2 ~M/ml Na + and i2i • 5.0 ~M/ml K +. These intracellular ion concentrations were not changed under the influence of 0.. This indicates that Na + takes the intercellular way. 2 min after the i.v. administration of 22Na+ the specific activity of mucosal Na + is unchanged whereas the specific activity of intraluminal Na + increases 2.5-fold under the influence of 0.. Therefore, it can be assumed that O. increases the Na + permeability of the tight junctions.

Dr. G. Nell, Institut f~r Pharmakologie und Toxikologie der Universit~t des Saarlandes, D-665 Homburg/Saar. (Supported by a grant of the SFB 38, membrane research at the University of the Saarland).

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FORMATION OF GUANOSINE-3~51MONOPHOSPHATE IN ISOIATED FAT CELLS OF RATS (Bildung yon Guanosln-3~5Lmonophosphat in isolierten Fettzellen der Ratte) E.-P. Neumann, S. Mocikat, E. BShme The activity of guanylate cyclase (GC), which catalyzes the formation of guanosine-3~5Lmonophosphate (Guo-3~5~P) from GTP, has been measured in rat epidydl- mal fat cells, isolated according to the method ogRodbell (J. Biol. Chem. 239, 375, 1964). Subcellular fractions were prepared according to the method of Angel (J. Lip. Res. ii, 420, 1970). Activity of GC has been d~ermined by measuring l#C-Guo-3~5LPf-ormed from 14C-GTP in the presence of Mn . A GTF regenerating system (creatinphosphate and creatinkinase) was added to the incubation mixture. The enzymatic degradation of 14C-Guo-3~5iP during th? incubation has been reduced by addition of Guo-3~5iP. The enzymatic degradation and the loss of l~C-Guo-3~S!P during the isolation procedure were determined by addition of 3H-Guo-3~5LP. 14C-Guo-3~5LP was separated from 14C-GTP and other labelled products by thin layer chromatography and precipitation by zinc carbonate. Subse- quently the GTP remaining after inkubation was determined. The activity of GC in homogenates of isolated fat cells in the presence of 5 mM Mn ++ is in the range of 20 to 30 pMol/mg Prot./min. The highest activity was measured in the presence of 2 mM Mn ++. The formation of Guo-3~5iP in the presence of Ca ++ or Mg ++ was lower, than with Mn ++. Ca ++ or Mg ++ reduced the effect of Mn ++. Fluoride has no effect on the activity of GC. 75 % of the total Guo-3~5iP formed is found in the I00,000 g sediment. Triton X-IO0 stimulated the formation of Guo-3~5!P in the homogenate and in the 50,000 g supernatant, but not in the sediment. Epinephrine, norepinephrine, ACTH, insulin and PGE 1 do not have any effect on the activity of GC. Formation of Guo-3~5!P is not altered by addition of theophylline or caffeine.

Dr. E. B6hme, Pharmakologisches Institut, D-6900 Heidelberg, Hauptstr, 47-51.

THE EFFECT OF k-STROPHANTINE AND DIGITOXIGENIN ON CONTRACTI- ON FORCE AND CELLULAR CALCIUM EXCHANGE IN ISOLATED GUINEA PIG ATRIA (Die Wirkung von~ k-Strophanthin und Digitoxigenin auf Kontraldi- onskraft und zellul~ren Calciumumsatz i so l i e r t e r MeerschweinchenvorhOfe) E. Noack, E. Heinen

Isolated, e lec t r ica l ly driven (12o b e a t s / r a i n ) a t r i a were incubated 3, 5, lo or 15 minutes in the p resence of cardiac glycosides, the total incubation t ime being always 45 minutes. The ex t race l lu la r space volume (ECS) of each isolated organ was encymatical ly es t imated by means of the i r inulin content. We found a mean ECS of 26.6 ~= o, 4 go (n=25). Non toxic concentrat ions (o. 8 #M) of k-strophanthine and digitoxigenin significantly dec rease the cel lular calc i - um content of isolated a t r ia by 33.5 go respec t ive ly 20.7 go and increase the exchangeable calcium fract ion up to 68.8 go respec t ive ly 48.9 go (control: 31.1 go) a f te r 15 minutes of incubation. Toxic concentrat ions (2.67 resp. 13.4 ~M of k-strophanthine or 2.67 pM of digitoxigenin) develop during the i r initial phase of action the same effect, but in the subsequent toxic phase the ce l lu lar calcium content again significantly increases and the non exchangeable calcium fract ion remains constant. With toxic concentrat ions the re la t i - ve specif ic 45Ca activity was found to be 59. 7 go or 45. o go(with 2.67 /~M of k-s t rophanthine or digitoxigenin)after 15 minutes of incubation. F r o m our r e - sults it may be concluded, that the effects of non toxic concentrat ions of ca r - diac glycosides on the cel lular Ca exchange are mediated by a net increase of Ca-efflux and the effects of toxic concentrat ions by a net dec rease or inhibition of Ca-efflux. Mooren- E. Noack, Pharmakologisches Institut der Universit~t, 4Dfisseldorf, s t r . 5

R83

THE EFFECT OF THEOPHYLL!NE ON CEREBRAL VASCULAR RESISTANCE (CVR) AND ITS REVERSAL BY ADENOSINE (Die Wirkung yon Theophyllin auf den Hirngef~widerstand (CVR) und ihre Umkehr durch Adenosin) G. 0berdSrster, R. Lan$, R. Zimmer

Experiments were performed on 10 canine brains which were isolatedly perfused by donor dogs according to the method described elsewhere (R. Zi~mer et al., Pfl~gers Arch. 328, 332-343, 1971). Arterial P02, pC02, pH and brain temperature were kept constant at normal values. Cerebral perfusion pressure was measured via the basilar artery and total cerebral blood flow (CBF) was moni- tored by cerebral venous outflow. I P - curves were determined with and without i.a. Theophylline infusions (2 rag/mAn). The autoregulation of the cerebral vessels was significantly decreased by Theophylline, whereat Theophylline decreased CVR more at higher per fusion pressures than at lower ones. Correspondingly, at normal perfusion pressure (80 ~ Hg) injected Theophylline (2 rag, i.a. ) showed a significant 15 % decrease in CVR, but at lowered perfusion pressure (40 ~ Hg) asignificant 19 % increase occured. During continuous infusion of Adenosine (I0-# mol/min), which decreased CVR by 23 %, Theophylline again increased CVR by 20 %. Simultaneously, since in all cases Theophylline dilated the extracerebral vessels leading to the brain, perfusion pressure chaoged. Thus the results were possibly influenced by autoregulation. After ex- cludir~ autoregulation, which was calculated from the Theophylline I P - curve~ the cerebral vasooonstricting effects of Theophylline at lowered per~usion pressure and during infusion of Adenosine were again quite similar. Thus the regulation of CBF might be influenced by Adenosine, which up to now has only been found in isohemic brains (B. Deuticke und E. Gerlach, Pfl~gers Arch. 292, 239-254, 1966).

Dr. G. Oberd~rster, Imstitut f~r normale und pathologisohe Physiologie der UniversitY% K~in. Present adress: Institut f~r Aerobio!ogie, 5949 Grafschaft

DOES A SOLVENT DRAG EXIST IN THE LIPOID PART OF THE CELL MEMBRANE? (Gibt es einen ' s o l v e n t drag' im L i p o i d a n t e i l der Zellmembran?) H. Ochsenfahrt and D. Winne

Exper imental data have shown tha t a p o s i t i v e water net f l u x (from lumen to blood) enhances and a negat ive one d imin ishes drug ab- so rp t i on in the ra t jejunum (H.Ochsenfahr t , D.Winne: Naunyn- Schmiedebergs Arch. Pharmak.266,414,1970). By means of a model the s i ev i ng f a c t o r r = I - ~ ( N ~ f s o n , A.A.Hakim: Amer .J .Phys io l .211 , 1137-1146,1966) f o r the ep i t he l i um was c a l c u l a t e d : amidopyr ine 1.94, a n t i p y r i n e 1.96 ( f o r comparison: HTO 1.18, urea 0 .90) . Whi le the r e s u l t s f o r HTO and urea s u f f i c i e n t l y agree w i th the values t h e o r e t i c a l l y expected, the s i ev ing f ac to r s f o r the more l i p o p h i l i c drugs do not. According to t h e o r e t i c a l cons ide ra t i ons (O.Kedem, A .Ka tcha lsky : J. gen. Phys io l .45 ,145-179,1961) a negat ive r e f l e c t i o n c o e f f i c i e n t o, i . e . a s i ev i ng f a c t o r r > 1, fo r n o n e l e c t r o l y t e s should be poss i - ble under the f o l l o w i n g c o n d i t i o n s : water content of the l i p o i d par t of the membrane > 0 and a cons iderab ly h igher a f f i n i t y of drugs than water to the membrane phase. These cond i t i ons being f u l f i l l e d our r e s u l t s are best exp la ined by assuming tha t a (per - haps min imal ) par t of water net f l u x do cross the l i p o i d par t of the membrane, thereby e f f e c t i n g a so l ven t drag on the drug mole- cu les. The s i ev ing f a c t o r can be i n t e r p r e t e d as the r a t i o of so lu - te to so l ven t v e l o c i t y , a s i ev ing f a c t o r > 1 means tha t the drug molecules cross the membrane phase f a s t e r than the water molecules.

Dr. H. Ochsenfahr t , Prof . Dr. D. Winne, Pharmakologisches I n s t i t u t der U n i v e r s i t ~ t , D-7400 TUbingen, Wi lhe lmstrasse 56

R 84

DIRECT AND INDIRECT ACTIONS OF OXYFEDRINE ON CARBOHYDRATE AND LIPID METABOLISM (Direkte und indirekte Wirkungen von Oxyfedrln auf den Fett- und Kohlen- hydratstoffwechsel) D. Onken, P. Neuvonen, E. Westermann

Oxyfedrlne given intravenously to male rats produced a dose-dependent rise in plasma free fatty acids (FFA), glycerol and glucose. On a molar basis oxyfedrine proved to be approximately as active in promoting llpolysis as noradrenallne while hyperglycema was induced only in lo-fold higher doses of oxyfedrlne.

Pretreatment with phentolamlne prevented hyperglycemia without affecting the lipolytlc action of oxyfedrlne. Pretreatment with 13-adrenolytlc agents (propranolol, KI 255, K~ 592), on the other hand, markedly inhibited the lipolytlc response t ~ oxyfedrlne as well as that to noradrenallne. Pretreatment with deslpramine or syrosingoplne markedly reduced the metabolic effects of oxyfedrlne but enhanced those of noradrenallne.

In isolated epidldymal fat pads of rats the llpolytlc potency of oxyfedrine (K A = 1.3 x lo -7 M; Ema x = 11.9 pEq FFA/g x hr) proved to be in the same range as that of noradrenaline. In the presence of 8-adrenolytlcs the oxyfedrlne-lnduced llpolysls in vitro was blocked in a competitive manner.

It is concluded that a significant part of the hyperglycemic action of oxyfedrine is mediated by a release of noradrenallne from adrenerglc nerve endings, while the llpolytlc action oF the drug could be explained by a direct stimulation of 8-receptors in adipose tissue.

Prof. Dr. s Westermann, Instltut fur Pharmakologle der Medizlnischen Hochschule Hannover, 3ooo Hannover, Roderbruchstr. lol

RENAL ELIMINATION OF EASY DIFFUSIBLE DRUGS DEMONSTRATED BY THE EXAMPEL OF SULPHALENE (Renale Elimination leicht diffusibler Pharmaka am Beispiel Sulphalene) H. O~wald

The renal excretion of sulphalene has been studied in dogs employing a single injection technique. S35-sulphalene was injected rapidly into one renal artery together with a glomerular marker (creatinine). Urine samples were collected at 20 sec intervals seperately from both ureters. The urine of the control kidney served for the correction of recirculation.

s3S-sulphalene appears in the urine in two fractions. The maximum of the first fraction appears about 60-100 see earlier than that of creatinine, whereas the maximum of the second fraction appears at the same time as creatinine. The percentage of the injected amount of S35-sulphalene excreted by the kidney is only a thenth of that of creatinine. The experiments in different states of diuresis indicates that sulphalene excretion depends on urinary flow rate and pH. It is assumed that the first fraction is excreted by diffusion and the second by glomerular filtration.

Dr. H. 0Bwald, Abteilung Pharmakologie der Med. Fakultat der RWTH Aachen, 51 Aachen, MTI II., Alter Maastrichter Weg I.

R 85

THE PRESENCE OF ACETYLCHOLINE IN STIPA TENACISSIMA LINNE (Espartograss and its toxicological significance.(Uber das Vor- kommen yon Azetylcholin in Stipa tenacissima Linn& (Espartogras) und seine toxikologische Bedeutung. S. Pallade und H. Antweiler.

The inhalation of dusts from espartograss leads in some exposed workers to dyspnoea and chest tightness.This may be due to the presence in espartograss of substances having bronchoeonstric- ting effects.-The effect of the watersoluble extracts was tested on the guinea-pig ileum.A rapid dose-related contraction,lasting a few minutes and returning to the base line was observed. There was no tachyphylaxia. Pretreatment with atropine abolished the contraction. We attribute these effects to the activity of acetylcholine (ACh)-like substances. The comparison of the log-dose- effect lines of esparto-extracts and ACh shows a similar steep- ness. From these lines a content equivalent to about 0,5/ug ACh per ml extract (corresponding to iO0 mg espartograss) ha@ been calculated.-Additional proof for the ACh content is furnished by the following facts:incubation of esparto-extract with rabbit- serum(cholineesterase)diminishes the contracting action. Treatment of the ileum with eserine leads to higher and prolonged contractions.-Both, esparto-extract and ACh-solution, having similar activity produced no contraction after being heated at iO0~ In industry, under the working condi tio~s with very high dust concentrations it seems possible that the persons who are sensitive to ACh can develop respiratory disorders. A non-specific irri- tant action of dust, with its bronchoconstricting or secretory effect may also contribute to dyspnoea appearing after inhalation of esparto-dust.

EFFECT OF LEUCINE AND ~-KETOISOCAPROIC ACID ON THE INSULINSECRE- TION OF ISOLATED PANCREATIC ISLETS (Wirkung yon Leucin und ~-Ke- toisocaprons~ure auf die Insulinsekretion isolierter Pankreasin- seln) U. Panten~ E. yon Krie~stein~ J. SchSnborn~ W. Poser~ A. Hasselblatt

Since the non-metabolized, but transported leucine analogue 2- aminonorbornane-2-carboxylic acid (BCH) stimulated insulin secretion, Christensen et al~ (Biochim. Biophys. Acta 241, 341 (1971) suggested that amino acids trigger insulin release by binding to transport molecules. In pancreatic islets of obese-hyperglycemic mice perifused without substrate additon of L-leucine (io mM) or (• (18,4 mM) induced insulin release with an initial over- shoot. Simultaneously both substances brought about a prompt twophase increase of fluorescence of reduced pyridine nucleotides (PNH). 5 mM glucose changed the kinetics of insulin release and PNH-fluorescence induced by L-leucine or (+)b-BCH in a strikingly similar way. io mM ~-ketoisocaproic acid (~-KIC) induced a marked stimulation of insulin release, which was accompanied by an over- shooting increase of the PNH-fluorescence.

These results are interpreted that metabolic events in the B-cell may be the first steps in the chain of events leading to insulin secretion.

Priv.-Doz. Dr. U. Panten, Institut fur Pharmakologie und Toxi- kologie der Universit~t, D 34 GSttingen, GeiststraBe 9

R 86

E F F E C T S OF BENZYDAMINE-HCI ON THE ADENYL CYCLASE SYSTEM (Beein- f lussung des Adenylcyc lase -Sys tems dutch Benzydamin-HC1). H . - D . Pe t e r s , V. Dinnendahl, K. Karze l , P .S . SchSnhSfer

Benzydamine-HC1 was found to reduce mucopolysacchar ide synthesis in f ibroblas t t i s sue cul tures and this reduct ion was par t ia l ly r e v e r s i b l e by d ibu tyry l -A-3 ' : 5 ' - M P . There fo re , the effect of this compound on the adenyl cyc lase sys tem of i sola ted epididymal fat ce l l s was tes ted. Benzydamine does not influence the A T P - l e v e l s of the intact ce l l s . Accumulat ion of A - 3 ' : 5 ' - M P is s t imula ted in a na r row dose range of 3 x 10 -4 M, even though l ipolysis e l ic i ted by 10 -6 M norepinephrine or 3 x 10-31VI d i b u t y r y l - A - 3 ' : 5 ' - M P is dose-dependent ly inhibited. The inc rease in A - 3 ' : 5 ' - M P accumulat ion can be ex - plained by a compet i t ive inhibition of phosphodies terase (K i = 1.1 x 10-3M) which appear to be s i m i l a r to that of theophylline (K i = 8.7 x 10 -4 M). The inhibition of l ipolysis appears to be due to effects on the protein k ina se / t r i g lyce r i de l ipase sys tem of the fat ce l l , even though the binding of A - 3 ' : 5 ' - M P to the regula tory s i te of musc le protein kinase was not affected by the compound at ant i - l ipolyt ic doses .

Dr . H . - D . Pe t e r s , Pharmakolog isches Insti tut de r Rheinischen F r i e d r i c h - W i l h e l m s Univers i t~t , D-53 Bonn, R e u t e r s t r a s s e 2 b

EFFECT OF METHOTREXATE ON THE ACTIVITY OF FOLIC ACID REDUCTASE IN RAT LIVER AND EMBRYOS (Effekt yon Methotrexat auf die Aktivit~t der Fols~urereduktase in Rattenlebern und Embryonen) pi~loslewicz, E. and K~h!er, E.

Despite the high rate of cell division an almost constant FH 2 reductase activity (5.4 * 0.5 m~moles NADPH+H § x mg protein -I x min -I) could be measured in embryonic Tissue during the late phase of organogenesis (day 11-14). So the FH 2 reductase does not seem to be rate limiting in DNA synthesis. Kinetic studies did not give any indication that the FH2 reductase has different properties in embryonic and adult tissues. The Ki value for methotrexate is 1.3 x 10 -11M for the embryonic enzyme as well as for the enzyme which is localised In the hyalo- plasmic compartment of adult livers. The amount of methotrexate reaching the embryo after pretreatment of mother animals apparently changes during the phase of organogenesis. Before the placenta starts functioning the FH2 reductase of the embryo is strongly inhibited by low doses of methotrexate, but rapidly resyntheslsed subsequently. The decidual tissue is even more sensitive to the drug and the FH 2 reductase of this tissue is resynthesised to a much smaller degree. After functioning of the placenta the trophoblast epithelium represents a barrier which can only be overcome by high doses of methotrexate (2.4 mg/kg and more). Before the FH 2 reductase shows any inhibition in embryonic tissue, the fetuses die due to a complete break-down of the placental function.

This work was supported by the Deutsche Forschungsgemeinschaft (SFB 29).

Prof. Dr. med. E. K6hler, Pharmakologisches Instltut der Freien Universit~t Berlin, Abt. "Embryonal-Pharmakologie", D-IO00 Berlin 33, Thielallee 69/73.

R87

THE IN YIVO INCOOPERATION OF RADIOLABELLED CHOLINE INTO THREE ACETYLCHOLINE FRACTIONS OF THE RAT BRAIN IN TOTO AND OF DIFFERENT BRAIN AREALS (Der in vivo Einbau von radiomarkiertem Cholin in drei Acetylcholinfraktionen des Rattenhirns in toto und in verschiedenen Hirnregionen) O. Pleul and H. Kewitz

I. v. administered 14C-Choline enters the acetylcholine (ACh) of the CNS within a few minutes. From the kinetics of the specific radioactivities of choline (Ch), free, labile bound, and stable bound ACh it can be concluded, that I. the transfer rates of the 3 ACh fractions are different, 2. at least 2 Ch fractions are present, which are labelled via the blood stream to different extends, 3. stable bound ACh is synthetisized from the higher labelled Ch pool free ACh from the lower labelled Ch pool and labile bound ACh originates from the compartment of free ACh, 4. the transfer rates of free and labile bound ACh are almost 20 times higher than that in the stable bound compartment, 5. the transfer rates of the fractions are equal in brain stem ganglia and cortex inspite of considerable differences in the size of the pools.

Dr. Otto Pleul, Institut fGr Klinische Pharmakologie der Freien Universit~t Berlin, D-IO00 Berlin 45, Hindenburgdamm 30

INHIBITION OF HEPATIC GLUCOSE PRODUCTION BY TREATMENT WITH GLI- BENCLAMIDE(HB 419,EUGLUCONn)(Hemmung der hepatischen Glucosebil- dung dutch Vorbehandlung mit Glibenclamid) W.Poser~J.SchSnborn, E.v. Kriegstein~U.Panten~A.Hasselblatt

Inhibition of hepatic glucose output after injection of hypoglyc- emic sulfonamides is well established. To further investigate this problem livers from fasting rats(80-100 g b.w.) were perfused without recirculation. The perfusion medium consisted in Krebs-Henseleit-buffer containing 10 mM L-lactate plus I mM pyruvate. After 30-40 minutes the~livers produced 3.6-4.2tumoles glucose x (100 g b.w.)- x min -~, When tolbutamide(0.5 m~) was added, a small(~5%) but reproducible decrease of glucose production was observed.

When donor rats had been treated for 12-14 days with glibenclamide(l mg/kg in the morning, 2 mgs/kg in the evening) their livers produced 20-43% less glucose and 42-57% less ketone bodies than those from controls. The tissue contents of malate, P-pyruvate, 2-P-glycerate and 3-P-glycerate were significantly(2P,0.05) lowered. These effects could be abolished by addition of albumin bound oleate(0.8 mM) to the perfusion medium. We therefore conclude that the observed effects are of minor importance in the living animal, when the liver is well supplied with fatty acids from adipose tissue.

Dr.W.Poser Institut fGr Pharmakologie und Toxikologie der Uni- versit~t 54 G~ttingen Geiststr.9

R 88

THE INFLUENCE OF INHIBITORS ON THE CONFORMATION OF ACETYLCHOLIN- ESTERASE (Einfluss von Hemmstoffen auf die Konformation der Ace~yl- cholinesterase) J. Preuner

Based upon the short report of R.J.KITZ and L.T. KREMZNER (Mol. Pharmacol. 4, 104-107, 1968), a detailed study of the conformation c~ acetylcholinesterase (ACHE) has been carried out measuring optical rotatory dispersion (ORD) and circular dichroism (CD). To stabilize the enzymatic activity all measurements were made in the presence of human albumin (0.i mg/ml) at pH 7.5, phosphate buffer 0.015 M. Under these conditions the enzyme displayed predominantely ~-heli- cal structure. Increasing the ionic strength over 0.6 altered the enzyme tertiary structure with two respects: the amplitude of the three ellipticity bands at 220,208 and 190 nlu decreased by a factor of 1.8 to i. 9 and the band at 208 r~u became more pronounced in comparison to the band at 220 m~. The enzymatic activity seems to be related to the discrete alteration of the band at 208 m~, probably indicating a particular side chain chromophore. As shown by Blout et al. (in"ORD and CD in Organic Chemistry",Ed. G. Snatzke, 1967, 224-300) the band at 208 n%u can reflect the superposition of a side chain chromophore. Independent of the original state, addition of DFP(IO-SM) transferred the enzyme into the monomeric form, indicated by a decrease of the amplitude. Furthermore a distinct reduction of the band at 208 nlu occured. The same "specific" reduction can be observed with edrophonium (3-10-#M~but without influencing the equilibrium between tetramer and monomer. The results suggest an interdependence of the ellipticity band at 208 m~ and the enzymatic activity.

Dr.J.Preuner, Institut f. Pharmakologie, D-23 Kiel, Hospitalstr.

EFFECT OF (4-CHLORO-O-TOLOXY)ACETIC ACID (IVEPA) C~ CATILE AND

RATS (Zur Wirkung yon (2-Methyl-4-chlorphenoxy)-Essigs~iure (MCPA) bei

Rindern und Ratten) W. Raake.

Es wird ilber eine Vergiftung yon 14, zum Tell graviden Kg~nnen im Alter yon 2, 5 Jahren und Untersuehungen an Ratten mit BANVEL ''~' (34% MCPA; 3% 3, 6-Dichlor-2-methoxybenzoes~ure) berichtet. Die weitgehend therapie- resistenten Vergiftungssymptome bestanden in Speicheln, Anorexie, Bewe- gungsstSrungen, Muskelsehw~ehe, Parese der Nachhand, blutig-sehleimigem Nasenausflu~, LungenSdem, Bradycardie, Pansen-und Darmatonie, Hiima- turie, St6hnen und progressiver Apathie. Auffallend war eine lange Rekon- valeszenzzeit mit 18-t~gigem Festliegen. MCPA ist naeh oraler Aufnahme wenigstens acht Tage fang im Rinderharn nachweisbar. Der Nachweis wird dureh Gefrieren des Hams (-20~ nicht gestSrt und erfolgt d(innschiehtehromatographisch (Methanol/Benzol 5/95; Brornphenolblau zur Entwicklung) und spektrophotornetrisch (Absorptions- maximum bei 278 nm) aus der sauren ~therfraktion naeh STAS-OTT(~_Im Rattenharn wurde sechs Tage naeh oraler Applikation von BANVEL-M'II)eir~ Linksverschiebung des MCPA-Spektrums urn 6 nm (Absorptionsmaximum bei 272 nm) beobachtet, die einern Metaboliten von MCPA zugesehrieben

wird.

Dr. W. Raake, Institut fiir Pharrnakologie, Toxikologie und Pharrnazie der Tier~rztlichen Fakult~it, 8000 Mfinehen 22, Veterin~rstr. 13.

R 89

METABOLIC AND VASCULAR RESPONSE TO IoA. ADENOSINE INFUSIONS IN DOG HIND LIMB (Stoffwechsel- und Gef~Btonusver~nderungen in Hund~ extremit~ten bei i.a.Infusionen von Adenosin)G.Raberger~M.Weissel

The investigations were carried out on 19 anesthetized mongrel dogs. In one set of experiments (8 dogs) femoral blood flow was measured electromagnetically and art. and fem. yen. 09-content (AO-Oxymeter),pCO2,PH,glucose~lactate,pyruvate,FFA~3-~ydroxybuty- rate,acetoacetate,and glycerine were determined before,during,and after a 5 mino infusion of ioo ~g/kg/min adenosine into the fem. art.In 9 dogs blood flow was measured electromagnetically and art. and fem~ ven. O^- and COg-content were measured volumetrically (Van Slyke).In ~ dogs bl~od flow was measured as femoven, outflow with a t-tube and the O~-content was determined using the AO-Oxy- meter. In all experiments the hind limb exhibited a reduced O^- uptake during the adenoslne infuslon together wlth a 5 to 6 fold increase in blood flow. At the same time an increased amount of CO 2 was released thus leading to RQ values up to 5. This amount of CO 2 cannot be derived from aerobic metabolism and is indicative of an additional intracellular formation of acid products.The increase in glucose uptake together with the increased lactate production indicate an enhanced glycolysis. But part of the decrease in Og-uptake could also be caused by a decreased FFA oxidation re flecked in a release of FFA. Since the flow changes parallel the changes in H + and CO^ release blood flow regulation in the canine hlnd l lmb seems to be primarily metabolically controlled.

Dr. G. Raberger, Pharmakologisches Institut der Ruhr-Universit~t Bochum, D 463 Bochum, Im Lottental.

ON THE TIME COURSE OF THE CHANGES IN ACTION POTENTIALS AND CON- TRACTILE AMPLITUDE AS INFLUENCED BY DIFFERENT CARDIAC GLYCOSIDES (Zeitliche Ver~nderungen der Aktionspotentiale und der Kontraktions- amplitude unter dem Einfluss verschiedener Herzglykoside) U.Ravens

In isolated papillary muscle of the guinea pig, 7 x IO-7M ouabain caused a maximum increase in contractile amplitude (CA) of 372 29% of the control value. The action potential, as recorded with conventional intracellular microelectrodes, decreased in duration from 149 ~ iO to 16 ~ 1 msec. The time course of the changes in each parameter was well described by an exponential function, however, the two half life times differed considerably. The increase in CA reached its maximum after 50 min., having a t 1/2 of 12,5 min., whereas the action potential duration (APD) continued to decrease until the end of a 120 min. period of glycoside action, yielding a t 1/2 of 20 min.

Furthermore, we studied digoxin, digoxigenin bisdigitoxosideand digoxigenin monodigitoxoside, of which the latter glycoside showed the most striking difference compared to ouabain. It caused the same augmentation in CA as ouabain, but the decrease in APD was smaller. Also, the half life times were different, i.e. 8 min. for the mechanical and 30 min. for the electrical activity. The results indicate a greater therapeutic range of digoxigenin monodigitoxoside as compared to ouabain.

Dr. Ursula Ravens, Institut f~r Pharmakologie der Universit~t Kie~ D-23OO Kiel, Hospitalstr. 4-6

R 90

NEURO- AND NEPHROTOXIC EFFECTS OF DICHLOROACETYLENE (Neuro- und nephrotoxische Wirkungen von Dichloroacetylen) D.Reichert, G.Brinke, P.Bannasch, G.Liebaldt

Using a dosage technique for dichloroacetylene (DCA) which was newly developed by us we successfully determined the cause of death and the functional and morphological pattern of lesions in experimental series of rabbits and rats following single (i hr. and 6 hrs.) and repeated daily exposures (up to 14 days). The neurotoxic action was in accord with clinical findings in man, with sensory nerve cells in particular exhibiting high-grade ti- grolysis with Nissl sintering and partly the state of primary ir- ritation. The individual brain nerve nuclei showed a characteristic pattern of lesions. The facial and oculomotor nerves, followed by the motor trigeminal nerve, were affected to a far lesser degree than was the sensory trigeminal nerve. Maximal damage was observed 2 weeks after exposure. Single 1-hr. exposure of rabbits to 200 ppm led to irreversible renal damage with extensive tubular necroses over the entire cortex. Clinico-chemical findings revealed a pronounced increase in nonprotein nitrogen values. Acute renal failure invariably ended in fatality. Fol- lowing sublethal doses only residues of necrotic tubuli and thickening of peritubular basal membranes were observed after 4 weeks, progressive scarring and marked increase of interstitial fibers appearing after 3 months. In toto renal damage appears to be clinically more significant than neurotoxic lesions.

D.Reichert, Institut f~r Pharmakologie und Toxikologie der Universit~t, D-87 Wfirzburg, KoellikerstraSe 2

THE INFLUENCE OF TEMPERATURE AND METABOLIC INHIBITION BY IODO- ACETIC ACID (IAA) ON THE SENSITIVITY OF THE ADRENERGIC r AND B-RECEPTORS IN DIFFERENT ORGANS D. Reinhardt~ H.J. Schdmann, J. Wagner

The influence of temperature and IAA, resp., on the effect of isoprenaline (IPN) and the rescrcin derivatives Th ll65a and orciprenaline (OPN) was studied on isolated rabbit ileum, electrically driven atrium and trachea of the guinea-pig. The experiments were^carried out in Krebs-Henseleit solution at 25, 33, 37 and 42 v C. The elevation of temperature shifted the dose-response curves of all B-agonists on ileum as well as on atrium to the right whereas on trachea the dose-response curve of IPN showed only a small shift to the right. The affinity -calculated as pD 2 values- of Th ll65a and OPN, resp., was not influenced. The shift to the right observed on ileum and atrium could be reversed by metabolic inhibition induced by IAA. On the tracheal B-receptors IAA was unable to change the affinity of the sympathomimetic amines. The e-receptors of the ileum remained unaffected either by temperature alteration or by IAA. These results show the different responsiveness of ~- and B- receptors to metabolic changes. The variable sensitivity of the B-receptors of different organs is possibly dependent on their metabolic rate.

Dr. D. Reinhardt, Pharmakologisches Institut der Ruhr-Universi- t~t, 43 Essen, Hufelandstr. 55

R 91

MODIFICATION OF THE ANTINOCICEPTIVE EFFECT OF MORPHINE BY DRUGS AFFECTING BIOGENIC AMINES IN RAT BRAIN. (Beeinflussung biogener Amine im Gehirn und antinociceptive Wirkung von Morphin an der Ratte). K. Reinhold t J. Bl~si~ and A. Herz. A number of investigations show that the antiuociceptive effect of morphine is changed by drugs which affect central biogeuic amines but the results are conflicting. However, the actual changes in amine content have been measured only in a few studies. In the present investigation, vocalization elicited by electrical stimulation of the tail root in rats was used as a nociceptive reaction. Noradrenaline (NA), dopsmine (DA) and serotonin (5-HT) in brain tissue were estimated fluorometrically. 75 % depletion of brain 5-HT by p-chlorophenylalanine as well as increase in 5-HT after administration of 5-hydroxytryptophan up to 200 % did not alter the morphine effect. Depletion of catecholamiues by C~-methyltyrosine (OC-MT) resulted in a sharp diminution of the morphine effect when NA content dropped below 30 %. Selective depletion of NA by diethyldithiocar- bsnmte and FLA-63 also resulted in a reduction of the morphine effect only when NA dropped below this level. Destruction of catecholamine-con- tainiug terminals by intraveutricular injection of 6-hydroxydopamiue also resulted in a reduced morphine effect. 5,6-dihydroxytryptamine which mainly destroys 5-HT-containing terminals did not apparently change the morphine effect. Intraventricular injection of NA in rats depleted of NA by O~-MT did not restore the morphine effect. The results suggest that NA is involved in the mechanism of the antinociceptive effect produced by morphine as tested by the vocalization reaction in rats. A role of DA cannot be excluded while the participation of 5-HT does not seem important. Dr. K. Reinhold, Max-Planck-Institut fGr Psychiatrie, D-8000 Mdncheu 23, KraepelinstraBe 2, Germany

METABOLISM AND PHARMACOKINETICS OF 4'''-METHYLDIGOXIN IN MAN (Stoffwechsel und Pharmakokinetik yon 4'''-Methyldigoxin beim Menschen) H. Rennekampt C. Rennekampr K.v. Ber~mann ~ U. Absha~en~ N. Rietbrock

In 10 patients without cardiac and renal deseases the pharmaco- kinetical behavior and the metabolism of 12~-3H-4'''-methyldigoxin (MD) after intravenous and oral administration were studied. About 15 minutes after oral administration maximal plasma c~centrations were observed which are equivalent to the plasma levels after intravenous administration. The biological half life of radioactivity in plasma averaged 43 hours. After 7 days 31.5 ~ 6.3 and 32.5 ~ 5.0 per cent of the dose were found in feces and '52.9

1.8 and 59.7 ~ 1.3 per cent were excreted in urine after oral and intravenous administration respectively. These results suggest a rapid and almost complete absorption of 4'''-methyldigoxiru

In man in contrast to rats the main metabolic pathway of MD is a demethylation to digoxin (D), which increases during the time of investigation. Apart from D only small amounts of bis- and mono-glycoside could be detected. The relative amounts of CHC13- insoluble metabolites in plasma and urine were highest during the first 8 hours and decrease rapidly, so that after 24 hours only minimal amounts of these metabolites can be found.

Prof. Dr. N. Rietbrock, Institut fur Klinische Pharmakologie der Freien Universit~t Berlin, D-IOO0 Berlin 45, Hindenburgdamm 30

R 92

CLEAVAGE OF GLYCOSIDIC BONDS AND FORMATION OF POLAR METABOLITES OF DIGOXIN AND ITS DERIVATES IN RATS (Spaltung der Zuekerkette und Bildung polarer Metabolite yon Digoxin und Derivaten bei der Ratte) N. Rietbrock t K. yon Ber~mann~ U. Absha~en Female rats weighing 200 g received DH-Digoxin (D), 3H-4'' '-acetyldigoxin (AD) and 3H-4"'-Methyldigoxin (MD) intraduodenally via bile duct. Bile and urine were collected for 12 hours, the glyc~ sides extracted with CHCI5 and chromatographed on formamide im~ hated cellulose thinlayer plates (solvent system CHCI~). After administration of D, digoxigenin-bis and mono-digitoxoside are excreted in bile and urine. No essential differences of metabol~ pattern in bile and urine exist between D and AD. The latter compound has not been completely desacetylated~ therefore small amounts of AD were detected in urine and bile (0o3-1,2 % of the dose). MD is slowly demethylated to D in the rat and excreted to small extent with bile, Therefore MD must be metabolized to the bis- and monoglycoside, which are rapidly eliminated in bile. About 70 % of CHCl3-soluble fraction consists of bis- and mono- ~lycoside, the remaining activity corresponds to D and MD. CHC13-insoluble metabolites as possible conjugates of glucuronic and sulfuric acid, were incubated with ~lucuronidase and sulfa- tase. After extraction with CHCI 3 the metabolites were separated by TCL on silica gel (solvent system Aceton/CHCl 3 1:1). In both cases D, bis-, monoglycoside and genin were detected. The CHCI~- insoluble fraction is observed mainly during the early period of elimination and possibly represents direct conjugates of the steroid moiety in 12 or 14 position. Prof. Dr. N. Rietbrock, Institut fur Klinische Pharmakologie der Freien Universit~t Berlin, D-1000 Berlin 45, Hindenburgdamm 30

EFFECT OF HALOTHANE ON THE HEPATIC DRUG METABOLIZING SYSTEM OF THE RAT (ZumEinflu~ yon Halothan auf das arzneimittelabbauende Enzymsystem in der Rattenleber) I.Rietbrock, G. Lazarus und E. Richter

Wiederholte einst~ndige Expositionen in 2 Vo1% Halothan an 5 aufeinanderfolgenden Tagen f~hrten bei 200 g schweren weiblichen Ratten 3 Tage nach der letzten Begas%mg zu einer maximalen Ver- k~rzung der Hexobarbitalschlafzeit yon 49 %.Verbunden war damit ein signifikanter Anstieg der Aktivit~t der NADPH-Cytochrom c Reduktase pro g Leber um 33 % (p<O,OOO5).Der Gehalt an Cytochrom PA~ pro g Leber blieb im Bereich der Norm.Wurde die Halothan- n~ose auf 5 mal je 2 Stunden tgl.verl~ngert,so war zwar eine weitere Erh6hung der NADPH-C.c R.-aktivit~t auf 60 % zu beobach- ten,jedoch keine st~rkere Verk~rzung der Hexobarbitalschlafzei~.

Dutch Vorbehandlung mit Trifluoressigs~ure ( 5 mal 15 mg/kg p.os) wurde bei weiblichen Ratten weder die Hexobarbitalschlafzeit verk~rzt,noch der Gehalt an Cytochrom P45o bzw.die Aktivit~t der NADPH-C.c R. pro g Leber ver~ndert. Eine anschliessende Halothanexposition yon 5 mal 2 Volt tgl.~ber 2 Stunden hatte zwar auch bier noch eine Schlafzeitverk~rzung zur Folge,die jedoch mit 22 % nicht soausgepr~gt war.Ferner war die Aktivit~t der NADPH-C.c R.nur auf halb maximale Werte angestiegen.

Demnach wird mSglicherweise die induzierende Wirkung des Halo- thans durch Trifluoressigs~ure gehemmt.

Dr.Ingrid Rietbrock,Abteilung f~r Anaesthesiologie der Univer- sit~t W~rzburg,87 W~rzburg,Josef-Schneider-Str.2

R 93

EFFECT OF N-BUTYL-BIGUANIDE ON SUGAR AND AMINO-ACID TRANSPORT IN INTESTINE AND KIDNEY (Die Wirkung yon n-Butyl-Biguanid auf den Dthandarm- und Nierentranspor• yon Zuckern und Aminos~uren) J.W.L. Robinson und A.L. Luisier

Der aktive Transport yon Aminos~uren und Zuckern in intestinalsn Ringen yon Mserschweinchen und Ratten wurde dutch hohe Konzen- trationen yon n-Butyl-Biguanid w~hrend langer Inkubationsperio- den gehemmt. Ebenso wie Strophanthin harts dieser Hemmstoff kei- hen Einfluss auf die Anfangsgeschwindigkei% des Transportes. Andererseits ftthrte die Vorinkubation des Gewebes in Anwesenhei% yon 8trophanthin sder Biguanid ~u einer Hemmung der Anfangs- geschwindigkei% yon Zucker- oder Aminosiureresorption wihrend einer nachfolgenden Inkubation. Diese Ergebnisse lassen vermuten, dass Biguanid eine Wirkung auf den Natrium-Gradiente%ha%.

In Nierenrindenscheiben yon Hunden wu~den die Zucker- und Amino- s~uretransporte teilweise dutch Strophanthin oder Ethacryns~ure, jedoch vSllig dutch eine Mischung dieser beiden S%offe gehemmt. Diese zwei Substan~en wirken anscheinend auf verschiedene Na- %rium-Pumpen. Auch n-Butyl-Biguanid steil% einen Teilhemmer dar, abet sein Einfluss vermehr% nut den yon Strophanthin, abet nich% den yon Ethacryns~ure. Diese Resultate kSnnten darauf hinweisen, dass Biguanid die e%haoryns~u~e-empfindliohe Na%rium-Pumpe beeinfluss%.

Dr J.W.L. Robinson, D~par%emen% de Chirurgie exp~rimentale, HSpi%al Cantonal, Lausanne, Suisse.

ANALYSES OF THE INHIBITION OF ADENOSINE UPTAKE BY CORONARY DILATING DRUGS (Analyse der durch Coronardilatatoren verursachten Hemmung der Adenosinaufnahme) H. Roos and K. Pfle~er

The uptake of adenosine into erythrocytes of guinea pigs takes place by two different mechanisms (H.Roos and K.Pfleger, Mol Pharmacol. July, 1972). One uptake mechanism is saturable and can be inhibited by dipyridamole, hexobendine and papaverine. The other mechanism occurs by diffusion, is proportional to the concentration and cannot be inhibited. Taking these two facts into consideration it is possible to characterize the type of inhibition, because interpretation of Lineweaver-Burk plot is only applicable when the diffusion component is subtracted from the total uptake. From the intercepts of the lines with the ordinate it is to conclude, that the inhibition of adenosine uptake by dipyridamole and hexobendine is competitive, but is non competitive for papaverine. These statements are statistically significant and ascertained by the method of least squares. Experiments performed with isolated guinea pig hearts led to the same results.

H. Roos, Institut fdr Pharmakologie und Toxikologie der Uni- versit~t des Saarlandes, D-665 Homburg/Saar~ (Supported by DFG)

R 94

TITRATION OF CATALYTICALLY ACTIVE CENTERS OF RAT HEPATIC MICROSOMAL CYTOCHROME P-450 WITH METYRAPONE DURING N-DEMETHYLATION REACTIONS (Titration katalytisch aktiver Zentren yon Cytoehrorn P-450 aus Rattenlebermikrosomen mit Metyrapon bet N-Demethy- lierungsreaktionen) I. Roots und A. G. Hildebrandt

Catalytically active centers (E+) of microsomal cytoehrome P-450 and their turnover number (TN) are titrdted with metyrapone (2-methyl-l,2-di-3- pyridyl-l-propanone) during N-demethylation reactions. A method of MYERS (Bioehem. J. 5_!1,303) can be applied provided that inhibitor and enzyme are present in similar amounts ("mutual depletion system", WEBB 1963) and that the reaction is noneompetively inhibited. Since under these conditions LINE- WEAVER-BURK kinetic does not apply�9 K. has to be obtained by determining

�9 "i . . .

the concentration of inhibitor which ylelds 50 % inhlbitlon (It.~o). Since It50 = K. + 0.5 x E~ the true dissociation constant of the enzyme iflhlbitor comi~lex,

I . . .

K. can be o~tamed but also E. and TN. Using amlnopyrme as substrate and mlcrosomes from phenobarbltal pretreated rats 1 mg of profeln contains 4.6 nmoles of E i.e. about 2.2 catalytically active centers per molecule of cyto-

�9 -l

chrome P-4~0. TN = 3.2 rain K. = 2.3 ,uM. Comparing PB-pretreated rats �9 �9 1 .

and controls reveals wlth ethylmorphme a~ substrate that K. and TN remain i .

constant but that E t raises 2.3 fold. The values obtained are in good agre~ent with spectrophotometrically obtained data and relate binding of metyrapone to rnicrosomal eytochrome P-450 to its inhibitory action (Arch. Biochem. Biophys. 145, 531 (1971)). I. Roots, Institut fiir Klinische Pharmakologie der Freien Universit~t D- I000 Berlin 45, Hindenburgdamm 30

ON A STRIKING PHAR/MACOKINETIC DIFFERENCE BETWEEN CHLORPHENTERMINE AND PHENTERMINE (0bet das unterschiedliche pharmakokinetische Ver- halten yon Chlorphentermin und Phentermin) E. Rossen, K.-U. Seiler and O. wassermann

After chronic treatment of rats with phentermine (Ph) and chlorphentermine (Ci-Ph) only Cl-Ph causes pulmonary hypertension and severe pathological alterations in the lungs of rats (enormous infiltration of macrophages "foam cells", G. Franken et al., Drug Res.2o, 416,1970). This finding prompted us to study the pharmacokinetics of Cl-Ph and Ph during chronic treatment (8 weeks) in rats (male Sprague-Dawley rats, 10 -4 moles Ci-Ph or Ph/kg, i.p., 6 x weekly, 3H-labelled).-Tissue-blood-quotients (t/b) were obtained for 8 organs. For Ph they were in the range of 3-7 and remained constant during 8 weeks.-After CI-Ph t/b-values were higher already in the first week. Thereafter, however, a sharp and unpro- portional rapid increase of t/b occurred particularly in lungs (40-160) and adrenals (20-85). To a lesser degree this tendency was visible in all organs.- Kinetically this process accounted for a Cl-Ph-induced formation of additional "binding sites" for CI-Ph. As to the lungs the time course of this phenomenon was closely correlated to the appearance of macrophages. These cells are extremely rich in phospholipids (lamellar bodies) and can provide a compartment of high binding capacity for Ci-Ph.- All the changes observed are reversible. According to our results Ci-Ph is able to induce a lipoid storage desease in rats and other species.

Priv.-Doz.Dr.O.Wassermann, Institut f[tr Pharmakologie der UniversitMt Kiel, D-23OO Kiel, Hospitalstr. 4-6

R 95

THE INFLUENCE OF ATROPINE ON THE RESPIRATION OF THE RAT IN DIFFE- RENT TYPES OF INTRACOLLICULAR DECEREBRATION. (Wirkungen yon Atro- pin auf die Atmung der Ratte bei verschiedenen Formen intracolli- cul~rer Decerebrierung.) Y. Sakai~ H. dal Ri~ G. Schmidt

The Impuls frequency of phrenic motoneurons of the rat was diminished after intravenous injection of 1.0 mg/kg atropine. This action was paralleled by a decreased number of impulses per cycle. In contrast the frequency of respiration was increased. There was no change of the described action of atropine when the carotid sinus was denervated. N-methylatropine, which hardly penetrates into the central nervous system, was without any effect. The results of experiments on decerebrated animals support the assumption that atropine influences a suprapontine cholinergic mechanism: When a total decerebration at the level of the colliculus superior was performed the effect of atropine was strongly diminished. When the medial and ventral parts where spared however, the effect of atropine was unchanged. Thus it seems to be possible that a suprapontine influence which promotes the discharge frequency of phrenic motoneurons is abolished by atropine.

G. Schmidt, Institut fur Pharmakologie und Toxikologie, Abto Neu- ropharmakologie, D-3400 G~ttingen, ~Geiststr.9

INACTIVATION OF LH-RH BY THE PLASMA OF VARIOUS SPECIES (Inaktivierung yon LH-RH dutch das Plasma verschiedener Tierarten) J. Sandow F. Enzmann T H.G. SchrSder~ and H.G. Vogel

The releasing hormone LH-RH is not species specific in its effect on LH and FSH-release. Isolation and purification of the natural hormone is a laborious and expensive procedure. The structure of porcine LH-RH has been elucidated by Schally and coworkers. There is no definite knowledge e.g. about the structure of ovine LH-RH, it is probably identiQal with cr very similar to the procine. To look for differences in the duration of the physiological effects of LH-RH, synthetic hormone was incubated with plasma of various species. The residual activity was tested in ovariectomized, steroid-pretreated rats. The inactivation curve was compared with the time course of plasma-LH following a single injection of LH-RH. A rapid inactivation phase of 2o-3o min. was found with plasma of the rat, dog, guinea pig, and after incubation in human plasma. Inactivation by plasma of proestrus sheep was less rapid. After up to 2 hours of incubation, there was still a considerable amount of residual LH-releasing activity as shown by the LH increase in 0EP-rats. This may be due to residual activity of LH-RH fragments, or to the fact, that LH-RH is not, as is the case with TRH and MIF (MIH), predominantly inactivated by plasma. The different inactivation rate in rat and sheep plasma could be related to the stage of estrus cycle in sheep, or to chemical characteristics of ovine LH-RH. There is a good coincidence between the delayed time course of LH-release in sheep, and the low inactivation rate.

Dr. J. Sandow, Farbwerke Hoechst AG., Frankfurt 80, Abt. fGr Pharmako- logie H 821.

R 96

LH-RELEASE BY A SYNTHETIC DECAPEPTIDE LH/FSH-RH (LH-Freisetzung durch ein synthetisches Decapeptid) J.Sandow~ A.V.SchallE~ T.W.Reddin~ W.Heptner~ and H.G.Vogel

The chemical nature of the releasing hormone of the luteinizing and follicle stimulating hormones (LH/FSH-RH) was elucidated in 1971 by Schally et al. The decapeptide corresponding to the natural hormone was synthetized by Geiger et al. at Farbwerke Hoechst AG. The effect of the synthetic decapeptide on the release of LH has been studied by in vivo and in vitro methods. (I) Release of bioassayable LH was induced in ovariectomized, steroid-pretreated rats, LH was assayed by ova- rian ascorbic acid depletion in immature rats pretreated with gonado- tropins (Parlow-assay). (2) Release of immunoreactive LR (RIA-LH) was measured by radioimmunoassay (double-antibody technique of Niswender et al.) after iv., sc., and im. injection of the decapeptide in 0EP- rats, immature and mature male rats. (3) In short term incubations and 5 days pituitary cultures, the in vitro-release of RIA-LH was shown. (4) Induction of ovulation was tested in virgin-rabbits by injection or infusion of LH-RH, and also in rats. In all assay systems, the biological activity of natural and synthetic LH-RH was identical. Release of bioassayable LH in the OEP-rat was stimulated by 8o-1oo ng LH-RH. The release of RIA-LH was stimulated dose-dependently over a range of I-Ioo ng in the 0EP-rat,and at 25-25o ng in the immature male rat. In the 0EP-rat, iv. injection was more effective than sc. injection. In male rats 3 ng LH-RH released LH by anterior pituitary perfusion.(Rea- gents for radioimmunoassay were kindly provided by the NIH and by Dr. Niswender). Dr.J.Sandow,Farbwerke Hoechst AG, 623 Frankfurt 8o~ Pharmakologie H 821

THE INFLUENCE OF RESERPINE ON THE ELECTRORETINOGRAM OF THE ISOLATED FROG RETINA (Der Einflu8 yon Reserpin auf das Elektroretino- gramm der isolierten Froschnetzhaut) H. Schalenbach, U. Borchard, R. Zim- mer Ale Beitrag zur Kl~irung der zentralen Wirkung yon Reserpin wurde dessen Einflu8 auf das Belichtungspotential der isoliert umstrSmten Froschnetzhaut untersucht. Unter der Wirkung yon Reserpin kam es nach ether anf~nglichen dosisabh~ingi- gen ErhShung der Potentialwerte zu ether stetig zunehmenden Erniedrigung des ERG. Der EiniluI~ des Rauwolfiaalkaloids erwies sich ale abh~ingig yon der Beleuchtungsst~irke. Bet kleinen Reizintensit~ten wirkte Reserpin st~irker po- tentialerniedrigend ale bet hohen, wobei hShere Dosen das Belichtungspoten- tial st~irker verminderten. Dies wird ale besonders empfindliche Reaktion yon neuronalen Summatiensprozessen auf Reserpin bet geringeren Beleuchtungs- st~irken gedeutet. Die Erholungslatenz nach einem Lichtreiz wurde im ersten Tell der Refrakt~irphase unter dem Einflu~ des Alkaloids dosisabh~ingig ver- l~ingert. Es beeinfluI~t daher vor allem den Absehnitt, in dem die Restitutions- vorg~nge normalerweise am schnellsten ablaufen. Der im Anschlu~ an die Re- serpinapplikation auftretende Potentialanstieg l~St etch, wie Kontrollversuche zeigten, mit der Wirkung yon Adrenalin, Noradrenalin, Serotonin und Dopamin auf die erregbaren Netzhautstrukturen deuten. Die sp~iter auftretende stetige Potentialabnahme wird ale Verarmung an den fur zentralnervSse Erregungs- vorg~inge notwendigen 4 biogenen Aminen diskutiert.

Dr. U. Borchard, Pharrnakologisehes Institut der Universit~it zu KSln, 5 KSln 41, Gleueler Str. 24

R 9 7

EFFECT OF TRYPAN BLUE ON THE TRANSFER OF SERUM PROTEINS INTO, THE EMBRYO OF THE RAT (Wirkung von Trypan-Blau auf den Transpor t von Serum-Proteinen in den Embryo der Ra t te ) . A. Schif fmann, R. Krowke Trypan blue (TB) is a tera togen which normal ly acts on ly at a specia l sta9e of embryogenesis. According to BECK and LLOYD (1967) an i n te r fe rence w i th h i s t l o - t roph n u t r i t i o n (hon.) takes place by an i n h i b i t i o n of p inocy tos is and of lyso- somal d iges t i ons w i t h i n the yo lk sac ep i the l i um ( y . s . e . ) . I t is almost impossib le to ob ta in biochemical data in the TB-sens i t i ve sta9e of embryogenesis. We t he re fo re have studied the uptake and metabol lsm of l ~ C - l a b e l - led serum p ro te ins on day 12 of ges ta t i on in v ivo , assuming t h a t the mechanism of h.n. is s t i l l ope ra t i ve in the y . s . e , a t t h i s stage of development apar t from the p lacenta l f unc t i on , 75 mg/kg TB were given s .c . 24 h p r i o r to the i . v . i n j ec - t i o n of 14C-compounds and the animals s a c r i f i c e d 3 h l a t e r . Uptake of lhC-macro- molecules and conversion in to acid so lub le substances and l i p i d s was measured in y ,soe, and embryonic t i s s u e , In con t ras t to l hC-g lobu l i ns the uptake of l ~C-a l - bumin in to the y . s . e , was decreased by TB. But no e f f e c t on the conversion of l~C-albumin in to ac id so lub le substances was found in the y . s . e . The ove ra l l i n - co rpo ra t i on of r a d i o a c t i v i t y in to the embryo was s l i g h t l y depressed. The re - su l t s are discussed w i th respect to BECK's theory ,

yo l k sac embryo acid so lub le acid inso lub le n acid so lub le acid inso lub le

14C-glucose . . . . 9 118 % • 18 1o9 % • 15

B+yglobulins 11o % • 23 92 % • 18 6 115 % • 25 99 % • 17

albumin 1oo % • 20 134 % • 30 5 1oo % • 9 6o % • 12

Dr. R. Krowke, Pharmakologisches Institut der Freien Universit~t Berlin, Abt. "Embryonal-Pharmakologie", D-1ooo Berlin 33, Thielallee 69/73

INVESTIGATIONS OF THE TRANSPORTCHARACTERISTIC OF NITROGLYCERINE AND BAY A 1040 ACROSS THE EVERTED RAT INTESTINE (Untersuchungen zur Transportcharakteristik yon Nitroglycerin und BAY a 1040 am Rattendarm in vitro) K.SchloSmann und K.MenK

Segments of everted rat intestine were incubated in buffer based substantially upon the Krebs-Henseleit solution (pH 7,4) in equilibrium with a gas phase of 95/5% 02/002. All studies were carried out at 30 ~ C. A perfusion apparatus was used which permits serial sample taking from the serosal medium.

The transepithelial mucosa/serosa and the serosa/mucosa-flux of nitroglycerine is a linear function of the concentration in the medium. The diffusion-kinetics of nitroglycerine is presumably of passive nature. The compound isnot accumulated in the tissue. The quotient (T/M) of the tissue concentration (T) to the concentration in the medium (M) is approximately I. - The mucosa/serosa-flux of BAY a 1040 is also an linear function of the concentration. The slope is not as steep as for nitroglycerine. BAY a 1040 is accumulated to a high concentration in the tissue relative to the medium, T/M = 6,0. The accumulation is the same, when the intestine is deprived of oxygen. The onset of action of both drugs is quickly. But the contrast to nitroglycerine BAY a 1040 acts for longer time, because this drug is bound in the tissue by physico-chemical factors.

Dr.K.Schlo~mann und Dr.K.Meng, Farbenfabriken BAYER AG, Institut fur Pharmakologie, 56 Wuppertal I, Postfach 130105

R 98

ON THE CATECHOLAMINE DEPLETING EFFECT OF O~RINE (Ueber die catecholamin- entspeichernde Wirkung yon Oxyfedrin) I. Schmid and Lo Maitre

Oxyfedrine caused a dose dependent reduction in the noradrenaline content of the rat heart and to a lesser extent in the noradrenaline and dovamine contents of the rat brain. When the substance was administered in a single oral dose of 60 mg/kg, noradrenaline depletion in the heart reached its maximum after about 24 hours. 1-Norephedrine produced effects identical with those of Oxyfedrine in equimolar dosage in the heart, but its activity in the brain was weaker and more prolonged. Oxyfedrine (6 and 60 mg/kg orally) inhibited the uptake and retention of radioactively labelled metaraminol into the heart, but these effects were minimal at the time of maximal depletion (24 hours). Oxyfedrine also exerts a dose dependent inhibitory effect on the uptake of radioactively labelled noradrenaline into isolated splenic nerve granules. In this experimental model Oxyfedrine was approximately 25 times more effective than 1-norephedrine. The sites of action of 0xyfedrine are presumed to be the nerve cell membrane and the storage granules.

Dr. I. Schmid, Biologlsche Forschungslaboratorien, Division Pharmazeutika CIBA-GEIGY AG, CH 4002 Basel (Schweiz).

STUDIES ON CHRONIC COPPER POISONING OF RU~[INANTS9 FORMATION 0r METHEMOGLOBIN BY Cu ++' IN THE BLOOD 0F MAN, HORSE, CATTLE, CALF, SHEEP, PIG, DOG, GUINEA PIG AND RAT (Untersuchungen Gber die chronische Kupfervergiftung der Wiederkiuer; Ferrih~moglobinbil- dung durch Cu ++ im Blut yon Mensch, Pferd, Rind, Kalb, Schaf, Schwein, Hund, Meerschweinchen und Ratte) A. Schmid~ K. Steiner

Bei 5 - 125 Minuten langer temperaturkonstanter (37 ~ Inkuba- tion von heparinisierten und h~molysierten (aqus dest.) bzw. nicht himolysierten (0,9%ige NaC1-LSsung) Blutpro~0en der e. g. Species in 0,78.10 -5 bis 50,36.10 -9 molaren Na-Azetat- (Kontrol- le) bzw. Cu(II)-AzetatlSsungen (Endkonzentrationen) wurden fol- gende Ergebnisse erhaltenl 1. Cu(II)-Ionen oxydieren in Konzen- trationen, wie sie bei der chronischen Kupfervergiftung im h~mo- lytischen Blut yon Wiederk~uern gefunden werden, freies H~moglobin gesunder Menschen, Pferde, Rinder, K~lber, Schafe, Schweine, Hunde, Neerschweinchen und Ratten zu Ferrih~moglobin (Absorp- tionsmaximum bei 626 nm). 2. Die kupferbedingte Ferrih~moglobin- bildung zeigt eine nicht lineare Zeitabh~ngigkeit. 3. Die 0xy- dationsempfindlichkeit freien H~moglcbins verschiedener Species gegen zweiwertiges Kupfer nimmt in der Reihenfolge Wiederkiuer = Pferd} Hund) Mensch = Schwein = Meerschweinchen) Ratte ab. 4. In nicht h~molysiertem Blut ist die kupferspezifische Ferrihimoglo- binbildung wesentlich geringer als in h~molysiertem.

Prof. Dr. A. Schmid, Institut fGr Pharmakologie, Toxikologie und Pharmazie der Tier~rztlichen Fakult~t, 8000 MGnchen 22, Veterin~rstr. 13

R 99

EFFECTS OF POLYCHLORINATED BIPHENYLS(PCB) ON RAT LIVER MICROSO- MAL ACTIVITY.(Einflu6 polychlorierter Biphenyle(PCB) auf die mikrosomale Aktivit~t der Rattenleber) A.Schmoldt, H.F.Benthe

Polychlorinated Biphenyls(PCB) are widespread environmental pollutants and the high chlorinated of them were detected in human adipose tissue(F.J.Biros et al.,Bull.Environ.Contam Toxi- col.~,317-323,1970).For studying the effects of low chlorinated PCB's on rat liver,male Wistar rats(200-230 g b.w.)were treated once with a mixture of Tetrachlorobiphenyle(AroclorR1248X500 mg/ kg b.w.,l+lw/v in pea nut oil ip.).The extend of induction of microsomal drug oxidizing enzymes attained a maximum after 4 days.The O-demethylation rate of p-Nitroanisole at this time was 485~35%(control rats=100%=6,72 nmoles/mg microsomal protein/10 min.).28 days after PCB-treatment the activity decreased to 225 • rel.liver weight(g/100 g b.w.)also increased(6,58~0,07% after 5 days)and did not return to normal(3,5~)after 4 weeks~ After the first day(136 ppm)the time course of PCB-concentration(for methods:HoFoBenthe et al.,in prep.)in liver showed a rapid decrease during 7 days and then a rather slow one.After 4 weeks the concentration was 7ppm whereas in adipose tissue it was 252~55 ppm.During stress(3~ and starvation up to 6 days) a redistribution could be observed along with fat mobilization~ The increased PCB-conc.in liver(32• ppm)caused again an induction of microsomal drug oxidizing enzymes (434~53% of stressed control animals).

Dr.A.Schmoldt, Pharmakologisches Institut der Universit~t Hamburg, 2 Hamburg 20, Martinistr. 52

KINETICS OF OXIDATION RATES AND MUTUAL COMPETITION OF SHORT CHAINY I~C-FATTY ACIDS AND 1"C-CARBOHYDRATES IN HEART MUSCLE CELLS. P. Schne ider , B. Ostheim, M. Siess (TUbingen/Marburg)

The o x i d a t i o n ra tes of exogenous 1"C- labe led g lucose, l a c t a t e , py ruva te ; and C6-, C8-, C9- and C1o - fa t t y acids were measured v ia 1"C02 in a t r i a of guinea p igs . (Siess et a l . , Drug research 18, 1245,1968) M ichae l i s -Menten k i n e t i c s of each subs t ra te could--b-e observed between a d i s t i n e t range of c o n c e n t r a t i o n s . This can be exp la ined by compet i t i on w i th endogenous subs t ra tes in o x i d a t i v e metabol ism. The K M values decrease from glucose ( I0 mM), l a c t a t e , pyruvate (1.2 mM), hexanoate (0.4 mM), octanoate and nonanoate (O.18mM) to decanoate (0.16 mM). The Vma x c a l c u l a t e d fo r the 02- consumption is 6-8 (~Mol/ lO0 mg w.wt. x h) f o r g lucose, l a c t a t e , pyruvate and hexanoate, but is cons ide rab l y lower f o r the C8-, C9-, and C 1 o - f a t t y acids (3.3 to 1.7 ~Mol/ lO0 mg w.wt . x h ) , which show less a c t i v i t y in o x i d a t i o n ra te i n s p i r e of t h e i r b e t t e r permeation through the c e l l membrane. The d i f f e r e n t a f f i n i t y of f a t t y ac ids , py ruva te , and l a c t a t e to Co-A is expressed by the mutual compe t i t i on of these s u b s t r a t e s . The Vma x depends on the card iac per formance, the endogenous glycogen concen t ra t i on and the a f f i - n i t y to the o x i d a t i o n cyc le . The C6- (3 mM), Ca- (1.6 mM) and C~o f a t t y acids (0 .4 mM) showed negat i ve i n o t r o p i c e f f e c t s . Decanoate (0.4 mM) increases in a r res ted a t r i a 1"C-glucose o x i d a t i o n ra te and much more the t o t a l 02-consumption presumably v ia an uncoup- l i ng e f f e c t . C8- and C~o- fa t t y acids are p r e f e r e n t i a l l y ox id i zed in a r res ted a t r i a . In beat ing a t r i a w i th enhanced performance a s h i f t to o x i d a t i o n of carbohydrates has been observed. Dr. P .Schne ider , Pharmakol. I n s t i t u t , 74 TUbingen, Wi lhe lms t r . 56

R 100

PROTEIN BINDING AND SUBCELLULAR DISTRIBUTION OF THE OPTICAL ANTI- PODES OF N-METHYLATED BARBITURATES (Proteinbindung und suhzellu- lira Verteilung der optischen Antipoden N-methylierter Barbi- turate) F. Schneider-Affeld$ W. Rummel und H. P. BUch

Investigations with N-methylated barbiturates have shown that the concentration in rat liver is up to 2 times higher than in the brain (H.P.Bdch et al.~ J.Pharmac.exp. Ther. i75, 709-716, 1970). With regard to the well known albumin binding of barbiturates it was supposed that their higher accumulation in the liver might be caused by protein binding. Knowledge of the subcellular barbiturate distribution in the liver is a prerequisite for proving this hypothesis.- The optical isomers of N-methyl-5- phenyl-5-ethyl-harbiturate (I) and of N-methyl-5-cyelohexenyl-5- ethyl-barbiturate (II) were injected i.v. (Wistar rats), and the binding by proteins in the particle free supernatant and the barbiturate concentration in other subfractions of the rat liver homogenate were measured. 3 min after 75 mg/kg of I the following subcellular distribution pattern was found: 30 % of the barbiturate are bound by the proteins of the cytosol, 40 % are contained in particle fractions (microsomes approx. I0 %~ nuclei and mitochondria~approx. 30 %)~ and only 30 % are unbound. No stereospecific distribution differences are detectable. II shows a quite similar distribution pattern in the liver. The conclusion was made that binding by proteins might be responsible for barbiturate accumulation in the liver.

Dr. F. Schneider-Affeld, Institut fur Pharmakologie und Toxi- kologie der Universit~t des Saarlandes, D-665 Homburg/Saar. (Supported by DFG)

EFFECTS OF ADH AND DOCA ON SODIUM AND POTASSIUM BALANCES IN RATS WITH HEREDITARY HYPOTHALAMIC DIABETES INSIPIDUS. (Einflu~ von ADH und DOCA aus die Natrium- und Kalium-Bilanzen bei Ratten mit here= dit~rem Diabetes insipidus.)

A. Sch~mi~ H. SchSmig-Brekner~ B. MShrins~ and J. MShrin~.

Treatment of male rats with hereditary hypothalamic diabetes insipidus (D.I.) with DOCA caused a positive sodium and a negative potassium balance. Vasopressin tannate induced potassium retention without affecting sodium balance, whereas in rats heterozygous for D.I., no change in either sodium or potassium balance was observed. After injection of both DOCA and vasopressin tannate to male D.I. rats, sodium retention was similar to that seen after administration of DOCA alone. However, the DOCA-induced negative potassium balance was prevented. These findings indicate that vasopressin might induce potassium retention without affecting sodium balance, even when sodium excretion is decreased and potassium excretion increased because of DOCA administration. According- ly, the effect of ADH on potassium balance does not seem to be mediated by aldosterone. It is suggested that vasopressin induces potassium retention via an increased water reabsorption in the distal tubules, thus decreasing distal tubular flow and distal transtubular sodium concentration gradient, which in turn decrease transtubular electrochemical driving forces for potassium excretion.

Dr. J. MShring, Department of Pharmacology, University of Heidel- berg, 69 - Heidelberg, HauptstraBe 47-51, Germany.

R 101

INDUCTION OF DRUG-METABOLIZING ENZYMES IN HUMAN LIVER (Induktion arzneimittel- abbauender Enzyme in der Leber des Mensehen) B. Schoene a R. A. Fleischmann t H. Remmer a and H. F. v. Oldershausen

In the homogenate of needle biopsy human liver samples the eyt. P-450 content (10.8 + 2.6 nmoles/g liver), and the activities of the NADPH-eyt.-e-reduetase (1.88 • 0.51 ~Amoles/g/mln), of the p-nitroanisole demethylase (83.6 + 23.8 nmoles/g/min), and of two enzymes not involved in drug-metabolism, glucose-6- ph0sphatase (11.8 • 2.8 ~lmoles/g/mln) and pseudocholinesterase (2000 • 570 nmoles/g/mln), were determined (normal values in parentheses). Patients with no drug intake and those treated with analgeties, antibiotics, benzodiazepines, glycosides and sulfonylurea showed no significant change in the activities of drug-metabolizlng enzymes. In 12 patients treated with diphenylhydantoin or with a combined therapy of rlfampicin (R), isoniazld, ethambutol and strep- tomycln, the P-A~30 content increased nearly 3fold, the activities of the cyt.- c-reduetase 50-100 %, and the demethylation rate of p-nltroanisole about 50 %, whereas the activities of the pseudocholinesterase and glucose-6-phosphatase did not differ from the controls. In two cases treated with R alone the P-450 content increased 2.5-fold, but no corresponding increase in the demethylation of p-nltroanisole in vitro or of aminopyrlne in vivo was seen. In 6 volunteers treated for 6 days with R, the amount of urinary 4-amlno-antipyrine after i.v. administration of aminopyrlne was only slightly enhanced 24 h after the last dose of R, but increased 50-100 % 3 - 4 days later. We conclude that R or a metabollte acts as an inhibitor of the microsomal drug oxidase while present in liver. 0nly after R or the metabolite is excreted from the liver, can the inducing effect of this drug be seen. This may be important in treatment of patients with both, R and anticoagulants.

Dr. B. Schoene, Institut fur Toxikologie der Universit~t, D-7400 Tdbingen, WilhelmstraBe 56

MECHANISM OF THE ANTILIPOLYTIC ACTION OF SALICYLATES (Zum Mecha- n ismus d e r ant i l ipolyt ischen Wirkung der Salicylate) P .S . SchSnh~Jfer, H.-D. P e t e r s , V. DinnendahI, K. Ka rze l

In i sola ted epididymal fat ce l l s sodium sa l icy la te (SA) and ace ty l sa l icy l ie acid (ASA) at concentra t ions Over 10-3M inhibit l ipolys is e l ic i ted by norepinephr ine or d i b u t y r y l - A - 3 ' : 5 ' - M P . Since t r i g lyce r ide l ipase is act ivated via A - 3 ' : 5 ' - M P , effect of both compounds on A T P - l e v e l s , A - 3 ' : 5 ' - M P accumulat ion and phosphodies te rase act ivi ty was tes ted . At anti l ipolytic concentra t ions SA d e c r e a s e s the A T P - l e v e l s in fat ce l l s while ASA has no effect . Both compounds, however , dec rea se the accumulat ion of A - 3 ' : 5 ' - M P in intact ce l l s and inhibit phosphodies terase act ivi ty in homogenates compet i t ive ly (SA: Ki = 8 x 10-3M; ASA: K i = 3,3 x 10-3M).

Since these r e su l t s do not explain the inhibition of d ibu tyrv l -A-3 ' :5"MP s t imulated t ipo[ysis , effects of both compounds on the A - 3 ' : 5 ' - M P binding prote in of musc le protein kinase was tes ted . At concentra t ions h igher than 3 x 10-4M a dose - dependent inhibition of the binding between A - 3 ' : 5 ' - M P and the binding prote in was observed with SA being m o r e potent than ASA. Analys is of the t~pe of inhibition shows compet i t ive pat tern for SA with an apparent K i = 3 x 10-~

There fo re , sa l icy la tes appear to i n t e r f e r e not only with the format ion, but also with the act ion of A - 3 ' : 5 ' - M P at i ts t a rge t sys tem.

P .S . SchSnhSfer, Pharmakologisches Institut de r Rheinischen F r i e d r i c h - W i l h e l m s - Universit~it, D-53 Bonn, R e u t e r s t r a s s e 2 b

R102

A N O B S E R V A T I O N ON T H E C H A N G E O F I N T R A M Y O C A R D I A L O X Y G E N

P A R T I A L P R E S S U R E A F T E R A C U T E C O R O N A R Y O C C L U S I O N A N D

A D M I N I S T R A T I O N O F C A R B O C R O M E N ( U n t e r s u c h u n g e n f i b e r d a s V e r -

h a l t e n d e s intramyocardialen S a u e r s t o f f d r u c k e s b e i a k u t e r K o r o n a r l i g a s

u n d V e r a b r e i c h u n g y o n C a r b o c h r o m e n ) J . S c h o k h o l s T . K o y a m a , R . - E . N i t z

E i n S e i t e n a s t d e s R a m u s d e s c e n d e n s n a r k o t i s i e r t e r H u n d e w u r d e f r e i g e l e g t

u n d a n g e s c h l u n g e n . I n n e r h a l b d e s v o n d i e s e m Gef~il~ v e r s o r g t e n G e b i e t e s

w u r d e n r a n d n a h e 2 P l a t i n e l e k t r o d e n e i n g e s t o e h e n . Die U n t e r b i n d u n g d e s

Gefa l~es f f i h r t e in 9 F a l l e n zu e i n e r d e u t l i c h e n M i n d e r u n g d e s R e d u k t i o n s -

s t r o m e s ( A u s g a n g s l a g e = 100~o). In 8 F a l l e n s a n k e r a u f N u l l . 5 - 6 M i n u t e n

s p a r e r b e w i r k t e d ie i n t r a v e n 6 s e Gabe v 0 n 4 m g / k g C a r b o c h r o m e n in f a s t

a l i e n F a l l e n e i n e n d e u t l i c h e n u n d a n h a l t e n d e n A n s t i e g de s R e d u k t i o n s s t r o m e s .

N u r in 3 V e r s u c h e n ( R e d u k t i o n s s t r o m a u f Nul l ) b l i e b d ie W i r k u n g a u s . I m

M i t t e l (N = 17) s a n k d e r R e d u k t i o n s s t r o m a u f 35% d e r A u s g a n g s l a g e a b .

C a r b o c h r o m e n n o r m a l i s i e r t e (95~0) d i e s e n W e r t Z0 M i n u t e n n a c h d e r I n j e k t i o n .

D r . J . S c h o l t h o l t , M e d i z i n i s c h e F o r s c h u n g , F a r b w e r k e C a s s e l l a M a i n k u r AG.

6-Frankfurt/Main-Fechenheim, H a n a u e r L a n d s t r a l l e 526

EFFECT OF SYNTHETIC LH/FSH-RH ON THE RELEASE OF FSH IN THE RAT (Wirkung eines synthetischen LH/FSH-RH auf die FSH-Freisetzung) H.G. SchrSder~ R.Geiger~ F. Enzmann~ W.Heptner~ and H.G.Vogel

The identity of the hypothalamic releasing hormone for LH and FSH has been proved by Schally, Arimura et al. Synthetic LH-RH~will release both hormones into the bloodstream. At a given dose, the ratio of LH/ FSH-release will be constant, but the time course is different. FSH- releasing activity was measured both by bioass@ys and radioimmuncassay (double antibody technique of Daane and Parlow'). Pituitary depletion was induced in intact male rats by injection of I-Io meg of LH/FSH-RH. The depletion effect was demonstrable from 45-90 min. after injection. The release of FSH from isolated pituitaries in vitro was assayed by HCG-augmentation of the incubation media. In some experiments, pituitaries of castrated male rats (Tx) were used. The time course of FSH-release into plasma was measured by RIA. In urethane-anesthetized male rats, after 1oo ng LH/FSH-RH a 3fold increase over preinjection levels was seen after 2 hours. In ether-anesthetized rats killed by decapitation, an increase of plasma-FSH was observed both after iv. and sc. injection of 250 ng LH/FSH-RH. A slight increase of FSH was also seen in ovariectomized, steroid-pretreated rats following sc. injection of I.o mcg LH/FSH-RH.

1(The gift of pituitary hormones by the NIAMD and of anti-rat-FSH-serum by Dr. A.F.Parlow is gratefully acknowledged).

Dr. H.G.SchrSder, Farbwerke Hoechst AG., 623 Frankfurt 80, Abt. f~ir Pharmakologie H 821.

R 103

POSSIBLE ROLE OF GLUTATHIONE REDUOTASE (EC 1.6.4.2.) FOR THE ACTION OF "DIRECT LYTIC FACTOR" (DLF) FROM COBRA VENOM ON ERY- THROCYTES OF VARIOUS SPECIES (Spielt die Glutathionreduktase eine Rolle bei der Wirkung des "Direkt Lytischen Faktors" (DLF) aus Cobragift auf Erythrozyten?) R. Scbroeter, P.G. Lankisch, L. Le~e, W. Volt

The activity of glutathione reductase (GR) is of importance for the resistance of erythrocytes against different oxidant drugs. As DLF binds reversibly to red cells and is assumed to react with SH-groups of membrane proteins, it seemed possible that GR also plays a part in the haemolytic action of DLF on washed red cells. The enzyme activities in haemolysates and red cell ghosts of the species: man, guinea-pig, rabbit, and rat were investigated. We found a correlation between GR-activity in red cell ghosts and DLF-sensitivity of the respective erythrocytes: red cells with high membrane enzyme activity can be attacked much better by DLF than cells with low enzyme levels. Further GR utilises DLF as substrate, probably by reduction of its disulfide-bonds. Reduced glntathione (GSH) which is known to inhibit GR in high concentrations, is able to protect washed guinea-pig red cells against the haemolytic attack of DLF alone and in combination with phospholipase A. These findings suggest that red cell membrane GR is essential for the baemolytic action of DLF.

Dr. R. Schroeter, Max-Planck-Institut fGr experimentelle Medizin, Abt. Biochemische Pharmakologie, D-3400 GSttingen, Hermann-Rein- Strafe 3.

INHIBITION OF ~-HCH-INDUCED LIVER CELL PROLIFERATION BY CFT 12ol AND

ACTINOMYCIN D. TIME OF ACTION OF THE INHIBITORS DURING THE REPLICATIVE

CYCLE. (Hemmung der ~-HCH-induzierten Leberze l lp ro i i fe ra t ion durch CFT 12ol und Actinomycin D. Zeitpunkt der Wirkung der Hemmstoffe wdhrend des Replikat ionszyklus.)R.Schulte Hermann, C.Leberl

~-Hexochlorcyclohexan(~-HCH) ibst nach einer initialen Latenzphase von

etwa 16 Std. in der Leber der Ratte eine Neusynthese yon DNS aus. Dieser Effekt kann durch CFT 12ol (B-Di~thylaminodthylphenyldial ly lessigsdure), einen Hemmstoff mischfunkt ionel ler Oxydasen der Leber, b lock ier t werden. Die Hemmung t r i t t nur dann auf, wenn CFT12o] mindestens 1-3 Std. vor dem Beginn der DNS-Synthese-Phase (=S-Phase) app l i z ie r t wlrd. Auch Actinomycin D verhindert dle DNS-Replikation, wenn es sp~testens 1-3 Std. vor dem Be- ginn der S-Phase gegeben wlrd. Die Hemmwirkung beider Inhlb i toren i s t dem nach im frUhen Abschnitt der prdsynthetlschen (G.-) Phase des Zel lzyklus

�9 . . |

l o k a l l s l e r t . Hi t dem Ubergang yon der Inhibztor-empflndl ichen zur Inh ib i - tor - res ls tenten Periode endet ein Abschnitt der G1-Phase , in dem bestimm- te zur DNS-Replikation er forder l iche Pr~zesse indOziert werden.

Dr.R.Schulte Hermann, I n s t i t u t fur Toxikologie und Pharmakologie der Philipps-Universit~t Marburg, 355 Marburg, Pilgrimstein 2.

R 104

ON THE MECHANISM OF THE LIPOLYTIC ACTION OF PURINE DERIVATIVES OF SYMPATHOMIMETIC AMINES (Zum Mechanismus der lipolytischen Wirkung von Purinderivaten sympathomimetischer Amine). Schulz, H.U., K. Stock, A. Aulich und E. Westermann

The compounds noradrenalino-ethyltheophylline (NAT) and norephe- drinoethyltheophylline (NET) respectively are distinguished by comprising two diFFerent types oF lipolytic stimulants. For that reason it was oF interest to Find out whether NAT and/or NET stimulate lipolysis in rat adipose tissue in vitro directly by stimulation os adenylcyclase (similar to NA) or by inhibition oF phosphodiesterase, similar to hydroxy-ethyltheophylline (HOT). The release oF Free Fatty acids (FFA) was measured and intrinsic activities (Emax=~Eq FFA/g.h) as well as apparent aFFinities (KA = moles/l) were calculated. The obtained values For Emax oF NAT and NET resp. were not diFFerent From those o9 NA or NE, although the KA-values proved to be 70-100 Fold higher. The lipolytic action oF NAT and NET was inhibited competitively by the adrenergic 8-blok- ker KL 255, the Ki (7 riM) being identical with that obtained with NA and NE resp..In contrast HOT was inhibited non-competitively (Ki=0.9 raM). NAT and NET inhibited PDE at a ~M-substrate range in a non-competitive manner (Ki=1.7 resp. 0.7 mM), as did HOT. Based on our results it seems to be unlikely that inhibition oF PDE con- tributes to the lipolytic eFFects os NAT and NET but is probably due to a stimulation oF adenylcyclase in adipose tissue.

Dr. H.-U. Schulz, Institut s Pharmakologie, Medizinische Hoch- schule Hannover, D-3000 Hannover, RoderbruchstraSe IO1, Germany.

PERIPHERAL MECHANISMS AFTER DRUG INDUCED DISTURBANCE OF THERMOREGULATION IN RATS OF DIFFERENT AGE (Periphere Mechanis- men bet pharmakogen gestSr%er Therrnoregulation yon Ratten versehiedenen Alters) G. Schulze, H. Romrnelspacher, I. Broermann, S. Sfrau~

A number of centrally active drugs influence the body temperature in small rodents. However, i% remains uncertain to what extent this effect is centrally or peripherally produced. The shift of the equilibrium of heat-production and heat loss depends on some external factors as well as the stability of the internal regulatory systems. The ability to react and the efficiency of some single peripheral mechanisms of the thermal regulation has been studied in 3- and 13-months-old rats. After 5 rng/kg d-amphetamine the course of body temperature (ambient temperature (a. t. ): 4 ~ 25 ~ 31~ and the evaporative heat-loss (ehl) was determined. As indirect factors of thermogenesis the concentration of corticosterone, NEFA, glucose (a. t. 4 ~ 25~ in serum has been determined. Amphetamine elicited a hyperthermia above +16~ a.t., which was accompanied by an increase of ehl, which raised sharply between 25 ~ and 31~ The hyperfhermia of old animals was more marked whereas ehl was significantly less. The observed hypothermia caused by amphetamine at 4 ~ a.t. reached its maximum at 90"p. i. whereby the concentration of NEFA (as an indicator of non shivering thermogenesis) and glucose has significantly increased in adult rats whereas corticosterone only seems to indicate a thermal stress. Remark- ably this effect is demonstrable also at 25~ a.t. In old rats this increase was not detectable, corresponding to the more pronounced hypothermia.

G. Schulze, Institut fiir Neuropsychopharrnakologie der Freien Universit~t Berlin. D-1000 Berlin 19, Ulmenallee 30

R 105

DAS-INDUCE D P L A T E L E T AGGREGATION AND ITS PREVENTION BY SP 54 (PENTOSANPOLYSULFATE) (DAS-induzier te T h r o m b o c y t e n a g g r e g a t i o n und i h r e V e r h i n d e r u n g dureh SP 54 (Pentosanpolysul fa t ) ) K._=A. S c h u m a e h e r , H.G. C l a s s e n

In ca ts the i . v . i n j ec t ion of a s u b s t a n c e en r i ched f r o m s t o r e d bovine s e r u m (DAS-dep re s so r ac t ive subs tance) led to an e x t r e m e i n c r e a s e in pu lmonary v a s c u l a r r e s i s t a n c e (PVR), and at the s a m e t ime to a s t e e p d e c r e a s e in p la t e l e t count in the a r t e r i a l and venous blood. The h is to logie examina t ion of lung t i s s u e - t aken at the t ime of m a x i m u m i n c r e a s e in PVR - r e v e a l e d an ex tens ive m i c r o e m b o l i s m of the t e r m i n a l pu lmona ry v e s s e l s by p la te le t agg rega t e s . The n u m b e r of p la te le t s of the t e s t an ima l s r anged be tween 100. 000 and 800. 000 m m 3 . The m a x i m u m i n c r e a s e in PVR was s t rong ly r e l a t e d to the p la te le t count" On the i . v . i n j ec t ion of 1 mg DAS, ca t s with a low content of p la te le t s showed only a s l igh t i n c r e a s e in PVR; on the o ther hand, PVR i n c r e a s e d e x t r e m e l y in an ima l s with a high p la t e l e t count. DAS was much m o r e e f fec t ive than an equal dose of ADP, f r o m which it i s d i s t i ngu i shed by be ing ve ry r ead i ly so luble ia c h l o r o f o r m and by leading to a t achyphylae t ic r e a c t i o n when in jec ted a s econd t i m e . The effect of DAS and ADP on the t h rombocy te s and PVR could be p r e v e n t e d by p r e t r e a t m e n t with SP 54 (pentosanpolysul fa te , MW about 2000)whereas Hepar in (MW about 16000) was inef fec t ive .

Dr . K.A. Schumache r , Ins t i tu t fi ir E x p e r i m e n t e l l e T h e r a p i e de r Universi t~it D-78 F r e i b u r g / B r . , Hugs te t t e r S t ra6e 55.

DIFFERENT EFFECTS OF LIPOLYTIC HORMONES AND INHIBITORS OF PHOSPHO- DIESTERASE ON CYCLIC 3' ,5'-AMP LEVELS IN ISOLATED FAT CELLS (Untem- schiedliche Wirkungen yon lipolytischen Hormonen und Hemmstoffen der Phosphodiesterase aus den 3',5'-AMP-Gehalt von fsolierten Fett- zellen) U. Schwabe and R. Ebert

Cyclic AMP levels in isolated fat cells of rats were increased about 40 fold by norepinephrine (I bM) within 4 min of incubation and had declined markedly after 10 min. The effect os lipolytic hormones on cyclic AMP levels was dose dependent over wide range os concentrations, ACTH and glucagon being most active on a molar basis. Among the adrenergic compounds tested isoproterenol exhibited the greatest response. This pattern was largely paralleled by the lipolytic effects of the hormones. Methylxanthines caused only a 3-5 fold elevation of cyclic AMP levels at concentrations which induced maximal lipolytic effects. Papaverine (0.5 mM) had virtually no effect on cyclic AMP level and did not potentiate the effects hormones on cyclic AMP production. On the other hand, theophylline (1 mM) caused nearly tenfold amplification of the effect of isoproterenol when added to 100 O00 cells per ml in the medium. At 20 000 fat cells per ml isoproterenol alone increased cyclic AMP levels 300 fold and was not potentiated by theophylline. Our results suggest that lipolysis induced by hormones is mainly mediated by an accumulation of cyclic AMP, whereas methylxanthines must have additional effects, possibly an inhibitory action on a hormone antagonist released by fat cells into the incubation medium.

Dr. U. Schwabe, Institut s Pharmakologie der Medizinischen Hochschule Hannover, D-SOO0 Hannover-Kleefeld, Roderbruchstr. 101

R 106

EFFECT OF TESTOSTERONE ON THE ACTIVITIES OF GLUTA~TE-PYRUVATE- TRANSAMINASE (GPT), GLUTAMATE-OXALOACETATE-TRANSA~INASE (GOT), AND GLUTAMATE-DEHYDROGENASE (GLDH). (Einflu~ yon Testosteron auf die Aktivit~t von Glutamat-Pyruvat-Transaminase (GPT), Glu- tamat-Oxaloacetat-Transaminase (GOT) und Glutamatdehydrogenase (GLDH) W. Schwarzlose

Compared with prepuberal 18 days old male white mice the activities (U/g wet wt.) of GPT, GOT and GLDH in the kidneys of post- pubertal 42 days old male were increased. In castrated animals the increase of the activities of GOT and GLDH was completely, that of GPT only in part prevented. A single dose of 16 mg/kg testosteronepropionate applicated to female white mice only enhanced the activity of GPT two days after application, whereas the weight of the kidneys, its protein concentration and the activities of GOT and GLDH remained unchanged. Two days after a dose of 100 mg/kg testosteronepropionate the activities of GPT, GOT and GLDH as well as the kidney weight and the protein concentration were increased. The enhancement of the activities of GPT and GOT caused by testosterone was nearly completely, that of GLDH to 70 percent prevented by actinomycin D (total dose 400/ug/kg). From these results it can be concluded, that the testosterone induced increase of the activities of GPT and GOT is the result of a neo-synthesis of enzyme protein. In the case of GLDH an additional allosteric hormone activation must be assumed.

Dr. W. Schwarzlose, Pharmakologisches Institut der Universit~t Erlangen-Nfirnberg, 8520 Erlangen, Universit~tsstr. 22

PURIFICATION AND CHARACTERIZATION OF PHALLOIDES HEMOLYSIN, A HIGH MOLECULAR WEIGHT TOXIN FROM AMANITA PHALLOIDES (Reinigung und Charakterisierung von Phalloides-H~molysin, einem hochmolekularen Toxin aus Amanita phalloides) R. See~er t H. Scharrer

A toxic and hemolytic protein was successfully isolated from aqueous extracts of Amanita phalloides. It was destroyed on extraction with methanol, the common procedure (Th. Wieland, Fort- schr. Chem. org. Naturstoffe 25, 214-250, 1967) for isolating known Amanita toxins. After precipitation from aqueous extracts with ammonium sulfate it was purified by ion-exchange and gel chromatography. The isolated hemolysin was uniform on disk electrophoresis at pH 4. Its apparent molecular weight, determined by gel chromatography, was 30 000. It was basic in nature, thermola- bile, acid labile and relatively stable to alkali. It was not inactivated by trypsin, chymotrypsin, subtilisin, pronase, nagarse, pancreatin and S-amylase. In 8 M urea it lost its hemolytic activity together with its toxicity and could not be reactivated e.g. by dialysis. It directly hemolyzed washed human erythrocytes and those of various animal species. The LDso for mice (i.p.) was 0.34 mg/kg. Its occurence in Amanita phalloides from various loca- tions was consistently demonstrated. 40 - 7o% of total Amanitas toxicity in mice (i.p.) could be attributed to this new protein. However, due to its thermolability and acid lability, it does not seem to be implicated in human Amanita phalloides poisoning.

Supported by the Deutsche Fcrschungsgemeinschaft. Dr. Ruth Seeger, Institut f~r Pharmakologie der Universit~t D-8700 W'~rzburg, Koellikerstrasse 2.

R 107

ON A BIPHASIC FORCE DEVELOPMENT OF THE GUINEA-PIG PAPILLARY MUSC- LE DURING POTASSIUM DEPOLARIZATION (~ber einen zweiphasigen Kraft- anstieg des Meerschweinchen-Papillarmuskels bei Kaliumdepolarisa- tion) K. Seibel~ M. Reiter

Eine Erh~hung der ~usseren+[K +] auf 22,5 bis 55 mM bei gleiohzei- tiger Erniedrigung der [Na ] unter 75 mM (abh~ngig vonder [K+]) ruft an isolierten Meerschweinchen-Papillarmuskeln (35~ zun~chst einen geringen Kraftanstieg hervor, dem nach einer Latenz yon meh- reren Minuten Dauer der Hauptteil der Kraftentwicklung folgt. Der Verlauf der Membrandepolarisation ist dabei yon der zweiphasigen Kraftentwicklung dissoziiert. Bei ErhShung der ~usseren Ca- oder st~rkerer Erniedrigung der Na-Konzentration tritt die zweite Phase der Kraftentwieklung frtther auf, ihre H~he und Steilheit nimmt zu, ohne dass die Membrandepolarisation beeinflusst wird. Eine ErhD- hung der iusseren [Ca 2+] w~hrend der zweiten Phase der Kontraktur ruft einen erneuten biphasischen Kraftanstieg hervor. - Ein zwei- phasiger Kraftanstieg wird auch ohne Depolarisation der ~usseren Membran in K-freiem N/hrmedium durch Reduktion der [Na +] auf 25 mM erzeugt. Die Befunde zeigen, dass das Kammermyokard des Meerschweinehens Strukturen hat, die unter den gew~hlten Bedingungen die Kraftent- wicklung steuern, ohne vom Membranpotential direkt beeinflusst zu werden. Dr.KoSeibel, Institut f.Pharmakologie u. Toxikologie d.Teehnisohen UniversitHt M~nohen, 8 M~nohen 23, Biedersteiner Str. 29

ONE OR TWO KININOGENS IN HUMAN PLASMA? (Ein oder zwei Kininogene in Humanplasma?) G. Seidel

Former studies of Vogt et al. (Naunyn-Schmiedebergs Arch. Phar- mak. u. exp. Path. 256,127-138,1967) indicated that in human plasma there are two ~inino~ens with different functional enti- ties. Corresponding results are presented by direct methods. The pseudoglobulins of non-contacted human plasma (I mg/ml hexadi- methrine) were separated. They were diluted with 0.02 M--K p~osphate ~uffer (pH 8, 40 mg/1 hd., I mg/ml o-phenanthroline)to the conductivity of 0.02 M K ph.b. (pH 8, 0.1M NaCl, 40 mg/1 hd.) and adsorbed onto DEAE-sephadex. Kg. was desorbed by 0.02 M K ph.b. with 0.3 M NaCI (40 mg/1 hd.). This fraction was passed through sephadex G 200 (0.2 M K ph.b., pH 8, 0.1 M NaCI, 0.1% protamin). 2 peaks of kg. were received. The same procedure wfth glass contacted plasma (O.1 mm beads, 170 mg/ml) resulted in low weight kg. (MW 50000) only, in contrast to plasma previously diluted tenfold with 0.9% NaCI and incubated with urokinase (40 plough-u./ml, 2h, 37~ in which only bigb molecular weight kg. (MW 110000) survived. The ratio of formed kinins from concentrat- ed fractions of both kgs. by trypsin and contact plasma kininogenase was much higher with low (> 3) than with high weight kg. labout 1.5) During gel chromatography with 0.1 M tris-HCl b. pH 8, 0.1M NaCI, 1 mg/ml hd.) the low weight kg. dimerized. -

It is assumed, that in human plasma both kgs. are functionally different distinct proteins, even if the high weight kg. (= kg. I) is only a dimer of the other kg. (= kg. If).

Dr. G. Seidel, Max-Planck-Institut fur experimentelle Medizin, Dept. of Biochem. Pharmaeol., D-3400 GSttingen, Herm.-Rein-Str. 3

R 108

CYCLIC ADENOSINE MONOPHOSPHATE (3':5'-AMP) IN PHAGOCYTIZING GRANULOCYTES AND ALVEOLAR MACROPHAGES (3':5'-AMP in phagozytlerenden Granulozyten und Alveolar- makrophagen) H. Seyberth, H. Schmldt-Gayk, I. Martiskalnen und E. Haekenthal

Based on conflicting results reported in the literature (B.H. Park et al., Nature New Biol. 229, 27-28, 1971; T.P. Stossel et al., J. Biol. Chem. 245, 6228-6234, 1970) the involvement of 3':5'-AMP in the process of phagocytosls was reinvestigated in hog polymorphonuclear leukocytes (PMN) and alveolar macrophages (AM) obtained from BCG treated rabbits. Both cell preparations when incubated in a modified Krebs-Ringer phosphate buffer with glucose rapidly phagocytize 0.481 ~ polystyrene latex particles (PMN 0. Ii mg/mg cell pro- teln; AM 0.13 mg/mg) and exhibit the expected burst in oxygen consumption (PMN + 260 %, AM + 26 %) in the first ten minutes following the addition of particles. The 3':5'-AMP content (estimated by the 3':5'-AMP dependent~protelnklnase mediated phosphorylatlon of histone) of PMN increased only slightly (4.6 -5,4 pmoles/mg protei~above controls (3.9 - 5.0 pmoles/mg protein) 30 seconds to 30 minutes following the addition of particles. In AM no difference in 3':5'- AMP content between phagocytizing and control cells (8.7 - 12.6 pmoles/mg protein) 5 - 60 minutes following the addition of particles was observed (no data for shorter periods). These results indicate that drastic changes in 3':5'- AMP concentration commonly observed upon hormone stimulation of appropriate target cells are not essential for the early events during phagocytosis, at least in PMN.

Dr. E. Hackenthal, Pharmakologlsches Institut der Unlversit~t, 69 Heidelberg, Hauptstr. 47 - 51

INFLUENCE OF CAFFEINE ON ETHANOL ABSORPTION IN RATS (Der Einflu8 von Coffein auf die ~thanolresorption bei Ratten) C.-P. Siesers~ O. Strubelt und G. Back

In rats, caffeine (i0, 20, 60 or i00 mg/kg) given orally 15 min before ethanol caused a dose-dependent depression of blood alcohol levels. This effect was demonstrated with different doses of ethanol (0.8, 2.4 or 4.8 g/kg), given orally as a 10, 30 or 60% solution, respectively, in saline. Intraperitoneal or subcutaneous application of caffeine (60 mg/kg), too, reduced blood alcohol level. However, caffeine (60 mg/kg i.p.) did not decrease blood alcohol levels if ethanol (1 and 2 g/kg) was injected intravenously. This indicates thatabsorption, but not elimination, of ethanol is impaired by caffeine.

The stomachs of rats treated with both caffeine (lO0 mg/kg p.o.) and ethanol (2.# g/kg p.o.) contained larger volumes and greater amounts of ethanol than the stomachs of rats treated with ethanol alone. In conclusion, caffeine in rats reduces ethanol absorption by delaying passage of alcohol from the stomach to the small intestine. In rat fundal strips

in vitro caffeine (lO -5 - 3 " lO -3 g/ml) caused a dose- dependent relaxation. Therefore, stomach emptying delayed by caffeine in rats must be due to gastric atony.

Dr. C.-P. Siegers, Institut fir Pharmakologie der Medizinischen Akademie LGbeck, D-2400 Ldbeck, Ratzeburger Allee 160

R 109

EFFECT OF AESCIN ON CAPILLARY FLUID EXCHANGE IN THE CAT HIINDLIMB ([Iber den Einfluf~ yon Aescin auf die Kapillarpermeabilit~t) S. Silber, J. Remien, W. Felix

The effect of the horse-chestnut saponin aescin was investigated in 30 cats anesthetized with chloralose (40 mg/kg oral). The hindlimb was blood- perfused at constant arterial flow and constant venous outflow pressure (W. Felix, J. Remien, K. H~llfritzsch, Pfliig. Arch. 329, 352-359, 1972). Recorded or calculated were: total volume of the hindlimb, venous volume, postcapillary resistance, effective capillary pressure (zero flow technique) and capillary filtration coefficient (CFC). The volume of the extremity, being isovolumetric before the application, was increased by i.v. and i. a. infusions of aescin (0, 02-1 mg/kg/min) in all animals investigated. Under condition of venous congestion edema developped more rapidly. The response of the resistance vessels initially changed with the animal and the applied dose but was always constrictive in the course of time. The effective capillary pressure was decreased, CFC was enhanced. Post- capillary resistance and venous tone was not influenced. The effect of aescin on fluid movement into the interstitial space could not be due to hemodynarnic influence on capillary pressure. There is much evidence that capillary permeability for proteins was enhanced.

Prof. Dr. reed. W. Felix, Pharmakologisches Institut der Universit~t, D-8000 Miinchen 2, NuSbaurnstr. 26

PHARNACOKINETIC STUDIES ON STRUCTURE-ACTIVITY RELATIONSHIP OF TREMOR-PRODUCING HARNALA AND IBOGA ALKALOIDS (Pharmakokinetische Untersuchung zur Struktur-Wirkungsbeziehung Tremor-erzeugender Harmala- und Iboga-alkaloide) G. Sin~bartl, G. Zetler and Lucie Schlosser

Differences in tremorigenic activity of substances given peripherally may be due to either different penetration into brain or different affinity to specific tremorigenic receptors. Therefore it was necessary to determine both the lipid solubility, and brain concentrations of the alkaloids at different intervals post injection. The alkaloids were extracted from brain in alkaline medium and their concentrations determined fluorometrically. In the kinetic experiments lO mg/kg of the drugs were injected intravenously into mice within lO sec, while the subcutaneous route was chosen in experiments testing tremorigenic potency. The moment of end of tremor was determined and the corresponding brain concentrations (TEC) of the alkaloids were interpolated from the time-concentration curves. The results indicate structure-activity relationships but no correlation between tremorigenic potency (after subcutaneous injection) and lipid solubility. However~ a correlation exists between tremorigenic activity and TEC. Thus, tremor-producing activity was much more influenced by chemical structure than by lipid solubility. This points to ~pecific receptors for indole compounds in tremorigenic brain structures.

Dr. med. G. Singbartl, !nsti~ut fur Pharmakologie der Medizinischen Akademie LNbeck, D 24 LGbeck, Ratzeburger Allee 160

Fi 110

THE POSSIBLE ROLE OF INTRACARDIAC RECEPTORS IN THE MECHANISM OF REACTIVE CORONARY HYPERAEMIA (Die Rolle cardialer Rezeptoren bei tier reaktiven coronaren Hyper~imie. Skolasihska K., A.Trzebski, E. Szostak~ Z. Religa

On 11 open-chest dogs the coronary sinus outflow was measured during and after 1 min long lasting ischaemia caused by closing the descending branch of left coronary artery. Latent periods of reactive hyperaem[a, period of time in which the maximum vasodilatation was reached and maximum changes in peripheral resistance were estimated and correlated in control series and after administration of 1)xylocaine 2) hexamethonium 3) atropine and 4) after epicardiactomy performed with local application of 80~ phenol solution on the epicardiac surface. The decrease of the peripheral resistance during the reactive hyperaemia was reduced after 1 and 2 about 30~, after 3 about 80% and absent after 4. 1,2,3 delayed the onset and the peak value of reactive hyperaemia. The results suggest the participation of a cholinergic reflex component in the reactive hyperaemia after short lasting myocardial ischaemia.

Dr. K. Skolasihska, Department of Physlology~ Institute of physiological Sciences, Faculty of Medicine, Warsaw, Krakowskie Przedmie~cie 26/28.

PECULIARITIES OF THE RESPIRATORY METABOLISM OF DIFFERENT TISSUES OF RAT EMBRYOS IN VITRO (Besonderheiten des Atmungsstoffwechsels verschiedener Gewebe von Rat- tenembryonen in vitro) Spielmann, H. and LUcke, I.

The respiratory activity of embryonic tissues was determined with the Warburg method (Spielmann and LBcke, Naunyn-Schmiedeberg's Arch. Pharmak. 270, 10, 1971) Whole naked rat and mouse embryos have identical respiratory rates during organogenesis when judgin 9 from the dry weight basis. The dry weight-dependent Qo 2 plot enables us to distinguish between a developmental retardation for hours and days and a significant developmental reduction on the basis of the two parameters. These conclusions in experimental teratology at this early stage of development cannot be drawn from an age-dependent Qo 2 plot. The respiratory capacity of whole mouse embryos on day 9 and of rat embryos on day 10 * 16 h is significantly lower than on the following day. Since whole rat embryos exceed the limiting on day 13 of gestation, we suggest performing measurements of the Qo 2 on day 14 and 15 with isolated organs (hearts and livers) and from day 16 to term with liver slices of rat embryos. The Qo 2 of isolated hearts and livers on day 14 to 15 and of liver slices from day 16 to term is at least 10 in a glucose medium, which is comparable to adult liver slices. The succinate-dependent respiratory activity can be measured with these embryonic tissue preparations on day 14 and later, it increases considerably on day 19 and reaches the values of adult rat liver tissue at term. These data are in good agreement with those of other investigators (de Vos et al., Biol. Neonate I__3, 83, 1968; Jacovcic et al., Biochem. J. 12._.!I, 341, 1971).

This work was supported by the Deutsche Forschungsgemeinschaft (SFB 29).

Dr. H. Spielmann, Pharmakologisches Institut der Freien Universit~t Berlin, Abt. "Embryonal-Pharmakologie", D-tO00 Berlin 33, Thielallee 69/73.

R 111

ALPHA-SYMPATHOMIMETiC INHIBITION OF THE RELEASE OF NORADRENALINE FROM THE RABBIT HEART (Alpha-sympathomimetische Hemmung der Noradrenalinfreisetzung aus den sympathischen Nerven des Kaninchenherzens) K. Starke

The hypothesis was tested that, in the rabbit heart, a-receptor- like structures mediate, on reaction with sympathomimetic agents, an inhibition, and on reaction with a-adrenolytics, a facilitation of the discharge of noradrenaline (NA) during sympathetic nerve stimulation. Experiments were done on isolated rabbit hearts with intact postganglionic sympathetic nerves. - Three sympathomimetics with predominant s-receptor affinity, phenylephrine, naphazoline, and oxymetazoline (0), cause a dose- dependent decrease of the stimulation-induced NA outflow. Ex- periments with labelled NA indicate that this decrease is due to an inhibition of the discharge of NA from the nerve terminals. Pretreatment with 0 specifically prevents the augmentation of the stimulation-induced NA overflow caused by phentolamine. Two results suggest that liberated NA shares the ability to depress, via s-receptors, the discharge of NA. l) There is an in- verse correlation between the stimulation-evoked outflow of NA before the infusion of O, an~4the degree of inhibition caused by O. 2-~--In hearts loaden with C-NA and subsequently perfused with cocaine, exoge~Rus unlabelled NA decreases the stimulation- induced outflow of ~C-NA; this decrease is prevented by phenoxy- benzamine. - The results are in accord with the hypothesis presented above.

Doz. Dr. K. Starke, Pharmakologisches Institut, Klinikum Essen der Ruhr-Universit~t, 43 Essen, Hufelandstr. 55

EFFECTS 0FBENZOCTAMINE AND DIAZEPAM ON ACID-BASE BALANCE, RESPIRATION RATE AND HEART RATE IN THE DOG (Einfluss yon Benzoctaminund Diazepam auf S~ure- Base~-Gleichgewicht, Atmungs- und Herzfrequenz des Hundes). J. Stepanek

Experiments were performed on conscious dogs with an implanted catheter in the abdominal aorta, to determine whether the changes in acid-base balance induced bythe two drugs were of respiratory or metabolic origin. 0ral administration of benzoctamine for four days in a dose of 3 mg /kg daily caused slight, compensated metabolic acidosis with respiratory a!kalosis; a dose of i0 mg /kg produced mixed, uncompensated alkalosis. Both doses slowed respiration, but only slight changes in heart rate were noted. The drug was well tolerated. The same doses of diazepam led to progressive metabolic acidosis, which was not compensated at the high dose-level. Respiration was accelerated and heart rate slowed. Repeated oral doses of i0 mg /kg diazepam were poorly tolerated by dogs with implanted catheters; in intact dogs, doses of 30 mg /kg p.o. were better tolerated and had a slight natriuretic effect. The tolerability of diazepam varies: it is well tolerated by healthy persons, but the same doses provoke toxic effects in patients with respiratory disease for instance, (Bfihlmann and Rossier: Klihische Pathophysiologie der Atmung, p. 200, Springer, Berlin, 1970) or in animals subjected to experimental procedures (Knorpp: Naunyn-Schmied. Arch. Pharmak. 267, 457-467, 1970).

Dr. Jo Stepanek, Biological Research Laboratories of the Pharmaceutical Division of CIBA-GEIGY Limited, Basle, Switzerland.

R 112

INFLUENCE OF LATRODECTUS TRIDECIMGUTTATUS TOXINE UPON THE MUSCULAR DYSTROPHY OF RATS (Einflul] des Latrodectus trldecimguttatus-Giftes (LTG) auf mus- kuldre Dystrophie (MD) der Ratten) P. Stern, K. Valjevac

Wir haben schon auf die ~,hnlichkelt der Botulinus-lntoxlkaHon und progressiven musku- I~ren Dystrophie aufmerksam gemacht. Da sich LTG als ein spezifischer Antagonist der Botulinus-lntoxikation zelgte (P. Stern, K. Valjevac : Arch.Toxikol. im Druck), haben wir dieses Gift auch bei MD Ratten (Methode nach Selye) angewendet. LTG erzeugt bei solchen Ratten Rigor auch der dystrophischen Hinterextremit~iten. Die tretierten Tiere h~ingen I~inger (8-10 Minuten) als nicht-trefierte (2-3 Minuten) an der unteren Seite eines Eisennetzes. Die Kontrollen h~ngen unbegrenzt lang. Dieser Effekt ist wahrscheinlich Resultat einer beschleunigten Freisetzung des Acetylcholins an Endigungen der motorischen Nerven hervorgerufen durch LTG.

Prof. Dr. P. Stern, Pharmakologisches Institut der Medizinischen Fakult~it, Sarajevo, Jugoslawien.

STUDIES ON ENERGY METABOLISM OF AN ISOLATED PERFUSED RAT BRAIN (Untersuchungen uber den Energlestoffwechsel des isollert perfundierten Rattenhirns) R. Stock, G. Krleglsteln, J. Krieglsteln

The object of the present work was to prove the sultablilty of the isolated pert:used rat brain prepared according to Andjus et al. (J.AppI.Physiol. 2_22, lo33, 1967) for studying cerebral energy metabolism. For this purpose substrate and metabolite levels were comparatively studied in rat brain ~n vlvo and in isolated perfused rat brain. Furthermore, the effects of phenobarbital and of an ischemic period of 5 rain on the isolated brain were investigated. Our results demonstrate that the concentrations of hlgh-energy phosphates as well as of substrates and metabolltes of the Embden- Meyerhof pathway in the isolated perfused rat brain do not differ markedly from those in rat brain in vivo examined in this work and from values known from the literature. Pyruvate and lactate are s~gnificantly increased, but the lactate/pyruvate ratio is unchanged. Phenobarbital (1.5 raM) causes statistically significant changes in the metabolic status of the isolated brain (accumulation of phosphocreatine, glucose; a decrease of pyruvate, lactate and cx-glycerophosphate) just as described for intact animals (O.H. Lowry et al . , J.bioI.Chem. 239, 18, 1964; L. Granholm et al . , Acta physiol.scand. 7...44, 398, 1968). After the ischemic period a breakdown of hlgh-energy substrates and an increase of catabolic products occur in the isolated brain in accordance wlth reports on brain in vlvo. Thus, the results presented demonstrate that the isolated perfused rat brain is an useful tool for studying drug effects on cerebral metabolism.

R. Stock, Pharmakologisches Insfitut der Universlt~it Mainz D-65oo Mainz, Obere Zahlbacher Str. 67

R 113

THE INFLUENCE OF DIFFERENT ANALGESICS (MORPHINE, PETHIDINE, PIRITRA- MIDE, FENTANYL) ON CONTRACTILITY OF THE ISOLATED CAT PAPILLARY MUSCLE. ((Jber die Wirkung verschledener Analgetlka (Morphln, Pethldin r Piritra- mid, Fentanyl) auf Herzmechanik und KontraktlJit~t). B.E. Strauer

The effect of morphine, pethidlne, pirltramide, fentanyl on myocardial contractility was studied in the isolated cat papillary muscle preparation. Isotonic afterloaded contractions were employed, the following mechanical parameters have been directly recorded: extent of shortening, velocity of contraction and relaxation, tension development, rate of rise and fall of muscle tension. Force velocity relations were obtained by plotting tension development and the peak velocity of isotonic shortening. At low doses both morphine (1 - 10 pg/ml) and piritramide (0,01 - 1 Ng/ml) exerted slight, but non sign~flcant increases in the extent of shortening, the velocity of shortening, the rate of tension development, cardiac work and cardiac power. No positive inotropic effects were detected for pethidine and fentanyl. At larger doses all analgesics caused dose-dependent decreases in these measures. Equal contractile depressant doses - measured in terms of the 50 % inhibition of the velocity of isotonic shortening - were for morphine: 2000 jug,/ml, for piritramide 1000 ;ug/ml for pethldlne: 100 jug/ml and for fentanyl 10~ug/ml. The results are discussed in relation to equi analgeslc potency.

Dr. B.E. Strauer, Medlzinische Universit~tskllnik D-3400 GSttlngen, Humboldtallee 1

RADIOIMMUNOASSAYS FOR THE DETERMINATION OF THE SERUM DIGOXIN LEVEL AFTER APPLICATION OF DIGOXIN AND DIGOXIN DERI- VATES IN MAN (Radioimmunologische Glykosidbestimmungen nach Gabe yon Digoxin und Digoxinderivaten beim Menschen) H. Strobach, Ko Greefi, F.A. Horster, w. Wildmeister

Durch Immunisierung yon Kaninchen erhielten wir Digoxin-AntikSrper, die den Nachweis einer Digoxinkonzentration ab o, 3 ng/ml Serum erlauben. Da- bei verhalten sich ;3-Acetyldigoxin, ~-Methyldigoxin und Lanatosid C wie Digoxin. Weitere Untersuchungen der Spezifit~[t des Testes ergaben, dass Digitoxin~ g-Strophanthin, k-Strophanthin, Cymaxin, Helveticosid, Convalla- toxin oder Proscillaridin in 5-fach h~herer Konzentration den Digoxinnach- weis nicht st~reno Bei Paiienten mit Herzinsuffizienz fanden wit bei tfiglicher Behandlung mit o, 5 mg Digoxin einen Serumdigoxinspiegel yon durchschnittlich 1, 3 ng/ml (n= 78). Bei t~glicher Behandlung mit o, 4 mg ~-Acetyldigoxin oder o, 3 mg fi-Methyldigoxin fanden wir Spiegel yon 1, 4 bzw. 1, 7 ng/ml (n= 81 bzw. 73). Die interindividuelle Streuung des Digoxinspiegels betrug o, 5 - 3 ng/ml.

Bei gesunden Versuchspersonen erreichte der Glykosidspiegel bei einmaliger ora ler Gabe von Digoxin (1, o rag) nach 4 Stunden und von ~-Methyldigo- xin (% 8 mg) nach 2 Stunden den hSchsten Weft.

H. Strobach~ Arbeitsgemeinschafs Klinische Pharmakologie, Universitfit Diisseldorf, 4 Diisseldorf, Moorenstr. 5

R 114

CALCIUM EFFLUX FROM CARDIAC SARCOPLASMIC RETICULUM FRACTIONS INTO CALCIUM- AND ATP-"FREE" MEDIA (Uber den Calcium Efflux aus Vesikeln des sarkoplasmatischen Reticulums in Calcium "freie" plus ATP "freie" Media). J. Suko

Sarcoplasmic reticulum fractions (SR) were prepared from dog hearts (12,000-40,000 x g fraction, additionally extracted with .6M KCI) and loaded with various concentrations of calcium at 37 ~ , pH 7.0 (medium in mM: 40 Histidine buffer, 100 KCI, 2 Mg, 2 ATP, 20xalat% 0.1 45CaCI~, 0.1EGTA, 0.1 mg protein/ml). The rate of calcium efflux folIowing the addition of EGTA (8mM final concentration) was about 0.03 ~moles/mg protein x min. Calcium efflux was increased 15 to 20 fold upon addition of 10 mg/ml Apyrase (Sigma, Grade I) plus 8 mM EGTA. The supernatant of boiled Apyrase plus EGTA did not increase the rate of calcium efflux above the calcium efflux observed with EGTA alone. The rate of calcium uptake and the calcium storing capacity of SR were unaltered by preincubation of SR with high concentrations of Apyrase. The velocity of calcium efflux in calcium- and ATP-"free"media depends strongly on the intravesicular free calcium concentration (varied by 2 to 10 mM oxalate in the medium) and follows Michaelis-Menten kinetics.

Dr. J . Suko, P h a r m a k o l o g i s c h e s I n s t i t u t d e r U n i v e r s i t ~ t Wien, 1090 Wien, W~hringer Strafe 13a.

N-HYDROXYLATION OF 4,4'-DIAMINODIPHENYLSULPHONE IN LIVER MICROSO- MES AND IN VIVO (N-Hydroxy l i e rung von 4 , 4 ' - D i a m i n o d i p h e n y l s u l f o n in Lebermikrosomen und in v ivo ) S. T a b a r e l l i and H. Uehleke

Frequent s ide e f f e c t s dur ing Dapsone ( 4 , 4 ' - d i a m i n o d i p h e n y l s u l - phone, DDS) therapy are o x i d a t i o n of haemoglobin, haemolysis and bone marrow depress ion . N -hyd roxy la t i on of sulphonamides is the cause of haemoglobin o x i d a t i o n (Thauer et a l . Arch.exp. Path. Phar- mak. 252, 291, 1965). N-hydroxy me tabo l i t es of DDS (DDS-NOH) were e x t r a c t e d i n to eh te r and est imated by complexing Fe 2+, formed by reduc t i on of f e r r i c i r o n , w i th ba thophenan th ro l i ne . N -ox ida t i on products were charac- t e r i z e d by t . l . c . ; d e t e c t i o n wi th AgN03 or w i th pentacyanoamine- f e r r a t e . A f t e r ora l doses of 50 mg/kg DDS female beagle dogs excre ted in the u r ine w i t h i n 24 h the f o l l o w i n g percentages of the dose: 0 - i % f ree and 4-8 % conjugated DDS-NOH. The u r i n a r y e x c r e t i o n of men (200 mg DDS o r a l l y ) in 24 h was: 1-3 % f ree and 8-25 % conjugated DDS-NOH. Exc re t i on cont inues s low ly a f t e r 24 h, During aerob ic i ncuba t i on of DDS ( i mM) w i th l i v e r microsomes ( I mg p r o t e i n / m l ) and an NADPH-regenerating system (0,36 mM) N-hydroxy me tabo l i t es were formed at the ra te of 3-4 nmoles/mg p ro te in in 1 min. The re- ac t i on was l i n e a r f o r 5-8 min. Microsomal N -hyd roxy la t i on of DDS was 70 % i n h i b i t e d by metyrapone ( i mM) and 40-50 % i n h i b i t e d by CO (2 % 02 + 98 % CO). I s o l a t i o n and i d e n t i f i c a t i o n of the N- hydroxy me tabo l i t es revea led the presence of 4 - h y d r o x y l a m i n o - 4 ' - amino d ipheny lsu lphone.

Dip l .B io-Chem. S . T a b a r e l l i , Prof . H.Uehleke, Pharmakologisches I n s t i t u t der U n i v e r s i t ~ t , D-7400 TUbingen, Wi lhe lmstrasse 56

R 115

ATPase ACTIVITY AND PHOSPHORYLATION OF THE MEMBRANE OF ADRENAL MEDULLARY STORAGE VESICLES(ATPase Aktivit~t und Phosphorylierung der Membran der Catecholamin-Speichervesikel des Nebennieren- markes. G.Tau~ner und F.John. Cleavage of ATP and phosphorylation of membrane structures has been correlated by Trifar6 and Dworkind(Mol. Pharmacol.~,52(197~) to an active release mechanism for catecholamine. The isolated membrane of catecholamine storage vesicles,however, regulary take up catecholamine in presence of ATP. 0nly,if essential SH-g~oups of the membrane are blocked by NEM whereby the ~TPase is partially blocked, the uptake is abolished and a release occurs in presence of ATP. ATP, added before the addition of thiol-reagent protects the transport functions indicating that ATP binding to membrane structures prevents thiol blockage. On the other hand, NEM blocks partially 3zP-ATP incorporation into the membrane. The number of SH-groups which are protected by ATP against thiol blockage and the number of bound 32 P-ATP are of the same order of magnitude. #gC-ATP binding to the membrane is somewhat lower. The transfer of P from ATP to ADP according to the reaction: ATP+E.----~EP+ADP, proves that during uptake of catecholamine phbs- phorylation of the membrane occurs. The rate of the ATP-ADP exchange is relatively slow, in comparison to52P-ATP incorporated into the membrane, indicating that both, phosphorylation and ATP binding to the membrane are essential for catecholamine storage. No relation, however, has been found between phosphorylation or ATP binding to the release of stored catecholamine.

G.Taugner, Max-Pla~ck-Institut(Physiologie),Jahnstr. 29 69 Heidelberg

INDUCTION OF A DNASE I WITHIN KIDNEYS OF mICE BY TOTAL-BODY X-IRRADIATION OR 2,4,6-TRIETHYLENEImINO-I,3,5-TRIAZINE (Induktion einer DNaSB I in m~u- senieren dutch ganzkSrper-R~ntgenbestrahlung oder 2,4,6-Tri~thylenimino- 1,3,5-triazin) K.Tempel

Specific in vitro-activities of the DNase I (EC 3.1.4.5) has been increased within kidney-homogenates of rats and/or mice, depending on the dose, after total-body X-irradiation (150-2340R, 70R/min)(TBI) or i.v. injections of 2,4,5-triethyleneimino-l,3,5-triazine (0,8-4,8 mg/kg)(TE~).~axi- mum values (+25 to +250%) have been reached within 3 (TBI) and 3-7 (TE~) days resp. - In vitro-activity of a (protein-like) DNase I-inhibitor, apparent K -values, degree of subcellular structural latency, and distribution of The DNase 1-activity (cortex:medulla=2:l) remained unaltered during the first three days after T81 or inj. of TEm, whereas the specific activity of mitochondria-bound DNase I decreased under the same conditions. - The increase of DNBBe l-activity in kidney-homogenates of TB!- and/or TEm-treated mice could be inhibited, depending on the doses, by actinomycin O (0,04-0,4 mg/kg i.v.), actidicne (0,5-4,0 mg/kg i.v.) or T81 (70-280R), when given 24 and 48 hours after the initial treatments. - DNA (calf thymus, mol.-weight 8.900.000, 12,5-50,0 mg/kg i.v. daily) increased DNase l-levels of the kidneys of mice by about 10-50%. This effect could be enhanced by about 50-400%, when DNA (12,5-25,0 mg/kg) was given to Tel- and/or TEm-treated animals. - In respect to the occurence of deoxyribopolynucleotides in the plasma, and the elevated DNA-content of the kidneys of irradiated and/or TEm-treated animals present results suggest, that induction of DNase I may be mediated by DNA (or DNA-frag- merits) coming out of damaged tissues to be metabolized within the kidneys.

Dr.K.Tempel, Univ.-Doz., Inst.f. Pharmakologie, To• und Pharmazie der Tier~rztl. Fak.d. UniversitGt mOnchen, 8 ~Onchen 19, KSniginstraBe 16

R 116

AN IMPROVED METHOD FOR SIMULTANEOUS MEASUREMENT OF HEAT PRODUCTION AND SOME PARAMETERS OF MECHANICAL FUNCTION OF ISOLATED HEARTS. (Eine ver- besserte Methode zur gleichzeitigen Erfassung der W~rmeproduktion und einiger Funktionsparameter isolierter Herzen.) M. Theisohn, K. G~ttler, W. Klaus und Io Theisohn-Schwedhelm Ausgehend van der Methode van McDonald (Am. J. Physiol. 22o:894-9oo, 1971) wird ein verbesserter Versuchsaufbau zur gleichzeitigen Messung der W~rmeproduktion und wichtiger Funkt• des isolierten Herzens vorgestellt. Das Herz wird volumenkonstant in einem Dewargef~ ~ber die Aorta perfundiert. Die auftretenden Temperatur~nderungen an Einflu~ und Ausflu~ werden mit NTC-Widerst~nden getrennt gemessen. Tem- peratur~nderungen van weniger als I/1oooo C (entsprechend 9oonV an der Br~cke) konnten nach Verst~rkung genau bestimmt werden, Der W~rmever- lust des Dewargef~es betr~gt o.43 cal/min/~ sder 2-3 % der produzier- ten W~rmemenge. Die Verz~gerung der Temperatureinstellung zwischen Ein- flu~thermistor und Ausflu~thermistor bei einem Durchflu~ van 25 ml/min betrug weniger als Io min. Durch die getrennte Messung der Einlauf- und Auslauftemperatur kann diese zeitliche Verz~gerung kompensiert werden. Die Verwendung eines offenen Perfusionssystems mit gr~eren Flu~ge- schwindigkeitenals bei Mac Donald erm~glicht eine bessere Versargung des Myokards und bietet f~r pharmakelogische Untersuchungen den Vorteil der leichten Variabilit~t der Versuchsbedingungen.

Inst. f. Pharmakologie der MHH, D-3ooo Hannaver, Raderbruchstr. 1oi

ON THE ACTION OF DIHYDRALAZINE AND RELATED HYDRAZINES ON MICROSO - MAL DRUG OXIDATION. (0ber die Wirkung von Dihydralazin und verwandter Hydra- zine auf die mikrosomale Oxydation von Arzneimitteln) R. Tiia and K.J. Netter

The antihypertensive substances dihydralazine and hydralazine as well as their congener 1-naphthylhydrazine were tested for their ability to inhibit microsomal mixed function oxidations. Dihydralazine and hydralazlne inhibit the oxidative dealkylation of p-nitroanisole in phenobarbital stimulated rat liver microsomes by about 60 per cent when substrates and inhibitors are present in concentrations of 10-~4M. The type of inhibition is competitive, K 1 is in the order of 2 .10-5M. In some experiments a mixed type of inhibition was observed. Naphthylhydrazine produces complete inhibition at 10-4M. Ana- lysis of the binding spectra of the three inhibitors reveals a concentration dependent formation of a type 11 spectrum. Spectral binding constants (Ks) for dihydralazlne and hydralazine are about 10-4M and thus do not coincide with the inhibitor constant, as has previously been shown to be the case with another competitive type II inhlbitor~ metyrapone. Upon reduction the difference spectra do not disappear. Instead they show a strong maximum at 443 nm. This applies also to naphthylhydrazine, which has a K I and Ks one to two orders of magnitude lower than the other inhibitors. Reduced binding spectra have so far been described only for metyrapone (c.f. Kahl and Kahll N.S.Arch. Pharmak. 270 R 68, 1971), CO as well as CCI 4 (Reiner and Uehleke, ibid. R 111), and metabolites of safrole (Lake and ParLe, Pers. Commun.). The spectral binding constant for dlhydralazine in clithionite reduced microsomes is approximately 10-5M. Thus the hydrazines show properties that are similar to those of metyrapone. This wo~ was supported by the Deutsche Forschungsgemeinschaft. Prof. Dr. K.J. Netter, Dep. of Pharmacology, University of Mainz, D 65 Mainz, Obere Zahlbacher StraBe 67.

R 117

REDISTRIBUTION OF NORADRENALINE (NA) AS A FACTOR INFLUENCING THE RATE OF RE- LAXATION OF AORTIC STRIPS (RGckverteilung yon Noradrenalin w~hrend der Er- schlaffung von Aortenstreifen nach einer Inkubation mit Noradrenalin) U. Trendelenbur$

Aortic strips from reserpine-pretreated rabbits were incubated with various concentrations of NA for various times; the rate of relaxation was measured during subsequen~ washing with amine-free solution. Block of COMT by U-O521 (18 ~g/ml throughout the experiment) slowed the monophasic rate of relaxation. Initial treatment with pargyline (P, IO0 ~g/ml for 30 mln) to bloek MAO produced a biphasic relaxation curve: after the usual quick relaxation a second slow phase of relaxation appeared. This second phase of relaxation became more prominent when the incubation time was increased from 5 to 30 to 90 m in. It was prevented if cocaine (IO ~g/ml) was present during the incubation with NA. The second phase of relaxation was accelerated if cocaine was added to the amine- free washing solution. The results are not consistent.with the hypothesis that the rate of relaxation is solely influenced by the rate of inactivation of NA (by uptake and metabolism). They support the view that NA enters neuronal (after P) and extraneuronal stores (after U-0521), and that the amine leaves the stores during washout to retard relaxation by acting on the receptors. The shape of the relaxation curve is determined by the rate of efflux which is high for extraneuronal and low for neuronal stores. Rabbit aortic strips show little or no "secondary sensitization", presumably because of the large neuro-museular distance.

U. Trendelenhnrg, Institut fGr Pharmakologie und Toxikologie, 87 W8rzburg, Koellikerstr. 2

THE INFLUENCES OF CA-ANTAGONISTIC COMPOUNDS ON THE ELECTRICAL AND MECHANICAL ACTIVITIES OF THE MAMMALIAN MYOCARDIUM MEASURED IN VOLTAGE CLAMP EXPERIMENTS. H. Tritthart t R. Kaufmann and H.P. Volkmer

The Na- and the Ca-dependent inward currents were measured simultaneously with the tension development in cat papillary muscles and trabeculae, emploing the double succrose gap technique. Verapamil and D 600 depressed selectively the Ca-inward current with noeffect on the Na-current (1,2). The mechanical threshold potential was found to be identical with that for the fast Na-inward current and remained accordingly unchanged during the negativ inotropic action of Verapamil or D 600. The first contraction (f.c.) after lm~re~ was compared with the steady state contraction (s.s.c.) reached after 2-10 subsequent beats. Depolarizing rectangular steps were applied beginning at the holding potential of about -70 mV over a range of 100 mV. The f.c. was essentially constant over the potent~l range studied, whereas the s.s.c, was increased at -40 to -30 mV, coincident with the onset of the Ca-current, and maximal around za~o potential. Low Ca, Verapamil and D 600 reduced both, the f.c. and the s.s.c, over the whole range of potential tested. The results suggest, that in addition to the depression of the Ca-influx dur~g excitation the compounds tested also diminuished the supply of activator calcium during rest. l) A. FLECKENSTEIN, H. TRITTHART, B. FLECKENSTEIN, A. HERBST u. G.GRUN

This journ. 264, 3, 227 (1969) 2) M. KOHLHARDT, B. BAUER, H. KRAUSE and A. FLECKENSTEIN

Experientia, 1972 (in press~ Adress: Physiol. Institut, D-78 Freiburg i.Br., Hermann-Herders~.7

R 118

INTESTINAL TRANSPORT OF QUATERNARY ANNONIUN CONI~ IN VIVO (Intestinaler Transport quatern~rer Ammoniumbasen in vivo) K.Turnheim and F.Lauterbach

A secretory mechamism of the isolated jejunal mucosa for monoquaternary ammonium compounds was found in vivo (Acta Pharmacol. Toxicol.29,Suppl.4:60,1971). In order to substantiate this observation in vivo amd to examine the relevance of secretion for absor~tion~ I nmole/g N-methyl-3H-scopolamine (NNScop), Nl-me - thyl-~4C-nicotinamide (NYLY), and tetraethyl-1-1~C_ammonium (TEA) were given i.v. to anaesthetized guinea pigs.After 75 min the concentration os the unmetabo!ized base was 5.73 and 4.71 times higher in the jejunal fluid than in the serum with NMN and NMScop~ respectively. The corresponding value for TEA was 2.27 after 480 min. The uphill secretion of NNScop was inhibited by the simultaneous i.v. administration of 400 nmoles/g NNN. The intestinal absorption of I nmole/g NNSeop was characterized by a rapid initial phase and a complete standstill after 45 min, although 80% of the dose given still remained in the intestine. This fact together with the constancy os the blood level after the 15 th min is interpreted by the establishmemt of an equilibrium between absorption and secretion. The simultaneous i.v. administration of 400 nmoles/g NNN increased the absorption of I nmoie/g NMScop by 80%, possibly by inhibiting the secretion of absorbed NMScop back into the gut lumen. Thus the peculiarities in the absorption of monoquaternary ammonium compounds are explained by an interference with a secretory process.

Dr.K.~urnheim,lnstitut fHr Pharmakologie und Toxikologie der Ruhr-Universit~t Bochum,D-4630 Bochum,lm Lottental,W.Germany

C- AND N-OXIDATIONS IN LUNG MICROSOMES, (C- und N-Oxidat ionen in Lungenmikrosomen) H. Uehleke, K.H. Hel lmer and O. Reiner

E a r l i e r i n v e s t i g a t i o n s wi th i s o l a t e d perfused l i v e r and lung of cats (Kiese u. Uehleke: Arch.exp. Path. Pharmak. 242,117(1961) and rabb i t s (Uehleke, Proc. Europ. Soc. Drug Toxic . I 0 ~ ( 1 9 6 9 ) demon- s t r a ted the high a c t i v i t y of lung mixed f u n c t ~ n o x i d a t i o n . Never- t h e l e s s , the s p e c i f i c a c t i v i t i e s of i s o l a t e d r a b b i t and ra t microsomes were cons iderab ly lower in lung microsomes.

With improved homogenization techniques the cytochrome concentra- t ions and the s p e c i f i c a c t i v i t i e s of microsomes (nm~es/mg p ro te in in i min) from r a b b i t lung ( l i v e r ) were as f o l l o w s : y i e l d of microsomes: 4.2 ( lO.7)(mg p r o t e i n / g organ); cytochrome P-450: 0.4 ( 1 . 6 ) ; cytochrome b : 0.26 (1.4)(nmoles/mg p r o t e i n ) ; N-hydro- x y l a t i o n of 4 - c h l o r o a n i l i n e : 2.0 ( 3 . 6 ) ; N-ox ida t ion of N-methyl- a n i l i n e : 3 . 0 ( 3 . 3 ) ; N -ox ida t ion of N , N - d i m e t h y l a n i l i n e : 5.2 ( 6 . 2 ) ; d e a l k y l a t i o n of N - m e t h y l a n i l i n e : 3.0 ( 3 . 0 ) ; d e a l k y l a t i o n of amido- py r i ne : 2.4 ( 0 . 9 ) ; p -C -hyd roxy la t i on of N - m e t h y l a n i l i n e : 2.3 ( 2 . 0 ) ; C -hyd roxy la t i on of 4 - n i t r o t o l u e n e : 2.4 ( 1 . 2 ) .

Phenobarbi ta l p re t rea tment of r abb i t s and ra ts produced no s i g n i - f i c a n t increase in the s p e c i f i c a c t i v i t i e s of i s o l a t e d lung microsomes. Metyrapone, d i s u l f i r a m and benzof lavone e f fec ted s i m i l a r i n h i b i t i o n in lung and l i v e r microsomes. The Vma x and the apparent K M of the s i ng le microsomal ox ida t i ons in lung and l i v e r were in the same range, but Vma x and K M were t en fo ld h igher in lung microsomes fo r the N-ox ida t ion of N - m e t h y l a n i l i n e .

Prof . Dr.med,Dipl.Chem. H. Uehleke, Pharmakologisches I n s t i t u t der U n i v e r s i t ~ t T~bingen, D-74 T~bingen, Wi lhe lms t r . 56

R 119

STUDIES ON SOME METABOLIC EFFECTS OF HALOTHANE IN RATS (Untersuchungen zur metabolischen Wirkungen von Halothan bei Ratten) H. Vapaatalo, A. M~keldinen and P. Nikki Anaesthetics depress thermoregulation and vasoconstr ic tor tone and resu l t in a f a l l of body temperature compensated by post-anaesthetic shiver ing (A. Hemingway, Physiol.Rev. 43,397-422,1963). In addit ion to shiver ing some peripheral mechanisms contr ibute to the return of normothermia. In the present stud~ these metabolic events were examined in halothane- anaesthetized rats (2% halothane for 30 min). Rectal temperature and post- anaesthetic shiver ing were recorded. Plasma FFA, g lycero l and blood glucose were measured at O, 30, 60 and 90 min from the s ta r t of anaesthesia. Temperature was lowered a f te r anaesthesia by 3~ normothermia returned in 30 min and shlver ing lasted fo r 15 min. The FFA leve l elevated by 50%, but blood glucose lowered s l i g h t l y . Propranolol (25 mg/kg) prolonged hypothermia and shiver ing and inh ib i ted the halothane-induced increase of FFA. Pretreatment of the rats with syrosingopine (0.5 mg/kg x 2) diminished, but did not abolish the elevat ion of FFA. Shivering was less in tens ive but prolonged as was hypothermia, too. Halothane induced l i p o l y s i s even a f te r adrenal demedullation and syrosingopine treatment. Rats kept for a week and then anaesthetized at 30~ developed nei ther hypothermia nor shiver ing. However, blood glucose and FFA values were increased a f t e r anaesthesia. In v i t r o studies ~sing rat epididymal fa t pads revealed a l i p o l y t i c e f fec t of halothane (10 uH-1 mH), blocked by propranolol (2 uH). The metabolic e f fec ts of halothane are associated with calor igenesis and they are obviously mediated through the sympathetic nervous system both in an ind i rec t and d i rec t manner. Grants: keiras Oy, The Finnish-Norwegian Foundation for Medical Sciences.

Dr. H. Vapaatalo, Dept. Pharmacol., Univers i ty of Oulu, SF-90 100 Oulu 10.

INFLUENCE OF VASODILAT!NG AGENTS ON OXYGEN 00NSUMPTION IN THE HINDLIMB OF THE CAT. (Ver~nderungen des Sauerstoffverbrauchs an der Katzenextremit~t in situ durch eine pharmakologisch ausge- l~ste Vasodilatation.) F. Vetterlein~ G. Schmidt

Vasodilating agents (papaverine, bradykinin, acetylcholine, co- baltous ions, and isoprenaline) when infused intraarterially increased the oxygen consumption of the resting hindlimb of the cat for about 30 sec. This rise was followed by a reduction of oxygen uptake, while blood flow was still augmented. The calculated oxygen consumption resulted in a negative balance. In contrast to this result, the inhibition of sympathetic constrictor tone was followed by a rise in oxygen consumption. This increase was paralleled by the augmented blood flow. Observations of the microcirculation in skeletal muscle vessels of the hindlimb revealed a stasis and partly disappearence of blood cells in the capillaries in the phase of diminished oxygen consumption. The capillary flow after sympathectomy was perma- nently increased. These results lead to the assumption that the tested vasodilating agents reduce the nutritive tissue perfusion.

F. Vetterlein, Institut fGr Pharmakologie und Toxikologie, Abt. Neuropharmakologie, D-3400 G~ttingen, Geiststr.9

R 120

THE Ca-Mg ANTAGONISM ON THE CONTRACTILITY OF THE GUINEA-PIG PAPIL- LARY MUSCLE IN DEPENDENCE ON THE SODIUM CONCENTRATION (Der Ca-Mg Antagonismus auf die Kontraktionskraft des Meerschweinchen-Papil- larmnskels in Abh~ngigkeit yon der Natriumkonzentration) W. Vierling, M. Relter und F. Ebner

2+ Mg (1-20 mM) wirkt am Meersehweinehen-Papillarmuskel (Krebs-

�9 v " " a

Henselelt-Lsg.35 C, I/see) negat~v inotrop. !m Gegens tz zu den Verh~itnissen am Frosehmyokard (H.Antoni et al.,PflHgers Arch.275, 507-525, 1962) besteht am Meersehweinehen-Ventrikel ein eindeuti- ger Ca-Mg Antagonismus. Die AP-Dauer, welehe dureh Mg 2+ am Froseh- myokard stark verkGrzt wird, ist am Meerschweinchenmyokard m~ssig verl&ngert. Die Ca-Wirkungskurve wird dureh Mg 2+ ~hnlieh wie dureh Na + konzentrationsabh~ngig naeh rechts verschoben. Die Analyse der kompetitiven Ca-antagonistischen Wirkungen von Mg 2+ und Na + in An- lehnung an Lineweaver u.Burk ergibt fur Na eine parabolische Be- ziehung mit einem Exponenten fHr die Frequenz i/see yon 1,7. FHr Mg 2+ besteht dagegen ein linearer Antagonismus zu der die Kontrak- tionskraft fSrdernden Wirkung des Calciums. I0 mM Mg 2+ heben bei 140 mM Na + die positiv inotrope Wirkung yon 1-2 mM Ca 2+ auf. Die negativ inotrope Mg2+-Wirkung wird dureh Reduktion der Na+-Konzen- tration im Bereieh zwisehen_140 und 70 mM etwa im selben Ausmass wie die positiv inotrope CaH+-Wirkung verst&rkt.

Dr.W.Vierling, Institut f.Pharmakologie u,Toxikologie d.Teehni- sehen Universit~t MHnchen, 8 MHnehen 23, Biedersteiner Str. 29

FURTHER STUDIES OF THE RELATIONSHIP BETWEEN THE INSTILLATION VOLUME AND CONCENTRATION OF DRUGS INTRODUCED INT0 THE INTESTINE ANDTHEIR TOXICITY AND EFFICACY (Weitere Untersuchungen zur Bedeutung des Instillations- volumens und der Konzentration fur Toxizit~t und Wirksamkeit yon Pharmaka bei Zufuhr in den Darm) G. Vo~el Experiments with hexobarbital have shown that when it is instilled into the gut its "efficacy" and "toxicity" are largely determined by the volume and concentration of the solution employed. In order to study how the absorption of drugs is influenced by instillation volume, toxicity and efficacy measurements were performed after instilling the following drugs into the stomach or small intestine: phenobarbital sodium, (n)-nic- otine, azeniaspiro-C3a-benziloyloxy-nortropan-8,1'-pyrrolidin~ chloride and atropine sulphate. These drugs were chosen because the first two are usually well absorbed and the latter two are relatively poorly absorbed. The results may be summarized as follows. On instillation into the stomach all the drugs tested were more "toxic" or "effective" when the instillation volume was big. Instilling the drugs into the small intestine they are more "toxic" or "effective" when solved in small volumes. When large volumes are introduced into the stomach almost the whole of the small intestine is filled and the absorbing surface is correspondingly large. When a small volume is instilled into the stomach the absorbing surface is quite small. If the drug is introduced directly into the jejunum its rate of absorption is a mathematical function of the slope of the concentration gradients.

Prof. Dr. G. Vogel, Biologisches Institut Dr. Madaus, 5 KSln 91, 0stmerheimer Str. 198

R 121

ON THE MECHANISM OF ANTICONVULSANT ACTION OF ~-MEXACHLORCYCLOHEXANE AGAINST PENTETRAZOL (Zum Mechanismus der antlkonvuisiven Wirkung yon ~- Hexachiorcyciohexan gegenuber Pentetrazoi) H.-W. Vohland und W. Koransky s Vorbehandlung mlt ~-Hexachlorcyclohexan (a-HCH) bewirk% eine etwa zehntdgige Resistenz der Versuchstiere gegenUber der krampfauslUsen- den Wirkung von Pentetrazol (PTZ). Um Einbl ick in den Mechanismus der ~-HCH-Wirkung zu erhalten, wurde der EinfluB yon ~-HCH auf den S• sel yon PTZ, auf die Permeation yon PTZ in das Gehirn und auf die Sensi- b i l i tY% des Gehirns fUr PTZ untersucht. PTZ und ~-HCH wurden gaschromatographishc gemessen. In v i t r o erhUh% =-HCH die Metabolisierung von PTZ durch Enzyme der keber- mikrosomen um etwa 140 %. In v ivo fUhrt es zu einer VerkUrzung der Halb- wertszei% von PTZ im Gehirn von etwa 3,8 Stunden auf etwa 1,3 Stunden. Die durch ~-HCH induzier%e Krampfresistenz t r i t% iedoch wesentlich frUher zutage als die dutch =-HCH bewirkte Verminderung der PTZ-Konzen%ration im Gehirn. Die Penetration von PTZ in das Zen%ralnervensystem wird dutch a-HCH-Vorbehandlung nich% beeinfluB%. Bei T i t r a t i o n der Pen%etrazol- schwelle und anschlieBender Bestimmung der Konzentration von PTZ und =- HCH im Gehirn zeigte sich, dab ~-HCH im Dosisbereich zwischen 50 und 300 mg/kg zu einer l inearen, konzentrationsabh~ngigen Anhebung der Pentetra- zolkrampfschwelle fUhr% und damit eine Uhnliche Wirkung aufweist wie z.B. Trime%hadion oder Phenobarbital. Dr. H.-W. Vohland Insti%u% fur Toxikologie und Pharmakologie der Phi l ipps Univers i tUt Marburg D-355 Marburg, P i lg r imste in 2

STUDIES OF THE BINDING OF 3H-ANGIOTENSIN I AND I~C-ANGIOTENSIN II IN SPONTANEOUSLY CONTRACTILE AND NON-CONTRACTILE VASCULAR SMOOTH MUSCLE (PORTAL VEIN AND VENA CAVA INFERIOR OF THE RABBIT) IN VI- TRO (Untersuchungen zur Bindung yon Angiotensin Iund II an einen autorhythmischen und einen nicht spontan aktiven glatten Gef~ss- muskel (Vena portae bzw. Vena cava des Kaninchens) D. Voth~ H. L~bcke und M.-L. F~rschler

Rabbit portal vein and vena c~va were incubated i~ha physiological salt solution containing )H-Angiotensin I or • sin II of high specific activity. The bound radioactivity in the vascular smooth muscle was estimated by subtracting from total tissue radioactivity that contained in the extracellular fluid volume determined by inulin diffusion. The reversibility of binding, the high affinity of the vascular smooth muscles to angiotensin I and II and the specifity of binding strongly indicate that the binding sites demonstrated here both for angiotensin I and II seem to be very important for the biological response and functional behaviour of the two smooth muscles. The binding sites especially fPr angiotensin I are correlated to the functional difference of the autonomously contractile vascular smooth muscle and those without this functional speciality.

Prof. Dr. D. Voth, Neurochirurgische UniversitRtsklinik Malnz, D-6500 M a i n z , Langenbeckstrasse I

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CORRELATION BETWEEN THE PHARMACOKINETICS AND THE HYPOTENSIVE ACTION OF HYDRALAZINE IN THE RAT (Korrelation zwischen der Pharmakokine- tik und der hypotensiven Wirkung yon Hydralazin bei der Ratte) J. Wagner, P.R. Hedwall

The correlation between drug concentrations and pharmacological activity - one important aspect of pharmacokinetics - was studied using hydralazine. The hypotensive action of this drug is general~ agreed to be due to a decrease in peripheral vascular resistance, probably resulting from a direct effect on vascular smooth muscle. In autoradiographic and distribution studies high concentrations of the drug could be demonstrated in the walls of the main arte- ries, and specific accumulation could still be shown 6 and 24 hours after intravenous administration. In non-anesthetized rats the course of the effect of intravenous doses of 1 or 5 mg/kg hydralazine on blood pressure was investigated together with the concentrations of drug plus metabolites (H) in blood, aorta and mesenteric artery. Finally, in studies in the isolated, perfused mesenteric artery, inhibition of the vasoconstrictor response to nerve stimulation and H concentrations in the perfusing solution and the artery were related. Threshold concentrations of H in blood or perfusion fluid were found to be in the range of 0,i pg/ ml, those in the mesenteric artery wall in the range of 5 Ng/g" Thus in both in vivo and in vitro experiments a definite relationship between H concentration and the pharmacological effect could be established. Priv.-Doz. Dr. J. Wagner, Chemical and Biological Research Laborato- ries, Pharmaceutical Division, CIBA-GEIGY Ltd., Basle, Switzerland

INFLUENCE OF TETRACYCLINES ON ISOLATED INTESTINAL MITOCHONDRIA (Zur Beeinflussung der Mitochondrienfunktion durch Tetrazykline) E.Weber, G.GroB, R.Horsch and K.-J.Hahn

Electronmicroscopic observations revealed grossly damaged mitochondria in intestine of humans and animals treated with tetracyclines in therapeutical doses (K.-J.Hahn et al.,Arch.Pharma- kol. 266, 347,1970). This study was undertaken to measure the effect of tetracycline,chlortetracycline and demethylchlortetra- cycline (DMCTC) on the respiratoryrate, the respiratory control index (RCI) and the P/O value in the presence of succinate, glutamate or ~ -ketoglutarate by an oxygen electrode in isolated mitochondria from small intestine of guinea pigs. The activity of cytochromoxydase was also determined. The tetracyclines (7,5-150/ug/ml) induced concentration depending changes in the uptake o~ oxygen (increase in state 4,decrease in state 3) with consecutive depression of RCI and P/O. The effects were enhanced if no Mg-ions were added to the incubation medium and were diminished by rising the Mg-concentration. Mitochondria from animals treated with 32 mg DMCTC/IO0 g b.w.by intragastri8 tube for five consecutive days showed a depressed oxydative phosphorylation. Addition of Mg-ions did not correct the impaired function. The activity of cytochrom-oxydase was slightly reduced. The described alteration of mitochondria might be due to the ability of tetracyclines to chelate with Mg-ions of these organelles. Prof.Dr.Ellen Weber, Medizinische Universit~tsklinik,Abteilung fHr Klinische Pharmakologie, 69 Heidelberg 1,Bergheimer Str.58

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STIMULATION OF RAT GASTRIC HISTIDINE DECARBOXYLASE BY SUBMAXIMAL DOSES OF PENTAGASTRIN. S t i m u l i e r u n g der H i s t i d i n d e c a r b o x y l a s e des Rattenmagens durch submaximale Dosen von P e n t a g a s t r i n . E. Weid le and K . - F r . Sewing

I t i s no t c l e a r whe the r g a s t r i n (G) or g a s t r i n l i k e p e p t i d e s s t i - mu la te g a s t r i c h i s t i d i n e d e c a r b o x y l a s e (HD) d i r e c t l y as suggested by Aures e t a l . ( A m e r . J . P h y s i o l . 219 ,214 -216 ,1970 ) or v i a a nega- t i v e feedback mechanism by r e d u c t i o n of g a s t r i c mucosal h i s t a m i n e (H) as proposed by Kahlson e t a l . ( J . P h y s i o l . ( L o n d . ) 174 ,400 -416 , 1964) . T h e r e f o r e in L a i - r a t s (Gut 5 , 327 -341 ,1964 ) g a ~ i c ac id s e c r e t i o n was s t i m u l a t e d by i . v , i ~ f u s i o n s of p e n t a g a s t r i n (PG) in doses c o v e r i n g the t o t a l dose response cu rve . A f t e r s t i m u l a - t i o n f o r I or 2 hr the an ima ls were k i l l e d and g a s t r i c H was de- t e rm ined f l u o r o m e t r i c a l l y and HD by f l u o r o m e t r i c e s t i m a t i o n of H formed o f L - h i s t i d i n e by mucosal homogenates, Ac id s e c r e t i o n rose in a dose dependent manner r e a c h i n g a maximum of 26 ~Eq H+/IO min at a dose of 2.56 ~ g . k g - ~ . m i n - I PG. HD was s t i m u l a t e d by about I00 % a f t e r 1 hr and by about 190 % a f t e r 2 hr of s t i m u l a t i o n at the h i g h e s t PG dose. The H c o n t e n t was not i n f l u e n c e d by PG what - ever the dose or d u r a t i o n of i n f u s i o n were. I t i s conc luded ( i ) t h a t the s t i m u l a t i o n of g a s t r i c HD is a p h y s i o l o g i c a l f u n c t i o n of G or g a s t r i n l i k e p e p t i d e s , (2) t h a t a r e d u c t i o n of g a s t r i c mucosal h i s t a m i n e by G or PG is a p h a r m a c o l o g i c a l r a t h e r than a p h y s i o - l o g i c a l e f f e c t , and (3) t h a t no n e g a t i v e feedback r e l a t i o n s h i p e x i s t s between g a s t r i c mucosal H and the a c t i v a t i o n of HD.

Doz. Dr. K . - F r . Sewing, Pharmako log isches I n s t i t u t der U n i v e r s i t ~ t D-7400 TUbingen, W i l h e l m s t r a s s e 56

THE EFFECTS OF INTRA-ARTERIAL INFUSION OF ADENOSINE ON SUBSTRATE LEVELS IN SKELETAL MUSCLE (Substratgehaltsver~nderungen im Skelet- muskel dutch intraarterie!le Infusionen von Adenosin) M.Weissel I Go Raberger

Experiments were carried out on 37 urethane-chloralose anaesthetized dogs. loo ~g/kg/min adenosine were infused into one femoral artery over a 5 min. period.Dogs infused with 0.9% NaCI into one femoral artery served as controls. At different time intervals during and after the infusion muscle specimens were taken simultaneously from each gastrocnemius muscle by means of Wollenberge~ clamps precooled in liquid nitrogen and the levels of adenine nucleotides and of glycolytic metabolites enzymically determined. The differences between the substrate levels in the adenosine infused and non-infused gastrocnemii were compared to the corresponding differences in muscle substrate levels in the control dogs. Mean arterial blood flow rose 4-5 fold immediately after adenosine infusion was commenced. Triosephosphates levels increased concomitantly with blood flow during the first 3 min. of the infusion period.The quotients malate/pyruvate,glyc.-1-P/TP,and lactate/pyruvate decreased significantly during this period.Towards the end of the infusion period G-6-P~F-6-P~glyc.-1-P~and malate contents rose substantially.The levels of energy-rich phosphate compounds showed no significant changes throughout the whole observation period. The here presented results indicate that adenosine stimulates glycolysis with a concomitant shift in the redox-potential towards the oxidized state in skeletal muscle.

Dr. M. Weissel~ Institut fHr Pharmakologie und Toxikologie der Universit~t Wien~ lo9o Wien, W~hringer StraBe 13a.

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PURIFICATION AND PROPERTIES OF A HUMAN PLASMA LOW MOLECULAR WEIGHT KININOGENASE (Reinigung und Eigenschaften einer nieder- molekularen menschlichen Plasmakininogenase) U. Wendel~ W. Vest, G. Seidel

Human plasma was stirred with quartz powder. The quartz eluate contained an active kininogenase which was further purified by ammonium sulphate fractionation, preparative isoelectric focus- ing and DEAE-cellulose chromatography. The activated enzyme was associated with non-inhibitory a-globulin. The combination of the two proteins had an apparent molecular weight of 180 000 (gel filtration) and pI 4.7. The enzyme proper was not absorbed from dilute buffer pH 8.0 by DEAE-cellulose and could be separated in this way from the anionic a-globulin. The separated enzyme (kininogenase) behaved like a uniform protein in electrophoresis (cellulose acetate, pH 8.6) and had a mobility like a fast y-globulin, a molecular weight of 36 000 anda pI value of 7.9 to 9.1. The enzyme had esterolytic activity for substituted arginine esters (spec. activity 20 000 mU/mg, 25 ~ ) and weak plasminogen activator activity. It generated kinin from partially purified human kininogen. The esterolytic activity was inhibited by DFP, Trasylol and SBTI, slightly by EACA and not by LBTI, ovomucoid and heparin.

Dr. W. Vogt, Max-Planck-Institut fur experimentelle Medizin, Dept. of Biochemical Pharmaaology, D-3400 GSttingen, Hermann- Rein-Strafe 3.

INFLUENCE OF HEAVY WATER ON THE NARCOSIS OF FISHES (Inhibierung der Narkosewirkung yon ,,Metaoain" bei Fisehen dutch D20)M.Wenzel~ P.E. Haeser, E. Lehr, F. Meyer und G. Werner)

B r i n g t man F i s c h e i n W a s s e r , da s 6 0 - 1 0 0 mg/L m - A m i n o - b e n z o e s ~ u r e - ~ t h y l e s t e r ( M e t a c a i n ) e n t h ~ l t , so w i r d i h r e F l o s s e n b e w e g u n g s i - s t i e r t , und s i e s i n k e n i n S e i t e n l a g e zu Boden; d i e K i e m e n a t m u n g b l e i b t j e d o e h e r h a l t e n . D i e s e N a r k o s e w i r d i n s t e i g e n d e m MaBe i n h i b i e r t , wenn man dem A q u a r i e n w a s s e r i n z u n e h m e n d e r Menge D20 z u s e t z t . D i e s e r S c h u t z g e g e n d i e , , M e t a e a i n " - N a r k o s e k ~ n n t e d u r c h den A u s t a u s e h yon W a s s e r s t o f f b r ~ c k e n g e g e n D e u t e r i u m b r d e k e n e r - k l ~ r l i e h s e i n , da l e t z t e r e e i n e g r ~ B e r e S t a b i l i t ~ t b e s i t z e n und d a d u r e h a u c h k S r p e r e i g e n e n M o l e k f i l e n e i n e e r h S h t e S t a r r h e i t und S t a b i i i t ~ t v e r l e i h e n ( E . L e h r e t a l . , N a t u r w i s s e n s e h a f t e n 5 7 , 5 2 1 , 1970; M. W e n z e l e t a l . , H o p p e - S e y l e r ' s Z . p h y s i o l . Chem. 351, 757, 1 9 7 0 ) . AuBerdem w i r d T r i t i u m - W a s s e r s o w i e d a s N a r k o t i k u m P r o e a i n - und d i e s e s b e s o n d e r s im G e h i r n - w e s e n t l i e h v e r l a n g s a m t a u f - genommen, f a l l s d i e F i s e h e s i e h i n 40%igen D20 b e f i n d e n . F e r n e r e r n i e d r i g t s i c h im D 2 0 - h a l t i g e n W a s s e r d i e A t e m f r e q u e n z und d e r 0 2 - V e r b r a u e h d e r F i s e h e .

P r o f . Dr . M. W e n z e l , B i o l . - C h e m . A b t . d e s P h a r m a z e u t i s e h e n I n s t i - t u t e s F r e i e U n i v . B e r l i n , D-1 B e r l i n 35, K S n i g i n - L u i s e - S t r . 2 - 4 . Dr . E. L e h r , M a x - P l a n c k - I n s t i t u t f u r H i r n f o r s e h u n g , A r b e i t s - g r u p p e N e u r o e h e m i e , D-6 F r a n k f u r t / M . , D e u t s e h o r d e n s t r . ~6

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A QUANTITATIVE ANALYSIS OF HEART RHYTHM CHANGES CAUSED BY NARCO- TICS ON MICE (Die quantitative Analyse der dureh Narkotika verursachten Herzrhythmusver~nderungen bei M~usen) G. Werner~ E. Lehr~ P. E. Haeser

Especially on mice tests were made with a new electronic method allowing measuring the ehanges of the heart rhythm after the giving of urethane, nembutal or chleralhydrate. Urethane causes bradyeardia in narcotics doses. But this heart periodicism shows a special arrhythmia. Heart frequency changes periodically every 7-8 min with a great regularity within a range of 2 o.p.s. The dimension of arrhythmia is changed correspondingly to the shifting of period durations. In the maximum frequency a minimum of arrhythmia and vice versa is shown. Nembutal and ehloralhydrate narcosis lead also to stronger bradyeardia; on the contrary however to mescaline (E. Lehr, G. Werner, Arzneim.-Forsoh. 20, 901- 9 0 3 , 1 9 7 0 ; N a u n y n - S c h m i e d e b e r g s A r e h . P h a r m a k o l . 2 6 6 , 3 8 9 , 1 9 7 0 ; E. L e h r , N a u n y n - S c h m i e d e r g s A r e h . P h a r m a k o l . 2 6 9 , 4 4 8 - - ~ 9 7 0 ) t h e r e a p p e a r s no h i g h e r a r r h y t h m i a , b u t t h e h e a r t r h y t h m g o e s on e v e n more r e g u l a r l y t h a n i n u n t r e a t e d a n i m a l s . E a c h o f t h e t h r e e n a r - c o t i c s shows i t s own c h a r a c t e r i s t i c s s l o p e i n t i m e o f t h e a c t i o n on t h e h e a r t .

P r o f . Dr. G. W e r n e r , M a x - P l a n c k - I n s t i t u t f f i r H i r n f o r s c h u n g , A r b e i t s g r u p p e N e u r o c h e m i e , D-6000 F r a n k f u r t / M . - N i e d e r r a d , D e u t s c h o r d e n s t r . 46

CONFORMATIONAL CHANGES IN MEMBRANES BY ~-SYMPATHOLYTIC AGENTS (Konformatlons~nderungen an Membranen dutch ~-Sympatholytika) G.~iethold,B.Lemmer,D.Hellenbrecht,H.Grobecker,D.Palm

The unspecific action of ~-sympatholytic agents can be described as a general impairment of membrane function. Several methods were used in order to obtain quantitative information on these effects. 1.) 1-anilino-8-naphthalene sulfonate (ANS) served as a fluorescence probe for conformational changes in human erythrocyte ghosts. Changes were indicated by the enhanced fluorescence intensity induced by seven ~-sympatholytics,which increased the number of ANS binding sites to a different degree (propranolol K1 255,alprenolol K8 592, oxprenolol,pindolol sotalol,practolol). The apparent dissociation constant remained unchanged. 2.)These conformational changes were well correlated with the pro- tective effect of the respective drugs in human erythrocytes against hypotonic haemolysis (membrane stabilisation,c.f. Seeman, Nature 231 (1971), 284-285). 3.)The s~patholytics investigated inhibited the active transport of H-serotonin in isolated human blood platelets. The potency of inhibition followed nearly the same order as in the experiments mentioned above. It is concluded that conformational changes of membranes may be the basic mechanism by which ~-sympatholytics impair~ membranes function.

G.~iethold,Pharmakologisches Institut der Universit~t D-6000 Frankfurt/Main,Theodor-Stern-Kai 7

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INFLUENCE OF BLOOD FLOW ON INTESTINAL ABSORPTION OF 3-O-METHYL- GLUCOSE (Der E i n f l u ~ der D u r c h b l u t u n g auf d ie e n t e r a l e Reso rp t i on von 3 - O - M e t h y l g l u k o s e ) D.Winne and B , L i c h t e n s t e i n

J e j u n a l loops of a n a e s t h e t i z e d r a t s ( u r e t h a n e ) were pe r fused w i t h 1 4 C - l a b e l l e d 3 - O - M e t h y l g l u c o s e (3-OMG) in b u f f e r e d s a l i n e s o l u t i o n at pH 6 . 8 ; c o n c e n t r a t i o n 37.8 ~M and 38.0 mM. The b lood f l o w , the d i sappea rance r a te f rom the i n t e s t i n a l lumen, and the appearance r a te i n the i n t e s t i n a l venous b lood were de te rm ined s i m u l t a n e o u s l y . At c o n s t a n t i n t e r m e d i a t e b lood f l o w (abou t 1 ml m i n - l g -~ wet t i s - sue w e i g h t ) the appearance ra te decreased from 18.4 to 13.7 nmole min-~g -~ resp . from 17.6 to 20.1 ~mole min-~g - I d u r i n g one hour . A r e d u c t i o n of b lood f l o w from 1.8 to 0.3 ml m i n - l g -~ caused a decrease of the appearance r a te from 20.4 to 5.9 nmole min-~g -~ resp . from 20.0 to 8.7 ~mole m in - "g - I . An augmenta t ion of b lood f l o w from 0.3 to 1.9 ml m i n - l g -~ was not f o l l o w e d by an i n c r e a s e of the appearance r a t e . On the c o n t r a r y i t decreased from 10.3 to 7.9 nmole min-~g -~ resp. from I 0 . I to 8.9 ~mole min-~g -~ . The d i f f e r e n t dependence of the appearance r a te on b lood f l o w in the expe r imen ts w i t h i n c r e a s i n g and d e c r e a s i n g b lood f l o w is ex- p l a i n e d by a decrease of the 3 -OMG- t ranspo r t w i t h t ime as w e l l as by an impa i rmen t o f t h i s t r a n s p o r t d u r i n g and a f t e r a pe r i od of low b lood f l o w .

P ro f . D r .D .Winne , B . L i c h t e n s t e i n ; Pharmako log isches I n s t i t u t der U n i v e r s i t ~ t TUbingen, D-7400 TUbingen, W i l he lms t raBe 56.

ANTITHYROIDAL PROPERTIES OF BUGLE WEED (LYCOPUb0-EXTRACTS (SchilddrUsen- wirksamkeit von kycopus-Extrakten) Hilke Winterhoff and F~ Kemper

In former investigations using histometric techniques koeser and Kemper have shown that extracts of kithosperm or Lycopus are able to diminish or abolish the effects of endogenous or administered thyrotrophlc hormone (TSH). Slight and moreover the interesting early antihormonal effects could not be estimated with these methods. - Thus in new series of experiments on plrbright white guinea pigs (2oo-25o g) the T4-serum contents using the in vitro 125 j "Res-O-Mat" test (Byk-Mallinckrodt) were measured under the influence of o.5 USP-U TSH (Thyreostimulin R Organon): A T 4 maximum (approx. + 3oo ~ compared with the control group) was found between 6 - 12 hi no more effects 96 h post inj. ( i .p.) - Using this experimental basis freeze- dried kycopus extracts of own preparation and plant cultivation were found to be effective in vitro as well in vivo. The dose-effect range in vitro was ~ lo and > 1 mg Lycopus --~ o.5 USP-U TSH; In vivo (injections on different sites) too mg Lycopus abolish the stimulation-effects of o.5 USP-U TSH on the thyroid gland.

Prof. Dr. Foil. Kemperl Institut fur Pharmakologie und Toxikologie der Universit~t MUnster, D-44oo MUnster, Westring 12

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HEPATIC MICR0SOMAL MIXED FUNCTION 0XIDASE AND BENZENE METABOLISM (Benzol-Metabolismus durch die mischfunktionelle 0xydase in der Leber) C_~.Witmer t L. Gonasunt J. Kocsls a and R. Snfder

14C-Benzene metabolism (I) was more vigorous in mouse liver microsomes than in those from the rabbit or rat. I was increased following benzene treatment (II). No change in K M was observed but there was an increase in Vma x. Neither cytochrome P-450 nor NADPH-cytochrome ~ reductase was increased after II. I was inhibited by aniline, metyrapone, aminopyrine, SEF 525A, cytochrome ~, and CO. The CO inhibition was reversed by visible light. I was not inhibited by KCN or aminotriazole. The interaction between benzene and microsomes was characterized by a Type I spectral change. II had no effect on KS but increased the magnitude of the spectral change (~ODmax388_418) by a factor equal to the increase in I following II.

Dr. R. Snyder, Department of Pharmacology, Thomas Jefferson University, 1020 Locust St., Philadelphia, Penna. 19107 (USA) (Instltut fur Toxikologie der Universit~t, D-7400 T~blngen, Wilhelmstr. 56)

RAPID DIAGNOSIS OF DRUG INTOXICATION BY GASCHROMATOGRAPHY AND NITROGEN SELECTIVE DETECTION ( S e h n e l l d i a g n o s e yon A r z n e i m i t t e l - i n t o x i k a t i o n e n d u r e h G a s c h r o m a t o g r a p h i e und s t i c k s t o f f s e l e k t i v e D e t e k t i o n ) H. K~ W i t z m a n n and K. P f l e ~ e r

R a p i d d i a g n o s i s o f d r u g i n t o x i c a t i o n i s p o s s i b l e by i d e n t i f i c a - t i o n and q u a n t i t a t i v e d e t e r m i n a t i o n o f u n c h a n g e d d r u g s i n s t o - mach c o n t e n t , b l o o d and u r i n e . A f t e r i s o l a t i o n w i t h w a t e r i n - s o l u b l e s o l v e n t s t h e e x t r a c t i s r a p i d l y c o n c e n t r a t e d by a d d i n g s m a l l a m o u n t s o f a h i g h b o i l i n g s o l v e n t c o n t a i n i n g d i m e t h y l - p h e n o b a r b i t a l a s i n t e r n a l s t a n d a r d and by e v a p o r a t i n g t h e l ow b o i l i n g s o l v e n t . A l i q u o t s o f t h e c o n c e n t r a t e d e x t r a c t s a r e i n - j e c t e d i n t o two c o l u m n s f o r g a s - l i q u i d c h r o m a t o g r a p h y . The d r u g s a r e s e p a r a t e d by t e m p e r a t u r e p r o g r a m m i n g u s i n g t h e r e t e n t i o n t i m e s and t h e r e t e n t i o n t e m p e r a t u r e s o f b o t h c o l u m n s f o r i d e n t i - f i c a t i o n . To e x c l u d e v o l a t i l e s u b s t a n c e s , w h i c h do n o t c o n t a i n n i t r o g e n , t h e s e p a r a t e d d r u g s a r e d e t e r m i n e d by a m o d i f i e d n i t r o g e n s e l e c t i v e f l a m e i o n i s a t i o n d e t e c t o r (N-FID H e w l e t t P a c k a r d ) . S i n t e r e d a l k a l i h a l i d e s w e r e s p e c i a l l y f o r m e d t o e x - c h a n g e them e a s i l y i n t h e c o l l e c t o r r i n g . T h i s m o d i f i c a t i o n e n a b l e s t o u s e t h e d e t e c t o r as n o r m a l FID o r N-FID w i t h v a r i o u s s e l e c t i v i t i e s (RbDr , CsC1, C s B r ) ~ The i d e n t i f i c a t i o n and d e - termination of amphetamine after oral administration of 50 mg a-phenyl-a-N-(i-phenylisopropyl)aminoacetonitrile (AN-i R) in blood and urimof human volunteers is given as an example of the effectiveness of the method.

H. K. Witzmann, Institut fur Pharmakologie und Toxikologie der Universit~t des Saarlandes, D-665 Homburg/Scar

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EFFECTS OF~-METHYL-DOPA, GUANETHEDINE AND RESERPINE ON BLOOD PRESSURE AND BEHAVIOR IN THE SQUIRREL MONKEY DURING AN AVOIDANCE EXPERIMENT (Zur Wirkung von~-Methyl-Dopa, Guanethidin und Reserpin auf Blutdruck und Verhalten des Totenkopfaffen im Avoidance-Versuch) W. Wuttke

Squirrel monkeys were trained to pressa key a fixed minimum number of times (fixed ratio schedule) w~ich turned off a light and prevented the delivery of an electric shock. Light periods alter- nated with time out periods, during which key pressinghad no scheduled consequences. Monkeys performing under such a schedule show episodic increases in mean arterial blood pressure during the light periods. (J.A. Herd et al., Amer.J.Physiol. 217, 24-29, 1969). ~-methyl-dopa, guanethedine and reserpine were found to affect key pressing behavior, mean arterial blood pressure level during time out periods and the elevated blood pressure during light periods in different ways. After~-methyl-dopa key pressing behavior and blood pressure level were decreasedwhile the episodic increases in blood pressure during the light periods persisted. Guanethedine decreased blood pressure level and diminished the blood pressure elevations during light periods but did not change key pressing behavior. After low doses of reserpine the episodic blood pressure increases were diminished, while after higher doses key pressing behavior, bleed pressure level and blood pressure elevations during light periods were decreased.

Dr. W. Wuttke, Inst.f.Pharmakologie der Farbenfabriken BAYER AG, 56 Wuppertal I, Friedr.-Ebert-Str. 217

HEPARIN CONTENT OF GUINEA PIG LIVER AND ITS CHANGE DURING ANA- PHYLAXIS (Heparingehalt der Meerschweinchenleber und seine Be- einflussung dutch den anaphylaktischen Schock) H.P. Zahradnik t R. ~iebi~r A. Jobke and F. Hahn

Heparin (H) was measured in guinea pig liver homogenates after lyophilization and tryptic digestion (L. Freeman et al., Arch. Biochem. Biophys. 70, 169, 1957), and extraction with phenol (~. Hahn et al., Arch. Pharmak. exp. Path. 256, 430-438, 1967), by determination of the metachromatic reacti-~with Azur-A (LOB. Jacques et al., Hey. cam. biol. ~, 740, 1947), inhibition of ri- bonuclease (B. Lorenz et al., Z. ges. exp. Med. 133, 144, 1960), hlstaminase release in normal guinea pigs (~. Ha~t al., Bio- chem. Pharmaeol. 15, 155-160, 1966), and anticoagulant activity (J. Meroier~ BullTSoc. Chim. Biol. 41, 1101, 1959). H "Novc" (ox heparin) served as standard. The Tollowing normal H values were found, in the given order of the methods used: 437.0 + 59.56, 330.1 + 5.03, 177.9 + 10.31, and 60.7 + 14.73, resp~, expressed by IUFkg body weight. In sensitized a~imals killed 5 min after challenging with antigen (10 mg ovalbumin/kg) the H contents were significantly lower (by 34.26, 28.64, 30.97, and 52~ resp.). The H content of the liver was not influenced by injecting either ovalbumin into normal or saline into sensitized animals. The decrease of the histaminase releasing capacity of the liver homogenates indicates a H release high enough to account for the anaphylactic histaminase release supporting the idea of a mediator function of H in anaphylaxis (F. Hahn and W. Schmutzler, Biochem. Pharmacol. 16, 1523-1531, 1967). • Prof. Dr. P, H~b~, Pharmakologisc-~esInstitut der Universi~ ~reiburg, D-7800 Preiburg, Katharinenstr. 29

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INTRACELLULAR STUDIES OF MICROELECTROPHORETICALLY APPLIED TETRODOTOXIN ON SPINAL NEURONES. (Intrazellul~re Untersuchungen Gber die Wirkung von Tetrodotoxin auf Neurone des RGckenmarks). W. Zie~l~nsber~er and E.A. Puil*. Tetrodotoxin (TTX) blocks electricml excitability of a variety of peripheral nerve and muscle cell membranes. The mechanism underlying this action is a specific blockage of the sodium conductance increase during electrical depolarization without alteration of the delayed potsssium conductance change. In the present study, TTX was applied extracellular- ly by means of the microelectrophoretic technique on spinal neu~ones which were recorded intracellularly. The onset and termination of the effect was comparatively slow e.g., application of 50 nA of TTX did not cause a distinct change in the rising phase of the spike (ortho- or anti-dromic) before 2-3 min. Reversion after a maximum effect took 5-25 min. The first sign of the TTX effect was the appearance of a pronounced AB-break. Secondly the B-spike failed sometimes, and finally only a, small depolarizing transient remained. During this time, the membrane potential and resistance were not changed, whereas the electrical excitability of the cell was drastically lowered. TTX did not affect fibers. The depolarization caused by glutamic acid is associated with an influx of Na +. There is evidence from literature that this mechanism can be influenced by TTX. However, in our experiments the depolarization caused by glutamic acid could not be blocked by TTX. In conclusion: the action of TTX on mebranes of spinal nerve cells is similar to that on peripheral excitable structures. The toxin obviously does not interfere with the depolarizing action of glutamic acid.

Dr. W. Zieglg~nsberger, Max-Planck-Institut ffir Psychiatrie, D-8000 M~nchen 23, Kraepelinstra6e 2, Germany. * Fellow of the Medical Research Council of Canada.

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