424 J. Physiol. (1966), 186, pp. 424-438With 15 text-figuresPrinted in Great Britain

FACTORS DETERMINING THE MAXIMAL RATE OF ORGANICANION SECRETION BY THE LIVER AND FURTHER

EVIDENCE ON THE HEPATIC SITE OF ACTIONOF THE HORMONE SECRETIN

BY E. R. L. O'MAILLE, T. G. RICHARDS AND A. H. SHORTFrom the Physiological Laboratory, the University of Liverpool

(Received 15 April 1966)

SUMMARY

1. Bromsulphthalein (BSP) was administered throughout the experi-ments at a constant rate well in excess of its excretory rate, to anaes-thetized dogs in which the common bile duct had been cannulated. Themaximal excretory rate ofBSP into bile (BSP Tm) obtained in this mannerwas greatly elevated by choleresis arising from the administration of bilesalt (usually taurocholate) at constant rate.

2. When bile flow rate was increased in stages by raising the tauro-cholate administration rate, successive increments in BSP excretion ratewere obtained up to a limiting value of about 3 times the original Tm.Beyond this point further increases in taurocholate administration ratecaused either no further enhancement of BSP Tm or a decline in the extentof enhancement produced at a previous lower rate of infusion.

3. When taurocholate maximal secretion was established first, thesubsequent administration of BSP at progressively increasing rates led toreduction in the taurocholate secretion rate.

4. Portal infusion of secretin at constant rate (usually 0x2 units/kg bodywt. min) which caused substantial increases in bile flow rate, had no effecton BSP Tm. Increases of bile flow rate of the same order following constanttaurocholate infusion produced marked elevation of the BSP Tm.

5. These findings are discussed and the following conclusions reached:(a) The limiting factor in BSP maximal transfer is the concentration of

BSP in bile; increased bile flow rate at the site of BSP excretion (canali-culi) produced by bile salt administration permits an increase in theoriginal Tm to occur without the limiting biliary concentration beingexceeded.

(b) There is excretory competition between BSP and bile salt but overa certain range of bile salt administration the facilitatory effects ofincreased bile flow rate outweigh the inhibitory effects due to competition.

BILIARY TRANSPORT MAXIMA

(c) Since secretin administration had no effect on BSP Tm, it is likelythat the hydrocholeresis it produces originates downstream from thecanaliculi, i.e. in the bile ductules or ducts; this supports previous evidenceobtained in a different manner.

INTRODUCTION

If BSP, an anionic dye with a high affinity for plasma albumin, isinjected into the blood stream it is removed almost exclusively by theliver and secreted into bile in high concentration. In these respects it issimilar to some naturally occurring substances, e.g. the bile salts andbilirubin and because of this and the fact it is easily measured in bodyfluids it has been used extensively in the study of hepatic physiology andpathology.

It is well established that when BSP is administered above a certainrate no further increase in the rate of excretion into bile occurs; it has alsobeen shown (Wheeler, Meltzer & Bradley, 1960) that this maximal transferrate from plasma to bile is limited by excretion from the interior of theliver cell into bile and not by uptake from the plasma. The situation inmaximal transfer of BSP may therefore be looked upon as a phenomenonof saturation of the excretory membrane (i.e. the membrane betweenparenchymal cytoplasm and canalicular bile). Maximal secretory rateshave also been determined for taurocholate (Wheeler, Mancusi-Ungaro &Whitlock, 1960; Sperber, 1965; O'Ma'ille, Richards & Short, 1965) andbilirubin (Arias, Johnson & Wolfson, 1961).

While abundant evidence exists that bile salt can displace BSP fromplasma albumin and hepatic intracellular protein (Andrews & Richards,1960), there is very little information on the interaction between bile saltand BSP at the level of the excretory membrane. In 1960 Wheeler,Mancusi-Ungaro & Whitlock reported in a brief note that the maximalexcretory rate of BSP (BSP Tm) was not reduced when it was administeredduring taurocholate infusion. This demonstration showed either (a) thatBSP and taurocholate did not compete with each other at all on the excre-tory membrane, or (b) that if competition existed it was modified in someway by other factors incident to taurocholate administration. In a veryearly stage of our investigation into the excretory interaction betweenBSP and taurocholate we found that administration of taurocholate atconstant rate could greatly increase the BSP Tm. Subsequent experimentswere carried out to gain some insight into the way in which bile salts liftthe barrier limiting BSP maximal excretory rate, with a view to extendingour knowledge of the factors determining maximal secretory rate ingeneral. The effect appears to be due to increased bile flow rate at the site

425

426 E. R. L. O'MAILLE AND OTHERSof BSP excretion (which results from bile salt administration) such thatincreased secretion of BSP into bile is possible without the concentrationof BSP in bile exceeding a certain limiting value.Recent work by Wheeler (1965) suggests that the hydrocholeretic action

of the hormone secretin is exerted on the bile ductules or ducts as opposedto the canaliculi. We have therefore tested the effects of increases in bileflow rate produced by secretin administration on the BSP Tm. On thehypothesis that the elevation of BSP Tm by bile salt administration is dueto the increased bile flow rate at the site of BSP excretion (the canaliculi)a hydrocholeresis originating down-stream from the canaliculi would notbe expected to have any influence on BSP Tm. Our results fully supportWheeler's deduction, which was based on a different kind of indirectevidence.Some of these findings have already been briefly reported (O'Maiille,

Richards & Short, 1966).

METHODS

All experiments were performed on adult mongrel dogs anaesthetized with pentobarbitone(Nembutal, Abbot Laboratories). The abdomen was opened by amedian incision, the commonbile duct cannulated and the cystic duct ligated. In some experiments a catheter was in-serted into a radicle of the splenic vein for the purpose of infusions into the portal system.When urine was to be collected, both ureters were catheterized intra-abdominally.

Sulphobromophthalein sodium (L. Light and Co., Colnbrook, Bucks.) was dissolved in0-154 m-NaCl to give solutions which ranged in concentration from 7 to 18 mM. In earlierexperiments bromsulphthalein supplied in ampoules by Hynson, Westcott and Dunning,Inc., Baltimore, was used after appropriate dilution with isotonic saline. Solutions ofsynthetic sodium taurocholate and sodium glycocholate (L. Light and Co.) were made upas follows: each was dissolved in water to give a 0-15 M solution, adjusted to pH 7-4 with0.15 m-HCl and brought to the final volume with isotonic saline. Sodium dehydrocholatewas prepared similarly but the pH of the solutions used was 76-7 8. Concentrations rangedfrom 14 to 73 mm. The secretin extract used (manufactured by Boots according to the methodof Crick, Harper & Raper, 1949) contained some bile salt but this was insufficient to con-tribute significantly to the bile flow rate changes produced. It was dissolved in isotonicsaline, 3-5-4-0 units/ml.BSP and bile salt solutions were administered systemically and secretin intraportally by

constant infusion pumps. Blood samples were withdrawn from a systemic vein not beingused for infusion, transferred to tubes containing 2-3 drops of heparin solution (1000 i.u./ml.,EvansMedical) and gently shaken. Alternatively, blood was receivedinto heparinized syringesand thence transferred to dry tubes. Bile and urine were collected in graduated tubes.

AnalysesFor analysis of BSP in bile, samples were diluted with distilled water and made alkaline

with m-NH40H (0-5 ml. diluted bile plus 7 ml. NH4OH). Each test sample was read againstits own blank (0.5 ml. diluted bile plus 7 ml. distilled water) at 580 m,u in an E.E.L.'Spectra' photometer. Urine was analysed for BSP in a similar manner. The method usedfor measuring the concentration of BSP in blood plasma has been described previously(Richards, Tindall & Young, 1959) as has that which was used for the determination oftaurocholate, glycocholate and free cholate in bile (O'Maille et al. 1965).

BILIARY TRANSPORT MAXIMA

RESULTS

General design of experimentsIn most cases BSP was administered systemically throughout at a

constant rate well above the expected maximal excretory rate. Failure ofthe output rate of BSP into bile to increase over a period of at least 60 minin the face of a supramaximal BSP infusion was taken as a satisfactorycriterion forthe observed BSP excretory rate being themaximum attainablein the particular circumstances; losses of BSP in the urine accounted foronly a small part of the deficit between input and output of BSP. Inseveral instances peripheral blood samples were taken to confirm that theBSP administration rate was indeed supramaximal; these showed asteadily increasing concentration with time. When this initial BSP Tmwas reached a choleretic, usually either sodium taurocholate or secretin,was administered at a constant rate until new steady bile flow and BSPexcretion rates were attained.

Elevation of initial BSP Tm by taurocholate administrationA representative experiment designed as described above is shown in

Fig. 1. Here BSP was administered throughout at 0 55 ,tmole/kg body wt.min-a rate well in excess of the excretion rate into bile at any time duringthe experiment; the initial maximal excretory rate of BSP was about0 14 ,tmole/kg body wt. min. After this had been established a systemicinfusion of taurocholate at a constant rate of 1-68 ,amole/kg body wt. mmwas given for 96 min; towards the end of this period a steady state existedwith respect to taurocholate (i.e. output rate into bile equalled infusionrate), but now the excretion rate of BSP into bile was about 0-29 ,tmole/kgbody wt. min-more than twice the initial BSP Tm. It can be seen (Fig.1)that this enhanced BSP excretion rate was still considerably below theBSP administration rate, i.e. it represented a new limiting value for BSPexcretion. The administration of taurocholate led to an increased bile flow(bile flow rate is linearly related to taurocholate output (Preisig, Wheeler& Cooper. 1962)), and in this instance the increase in BSP Tm was directlyproportional to the increase in bile flow rate, so that the BSP concentra-tion in bile was unchanged. When taurocholate infusion was discontinuedthe BSP Tm declined to its original value or sometimes below it. In someexperiments when taurocholate was infused at constant rate together withBSP from the beginning a high excretory rate of BSP was achieved whichfell to much lower values when the taurocholate infusion was stopped, thusresembling the second and third phases ofthe experiment depicted in Fig. 1.In all eight of these two types of experiment the BSP maximal excretoryrate was much greater during taurocholate choleresis than in its absence.

427

E. R. L. O'MAILLE AND OTHERS

BSP in I--

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Time (min) from beginning of BSP infusion

Fig. 1. The initial BSP maximral rate of excretion into bile was approximatelydoubled by taurocholate ion. Bars represent the meanirates over thetimes indicated. BSP in---- BSP was administered throughout at a constant rate of0.55 tmole/kg body wt. min. Taurocholate was infused at a constant rate of1-68etmole/kg body wt. min. from 147-243 mn. BSP out-the excretory rate ofBSP into bile. TC out-biiary excretion rate of taurocholate dividedby 15. B.F.R.-bile flow rate. Plasma BSP-wconcentration of BSP in plasma.

The effect of progresiVe increases in taurocholatte administrationrate onBSP Tm

If taurocholate enhances the BSP Tm by its choleretic effect as seemsthe probable explanation, one would expect progressive increases in BSPexcretion rate to be obtained by increasing the bile flow rate in stepwisefashion. A-n experiment which shows the results of this procedure is illu-strated in Fig. 2. BSP was administered throuighout at a supramaximalrate of 0-62 ,imole/kg. -mini; taurocholate was adminiistered at four constantrates (0.73; 1-47; 2-20 and 2-99 /tmole/kg . mini) each stage lasting about70 min towards the end of which time bile was collected. In each of thesestages a steady state existed with respect to taurocholate. In Fig. 2 it can

428

BILIARY TRANSPORT MAXIMA 429

be seen that the first two stages of choleresis elevated the initial BSP Tmto two limiting values, but further increases in taurocholate administrationproduced either no further enhancement (third stage) or an actual reduc-tion in the degree of enhancement obtained previously (final stage). Whenthe maximal excretory rates of BSP were plotted against bile flow rate infour experiments of a design similar to the one that has just been described,it was evident that the increment in BSP Tm per unit increment in bileflow rate (A BSP Tm/Abile flow rate) was greatest at the lower end of theBSP Tm-taurocholate choleresis curve.

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Excretory competition between bile salt and BSPThe fact that after a certain stage further increases in taurocholate

administration either failed to produce a further enhancement of BSPmaximal excretion rate or caused a reduction in the enhancement attained

E. R. L. O'MAILLE AND OTHERS

previously indicated the presence of a second phenomenon namely com-petition between BSP and taurocholate for sites on the excretory mem-brane. Thus when the converse of the experiment depicted in Fig. 2 wastried (taurocholate maximal secretion was established first and thenBSP administered at progressively increasing rates), the administration ofBSP (Fig. 3) led to reduction in the maximal secretory rate of taurocholate.The administration of BSP also caused a fall in the maximal secretoryrate of free cholate. The maximal secretory rate of free cholate was pro-duced after acute taurine depletion (see O'Maille et al. 1965).

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Fig. 3. Notation as in Figs. 1 and 2. The administration of BSP caused a reductionin the maximal rate of taurocholate secretion.

Effects of glycocholate and dehydrocholate on BSP TmThough taurocholate, which is the principal bile salt in natural canine

bile, was the most frequently used choleretic in these studies, the abilityto elevate the BSP Tm is probably common to all the bile salts. Thus whendehydrocholate was administered at constant rate (6-S4.1 ztmole/kg bodywt. min) in three experiments of similar design to those described above,the initial BSP Tm was markedly elevated in all cases. Infusion of glyco-cholate at a constant rate of 2 1 ,umole/kg. min produced a qualitativelysimilar effect.

Summnary of effects of bile salt admninistration on BSP Tmand on BSP concentration in bile

In all experiments of the class described bile salt administration led toan increase in the maximal rate at which BSP could be excreted. The

430

BILIARY TRANSPORT MAXIMA

concentration of BSP in bile during bile salt infusion was either the sameor less than that present during the course of the initial BSP Tm (with theexception of one experiment). Taurocholate administration at or aboveabout 2 ,tmole/kg. min during maximal BSP excretion always led to a fallof the BSP concentration in bile, but even at fairly high rates of tauro-cholate infusion (relative to its secretory maximum-see Discussion) theinitial BSP Tm was still enhanced due to the fact that the increase in bileflow rate was proportionately greater than the fall in BSP concentration.In a representative experiment, for example, the BSP concentration of bileduring initial maximal secretion was 17-2 mm, which fell to 9X43 mm whentaurocholate was administered at a constant rate of 5.1 ,tmole/kg .min;however, the bile flow rate increased from 9 54 to 45 2 ,ld/kg body wt. minand therefore the excretion rate of BSP from 0O 16 to 0 43 Itmole/kg .min;a steady state existed with respect to taurocholate. The relation betweenthe concentration of BSP during the initial maximal excretory rate andthat obtained during the elevated BSP Tm in the course of taurocholatecholeresis is shown in Fig. 4. The concentration of taurocholate in bile(following its administration) always increased to a value of at least5 times that present during the initial BSP Tm.

Secretin experimentsAs in previous experiments BSP was administered throughout at a

supramaximal rate and, after the excretion rate ofBSP into bilehad reacheda maximum, secretin was infused intraportally at a constant rate (usuallyabout 0-2 units/kg body wt. min) for periods up to 80 min, bile beingcollected continuously at suitably spaced intervals. Observations werecontinued until bile flow and BSP excretion rates had remained steadyover a period of at least 40 min. In Fig. 5 (a), which depicts one of these-experiments, it can be seen that after expulsion of dead space bile in thefirst collection period following secretin administration (labelled D, inFig. 5a), there was no significant change in BSP excretion rate comparedto the period before secretin was given. This was in marked contrast to anincrease of 55 % in the initial BSP Tm produced by taurocholate admini-stration at a low constant rate (0 87 ,tmole/kg . min) later in the experiment.The difference between the effects of taurocholate and secretin cholereses

is shown strikingly in Fig. 5 (b). This experiment had the same design asthat depicted in Fig. 5 (a) except that two constant rates of taurocholateadministration were used instead of one. BSP was infused throughout ata rate of 0-48 ,tmole/kg. min which at all times was well above the excre-tion rate of BSP into bile. Point 1 on the ordinate was the initial maximalrate of excretion of BSP. After this had been established a constant infu-

431

432 E. R. L. O'MAILLE AND OTHERS

sion of secretin (0-20 units/kg. min) was given portally for 60 min and a seriesof time bile collections were taken; point 2 was the rate of BSP excretionin the last of these collections after the bile flow and BSP excretion rateshad remained constant for 40 min. The secretin infusion was then stoppedand in a short time the bile flow rate returned to its original value. Tauro-

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Fig. 4. To show the relation between the concentration ofBSP in bile during initialmaximal BSP excretion and that obtained during elevated BSP excretion producedby steady-state taurocholate infusion. BSP was infused at a supramaximal ratethroughout the experiments. Ordinate: BSP conen. during the period of constanttaurocholate infusion divided by that which obtained during the initialBSP Tm.Abscissa: bile flow rate during constant taurocholate infusion divided by that whichobtained during the initial BSP Tm. Data drawn from ten expts. The fall in BSPconcentration which occurred as bile flow rate was raised by taurocholate infusionwas very much less than would have taken place if the effect of the choleresis weremerely to dilute the bile formed during the initial BSPdTm without affecting BSPexcretory rate (indicated by the interrupted curve). In those expts. in whichsecretin had been administered at an earlier stage, the concentration ofBSP in bileduring subsequent taurocholate infusion was significantly greater than that whichobtained during the initial BSP Tm (filled triangles).

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BILIARY TRANSPORT MAXIMA 433

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4E. R. L. O'MAILLE AND OTHERScholate was then administered peripherally at a constant rate of 1X 13 /tmole/kg. min for 80 min and this produced an elevation of the BSP excretionrate to a new limiting value which remained constant for the final 38 minof infusion (point 3). Subsequently the taurocholate infusion was reducedto a constant rate of 0 73 ,tmole/kg. min, which was continued over aperiod of 93 min; BSP excretion rate fell to a lower limiting value whichremained constant for the final 45 min of taurocholate infusion (point 4).A steady state existed with respect to taurocholate at both rates oftaurocholate infusion. The increase in bile flow rate resulting from secretinadministration did not cause any significant change in the initial BSPmaximal rate of excretion (Fig. 5, b), whereas a choleresis of the same orderof magnitude produced by taurocholate administration led to a greatelevation of the initial BSP Tm. Secretin was administered in the mannerdescribed in four experiments and in no instance was there a significanteffect on BSP Tm.When taurocholate was administered after the secretin effect had worn

off the concentration of BSP in bile was significantly greater than thatduring the course of the initial BSP Tm-in one instance up to 30% greater(see Fig. 4).

DISCUSSION

When a maximal rate (a rate which is no longer influenced by increasedpresentation of substrate) can be demonstrated for a transfer process, it isusual to infer that the process is mediated by some structure, i.e. carrier,present in limited quantity in the system. The inference itself is mostprobably a sound one, but the maximal rate so demonstrated may not bedetermined solely by the quantity of mediator present. The findingsreported here indicate that other factors may exert very substantialeffects. A previously accepted BSP Tm has been found to increase up tothree fold during simultaneous transfer of bile salt, even though the latterwas also shown to be a competitor with BSP for excretion.

Since, in the course of BSP transport, the concentration of BSP in bileis very much greater than that in plasma (or liver cell) it seems almostcertain that its secretion is an active process. Even though the true(electro-chemical) gradient against which BSP is transferred cannot bestated because the value of the electrical term involved is not known, it isstill possible to make some valid qualitative observations about the natureof the limitation present in BSP maximal transfer. The following possibili-ties will be considered:

(a) that transfer of BSP from the liver cell interior to bile is limited bythe concentration ratio between these compartments;

(b) that transfer is limited by the concentration of BSP in the bilecompartment alone.

434

BILIARY TRANSPORT MAXIMA

(a) In these experiments the BSP Tm was determined in the course ofa supramaximal infusion, a time during which the concentration of BSPin plasma and liver cell was rising while that of bile remained constant.Since the osmotic work done in transferring BSP from liver cell to bile ata given rate is directly proportional to

In free BSP concn. in biletfree BSP concn. in cell)

a term decreasing in value with time, it would appear that maximaltransfer ofBSP is not fixed as a result ofenergy limitation for doing osmoticwork when calculated in this manner.

(b) The choleretic effect of taurocholate is such that increased transferof BSP can take place without a concomitant rise in its concentration incanalicular bile. This would seem to indicate that the initial BSP Tm waslimited by the canalicular concentration ofBSP. If transfer ofBSP throughthe excretory membrane is considered to be carrier-mediated, then in theseexperiments full saturation of carriers on the cytoplasmic surface of themembrane was at all times assured by an infusion of BSP well above itsrate of excretion. If the permeability of the membrane to the carrier-substrate complex is assumed to remain constant during the experiment,the only step in the transfer of BSP open to modification is the rate ofdissociation of the carrier-BSP complex at the biliary surface of the cell.This rate of dissociation was capable of great increase as evidenced byincreased BSP maximal transfer during taurocholate choleresis. Uphilltransport by a carrier system requires energy injection at some point.These experiments show that increased rate of energy supply, necessaryfor the increased rate of cycling of carrier (which raised BSP transfer mustinvolve) is available, provided the concentration of BSP in bile is kept ator below a certain limiting value. Because of this flexibility in the availablerate of energy supply, the limitation in BSP maximal transfer is likely tobe set by the molar energy requirement for dissociation of BSP from thecarrier-BSP complex. The magnitude of this energy requirement will berelated to the difference between the high concentration of BSP incanalicular bile at Tm and the very low BSP concentration which would bejust sufficient (if no metabolic energy were supplied) to saturate the sameamount of carrier on the biliary surface of the membrane. It is thereforesuggested that the osmotic work requirement for dissociation of one moleof BSP-carrier complex is given by the expression:

RT1ln BSP concn. in bile at Tm(minimal saturation BSP conen.!'

where R is the Gas constant and T the absolute temperature.

435

28 Physiol. I86

E. R. L. O'MJfILLE AND OTHERS

At a Tm this quantity will have reached a maximal value. The size ofthis critical energy barrier may be determined by the available energy ofa single high-energy bond. The energy-utilizing mechanism of the carrieris therefore considered to set an intrinsic limit to the achievable biliaryconcentration, and choleresis to enhance the Tm by permitting increasedtransfer within that limit.

It must be noted that the bile collected is not necessarily identical incomposition with canalicular bile. While it is possible that the undamagedbiliary tract is effectively impermeable to large anions, it is likely to beable to re-absorb fluid and electrolyte, at least to a modest extent.Wheeler (1965) discusses some of the indirect evidence for such re-absorp-tion. We feel that our findings concerning the concentration changesrelative to the initial Tm secretion (Fig. 4) are consistent with the existenceof such re-absorption, its being more likely to affect the concentration ofBSP of collected bile at low bile flow rates, and when bile salt had beenadministered after secretin, when bile duct permeability may be increased.

It has been pointed out that A BSP Tm/A bile flow rate was greatest atthe lower end of the BSP Tm7taurocholate choleresis curve, and that bilesalt administration above a certain rate caused either no further elevationof BSP Tm or a decline in the BSP maximal excretory rate produced bya lower rate of bile salt administration. The maximal secretory rate oftaurocholate was also reduced by the subsequent administration of BSP.These phenomena are interpreted as indicating the existence of competi-tion between BSP and bile salt for carrier sites on the excretory membrane.Thus the initial maximal rate of BSP excretion was O17 +O05 (S.D.)/tmole/kg body wt. min (fifteen experimental animals); this situationwould represent complete saturation of all sites on the excretory membrane(neglecting the very low outputs of endogenous bile salt). The maximalsecretory rate of taurocholate in similarly prepared dogs was 8*5 + 1-6 (S.D.),tmole/kg body wt. min (O'Ma'ille et al. 1965)-about 50 times the BSP Tm.Thus at low rates of taurocholate administration only a small fraction ofthe membrane sites would be occupied by taurocholate and if all sites onthe membrane were common to both taurocholate and BSP the enhancedBSP maximal excretion rates would be linearly or near linearly related tothe increases in bile flow rate produced. However, at higher rates oftaurocholate infusion, with greater encroachment on membrane area, theenhancing action resulting from increased bile flow rate would be to anincreasing extent offset by the reduction in membrane area left over forBSP excretion.

If a choleretic is to enhance BSP maximal secretion according to theabove hypothesis, it is clear that the site at which water is added to thebiliary tree must either be the same or anatomically above that at which

436

BILIARY TRANSPORI MAXIMA

BSP is excreted into bile. The evidence for both BSP and bile salts beingexcreted by parenchymal cells into canalicular bile is overwhelminglystrong (Armstrong, 1947; Mendeloff, 1949; Krebs & Brauer, 1949; Hanzon,1952). The failure of secretin infusion to influence BSP Tm must thereforemean that the hydrocholeresis which it produces originates downstreamfrom the canaliculi, i.e. in the bile ductules or ducts. These results fullysupport a deduction on the hepatic site of action of secretin made recentlyby Wheeler (1965). He found (a) that the volume of 'dead space' bileexpelled following secretin administration was significantly less than thatexpelled after taurocholate infusion, and (b) that secretin administered bythe arterial route (predominantly supplying the duct system) was morepotent in promoting bile flow than when administered into the portal orperipheral veins. The ineffectiveness of secretin hydrocholeresis on BSP Tmmay also explain the observation of Wheeler, Meltzer & Bradley (1960)that spontaneous bile flow rate changes which occurred in their consciousdogs did not affect the BSP Tm, since it seems likely that they resulted,at least in part, from secretin liberation (Preisig et al. 1962; Wheeler, 1965).Another possible component in these spontaneous bile flow changes,namely reflex excitation of hepatic parasympathetic nerves (Tanturi &Ivy, 1938), may also act in a manner similar to that of secretin.The fact that BSP is partly conjugated before being excreted into bile

(Meltzer, Wheeler & Cranston, 1959), does not materially affect the presentdiscussion, since increase in bile flow rate produced by bile salt administra-tion would enhance the transfer of both conjugate and unchanged BSPfrom liver cell to bile.The possibility that these results could be due to hepatic blood flow

changes following bile salt administration must be considered. However,during supramaximal BSP infusion it is unlikely that BSP excretory ratewould be very sensitive to physiological changes in blood flow. Grodins,Osborne, Ivy & Goldman (1941) studied the effects of various bile salts(mostly given by single injection) on hepatic blood flow. In general therewas no indication in their studies that the bile salts had any consistenteffect on total hepatic blood flow. In one experiment in which we estimatedtotal hepatic flow by the Fick principle using BSP as the indicator, wefound that taurocholate administration, at rates shown in previous ex-periments to be sufficient to elevate the BSP Tm, did not cause any signi-ficant change in blood flow.

It is clear that the study of the elevation of the maximal rate of excre-tion ofa substance would be made simpler ifno competition existed betweenthe choleretic agent and the other substance being transferred. There isevidence (Schanker, 1965; Sperber, 1965) that competition does not existbetween organic anions and organic cations concentrated in bile. We are

28-2

437

438 E. R. L. O'MIILLE AND OTHERSat present looking for a suitable non-toxic cationic substance which is con-centrated in bile, with a view to studying the effects of taurocholate-induced choleresis on its maximal excretory rate and the bile concentra-tion associated with maximal transfer. It is hoped that a more detailedpicture of the secretory apparatus of the liver may be built up by thisapproach.We are grateful to Mr A. Liddy and Mr G. Nevins for invaluable technical assistance.

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