S a m m e l r e f e r a t

T H E P H Y S I O L O G I C A L G R A D I E N T S

C. M. CHILD (From the Zoological Laboratory, University of Chicago)

Received for publication, June 17, 1928

That local or regional differences of some sort in the protoplasm con- cerned constitute the primary factors in the pattern of the individual organism appears to be beyond question. If this is true the problem of the nature and origin of such differences is of fundamental significance for biology. In

the simplest organisms regional differences are perhaps limited to those between surface and interior and all radii are essentially similar, except as local environmental action may determine local reversible alterations. Such surface-interior pattern appears in one form or another in all organisms, but in all except the simplest other orders or patterns appear in definite direc- tions and these directions are regarded as axes of the particular orders con- cerned. Such axiate pattern may exist, not only for the individual as a whole, but for particular organ complexes, organs and cells, and axes may extend in all possible directions within the same individual. We commonly speak of such patterns as expressions of physiological polarity and symmetry, that is, we conceive polarity and symmetry as ordering factors of some sort.

The question of the nature of such factors has long been a matter of discussion and speculation and in the past attempts have often been made to account for polarity and symmetry in terms of stereochemical or Other structure which was assumed to be a fundamental inherent property of the protoplasm concerned (e.g., DR1ESCH, 1908; F. R. LILLIE, 1908; MORGAN and SPOONEa, 1909). Not infrequently it has been suggested that such structure is analogous to crystalline structure and even recently interpretations of polarity and symmetry or asymmetry have been suggested in terms of right and left handed crystals (HARRISOS, 1921) or of a space lattice (PRzmRAM, 1921). Irrespective of the fact that no evidence for such structure as a general characteristic of protoplasm has been discovered and that it has never been shown satisfactorily how, if it were present, it could determine the

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localized regional differentiation at different levels of an axis, we find that under experimental conditions polarity and symmetry do not seem to accord with the requirements of such a structure. We should expect the structure of a particular species-protoplasm to be as stable as other species characteristics, but polarity and symmetry show no such stability. They can be reversed, obliterated or established de novo, radial organisms can be made bilateral and vice versa and partial axes are readily induced. Moreover, the experimental conditions which determine these changes are in general merely factors which inhibit or accelerate physiological processes and there is not a scintilla of evidence to indicate, either that they alter a fundamental physical proto- plasmic structure, or that such a structure determines the differences in pattern which appear (CHILD, 1923b).

On the other hand, the conception of polarity in terms of the flow or gradation of formative substances (e. g., SAcHs, 1880; MOR~A~, 1904a, 1905; LOEB, 1924, &c.) requires, either some hypothetical inherent protoplasmic factor which is itself the real basis of polarity and determines the flow or gradation of substances, or a definite relation to environmental factors, such, for example, as gravity, which is not realized in fact, or even if realized, is at best only very rarely effective. Physiological gradations of some sort and often visible structural gradations are very general characteristics of physio- logical axes in both plants and animals, as is evident both from the data of ontogeny and of reconstitution. MOaGA~'s hypothesis of polarity as a gradation of formative substances (MoRoAN, 1904a, 1905, 1907, chap. XVII) recognizes the existence of such gradations, but translates them into terms of hypo- thetical head-forming or hydranth-forming and tail-forming or base-forming substances and fails to account for the origin of the gradations. These axial gradations have impressed various other investigators. BOVER~ (1910), for example, suggested that polarity might be a gradation of some sort. DELLA VALLE (1913) maintained that polarity is not an inherent property of proto- plasm and compared it to gradations arising in chemical systems under the differential action of external factors. On the basis of his studies of echinoderm development RUN~STaO~ (1914, 1915, 1917, 1925) has suggested that polarity and symmetry are essentially concentration gradients of substances. HA~tRISON (1921) admits the existence of the gradients, but regards them as expressions of the fundamental protoplasmic structure. During the course of experimental studies of reconstitution and development the writer became increasingly impressed by the accumulating evidence, both structural and physiological, for the existence of quantitative gradations of some sort along physiological axes. On the basis of this evidence and of the many facts which indicate that polarity and symmetry are fundamentally similar phenomena in different organisms, the hypothesis that the various observed gradations must have a common physiological basis and that this basis must involve metabolic as well as structural factors originated.

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Occurrence and Nature of the Gradients

NORMAL DEVELOPMENT AND RECONSTITUTION. The very general presence of a gradient of some sort, particularly in the polar axis, is indicated by the cytoplasmic structural gradation in many eggs and embryos, both plant and animal, by gradation in rate of cleavage, size of cells and order of growth and differentiation. In reconstitution also graded differences of some sort usually if not always appear at different levels of the body: for example, in various hydroids a difference in rate of hydranth formation, in size of hydranth developed, or in character of form produced by pieces from different levels1; in flatworms, a difference in rate of head formation, in size and character of head, or in ability to develop a head (CHILD~ 1906, 1911a; LILLIE, 1901; S~vlCKIS, 1923 &c.). The differences in character of form usually appear, not as sudden change in character at a particular level, but as a gradation in the frequency of particular forms at different levels. In Tubularia, for example, stolon frequency increases basipetally (CHILD, 1907a), in Corymorpha the frequencies of unipolar, bipolar and multipolar forms change progressively with change in level of body (CmLD, 1926b). In Ptanaria dorotocephala the frequencies of normal head forms and forms more or less inhibited in the median region differ characteristically with body level (CmLD~ 1911a, 1921b). But that such frequency differences are not expres- sions of fixed, inherent characteristics of the protoplasms concerned is clearly indicated by the fact that all of them can be altered and controlled experi- mentally 2. In a few cases among p]anarians and annelids pieces from the more posterior levels do not develop heads (LILLIE, 1901) or develop posterior ends in place of heads (e.g., MOaGAN, 1902, 1904b). However, the fact that it has been possible to determine experimentally by means of different con- ditions which produce similar effects on development whether a hydranth or a basal end shall develop from a given cut surface or region in hydroids (CHILD: 1923a, 1926b, 1927a, b) and whether a head or a posterior end in Planaria (RusTIA, 1925)~ makes it probable that even the factors concerned in determining these different parts are primarily quantitative rather than qualitative. Graded axial differences appear in the development and recon- stitution of axiate organs as well as in the whole organism. In fact the so- called law of antero-posterior development is simply a special case of the more general rule that development and reconstitution follow a definite course with respect to a physiological axis.

t CHILD, 1907b, c, d, e, 1926b; DRIESCH, 1899; GARCIA-BANUS, 1918; (CAST and t~ODLEWSK[, 1903; HYMAN, 1920a; MORGAN, 1901, 1905, 1906, 19081 ]~ORGAN and STEYENS, 19041 WEIMER, 1928.

~BEHRE, 1918; BUCI~ANA~, 1922 ; CHmD, 1916 b, 1920 a, 1921b, 1926 c, 1927 a, 1927 b. Protoplasma. V 9~9

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All these various lines of evidence are at best merely indicative or suggestive of the existence of a physiological gradient or differential of some sort along at least the polar axis, but it is important to note that they indicate or suggest a differential primarily quantitative in character, rather than localized specific or qualitative differences at different levels of the polar axis as the essential factor underlying the axial differences observed. Moreover, the modifications of these differences by experimental conditions indicate, though they do not prove, that polarity and symmetry depend, not on inherent protoplasmic structure, but on graded differences in physiological condition which, like other physiological factors in development and recon- stitution, are influenced by environment.

DIFFERENTIAL SUSCEPTIBILITY. Differential susceptibility was first noted by the writer in the differences in survival time of different body levels and of yonng and old individuals of Planaria (CHILD, 1913a, 1913b, 1914b) in certain chemical agents. Further investigation indicated that these differences paralleled more or less closely differences in the rate of respiratory metabolism and in the light of this evidence it seemed probable that study of susceptibility to KCN or other cyanides or HCN in different organisms and in different physiological conditions, particularly in eggs, other single cells, embryos and other small organisms might afford data of interest and perhaps serve to indicate physiological differences which could not be directly determined in these small forms. Since these agents are powerful inhibitors of most physiological oxidations it might be expected that differences in susceptibility would show some relation to differences in respiratory rate. The use of KCN showed that axial differential susceptibility to this agent, as indicated by cytolysis, disintegration and death in a certain range of con- centration, different for different species, is at least a wide-spread characte- ristic of physiological axes, particularly in the earlier stages of embryonic development and in many of the simpler organism, both plants and animals, throughout life 1.

In the attempt to throw more light on the question of the relation between differential susceptibility to KCN and differences in metabolic con-

1 The following references include most of the published data on differential susceptibility to lethal and highly toxic concentrations of KCN. Algae: CHILD, 1916c, e, 1917a, b, 1919f. Protozoa: CHILD: 1914a; CHILD and DEYINEY, 1925; HYMAN, 1917; B. L. LUND, 1918. Coelenterates; CHILD, 1918, 1919b, 1926a; CHILD and HYMAN, 1919; [-~UXLEY and DE BEER, 1923, WEIMER, 1928. Ctenophores: CHILD, 1917c. Turbellaria: BEnRE, 1918; CHILD, 1913a, b, 1914b, c; SIVlCKIS, 1923. Echinoderms: CHILD: 1915a, 1916a; (~ALIGHER, 1921a; HUXLEY, 1922; MACARTHUR, 1924. Annelids: CHILD, 1917d; GALIGHER, 1921b; HYMAN, 1916a. Ascidians: HUXLEY, 1921. Fishes: HYMAN, 1921. Amphibia: BELLAMY, 1919, 1922; BELLAMY and CHILD, 1924; BUCHANAN, 1926a; CANNON, 1923. Birds: BUCHANAN, 1926b; HYMAN, 1927a, b.

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di t ion the resp i ra tory metabol ism of various inver tebra tes and the effect of KCN on i t were studied, not merely in different regions of the individual , but in different individuals in different physiological condit ion. Some of the results of these invest igat ions led to criticism, both of the hypothesis tha t differences in suscept ibi l i ty to KCN might serve as a rough indicator of dif- ferences in rate of respi ra tory or oxidat ive metabol ism and of the g rad ien t hypothesis in genera l 1. Some of these cri t icisms were due to misunder- s tanding, some to the fact tha t differences in suscept ibi l i ty concern p r imar i ly the body wall in such forms as Planaria while de terminat ions of resp i ra tory ra te include all organs. In spite of the crit icisms the data at hand show or indicate tha t differences in susceptibi l i ty to KCN are very general ly para l le led by quant i ta t ive metabol ic differences.

The question of the physiological significance of differential suscept ibi l i ty to KCN alone soon became relat ively un impor tan t for the g rad ien t t h e o r y ,

i According to LUND oxygen consumption is not decreased by KCN in starved Paramecia~ even in gradually lethal concentrations, and is independent of oxygen con- centration (E. J. LUND, 1918, 1921@ On the basis of these findings LUND criticizes the conclusions from differential susceptibility and the gradient theory. As a matter of fact, however, differential susceptibility ill Para,mec4um concerns the ectoplasm alone while the oxygen consumption of the whole animal is deternlined, i. % the data given by the two methods are not directly comparable. Moreover, according to unpublished work of ~-[YMAN which I am permitted 4o mention here~ KCN brings about a very marked decrease in oxygen consumption in _Paramecium under certain conditions.

C~. D. ALLEN (1919a), HYMAN (1919a), IE. J. LUND (1921b) all found that oxygen consumption in Planaria is decreased by KCN and LUND also found that it is decreased by decrease in oxygen concentration. On the basis of a study of oxygen consumption in Planaria during starvation and of C0~ production in pieces ALLEN (1919b: 1920a) criticized the inferences from differential susceptibility and the gradient theory. ItYMAN (1919b, 1920b) showed, however, that with a longer starvation period than ALLEN used the respiratory changes in P. agilis, the species used by him, parMlded the changes in P. dorotoeephala. HYMAN also found that the differences in oxygen consumption in relation to reconstitution, physiological age, and size and level of piece paralleled the differences in susceptibility (HYMAN, 1919b, d, 1923). CHILD (1919C) found for Planaria that the changes in C0~ production during starvation, as estimated colorimetrically, paralleled the changes in oxygen consumption as recorded by ItYMAN, that susceptibility to lack of oxygen during starvation paralleled susceptibility to KCN and also (CHILD, 1919d) that C0~ production is decreased by KCN and that KCN and lack of oxygen are additive in their effect upon survival time. ~:{,OBBINS and CHILD (1920) showed that differences in C0~ production in pieces from different levels of P. dorotoeephala and at different stages of reconstitution paralleled the differences in oxygen consumption and susceptibility. HYMAN (1916b, 1919a, d, e), GALIOHER (1921b) and ALLEN (1923) also investigated the action of KCN on oxygen consumption in various other invertebrates and found in all cases a decrease with the higher concentrations, but in some forms a primary increase with low concentrations.

29*

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for it was found on further investigation that axial differential susceptibility to other agents, both chemical and physical, showed in general the same relation to the axes as susceptibility to KCN. In the light of these facts the question of the significance of differential susceptibility became a broader one. At present it appears tha t differential susceptibility to chemical and physical agents, within certain limits of concentration or intensity, as indicated by the course of cytolysis, death or other evidences of toxic action and in many cases also by ability to acclimate or acquire tolerance to lower con- centrations or intensities or to recover after temporary exposure, is a very general, if not a universal characteristic of physiological axes, at least in the earlier stages of development and in many of the simpler organisms throughout life. Moreover, except where there is a considerable degree of differentiation, the differences in susceptibility at different levels are not specific for different agents, but show in general the same relation to the axis, though they may differ in degree. I t must be emphasized that this non-specific susceptibility appears when the degree of differentiation along the axis is not great. With the progress of differentiation particular regions or levels may become speci- fically susceptible to particular agents. Even in eggs wii~h a large amount of yolk more or less definitely localized the yolk region may be specifically more susceptible than other regions to certain agents, e.g., fat-soluble agents, in certain concentrations, while other regions are more susceptible to other agents.

Differential susceptibility appears not only as differential death or dif- ferential inhibition, but often also as differential acclimation, differential recovery and differential acceleration. In differential acclimation the regions of the axis which are most susceptible to the gradually lethal or highly toxic concentrations or intensities acclimate or acquire tolerance most rapidly or most completely to a certain lower range of at least many agents. In certain concentrations the death gradient may be completely reversed, the regions originally most susceptible dying last or remaining alive indefinitely, while regions originally less susceptible die or die earlier because they are less able to acclimate (e.g., CHILD, 1913b, 1914b, 1916d; MAcAItTttUI~, 1920) 1. Similarly, on return to the normal medium after exposure to a certain range

1 In differential acclimation of Planaria to low temperature of proper intensity this reversal of the death gradient appears with .~-reat clearness. Death begins at tile posterior end of the first zooid and progresses in the anterior direction instead of pro- g'ressing from the head posteriorly, but as the anterior region becomes acclimated the forward course of desintegration ceases, the end heals and finally a new posterior end may develop in the same temperature which broug'ht about the death of the original posterior region. In other eases the anterior end may be unable to acclimate to a suf- ficient degree to remain alive and may finally die" after perhaps two or three weeks (unpublished, CItILD).

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of concentration or intensity for a certain period the region originally most susceptible recovers most rapidly or most completely, while other regions recover less rapidly (CHILD, 1916d). And finally, with agents, concentrations or intensities whose action is primarily or chiefly acceleratory the most sus- ceptible regions may show a greater acceleration than the less susceptible (BEHRE, 1918; BELLAMY, 1919; HmRIG~S, 1924a).

If these different expressions of axial differential susceptibility actually occur it is evident that differential susceptibility may serve as a basis for the modification and control of embryonic and reconstitutional development. Moreover, we may expect to find that modifications depending on differential susceptibility are not specific for particular agents. I t has long been recog- nized that most, if not all developmental modifications result ing from the general exposure of early stages to external agents are not specific for parti- cular agents. As regards the two most notable cases which were at first supposed to be specific, viz., HERBST'S exogastrula (HERBST, 1892, 1895a, b) or lithium larva and STOCKARD'S MgCl,~ cyclopia in fishes (STocKARD, 1907, 1909, 1910a, b) it has been shown by later work that both the lithium larva (HERBST, 1895; MAcA~TIIUa, 1924 &c.) and the fish cyclopia (Mc CLENDON, 1912a, b; NEWMAn, 1917; HINRICHS, 1925) can be produced by various other agents. At present we know of no other modifications resulting from general exposure of the earlier developmental stages which are specific in character for particular agents and it seems highly probable that quantitative differences in susceptibility along the axes are the primary factors in all such modifi- cations. Four groups of modifications may be distinguished (CHILD, 1916d, 1917d, 1924a). In the higher ranges of concentration or intensity differential inhibition occurs, the most susceptible regions being most inhibited in develop- ment. With a certain lower range of concentration or intensity the region originally most susceptible acclimates most rapidly and in later stages is least inhibited, i.e., the differential acclimation appears in proportions of parts differing from the normal in the opposite direction from those charac- teristic of differential inhibition. On return to the normal medium after exposure to the proper concentration or intensity for the proper period, the region most susceptible to the higher range recovers most rapidly. Lastly, with accelerating agents, concentrations or intensities the most susceptible region may be most accelerated. The forms resulting from differential reco- very and differential acceleration resemble in general those of differential acceleration, but in differential recoveryi:as in differential acclimation there is always more or less primary inhibition, development is slower and the individual smaller than normal, while in differential acceleration develop- ment is more rapid and the individual larger than normal, at least in all cases thus far observed. By varying the concentration or intensity or the period of exposure, or in consequence of individual differences in susceptibility

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in mass experiments it is often possible to obtain combinations of differential inhibition and acclimation or recovery, as indicated by the proportions of different levels of the axes (CHILD, 1916d).

Axial differential susceptibility has been found in all forms thus far examined for this purpose, at least in early developmental stages and often in the simpler organisms throughout life. Up to the present more than two hundred species have been examined, representing the larger animal groups except the arthropods and some fifty species of algae. The following agents have been used to show differential susceptibility: KCN, HCN, various an- esthetics, ethyl alcohol, ethyl ether, chloroform, chloral hydrate, chloretone, ethyl urethane; formaldehyde; weak and strong acids, COe in water, acetic, HC1, H2SOt; weak and strong bases, NH~OH, NaOH; various other electro- lytes, LiC1, the salts of Ringer's solution singly and in various mixtures, KMnO4, HgC12, CuSOt; the alkaloids, caffein, strychnin, pilocarpin, atropin; the basic dyes, toluidin blue, Victoria blue, crystal violet, methylene blue, brilliant cresyl blue, Janus green, neutral red, nile blue sulphate; old culture water, crowding in culture; hypertonic and hypotonic sea water with marine forms; ultraviolet radiation, visible light after sensitization by cosin or other photodynamic dyes, extremes of temperature, lack of oxygen 1.

1 The following references give data on the use of other agents than KCN in differential susceptibility, including both differential death and differential modification of embryonic and reoonstitutionM development. BEHRE, 1918, modification of planarian head, temperature. BILLS, 1924, death gradients in Parameciu~n, various alcohols. BOVIE and BARR, 1924, death gradients in Ameba, ultra violet radiation. BUCHANAN, 192% modification of planarian head, various anesthetics. BELLAMY, 1919, 1922, death gradients and modification of development in frog, CH s O, K Mn 04, HFCI~, Mg Cls, sodimn butyrate, ethyl alcohol. CHILD, 1913b, death gradients in Planaria, ethyl alcohol; 191~d, death gradients in hydroids, ctenophore, polychete annelids, crustacean nerve, ethyl alcohol, ethyl ether; 1916 d, modification of echinoderm development, ethyl alcohol, H C1, CHs C00H, Na0H, NH,0H; in unpublished work HgCl~, CuS0~, LiCl have Mso been used with the same results; 1916c, e, 1917a, 1919f, death gradients in algae, HgCls, CuS04, HCI, ethyl alcohol, neutrM red; 1917d, death gradients and modification of development in polyehete aunelids, HgCl~, CuSQ, NaOH; 1919b, death gradients and differential accli- mation in hydroids, Li C1, Mg SOt, H Ct, ethyl alcohol, ethyl ether, ethyl urethane, neutral red, methylene blue; 1923a, modification of reconstitution in hydroids, MgS04, LiCl, HC1, ethyl urethane, neutral red; 1925b, nmdification of development in hydroids, HCI, LiC], ethyl urethane, neutral red; 1926a, 1927a, b, death gradients and nmdification of polarity and symmetry in hydroid Corymorpha, NaOH, NH~OH, HC1, Ntt4C] , LiC1, ethyl alcohol, ethyl ether, ethyl urethane, chloretone, strychnin sulphate, nicotin, caffein, methylene blue, hypotonic sea water, sunlight after sensitization by eosin. CHILD and DEVINEY, 1925, death gradients iu Paramecium and other protozoa, ~H40H , NH4C1, NaOH, CH s C00H, NaHCOs, NaI-IC0a -~ COs, H C1, Hs S0,, neutral red, methylene blue, ultraviolet radiation, visible light after sensitization by eosin, lack of oxygen. CHILD and HYMAN, 1919, death gradients in Hydra, ethyl alcohol, ethyl ether, neutral red,

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With some of the agents used in ]ethal concentrations the death changes are not readily visible macroscopieally, but in such cases the use of a con- siderable number of individuals and return of samples to water at regular intervals usually serves to show which parts are dead or beyond recovery and which are still intact. In other cases low concentrations of eosin have been used to indicate the dead or dying parts~ these staining rapidly while other parts do not stain.

The studies of head frequency in Planaria in relation to length of piece and level of body and to nutrition, motor activity, age &c. and its modi- fication by external agents indicate that physiological conditions and external agents may show the same differential in their action on head development (BEMIRE, 1918; BUCttANAN, 1922; CItiLD, 1911 a, 1916b, 1920a, 1921b; I-I~NR[CttS 1924a, S~V~CKIS, 1923). The studies on form frequency in the hydroid Cory- morpha in relation to length of piece and level of body and its alteration by external agents (C~uLD, 1926b, 1927 a, b) and the differential modification of echinoderm development by radiation of the sperm (HI~Rm~S, 1926b) and earlier work cited by H!zqamns on differential modification of development by treatment of sperm with various agents, all point to the same conclusion. Moreover, NEWMAN (1917) has shown that monsters resulting from hybridization between different species genera &c. of fishes are similar to those produced by chemical agents and represent in general differential inhibition.

The purpose underlying the use of the apparently miscellaneous assort- ment of agents listed above was in part that of testing susceptibility to agents of widely different constitution and supposedly different method of action on protoplasms, agents which penetrate readily and without producing

methylene blue, Janus ,a'reen. GALIGHER~ 1921a death gradients in echinoderm develop- ment, NH4OH. HINRICHS, 1924a, death gradients in Planaria, eafl~in; 1924b, death ,e'radient.s in P~ra.meciu~n, Ify(lr(r> Pla~taria, oligochete annelids, ultraviolet radiation; 1925, 1926a, b, 1927, modification of development in teleosts, echinoderms and chick. HUXLEY, 1922, t{UXLEY and DE BEER, 1923, lnodifieation of development an(] dediffer- entiation in echinoderm larvae and hydroid, HgCI~. HYMAN, 1920a, modification of rate of reconstitation iu Tubu, laric~, ethyl ether; 1921b, death gradients in telcost and Petvomyzon embryos, NH, OH, CHsCOOH; 1927a, b, death gradients in chick embryos and embryonic hearts, NH~ 0It, NaOH. MACARTHUR, 1920, death ~-radients in Plano~ia~ HC1, NaOH, 1921, death gradients in P~trameeius% Dileptus, Hydra, Planaria, various rbabdocoels and mierodrflous oligochetes, various bases and soIne aeid dyes; 1924; M.di- fication of development, particularly exogastrulation in echinoderms, LiC], CuSO~, HgCls, HC1, CaCls, NaCI, hypotonic and staling sea water. RUSTrA, 1925, modification of biaxial head frequency in Pla~a~'ia~ H C1, ethyl ether, chloretone, low temperature, chloral hydrate, ethyl aleohol~ caffein. A cousiderahle volume (ff data obtained with various agents is still unpublished and in the literature of experimental modification of develop- ment by chemical and physical agents and the literature of physiology there is much further evidence for differential susceptibility.

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appreciable injury and those which penetrate only slowly or only after injury to the surface and to physical agents and conditions as well as chemical agents. The list also includes general protoplasmic poisons and agents which act more or less specifically on particular organs of the higher animals, at least in certain concentrations. Not all of the agents listed have been used on any one form, though on some forms, such as Paramecium (CmLD and DEVINEY, 1925; MACARTHUR, 1921), Corymorpha (CHILD, 1926a), Planaria (BEHRE 1918; BUCKANA~, 1922; CHILD, 1913b and work with various other agents, unpublished; MACARTHUR, 1921) and developmental stages of echinoderms (CHILD, 1916d, Hmaicns, 1925, 1926a, MAcAI~TItUR, 1924) a large number of them have been used and in almost all cases several agents of different con- stitution and supposedly different action have been used.

With all agents used thus far it has been possible to find concentrations or intensities which give differential susceptibilities showing the same relation to the axis concerned, though the differences in susceptibility at different levels may differ widely with different agents and concentrations. The fact that the axial differences in susceptibility are not specifically different for different agents suggests that a general quantitative physiological dif- ferential of some sort is the factor primarily concerned in determining them. Moreover, the general parallelism between differences in susceptibility and in metabolism shows that quantitative differences in metabolism con- stitute a factor in this differential and other methods to be discussed below add further evidence in support of the same conclusion. On the basis of these and many other facts the axial gradients have been called metabolic gradients, but this does not mean that quantitative differences in metabolism are the only, or in all cases the primary factors. The metabolic differences cannot be independent of other differences in protoplasm. The alteration by gravity of polarity in the frog's egg in reversed position (ScHVLTZE, 1894; H_&MMERLING, 1927; PENNERS und SCHLEIP, 1927--28) and the determination of the dorsoventral axis in the sea urchin by centrifugal force suggest that in some cases differences in distribution of yolk or other substances may determine the metabolic differences. Doubtless other protoplasmic differences are also characteristic of the gradients. I~ I~KA (1927) for example has reported a gradient in colloidal dispersion corresponding to the susceptibility and other gradients. A permeability gradient seems also to be a charac- teristic feature of the axial gradient, but, as will appear below, differential permeability is not the only nor the primary factor in differential susceptibility. Further investigation will probably bring to light other axial differentials.

The general conclusions from the data of differential susceptibility are as follows. Axial differential susceptibility, so far as it is not specific for particular agents, indicates the existence of an axial quantitative differential in physiological condition, involving metabolic, as well as other factors. To

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concentrations or intensities of agents which are above the limit of tolerance but not immediately lethal for the form or stage concerned the most sus- ceptible regions are in general the most active physiologically and suscep- tibility decreases with such activity along the axis. The rate or degree of acclimation to a lower range of concentration or intensity and the rate or degree of recovery after temporary exposure to a certain range of concen- tration or intensity and for a certain time also vary with the physiological activity along the axis. The indefinite term, "physiological activity" is used here because; although the facts indicate that we can substitute for it in many cases "respiratory metabolism" or perhaps "oxidative metabolism", it is not certain that this will always hold true.

The susceptibility method alone demonstrates nothing but differences in susceptibility as indicated by cytolysis, death; inhibition acclimation, reco- very; acceleration. I t is at best merely indicative or suggestive. As already pointed out other methods of investigation are necessary for interpretation of the data of susceptibility. But the fact that the relation between suscep- tibility and physiological condition is; within certain limits; similar for the large number of agents employd and for so many different organisms is of considerable interest and requires consideration. I t cannot be assumed that all the different agents in the concentrations or intensities used act in the same way on all the different protoplasms and certainly the protoplasms of different species are specifically different in constitution. Obviously the similarity in the susceptibility gradients observed with different agents must be essentially independent of the particular method of action of particular agents and also independent of the specific protoplasmic constitution in the different species. How then are we to account for this similarity. At present the only interpretation which appears possible in the light of all the facts is in terms of dynamic equilibration and disturbance (CKILD; 1923c). The living protoplasmic system is commonly regarded as a system undergoing continuous dynamic equilibration or perhaps sometimes in or near dynamic equilibrium. The more rapid the changes characteristic of such a system; the more rapid its reaction to any disturbance sufficient in degree to bring about disruption or irreversible or long persistent change in its equilibrium. And on the other hand, the more rapid the change in such a system; the more rapid is its equilibration to slight continuous or ~emporary disturbance. This relation between equilibration and disturbance is characteristic, not only of living protoplasms, but of other dynamic equilibrating systems as well. I t holds for a flowing stream and a man-made machine, as well as for organisms and irrespective of the nature of the disturbance.

Only in these terms does it appear possible at present to interpret the facts of non-specific axial differential susceptibility. According to this view the axial differences in susceptibility are dependent on the difference in rate

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of change characteristic of different levels. Differential death and differential inhibition represent the effects of disturbance of any kind which is sufficent in degree to bring about irreversible or long perisistent changes in the system. Differential acclimation and differential recovery represent the effects of less extreme disturbances and consist in approach or return to the original con- dition. If this interpretation is correct, there is good reason for calling the axial differentials metabolic gradients for the changes which constitute meta- bolism are fundamental factors in the dynamics of living systems.

With increasing differentiation of parts, both in the individual and in the higher groups, specific susceptibilities of particular parts to particular agents may make their appearance, but even an organ with specific suscep- tibility to certain agents may exhibit non-specific differential susceptibility along its axis. In some cases also more or less specific susceptibilities of particular parts appear with low concentration or intensities, while with a higher range the rate rather than the kind of change apparently becomes the important factor and non-specific differential susceptibility appears. In such cases it seems probable that the specific differences of different regions are not sufficiently great to overbalance the quantitative differences. HU'(Lr3Y (1922) has suggested that the amount of surface exposed to the agent may be a factor in the susceptibility of individual cells. This is doubtless true for certain agents which penetrate readily and accumulate in the cell, but for those which penetrate only after toxic action on the surface it cannot be the fundamental factor and for physical agents such as temperature and the penetrating radiations the amount of surface exposed is of little impor- tance. Moreover, most of the data on axial differential susceptibility con- cern stages of forms in which regional differences in exposed cell surface are not great.

AXIAL DIFFERENTIAL PERMEABILITY. An axial differential permeability to NH4OH has been observed with very low concentrations in Paramecium (CH[LD and DEVINE',', 1925) and in various hydroi, ls (CmLD, 1926a and unpublished data) after staining with neutral red, and less clearly a similar differential to acetic acid. The rate of penetration decreases from the apical end basipetally. The concentrations which show this gradient in penetration, however, are only slightly or not appreciably toxic and with the higher, toxic concentrations of weak bases and acids which show a lethal gradient penetration is intantaneous or almost so at all elvels. On the other hand, strong bases and acids, even in toxic concentrations, do not penetrate in appreciable amounts until injury of the surface occurs. Obviously an axial differential susceptibility of the cell surface exists but this is just as obviously not simply a matter of permeability. Chemical agents which penetrate very readily, those which do not penetrate except after injury to the surface and physical agents whose action does not involve permeability, show similar

Sammelreferat 459

susceptibility gradients, and even within single cells, such as the elongated cells of monosiphonous algae (CHILD, 1916C, e, 1917a, b, 1919f, Parameeiu~

(CHILD add DEVINEY, 1925) and various eggs a marked axial differential sus- ceptibility appears, e.g., with basic dyes, even when differential penetration is not evident within the cell. With low concentrations of the basic dyes, neutral red, methylene blue, Janus green &c. a marked gradient in rate of staining is characteristic of axes (MAcARTnUR, 1921): with higher concen- trations this may be absent, so far as can be determined, but the differences in susceptibility still occur. And finally, differential acclimation and recovery evidently depend on other conditions than permeability, for it is the most permeable regions which acclimate or recover most rapidly or most completely. The various data apparently iustify the conclusion that while differential permeability is, or may be one expression of the physiological axial differences, it is not the primary factor in determining differential susceptibility. With some agents or concentrations it may perhaps make the differences in sus- ceptibility greater than they would be otherwise, but it alone does not account for them.

AXIAL DIFFERENTIALS IN RESPIRATION. Respiratory gradients, as indicated by oxygen consumption or CO 2 production or both, have been found in the forms examined for this purpose by determining respiratory rate in pirces from different levels. These include the sponge, Grantia

(HYMAN, 1925). the hydroids, Cory,rnorpha (CHILD and I-IYMAN, 1926) and Tubularia

(ICIYMAN, 1926a), Planaria (HYMAN, 1923) ROBBINS and CHILD, 1920), Nereis and Lumbriculus (HYMAN and GALmHER, 1921) the earthworm and the chick embryo (S~EARER, 1924). HYMAN used the WmKLEa method for determining oxygen consumption, CO~ production was estimated in Corymorpha and Planaria by a colorimetric method and SHEARER obtained his results with a mierorespirometer and KJELDAItL determinations on the pieces following the respiration period, determining the oxygen consumed per co. or per 100 cc. of NH 3 liberated in the KJELDAHL determinations. He also determined in the same way the oxygen consumption of acetone powder of the anterior and posterior regions and found a difference similar to that in the living pieces. As regards the existence of a primary gradient of decreasing respiration from the apical or anterior end all the data agree, but SHEAREWS data show no evidence of the secondary posterior region of high respiratory rate found by HYMAN and GALIGHER in the annelids and indicated as a region of high metabolism by the susceptibility method in both annelids (HYMAN, 1916) and the chick (HYMAN, 1927a, HINRICHS, 1927), as well as in other vertebrates (BELLAMY, 1919; CHILD, 1925c; HYMAN, 1921, 1926b) and in aseidians (CmLD, 1927d) ~.

Only further investigation can determine the reasons for these differences in result, but several possibilities suggest themselves. S~EARERS earthworms were starved for three weeks before the experiments and the low functional metabolism, of the intestine

460 Sammelreferat

All methods of de termining respira t ion in pieces are open to cer tain possible objections, so far as the gradients are concerned. In the first place the separat ion of the organism into pieces may br ing about a change in respi ra tory metabol ism as i t apparent ly does in 2Planaria (HYMAN, 1923 ; ROBBINS and CHILD, 1920). Second, in organisms with a considerable degree of different iat ion of organs the or iginal gradients of the embryo may be highly modified and different in direct ion in different organs. In pieces consist ing of a complex of organs, e . g . , body wall, in tes t ine etc. differences in one direction in one par t of the complex may be masked or over-compensated by differences in the other direct ion in other parts. The methods of differential suscept ibi l i ty have at least the advan tage of work ing with the in tac t individual and of indica t ing regional or local, apparent ly quant i ta t ive differences in physiological condit ion, a t least in the body wall, i r respect ive of differences in in ternal organs. In many cases these methods can be applied to the in ternal organs also, though this usually involves injury to the individual. Third, in the l ight of recent work of WARBURO and others and of the present s ta te of our knowledge concerning susceptibil i ty, the question must be ra ised whether respi ra tory metabol ism is in all cases an adequate or even a correct measure of g rad ien t differences. Four th , the question must also be raised whether any method at present avai lable for de te rmining or es t imat ing resp i ra tory ac t iv i ty can always be t rus ted to give a true picture of physiological

in the posterior pieces may have nlore than compensated the higher metabolism of the posterior growing region. In the case of Planaria the total oxygen eonsmnption is almost immediately and very gxeatly increased by feeding after a period of starvation (I-IYMAN, 1919b: 1920b). I t is also possible that the posterior growing region of the earthworm becomes less active or disappears as full size is attained, as it does in vertebrates and probably in arthropods, though in many of the microdrilous oligoehetes it persists throughout life. Respiration in earthworms of different size has not yet been determined. In the case of the chick embryo the only data on oxygen consumption are those of SHEARER. The high susceptibility of the posterior growing reg'ion indicates, but certainly does not demonstrate high metabolism, but it is possible that oxygen consumption is not an adequate measure of the metabolism of these embryonic stages. On the other hand, in consequence of its high susceptibility this reo-ion may be inhibited by the experimental conditions and its oxygen consumption may fall rapidly. SHEARER speaks of the occurrence of disintegration of the pieces in some of his experiments. And finally, the question must be raised whether the determination of oxygen consmnption per unit vohme of NHa set free in KJELDAHL determinations is entirely satisfactory. Oxygen consumption in a given organ or tissue certainly varies widely without corresponding variation in proten or nitrogen content, and there seems to be no good reason for assuming that it is the same in a part more or less advanced in differentiation, like the head region of the chick embryo, and a rapidly growing, more embryonic region, like the posterior end. This discussion indicates stone of the difficulties involved in obtaining correct and adequate data concerning the gradients.

Sammelreferat 461

condition. In some cases only a few cells of the region or mass may be active and in the respiration of the whole mass, however determined, their activity may be completely masked. Also respiration may vary within wide limite for the same weight of tissue and for the same nitrogen content. The demonstration of respiratory differences along an axis, or of the absence of such differences does not constitute the final and conclusive evidence for or against the existence of a gradient. In most of the cases where the methods of determining or estimating respiration can be used at present, various unknown and often indeterminable factors complicate the observed results.

AXIAL DIFFERENCES 1N OXIDATION-REDUCTION REACTIONS. An axial differential in the reduction of K M n Q has been observed in many forms (CHILD, 1919a, f, 1921c, 1924b p. 87--8, 1925a, 1926a, 1927d; GALIGHER, 1921a), chiefly protozoa, eggs, embryos, hydroids and some algae. A marked axial differential is found in rate of reduction, as indicated by appearance and depth of the brown or blackish color of the oxides in the cells, apical regions showing the color first and later greater depth of color. Although, even in the low concentrations used KMnO 4 apparently penetrates rapidly with injury to the surface and is highly toxic, the axial differential appears only gradually. When the reaction is allowed to continue to completion in an excess of KMnO 4 the organisms are usually opaque black, but some forms, e. g., hydroid planulae (CHILD, 1925), become translucent after dehydration and clearing and in such cases an axial differential in intensity of color appears clearly. The same differential has been observed in sections (GALIGHEH, 1921a). If the animals are killed before exposure to KMnOt they show either no reduction gradient or in some cases for a short time vestiges of the earlier gradient (CmLD, 1919a, 1927d). In all cases observed thus far the reduction gradient has been found to correspond to the susceptibility gradient.

In low concentrations of methylene blue and brilliant cresyl blue the apical regions (Paramecium, various embryos) which stain first in higher concentrations decolorize more rapidly than lower levels or may even remain unstained while basal regions stain (CHILD and DEVINEu 1925 and unpublished data). In low oxygen this decoloration occurs more rapidly.

The indophenol blue reaction (a-naphthol and dimethylparaphenylene diamine) has also been used with various protozoa, embryos and smaller forms and shows a very distinct axial differential in the living organism, but none or only traces for a short time if the organisms are first killed by other means (CHILD, 1915a, 1926a; CHILD and DEVINEY, 1925, also unpublished data). Exposures of the organisms first to one of the agents, then to the other, show that both penetrate readily, but the axial differential in the appearance of the indophenol blue is nevertheless strongly marked, often grading from deep blue apically to slight traces of color basally. The reaction is apparently

462 Sammelreferat

catalyzed to a much greater degree in the apical regions than in the basal and a gradation between the two exists. Discussion of the literature concerning the intracellular conditions which catalyze the reaction is unnecessary here. It needs only be noted that the reaction indicates a quantitative difference in physiological condition along the axis and that this difference coincides as regards region and direction with that indicated by susceptibility, KMnOa and the reduction of dyes.

In Corymorpha the nitroprusside reaction shows a slight color gradation, indicating a glutathione gradient (CHILD, 1926a).

AXIAL ELECTRIC POTENTIAL DIFFERENCES. Such differences have been recorded for various forms by a number of observers and that electric potential gradients are characteristic features of physiological axes seems evident, but at present different investigators do not agree concerning the direction of axial potential differences and the reasons for the lack of agreement are not yet entirely clear. Under these conditions the only inference justified by the facts is that axial potential differences are in some way associated with the metabolic and other Observed differentials. Leaving out of consideration the earlier literature of bioelectric phenomena, only the following citations need be made here. MATHEWS (1903) found distal regions of certain hydroids externally electronegative to proximal regions. According to HYDE (1904) the animal pole of the turtle egg is externally negative to the vegetative pole and an electric polarity is also present in the blastodisc. In the egg of Fundulus an axial potential difference is present but undergoes periodic reversal at certain stages following fertilization. MOaGAN and DIMON (1904) found both ends of the earthworm externally electronegative to the middle. According to HYMAN, (1920a) distal levels of the Tubularia body are externally electronegative to proximal, within the limits of the individual, but in the longer stems the more proximal regions often represent new individuals and show an increase in external negativity over levels just distal to them. Similarly in Corymorpha external negativity was found to decrease basipetally from the apical region with a secondary increase toward the basal holdfast region (CHILD and HYMAN, 1922). HYMAN and BELLAMY (1922) have recorded axial potential differences for various forms, including sponges, hydroids, medusae, a ctenophore, Planaria, Nerds and frog tadpoles. They found that the regions indicated by differential susceptibility and other methods as regions of high metabolic activity are usually electronegative externally to other regions and that most of the motile forms examined orient in an electric current with regions of high external negativity toward the cathode. Lush, on the other hand, finds distal levels of the hydroid, Obelia~ externally electro- positive to proximal down to a certain level, proximal to which an increase in positivity appears (E. J. LUND 1922, 1925). Exposed to a transverse current

Sammelreferat 463

the new outgrowth at the cut end of the Obelia stem is deflected toward the anode, according to LUbtD, because growth is stimulated on the side toward the anode (E. J. LUND, 1925). Usually, however, the deflection of a growing axis is away from the side on which growth is stimulated and LUND has not explained how the opposite effect is brought about in this case.

In plant roots Lu~I) and KENYON (1927) find the region of active cell division at the tip externally electropositive to other levels and some roots show a second region several millimeters from the tip which is externally negative to more basal regions. Earlier investigators have usually found root tips externally negative to more basal levels. The regions of highest positivity correspond to the regions of greatest reduction of methylene blue.

In the hydroid, Obelia, LUND also finds the external surface of the coenosarc electropositive to the inner entodermal surface. OSTERHOUT, DAMON and JACQUES (1927) and OSTERHOUT and HARRIS (1928) state that in single cells of the algae, Valonia and Nitella the inner surface of the protoplasm adjoining the vacuole is positive to the outer surface.

That electric potential differences are present along physiological axes, at least in many of the simpler organisms, seems to be demonstrated, but in view of the present disagreement concerning their direction further investigation is necessary before anything definite can be said concerning their relation to other characteristics of the axes.

The Gradients in Development

THE RELATION OF THE GRADIENTS TO POLARITY AND SYMMETRY. It has long been known that polarity and dorsoventrality can be determined in various plants by differential illumination. Polarity has been determined by the electric current in pieces of the hydroid, Obelia

E. J. LUND, (1921C) and in the alga Fucus (LuND, 1923). GILCHRIST, (1928) has determined a new polarity in the amphibian embryo by means of a temperature gradient. Alteration of polarity in the frog's egg has been brought about by reversal of position with respect to gravity (SCHULTZE, 1894; I-IAMMERLING, 1927 ; PEbINERS and SCHLE[P, 1927--28) and according to RUNNSTROM (1925) dorsoventrality may be determined in the seaurchin egg by centri- fugal force.

It is well known from the work of many investigators that in Tubularia

and other hydroids, in various turbellaria and in certain cases in other forms conditions arising at the proximal or posterior cut ends of isolated pieces as the result of section determine new polarities opposite in direction to the original. It has also been possible to localize new polarities in other directions with respect to the original axis by the growth resulting from local injury (CHILD, 1909, 1910b, 1927c). According to TAYLOR and TENNENT (1926),

464 Sammelreferat

TENNENT (1926) and TAYLOr, TENNENT and WHITAKER (1926), the definitive polarity in echinoderm eggs cut by micrurgic methods is vertical to the cut surface, the cut surface becoming the basal pole. In nature new polarities very frequently arise as localized regions of more active growth, e. g., buds, many axiate organs. Such regions are in general not sharply delimited, but their activity decreases from a center radially, i. e., they show a radial gradient. In consequence of more rapid growth of the central portion, the radial gradient becomes an axial gradient. In the hydroid, Corymorpha, polarity and dorso- ventrality in place of radial symmetry have been determined by differential exposure of isolated pieces, i. e., by the differential between region in contact with a solid surface and a region freely exposed to the water, either with or without previous decrease or obliteration of the original polarity (CmLD, (1926b, 1927a, b, 1928c) and in the early developmental stages of another hydroid polarity has apparently been determined in the same way (CHILD, 1925b). The differential in such cases appears to be chiefly or wholly a matter of oxygen supply and CO~ diffusion.

In consequence of the differential susceptibility of the different levels of the axes of polarity and symmetry it has been possible, by means of various external agents, to obliterate more or less completely all distinguish- able indications of the normal symmetry or of both symmetry and polarity (BELLAMY, 1919; CHILD, 1916d, 1925b, 1927a, b). In this way forms normally bilateral may be made completely radial or even completely apolar, so far as can be determined. In such forms the gradients characteristic of the axes concerned are also obliterated and further development along such axes cannot take place unless new differentials or gradients are determined in them. Such new differentials may have any direction with respect to the original axis and under proper conditions multiple polarities may develop in place of the original single polarity, and dorsoventrality in place of radial symmetry (CHILD, 1927a. b). A new gradient is characteristic of every such new axis.

In connection with this brief and incomplete survey of these develop- mental aspects of the gradient problem, the following points may be noted. First, the facts at hand show that polarity and symmetry may be altered, obliterated or determined by environmental conditions which affect the physiological activities of the protoplasms concerned, i. e., polarity and symmetry appear to be associated with physiological conditions in protoplasm rather than with inherent structure. Second, these experiments indicate that quantitative differentials, rather than specific or qualitative differences at different levels, are sufficient to determine physiological axes in many if not in all cases. Third, conditions which obliterate physiological axes are such as decrease the quantitative differences in different regions. Fourth, when the axes are obliterated, the corresponding gradients are no longer present, and, on the other hand, new axes are indicated by new gradients. In short,

Sammelreferat 465

the gradients appear to constitute an adequate physiological basis for the phenomena of polarity and symmetry and afford no support to the hypothesis of a stereochemical or any other structural basis of polarity and symmetry than quantitative regional differences in protoplasmic condition associated with quantitative metabolic differences along the axis.

THE GRADIENTS IN RELATION TO DIFFERENTIATION. The experiments on modification of development and on obliteration and determ- ination of polarity and symmetry show beyond doubt that different parts arise at different levels of the gradients. If polarity and symmetry are primarily quantitative physiological differentials, the problem of the origin of specifically or qualitatively different parts from different levels of a gradient is of considerable interest. We know little concerning the processes of differentiation, but it seems probable that differences in concentration of substances at different levels may constitute the first step toward specific or qualitative difference. In vitro differences of concentration of reacting substances may determine different products of reaction and in so complex a system as protoplasm the possibilities of this sort are far greater. Such differences in concentration must result from the quantitative metabolic and protoplasmic differences in relation to intake of oxygen and nutritive material and output of CO 2 and products of metabolism and the accumulation of various substances in cells, e. g., fat, as a differentiation apparently depends on relations of this sort. In regions of intense metabolism the products which remain as constituents of the protoplasm are different from those in regions of low metabolism.

In th~ protozoa, so far as examined, the gradients are present only in the ectoplasm (HY~AN, 1917; CHILD and DEVINEY, 1925) and this is the only part of the body in which a persistent morphological differentiation takes place. In various animal eggs also the gradient appears to be present only in the superficial cytoplasm, at least in certain stages. In such forms other parts of the cytoplasm are not sufficiently stable in position and perhaps not in chemical constitution to permit the persistence of a gradient and the occurrence of differentiation in relation to it.

MODIFICATIONS OF THE GRADIENTS DURING DEVELOPMENT. In some of the simpler organisms the primary gradients persist throughout life, but in many forms, particularly in the higher animals, they undergo extensive modification. The original polar gradient may disappear or undergo reversal; one sort of symmetry may be replaced by another; new gradients m a y be determined by the localization of growing regions or otherwise. Each tentacle of the coelenterate and other sorts of appendages exhibit well marked gradients, at least during growth. Axiate organs develop as gradients

Protoplasma. V 30

466 Sammelreferat

and the functional relations of later stages apparently originate in the developmental gradients. The ctenophore plate row (CIIILD, 1917c, 1921d, pp. 212--220), the alimentary tract (ALvAI~EZ, 1928), the heart (HYMAN, 1927b, ALVAaEZ, 1928, pp. 59--61 and much other work) and various other axiate organs are good examples. Such organ gradients sometimes coincide in direction with the primary body axis, but may make any angle with it.

In the case of the alimentary tract of the mammal ALVAREZ has demonstrated a gradient in rhythm, in force of contraction, in latent period, in tone, in rhythmic tendency and in irritability, in catalase content, in COe production and in oxygen consumption, all of which correspond in direction (ALvAREZ, 1928). This is the most extensive study of the various expressions of an axial gradient that has been made. The literature of physiology contains many other facts which indicate the existence of a quantitative differential of some sort along the axes of various organisms and organs.

One modification of the polar gradient which occurs very widely is the appearance at the basal or posterior end of a secondary region of high susceptibility, high respiration and often rapid growth. This gives rise t o a second polar gradient opposite in direction to the original, but does not usually develop as a new apical or anterior end, though in some cases it may do so (CItlLO, 1925a). In various hydroids it becomes a stolon or a new apical end according to conditions, in the turbellaria it is a posterior growing region. In segmented animals generally, both invertebrates and vertebrates, it is the segment-forming region and gives rise to most or all of the trunk. All such animals thus far examined show a double polar gradient from early developmental stages on, at least through the embryonic period and in many of the annelids throughout life ~.

In the amphibia the secondary region of high susceptibility is the region about the dorsal lip of the blastopore (B~LLAMV~ 1919), that is, the region of the "Organisator" of SPEMANN.

Physiological Dominance, Subordination and Isolation

This aspect of the gradient problem concerns the foundations of physiological control, but can be dealt with only very briefly in the present,

1 In the annelids the presence of this double gradient has been indi,ated by lethal differential susceptibility and developmental modification (HYNAN, 1916; CHILD, 1917d), by potential difference (MORGAN and DIMON, 1904) and by respiratory differences (HYMAN and GALIaHER, 1921); in aseidians by developmental modification and KMn04 (CHILD, 1927 d); in fishes by lethal susceptibility (HYMAN, 1921, 1926b) and developmental modifieati on (STOCKARD, MC CLENDON and others; HINRICIIS, 1925); in amphibia by lethal susceptibility and developmental modification (BELLAMY, 1919, 1922); in birds by lethal susceptibility and developmental modification (HINRICIIS, 1927 ; HYMAN, 1927 a).

Sammelreferat 467

paper 1. The physiological dominance of the growing tip of the plant axis has long been known and during recent years it has been shown by various lines of evidence that the "high" region of a gradient exercises a certain control or dominance over other regions within a certain distance from it, at least in the simpler organisms and in earlier developmental stages. Since this dominance is limited in range before the nervous system arises, physiological isolation of a region or part may result, (a) from growth in length of the axis concerned; (b) from decrease in range of dominance through inhibition or other changes decreasing the activity of the dominant region; (c) by blocking the influence of the dominant region in its passage (CHILD, 1920b, 1921a; CHILD and BELLAMu 1919); (d) by directly stimulating the subordinate part and so making it insensitive to the influence of the dominant region.

In the earlier developmental stages of many forms and in plants and many of the simpler animals throughout life the effect of physiological isolation of a part is essentially similar to that of physical isolation, that is, the isolated part loses more or less completely its characteristics as a part, undergoes reorganization and becomes or approaches a whole, in other words a process of reproduction of a new whole or a repetitive part occurs. I t has been pointed out that the various processes of agamic reproduction in axiate organisms are very generally if not always the result of such physiological isolation. I t has also been suggested that the appearance of a secondary region of high developmental activity in the posterior region o f early developmental stages of many animals results from partial physiological isolation (BELLAMY~ 1919, CHILD, 1917, i921 d, chap. VIII), as does the appearance of a new zooid in the posterior body region of Planaria (CHiLl), 1910a). In the segmented animals particularly the course of development suggests that the dominance of the extreme anterior region is weak in embryonic stages, consequently the region of the embryo most distant from the head sooner or later becomes partially physiologically isolated, increases in activity and becomes itself a secondary dominant region which behaves somewhat like a growing tip and gives rise to most of the body.

Extended discussion of SPEMANN'S "Organisator" is impossible here and SPEMA~N'S general papers (SPEMANN, 1925, 1927) and DE BEER'S review of the subject make such discussion unnecessary, but it may be pointed out that the organizer seems to represent essentially the secondary region of dominance characteristic of segmented animals. According to the more recent work there is at least a labile determination of the extreme anterior embryonic region independently of the organizer (GOERTTLER, 1925, 1927; LEHMAN~1928)

1 For more extended discussion and references to literature up to 1924 see CHILD~ 1924b, chap. X and literature there cited and for earlier discussions see CHILD, 1911b, 1915b, chaps IV, V.

30*

468 Sammelreferat

and LEttMANN maintains also that the degree of determination decreases and so the influence of the: organizer in determination increases posteriorly. Moreover, GOERTTLER (1927) has suggested that the difference between organizer and other parts o f the embryo is primarily quantitative and the fact that :the organizer is not species-specific in its action has been shown by transplantation into other species, genera and even subclasses with induction of primordia. This result of transplantation particularly suggests that its action depends on quantitative, rather than specific differences from the parts affected. And finally GILCHRIST (1928) has shown that a new embryonic primordium can be induced at the high end of a temperature gradient. In this case it appears that ~he differences determining the new polarity must be primarily quantitative. In short, at present the influence of the amphibian organizer appears to be a special case of physiological dominance associated with a gradient and the facts at hand suggest that its dominance depends primarily on a quantitative difference in physiological condition and activity from the parts influenced by it, in other words, it is dominant and an organizer because it is more active. I t may be suggested that the induction of an embryonic primordium by a piece of organizer or of medullary plate occurs because the active tissue increases the activity of the tissue about it or above it to the level at which it develops as medullary plate. GILCHRIST'S results with a temperature gradient are particularly significant in this connection. If this suggestion is correct, any other sufficiently active tissue or any means of making a certain ectodermal region sufficiently active should have the same effect as the organizer. If the data on susceptibility are of any value, the organizer region is not a particularly active region at the beginning of development, but becomes active some time before gastrulation (BELLAMY, 1919). If this is true the amphibian resembles the bird, the fish, the ascidian and the annelid (probably also the arthropod) in developing relatively early a secondary dominant region which forms most of the body and is carried posteriorly by fur ther development. In the amphibian the primary anterior dominance is apparently weak during the earlier stages of develop- ment and the secondary posterior dominant region becomes the more impor- tant determining region. In the chordates generally the secondary posterior dominance disappears sooner or later, but in some of the annelidS it may persist throughout life.

Organizers are not peculiar to amphibian development, but are charac- teristic of axiate organisms in general. For example, the growing tip of Che plant~ the developing head on a piece of Planaria (CHILD, 1911C), the active regions at the cut ends of hydroid stems, the local injury which determines a new axis in Corymorpha (CHILD, 1927C) have all been shown beyond doubt to be organizers. In fact, is not every more active region dominant in some way and to some degree over less active regions? SPEMA~ (1925, 1927)

Sammelrefera/ 469

has suggested that organizers of various rank are undoubtedly concerned in development and this suggestion is in complete agreement with the conception of dominant regions in organ axes etc. Even within a specifically determined tissue the most active region may act as organizer. From the standpoint of the gradient theory the organizer is essentially a developmental pacemaker for a particular axis, whether of the whole organism or a particular organ. Moreover, from this standpoint amphibian development does not differ in principle from that of the lower invertebrates. Not only is there no conflict between the facts at hand concerning amphibian development and the gradient conception which has developed largely from work on the simpler organisms, but the investigators in the field of amphibian development ~re suggesting interpretations essentially in terms of quantitative differentials or gradients.

Moreover the concept of the gradient with its relation of dominance and subordination, whether the dominant region be the center of an area or the end of an axis, does not differe essentially from the concept of the ,,Wirkungsfeld", the ,Organisationsfeld", or the ,,Determinationsfeld" (WEiss, 1926). The gradient is such a field in its simplest and most general terms and the gradient concept is an attempt to define the primary character of the field. I t is encouraging to find general conceptions concerning the physiology of development approaching even within hailing distance of each other, particularly when they have developed from the work of different investigators in different fields.

Concerning the nature of dominance in its most general form definite conclusions are not yet possible, but many facts indicate that the influence of a dominant region is primarily transmissive, rather than transportative in character, that is, that it consists in some form of energy transmission in protoplasm or along l imit ing surfaces, rather than in mass transport of substance. Quantitative differences along an axis do not appear to provide an adequate basis for definite and specific chemical correlation as the primary form of correlation. Such correlation is possible only after some degree of differentiation has taken place. The definite range of dominance in the simpler organisms indicates transmissive rather than transportative correlation. Similarly, the fact that dominance has been shown in various eases not to be species-specific argues against its chemical or transportative character. And finally, the fact that the organ of dominance par excellence, the central nervous system, is in general the first definitive organ to develop and arises from the dominant regions of earlier stages is at least highly suggestive. I t seems probable that a dominant region exercises primarily some more or less continuous effect over a certain distance in protoplasm. This effect perhaps resembles the influence producing tonus. The limited range of dominance in the absence of nerves suggests a decrement, that is, the dominant region

470 Sammclreferat

determines or maintains a gradient and so const i tutes the primary factor in establishing and main ta in ing physiological axes. From this point of view the establishment of new axes by differential i l lumination, by the electric current, by differential exposure of free surface and surface in contact, by ]ocM high temperature, by local injury, by graft ing a piece of the organizer etc., are all essentially similar processes in that they determine quant i ta t ive physiological differences in a certain direction or in certain directions and establish a basis for physiological dominance or control.

LITERATURE

ALLEE~ W. C.~ The F~ffect of Potassium Cyanide on Metabolism in two Fresh Water Arthropods. Amcr. Jonrn. Physiol. LXIII, 1923.

ALLEN, (~. D.~ Quantitative Studies on tile Rate of Respiratory Metabolism in Planaria. I. Amer. Journ. Physiol. XLVIII, 1919a.

Quantitative Studies &e. II. Amer. Journ. Physiol. XLIX, 1919b. - - The rate of Carbon Dioxide Production in Pieees of Planaria in Relation to th~

Theory of Metabolic Gradients. Proe. Amer. Soc. Zool., 1919; Anat. Rec. XVII, 1920.

ALVAREZ, W. C., Tile Mechanics of the Digestive Tract. Second Edit. New York 1928. BEHRE, ELINOR H.~ An Experimental Study of Acclimation to Temperature in Plana~ia

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