Physiological Foundations of Behavior

(Hinrichs, ’23b) and I have obtained further data alon forms, both plant and animal (unpublished). In all forms thus far examined the results obtained by photolysis are similar to those obtained with other agents. _ follows: To a certain range of concentrations or intensities of agents which are experimentally determined to be above the limit of tolerance of the organism concerned and therefore strongly toxic or lethal in their action the susceptibility varies directly with, though not necessarily proportionally to, the general protoplasmic activity or rate of metabolism. To a certain lower range of concentrations or intensities, also experimentally determined for each species, the rate and degree of acclimation or acquirement of tolerance varies directly with, though not necessarily proportionally to, the general protoplasmic activity and the rate and degree of recovery after temporary exposure to a certain range of concentration or intensity varies in the same way. General protoplasmic activity is a vague term, but is used in order to avoid the implication that any particular component factor in protoplasmic condition is necessarily regarded as primary. Protoplasm is a system in which the dynamic changes are so closely correlated and integrated that we cannot point to any one as primary, moreover, it is possible that the initiatory change in a complex physiological process such as excitation is not always necessarily the same.

The data on susceptibility constitute some of the strongest evidence for the existence of such integration in the protoplasmic system. In the simpler organisms and the earlier stages of development of higher forms the susceptibility relations are in general similar for a great number of agents of different chemical constitution, e. g., cyanides anesthetics, acids, alkalies, various neutral salts, vital dyes, etc., and for physical conditions such as extremes of temperature.! This cannot possibly mean that all these agents and conditions act on living protoplasm in the same way or alter primarily the same factor in the system. Certainly the effects of the different agents and conditions are exerted primarily or chiefly upon different factors of the protoplasmic system, and the results show clearly enough that when any essential factor is sufficiently altered, the system as a whole is altered. In the less highly specialized protoplasms, therefore, we find little or no indication of regional differences in susceptibility specific for

been in part disposed of by Hyman’s later work and other data soon to be published will dispose of others. As is evident to those familiar with the gradient conception, these criticisms appear to be due largely to misapprehension and in a number of cases are directed against views quite different from those advanced by us; consequently they require no consideration here. 1 Recent work, in large part not yet published, has added to this list as follows: formaldehyde, various alkaloids, lack of oxygen, ultraviolet radiation and sunlight after photochemical sensitization.

particular agents. Among the lower invertebrates the differences in susceptibility to acids and alkalies seem to be more nearly specific than any other differences yet discovered in these forms (Child, ’20 c, p. 158, J. W. MacArthur, ’20), but these differences appear during the course of development and are not present in the earlier stages. And in the higher animals, even the vertebrates, the apparently specific relations between particular tissues or organs and particular agents are largely or wholly absent in the early stages. In general, the susceptibility gradients of the simpler organisms and the earlier stages of development are non-specific in relation to particular agents and conditions. This being the case, we may expect to find a relation between susceptibility aud rate rather than kind of dynamic change in protoplasm. The chemical reactions of metabolism are essential factors in the activities which we call life, and among these the oxidations are regarded as of fundamental importance. With certain qualifications and limitations it appears to be true that the rate of oxidation is in some degree a measure of the rate of living. It has also been shown that in the less highly specialized protoplasms susceptibility to certain ranges of concentration or intensity of external agents is an indicator of rate of oxidation and may be used as a rough / comparative measure of differences in rate. There is therefore a real experimental basis for the statement, that susceptibility is in general a measure of rate of metabolism or more particularly oxidation. In highly specialized organs and tissues specific susceptibilities to particular agents may appear and these may in some cases be related to qualitative factors of constitution rather than to rate of oxidation.

Nevertheless the fact remains that in the less highly specialized protoplasms susceptibility is, within certain limits of concentration or intensity of agent and with proper technical pro- © cedure, a rough measure of physiological condition and more particularly of rate of oxidation. The existence of this general non-specific relation between susceptibility to the action of external agents and metabolic condition in protoplasm seems to me to be merely a special case under general physico-chemical laws which may be stated in more general terms as follows: The greater the velocity of changes concerned in the maintenance of, or approach to, dynamic equilibrium in a system, the more rapid and extreme the effect of sufficiently powerful external agencies in altering or destroying that equilibrium, and the more rapid the equilibration to, or recovery from, slight or temporary disturbances. In other words, the more active the system, the more sensitive it is to gross disturbances and the more rapidly

Fies. 35-41.—Susceptibility gradients as indicated by the course of disintegration in various animals: Figs. 35, 36, early and later stage of disintegration in the infusorian Spirostomum in KCN m/200 (similar in 40 Apical end uppermost, body contracted; Fig. 37, disintegration of unfertilized egg of sea urchin, Arbacia in neutral red. Intact regions are stained deep red and decoloration accompanies disintegration; Figs. 38, 39, early and later stage of disintegration of blastula of medusa, Phialidium in KCN, m/100; Figs. 40, 41, early and later stage of disintegration of blastula of sea urchin, Arbacia, as observed in many different agents. The difference in rate of basipetal progress of disintegration in different meridians shown in Fig. 41, probably indicates an early stage of the differences between anterior and posterior in later larval stages. In each figure the arrows indicate the direction

it equilibrates to slight disturbances. I believe the relation between susceptibility and metabolism in its simplest form is nothing more than this.! Differences in susceptibility along an axis may be made evident in various ways. In highly toxic concentrations or intensities a definite gradient in the course of death and disintegration may appear and this may be preceded in motile forms by a gradient in loss of motility. In Figs. 35-44 the course of death and disintegration along the chief axes of various organisms is indicated. In each case the same death and disintegration gradients have been observed with various agents. The portions figured as intact are still alive at the stages indicated: in ciliated forms, such as Spirostomum (Figs. 35, 36) and the hydroid and sea urchin blastule (Figs. 38-41) the cilia of the intact portions may continue movement until disintegration begins and in forms with muscular differentiation (Figs. 42-44) movement may occur almost up to the moment when disintegration begins. If the organisms are returned to the normal medium before the toxic action has proceeded too far, all, or nearly all of the intact portion of the body will recover. Numerous other figures of both plants and animals might be added.

When early developmental stages are exposed to toxic but nonlethal concentrations or intensities differential susceptibilities along an axis may appear in a differential modification of development. In inhibiting agents the most susceptible regions are most inhibited, but in a certain range of low concentrations or intensities they show the most rapid and most complete acclimation or development of 1 Huxley (722) has advanced the view that the susceptibility of cells may also vary with amount of surface exposed to the agent. This is undoubtedly true, but it may be noted that differences in exposure of cell surface in a particular body layer, or in the body surface, sufficient to affect susceptibility to any marked degree, usually arise secondarily in the course of differentiation. I have pointed out repeatedly that with the progress of differentiation various factors appear which alter the general relation between susceptibility and metabolic rate. Un- questionably difference in exposure of cell surface is one of these factors.

In Huxley’s work the index of susceptibility appears to be change in behavior of the living cell as a whole, usually change in form. It cannot be assumed, however, without further evidence, that because a flattened cell shows marked change in form and a cuboidal or rounded cell little or none in a certain concentration of agent, the toxic action of the agent is greater upon the flattened cell. It is possible that change of form may occur in the flattened cell with very slight toxic action, while a greater degree of toxic action is necessary to produce visible change in the cuboidal or rounded cell. In other words, the change in form of the flattened

cell does not necessarily mean a higher susceptibility in the sense in which I have used the word. Fics. 42-44.—Susceptibility gradients in flatworm and annelids: Fig. 42, a stage of disintegration of the rhabdocoel, Stenostomum in KCN, m/1000 (and many other agents). The figure represents a chain of zooids. The head of the anterior zooid, as most active region is most susceptible, that of latest formed, shortest zooid least susceptible. Ventral regions are more susceptible than dorsal; Fig. 43, a disintegration stage of obligochete, Dero limosa, showing primary antero-posterior, and secondary postero-anterior gradient, the latter resulting from development of new segments anterior to anal segment; Fig. 44, disintegration stage of Limnodrilus claparedianus, showing the double gradient (Figs. 43, 44 from Hyman, ’16 a).

tolerance and after certain ranges of temporary exposure they recover most rapidly and most completely. In exciting or accelerating agents a differential acceleration of development may also occur at different levels of a gradient. All of these differentials appear as differential effects on rate of development, and in this way on size and proportion of different parts. Four sorts of modification in two opposite directions occur as the result of differential susceptibility (Child, 716 d, 17d). In differential inhibition the most susceptible regions are most inhibited and therefore least developed. In differen-

Fies. 45-47.—Differential inhibition in early developm Fig. 45, normal early stage of elongation of blastula and Immigration of cells from basal ithout visible polarity. Excessive formaof enteric cavity. Arrows in Figs. 45 and perical planula of Fig. 47 is incapable of 1m; (en), entoderm. tial acclimation they are at first most inhibited, but later show more acclimation and so are finally less inhibited than the less susceptible regions. In differential recovery the modifications are essentially like those of differential acclimation but they occur-after return to the normal medium. In differential acceleration the most susceptible re- ‘gions are most accelerated and therefore relatively larger than normal.

_ Developmental modification through differential susceptibility is illustrated here by a few examples. Further data are presented in the following chapter in another connection. Cases of differential inhibition are shown in Figs. 45-52. In normal hydroid development immigration of cells to form entoderm (Fig. 45) and elongation of the blastula into the planula occurs (Fig. 46). In differential inhibition the original polarity may completely disappear, 7. e., the

Frias. 48-52.—Differential inhibition in larval development of sea urchin, Arbacia: Fig. 48, normal pluteus larva, basal view; Fig. 49, normal pluteus, side view; Fig. 50, differential inhibition by action of KCN throughout development, side view. Apical and anterior regions more inhibited than basal and posterior; Fig. 51, differential inhibition by temporary exposure to KCN, basal view. Anterior and median mere inhibited than posterior and lateral; Fig. 52, greater degree of differential inhibition by temporary exposure to KCN, basal view. Median region so completely inhibited that a single median arm and skeletal rod develop instead of paired lateral structures.

apical region is most inhibited, the basal least and with a certain degree of inhibition the axial gradient is reduced to a level. In such cases the larva remains spherical, loses its definitely directed movement (Fig. 47) and does not develop further unless a new gradient arises in it (Chap. TX). Figures 48-52 show differential inhibition in sea-urchin development. Figures 48 and 49 show normal pluteus larve in lateral and basal aspects. Figure 50 is a differentially inhibited larva in side view, and it is evident that the apical region, the oral lobe, is more inhibited than the basal region. Figure 51 is a basal view showing

that anterior and median regions are more inhibited than Posterior and lateral, and Fig. 52 is a more extreme inhibition in which the median region is completely inhibited so that the lateral arms arise as a single structure in the median line. In differential acclimation, recovery and acceleration the modification of form is opposite in direction to that occurring in differential Figs. 53-56.— Differential acclimation in Arbacia: Figs. 58, 54, basal and lateral views of a slight degree of differential acclimation; Fig. 55 (A), basal, (B), lateral view of more extreme degree of differential acclimation; Fig. 56 (A), basal, (B), lateral view of still more extreme modification. In all these cases apical, anterior and median regions are disproportionately large, as compared with basal, posterior and lateral.

inhibition. Figures 53-56 showing different degrees of differential acclimation in the sea urchin will Serve as examples. Comparison of these figures with the figures of normal plutei (Figs. 48, 49) shows that in the stages figured of these differentia] acelimations gions are more inhibited than a posterior more than anterior. Fic. 57.—Differential acceleration in a fish, Macropodus viridi-auratus: (A), control developing under standard normal conditions; (B), exposed to atropin sulphate 134 hours during early cleavage. Rate of development and of heart beat more rapid than in control (by permission of J. N. Gowanloch from unpublished work).

reverse was the case, but apical, anterior and median regions have undergone acclimation more rapidly and to a greater degree than basal posterior and lateral regions, that is, differential acclimation is preceded by more or less differential inhibition. The changes in differential recovery and differential acceleration are similar in direction to these, and differential recovery, like differential acclimation, is preceded by differential inhibition. In differential accleration, — however, there is no inhibition or retardation, but development is accelerated, from the beginning. Bellamy (’19) has been able to produce differential acceleration in amphibian development and Mr. J. N. Gowanloch has accomplished the same result with fishes. Figure 57 shows a case of differential acceleration (B) and normal control (A) obtained by Mr. Gowanloch with a low concentration of atropin.!

These experimental modifications of development through differential susceptibility afford a physiological basis for interpretation of a wide range of teratological forms in nature. In fact, differential susceptibility appears to be a fundamental teratogenic factor in all cases where the modification results from exposure of the embryo as a whole to the external condition concerned and not from direct mechanical or other injury to some part. It is perfectly evident that the types of developmental modification are non-specific and essentially quantitative both in their relations to the axes and to the different protoplasms and external agents. Similar external agents and conditions produce the same sorts of modification in flatworms (Child, ’16 b, ’20 a, ’21 ¢, Buchanan, ’22), echinoderms (Child, ’16 d), annelids (Child, 717 d), fishes (Stockard, ’07, ’21), and amphibia (Bellamy, 719, ’22). Similarly in teratological forms of fishes produced by Werber (’16, ’17) the indications ‘of differential susceptibility are evident. And finally Newman (’17 a, ’18) has shown that monsters resulting from hybridization in fishes may be interpreted in terms of the same differentia] susceptibility, ?. €., aS might be expected from what we know of differential susceptibility elsewhere, a differential susceptibility of one protoplasm to the other or of each to the other exists.

The most important point for present purposes is that all the data on differential susceptibility indicate very clearly that each physiological axis or direction of order exists primarily as a quantitative gradient in physiological condition in which differences in rate of oxidation and associated differences in protoplasmic condition are fundamental factors. As will appear later, these differences in physiological condition along the axes are of fundamental importance for the behavior of the organism as a whole.

Penetration. — Axial gradients in “permeability ” or rate of penetration of various substances have been demonstrated in many forms *Mr. Gowanloch has very kindly permitted this use of figures and data from his unpublished work. both animal and plant. The regions more susceptible to the higher concentrations of external agents show in general a higher rate of penetration of such agents as vital dyes than less susceptible regions. t& indicate thdt permeability of living cells is itself closely associated with physiological condition and with metabolism as a factor in such condition. It has been pointed out elsewhere (Child, ’21 a, pp. 40-42) that the susceptibility gradients cannot be interpreted in terms of permeability of cell surfaces as a physical condition independent of metabolism. There is no reason to believe that the permeability of living cells is independent of metabolism, moreover, a certain parallelism exists between rate of penetration of various substances and rate of oxidation. And finally the results of differential acclimation are opposed to the differential permeability, 7. €., it is the regions of higher permeability which acclimate most rapidly and to the greatest degree.

Oxidation-reduction reactions. — Certain oxidation-reduction reactions render the physiological gradients directly visible as color gradients resulting from the deposition of the products of oxidation or reduction at different rates or in different amounts at different levels of a gradient. For example, the different levels of a physiological gradient reduce potassium permanganate at different rates and in different amounts and so become visible as gradients in depth of

brown or blackish color due to deposition of MnOz, or other oxides in the protoplasm (Figs. 58-60). This reaction has proved to be a very delicate method for demonstrating the physiological gradients in many cases.!_ The indo-phenol reaction, an oxidation, has also been used in certain cases, the physiological gradients appearing as gradients in depth of blue resulting from deposition of the blue indophenol in the protoplasm (Child, ’15a). The gradients have also been demonstrated in various plant and animal forms after staining with methylene blue as gradients in rate of reduction of the dye in the protoplasm.

Electrical potential. — The physiological gradients are also characterized by differences in electric potential. Observations have been made on many different forms, both animals and plants.” In general, in animals the higher levels, 7. e., the levels of more intense activity, of higher rate of oxidation, are electro-negative, galvanometrically, Fic. 58.—Reduction gradient in wall of sycon sponge split longitudinally. Fie. 59A-C.—Reduction gradients in developmental stages of hydromeduse: (A), in superficial cytoplasm of ovarian egg of medusa, Stomotoca atra; (p), area of attachment of egg in gonad; (B), in two cell stage of medusa, Phialidium: (C), a stage of reduction gradient in ectoderm of planula of Phialidiwm.

Fie. 60A—B.—Reduction gradients in sea urchin development: (A), in optical section of ovarian egg; (B), in optical section of early blastula. to lower levels. In the alge thus far examined the higher levels of a physiological gradient are usually electro-positive to lower levels. This difference of sign between animals and algse may be connected with the fact that in the latter the reduction and syntheses connected with photosynthesis and its products overbalance the oxidations as

~ factors in potential difference. Further work is necessary, however, to determine whether this suggestion is correct. The existence of characteristic differences of electric potential associated with the physiological gradients suggests the probability that the differences in rate of metabolism or oxidation along the gradient play an important part in determining these potential differences. Moreover, the fact that the differences of sign at different levels are such as we should expect from the differences in metabolic rate known to exist increases this probability. The evidence for this view is presented in the recent discussion of this question by Hy- man and Bellamy (’22). It should, however, be stated with some emphasis that this view by no means involves the assumption that differences in metabolic rate are the only source of potential differences in organisms. Unquestionably there are many other sources of such difference, and other factors than metabolism may be concerned in the axial potential differences, but many lines of evidence indicate that the metabolic factor is of importance as a source of potential difference.

Galvanotaxis. — A relation of some sort between the galvanotactic reaction and the gradients exists, et least in many of the simpler animals, as Hyman and Bellamy (’22) have shown. So far as observations go at present, forms with a simple polar gradient tend to orient themselves in the electric current so that the high end of the gradient is toward the negative pole. The same reaction appears in various axiate organs such as ccelenterate tentacles. In annelids with a second region of high metabolic rate at the posterior end, the two ends of the body are turned toward the negative pole so that the body takes U-shape. In the arthropods and vertebrates the situation is commonly more complex, as might be expected in consequence of the high degree of modification of the primary gradients and the neuromuscular complexity.

At present any attempt at interpretation of this apparent relation between the gradients and the galvanotactic reaction can be little more than surmise. Hyman and Bellamy suggest that the orientation is in some way connected with the fact that the region of highest metabolic rate is positively charged in relation to other parts, but they do not attempt to show how the positivity of this region determines orientation. A. R. Moore (’23) has shown that in the earthworm a reaction very similar to, if not identical with, the galvanotactic reaction oecurs when the current is passed transversely through a few body segments, as well as when the whole body is exposed to it,

and that the nerve cord is necessary for conduction of the stimulus which brings about orientation of the parts not directly affected by the current. On the basis of these experiments he maintains that the galvanotactic reaction in the earthworm results from the action of the current on the nervous system. His experiments and interpretation do not, however, alter the fact that a relation of some sort exists between the polar gradient and the orientation, nor will it hold for some of the simpler organisms which have only a rudimentary nervous system or none. At present only the apparent fact of the relation between the gradient and the galvanotactic reaction need be noted: further work is necessary for any general interpretation of the mechanism of orientation.

Respiration. —- And finally it has been possible in various cases to determine directly differences in rate of oxygen consumption and CO production at different levels of physiological gradients.! This method of direct determination of respiratory exchange of different body levels meets with various difficulties, e. g., the effects of separating different body regions, the fact that it gives only total oxygen consumption or CO, production of all organs and does not afford any means of distinguishing between different organs or regions in which the gradients are not necessarily the same or in the same directions. For this reason the method has been used chiefly for simple organisms.