Comparative Electro-Physiology: A Physico-Physiological Study

hitherto horizontal base-line. This bifurcation causes the first contractile response of the now growing organ—sixth of tonic condition is gradually established by the absorption of energy and the molecular mobility of the responding organ is increased, the contractile response becomes larger, and growth goes on at a certain steady rate. This constitutes an instance in which stimulus, so far from lowering the energy of the responding system, has actually raised it above par.

It would thus appear that while the theory of assimilation and dissimilation is insufficient for the explanation of the various characteristics of response, the difficulties there en- countered are, on the contrary, satisfactorily explained, on taking full account of the influence on response of the molecular condition of the responding substance. From the chemical hypothesis of an explosive molecular change, with its attendant dissimilation and run-down of energy, it would follow that previous stimulation should always induce a depression of the subsequent responses. Instead of this, however, it is found that previous stimulation sometimes exalts, and at other times depresses, the subsequent re- sponses. This apparent anomaly we have seen to be ex- plained by the consideration of molecular transformation. From the sluggish condition A, we have seen tissues trans- formed, by the impact of moderate stimulus, to condition B, with its greater excitability. It is only when the molecular condition has been brought to D or E, that the responses undergo a diminution or reversal.

The molecular condition, then, undergoes a continuous transformation, in consequence of the action of stimulus, from the extreme of sub-tonicity A to the overstrained molecular conditions D and E. In the A stage, there is no true ex- citatory expression, response to stimulus being here by the abnormal positive variation. The substance is next trans- formed into stage B, where response exhibits a staircase character. In the next stage C, the responses are uniform.

Under over-stimulation, the stages D and E are reached, characterised by diminished amplitude of response, or actual reversal into positive. There are thus two conditions under which we obtain abnormal positive responses. One of these is that of sub-tonicity, and the other, the reversal due to fatigue. There is, again, no tissue which is exclusively characterised uniform, and fatigue—will occur in muscle, nerve, plant, and even inorganic matter, under certain definite and ap- propriate conditions. In a future chapter, we shall study in detail the characteristic molecular curve, from which light will be thrown on the internal molecular condition of the tissue, and the influence of that condition on response.

Anomalies in mechanical and electrical response—Resultant response determined by differential excitability—Responsive current from the more to the less excitable—Laws of response in anisotropic organ—Demonstration by means of mechanical stimulation—Vibrational stimulus—Stimulation by pressure— Quantitative stimulation by thermal shocks. IT has been customary, as we know, to ascribe the varied movements of plant-organs under external stimulus, to the presence of different specific sensibilities ; and, indeed, it would seem at first sight impossible to reduce such highly complex and apparently unrelated phenomena, to the terms of a single fundamental reaction, common to all alike. There is no denying, for instance, that certain plant-organs, when acted on by light, bend towards it, and others away. I have elsewhere shown,' however, that all these diverse movements are clearly traceable to one fundamental excitatory reaction, and that the different effects observed are due merely to the differential excitabilities of various parts of the structure ; and that the resultant movement is in all cases brought about by the greater contraction of the more excited side. Passing next to the electrical response of living tissues, animal and vegetable, we encounter many anomalies. Not

inconsistencies are often due, as we shall find, to the dfferentzal excitability of anisotropic structures—a factor in the problem which has not hitherto been recognised. An investigation on 1 Bose, Plant Response. this subject, then, demands that we first discover some means of determining the relative excitabilities of different parts of a tissue. As the simplest example of an anisotropic structure, we may take a compound strip of ebonite and stretched india- rubber, glued firmly together throughout their length. Of these, the india-rubber is the more contractile, and when the strip as a whole is subjected to periodic thermal stimulation, response takes place by the greater contraction induced in > the india-rubber. Ifthe strip be held, with the india-rubber below, response will be by the induced concavity of the lower side. -In fig. 77 is shown a series of these responses of the compound strip, taken on a smoked surface by means of a recording lever.

In anisotropic motile organs, such as the pulvinus of J/zmosa, response takes place by differential contraction, the more excitable side being that which under diffuse stimulation becomes concave. If we apply very moderate stimulus AG 21. DiRerenge locally on the upper half of the pulvinus, sponse of Artificial we shall find that, by the excitatory con- sh traction of this half, the leaf is raised. A. similar contractile effect, though of greater intensity, is induced when the lower half of the pulvinus is stimulated locally, the leaf in this case undergoing a depression. When both upper and lower halves, then, are excited simultaneously, the resulting fall of the leaf shows that the contraction of the lower half must in this case be the greater, or, in other words, that this half is the more excitable of the two. This experi- ment may be carried out very easily by using the stimulus of light. Fig. 78 gives the results observed (a), showing the up movement consequent on stimulation of the upper half; (¢) that caused by equal stimulation of the lower half; and (c) the resultant fail when the two are excited simultaneously. In the case of mechanical response, then, we find it true that response is by the greater contraction of the more excitable.

We shall next observe what is the electrical mode of response for.a tissue which is anisotropic, or unequally ex- citable on two sides. For this purpose we may again take the pulvinus of A/zmosa, and make electrical connections at two diametrically opposite points on the upper and lower halves of the pulvinus respectively. It is to be remembered that electrical response takes place on excitation, whether the leaf be free to move, or physically restrained. We may, therefore, hold it in a fixed position; and indeed this is advisable, in order to avoid that shifting of the electrical contacts which might possibly take place if it were allowed to fall.

The two contacts are made with two fine straws filled with kaolin paste, moistened in normal saline. On now applying a series of thermal stimuli, on the petiole, near the pulvinus, I obtained the responses given in fig. Fic. 78. Responses of AZ/mosa to Sunlight of ; not too long Duration 79. It will be seen : (a) Light acting on pulvinus from above ; (64) light that the responsive acting on pulvinus from below ; (c) light acting current flows in the simultaneously from above and below. Dotted

3 line represents recovery on cessation of light. tissue from the rela- tively more excited lower, to the less excited upper, half of the organ. We thus arrive at a comprehensive law of the mechanical and electrical response of anisotropic organs: Diffuse stimulation induces greater contraction and galvanometric negativity of the more excitable side. The laws of electric response in the anisotropic organ may then be detailed ‘as follows :— 1. On simultaneous excitation of two points, A and B, the responsive current flows in the tissue from the more to the less excited.

2. Conversely, if under simultaneous excitation the responsive current be from B to A, then B is the more excitable of these two points. These form only an instance of the general law that the responsive current always flows from the more to the less excited. For when a point, B, is excited locally—-this point, that is to say, being the more excited—the responsive current is found to flow away from it to a neutral or in- different point, A, for which any distant point will serve, provided the tissue be non-conducting. Should it be con- ducting, the neutrality of A is maintained by interposing a

The petiole is securely held to prevent movement, and diametric electric contacts made in the upper and lower surfaces of pulvinus. Re- sponsive current is from lower to upper surface. block. Should the stimulus, however, not be local, but diffuse, a resultant response may still be obtained by injuring or killing the point A, and thus diminishing or abolishing its excitability. On stimulation, the point B is now necessarily the more excited, and the responsive current is still away from B, towards A. And finally, owing to physiological anisotropy, B may be naturally more excitable than A, and 6n stimulation the responsive current will then be found to flow from the more excited B to the less excited A.

The comparison of the excitabilities of the two points A and B, therefore, reduces itself to the application of similar stimuli to the two points simultaneously, and then ascertaining the direction of the responsive current. For this purpose we might employ any form of stimulus, and it is extremely interesting to find that, however diverse the. stimuli, the results obtained by them are always identical And here we have not merely a means of qualitative demonstration, but in some cases one of quanti- tative also. i

If we take an erect stem of Cucurbita, it being radial and isotropic, all its flanks will be found equally excitable. Hence, if two diametrically opposite contacts are made, there Fic. 80. Diametric Method of Stimulation of an Anisotropic Organ Diametrically opposite contacts are made at A and B, and tissue subjected to vibrational stimulus. will, on diffuse stimulation, be no resultant response. But when such a stem becomes recumbent, the upper side, being now constantly exposed to light, becomes fatigued by over- stimulation, with consequent diminution of its excitability. This is true only when the stimulus has been excessive and long continued ; for we have seen moderate stimulus may sometimes enhance the excitability. By the unilateral action of light, then, the organ has been converted from radial into anisotropic, the lower side being that which we shall expect to find the more excitable.

On mounting such a stem in the vibratory apparatus (fig. 80), and making diametrically opposite contacts on the two anisotropic surfaces, we find that on applying vibration both sides are subjected to similar stimulus simultaneously ; and the responsive current is now found to flow across the tissue, from the lower to the upper side. The lower is thus, as we expected, the more excitable. Since we can by means of vibration apply measured stimuli, it will be seen that we have here a quantitative method of investigation. Moreover, as the stimulus is applied directly, it is applicable not only to conducting but also to non-conducting tissues.

slit it longitudinally, we obtain, in either of the halves, a specimen having an inner and an outer surface. As one of these has been exposed to light and the other protected from it, we should expect to find, on examination, that there has been an induction of physiological anisotropy. As such a specimen is not very well adapted for vibrational stimula- tion, we may use that of pressure. Two moistened rags, in _ connection with non-polarisable electrodes, pass through two pieces of cork, adjusted on the two surfaces—outer and inner—at diametrically opposite points. When the inter- posed tissue is now subjected to sudden pressure its two surfaces are excited simultaneously, and the responsive - current is found to flow from the inner concave to the outer convex surface, proving that the former was the more excitable. |

We might again use the chemical form of stimulation, and the results obtained by this method will be described in the course of the next chapter. But these forms of stimulus —by pressure, or by chemical means—are not capable of exact measurement. For quantitative observations, then, it is necessary to employ some other form of stimulus, and the electrical offers us in this respect many advantages. There are, however, in this case many possible disturbing influences to be considered, all of which must be carefully eliminated before the method can be used without misgiving. How this may be done will be shown in a future chapter. For the present I shall describe another method of stimulation which I have been able to bring to great perfection, by which

two points of an anisotropic organ may be simultaneously excited, under a series of stimuli of uniform or increasing intensity. This mode of excitation, by thermal shocks, will be found in every way satisfactory and convenient. The Thermal Variator, by which stimulation is effected, consists of a spiral of german-silver wire, the diameter of the spiral being about 3 cm. The electrical circuit, through which the heating- current is sent, is closed periodically for a definite length of

The anisotropic tissue-petiole of J/wsa is held in ebonite clip, c. 8, &’, electrodes connected with opposite sides. Specimen after adjustment pushed inside heating-spiral, T, by slide, s. Spiral heated periodically by closure of electric circuit by metronome, M. time, by means of a metronome (fig. 81). The thermal variation within the coil can be controlled by a suitable adjustment of the battery-power, or by the duration of closure, or both. The experimental tissue is held in an ebonite clip, C, fixed on a slide, S, on the same stand as the heating-spiral. This slide is pulled out for the purpose of adjustment. Square or circular pieces of wetted muslin make contacts with equal areas on two opposite sides of the experimental tissue, these pieces of cloth being connected with non-polarisable electrodes, E and E’ After the adjust-

ment is made, the slide is pushed in, till the tissue is well in the centre of the coil. When the circuit is completed, for a brief period, both the sides A and B are subjected to the same sudden variation of temperature, which, as we know, acts as a stimulus. As the two contacts are thus in practice raised to the same temperature, there will be no thermo-electrical disturbance. The responsive current, therefore, will be determined by any difference of excitability which may exist

tion, which acts as a contributory stimulus. That it is the thermal variation, and not the temperature, which acts as the efficient external stimulus, is seen from the fact that when the tissue is subjected to the higher temperature con- tinuously, the galvanometric deflection obtained is opposite in direction to that induced by the thermal shock. This is because the absorption of heat, as such, increases the internal energy, and thus induces an electrical effect opposite to that caused by external stimulation.

As experimental tissue, we may use the sheathing petiole of Musa. The required piece is cut and mounted in the apparatus, the concave surface being taken, say, as B, and the convex as A. I have mentioned J/usa as suitable for this purpose, because I find it, when fresh, to show practically no sign of fatigue in its responses. There are many other sheathing petioles, which would doubtless answer the same purpose more or less perfectly. | In obtaining records with this specimen, it is found that the responsive current flows across the petiole, from the inner concave surface B to the outer convex surface A, showing that it is the inside which is more excitable. Uniform stimuli of short duration were applied at intervals of one minute, and the responses obtained are seen to be fairly uniform (fig. 82). The specimen was next subjected to the anesthetic action of chloroform. This, it will be seen, in- duced a very great depression of the response.

It has thus been shown that just as the greater contrac- tion and concayity of a motile organ enables us to discrimi- nate which side of two is the more excitable, so here also the more excitable side is that which, on diffuse stimulation, exhibits galvanometric negativity relatively to the other. From this it becomes possible to determine the relative excitabilities of any anisotropic organ, even though it be non-motile, and therefore incapable of exhibiting any con- spicuous mechanical response. The difficulty of applying equal and quantitative stimulus on two sides simultaneously

Fic. 82. Responsive Current in Petiole of Musa from Concave to Convex Side First series, normal ; after application of chloroform subsequent depression. has now been overcome by vibrational stimulation, and by the perfection of the method of thermal shocks. Thus a definite resultant response has been shown to be determined by the differential excitabilities of two parts of an experi- mental tissue. And that from this consideration it becomes further possible to resolve many of the remaining anomalies of electrical response will be fully demonstrated in a sub- sequent chapter.

Natural current in anisotropic organ from the less to the more excitable—External stimulus induces responsive current in opposite direction—Increase of internal energy induces ‘positive, and decrease negative, variation of natural current— Effect on natural current of variation of temperature—Effect of sudden variation—Variation of natural current by chemical agents, referred to physiological reaction—Agents which render tissue excitable, induce the positive, and those which cause excitation, the negative variation—Action of hydrochloric acid—Action of Na,CO,—Effect modified by strength of dose— Effect of CO, and of alcohol vapour—Natural current and its variations— Extreme unreliability of negative variation so-called as test of excitatory reaction— Reversal of natural current by excessive cold or by stimulation — Reversal of normal response under sub-tonicity or fatigue.

WE have seen that when the pulvinus of A/zmosa is excited by an external stimulus, there is a relatively greater expulsion of water from the more excitable lower half, with a con- comitant greater contraction. Conversely, the lower half of the pulvinus is capable of absorbing more water, and of expanding to a greater extent, than the upper. Increased internal energy, in contrast to the action of external stimulus, has the effect of causing a greater expansion of the lower half of the pulvinus, and thus raising the leaf. This we saw exemplified when the plant was subjected to a gradually rising temperature, so as to increase its internal energy, its leaves being thereby made to show increased erection (p. 72), Hence the more excitable tissue in the pulvinus of Wzmosa is characterised, both by greater power of absorption and by greater emission of energy, according to circumstances. In this we see a close analogy to the action of inorganic bodies, in which also we find the greatest power of emission to be associated with a correspondingly great power of absorption of

We have thus seen that in order to maintain a high state of excitability, absorption of energy is necessary. On excitation, emission of energy occurs. In this latter case, of emission, we observe a concomitant galvanometric negativity of the more excited lower side. Since to have maintained its excitability the opposite process of absorption would have been necessary, it follows that the more excztable lower side must under normal conditions be galvanometrically positive. This is found to be the case. For when the leaf is in an excitable condition, there is an electro-motive difference between the upper and lower halves of the pulvinus, in con- sequence of which a current flows across the tissue, from the less excitable upper to the more excitable lower half, which is thus galvanometrically positive, in relation to the upper. We have here, then, an additional instance of the opposite effects of internal energy and external stimulus. Internal energy, maintaining a greater excitability of the lower half of the pulvinus, induces in it a relative galvanometric positivity. External stimulus, on the other hand, gives rise to precisely the opposite effect—namely, the relative galvanometric negativity of the lower half. Under typical conditions, then, we may expect the more excitable point to be galvano- metrically positive; and the more excited to be galvano- metrically negative.

Turning next to non-motile tissues, we find the same conclusions to hold. good. We saw that in the case of the sheathing petiole of M/usa, the concave was more excitable than the convex side. The concave is thus normally positive to the convex side, and the natural current flows across the tissue from the convex to the concave. While the natural current flows from the more excitable to the less excitable, external stimulus gives rise to a responsive current in the opposite direction, from the more excited to the less excited, constituting a negative variation of the current of rest. .

Let us next consider what would be the effect of an increase of internal energy on the natural current. Since the stimulus, we should expect it to induce a positive variation of the natural current. Diminution of internal energy on the other hand might be expected to cause a negative variation. These effects are diagrammatically represented in fig. 83, which also exhibits the parallelism between the electric responses of motile pulvinus and non-motile anisotropic organ. :

increased or diminished internal energy on the natural current to experimental verification. As regards the increase ~ of internal energy, we have already seen that this can be secured by a gradually id Than rising _ temperature, its ¥ diminution being, con- eas 2G, trariwise, secured by a 2 falling temperature. In Fic. 83. Parallelism of Natural Current . in Pulvinus of Mimosa and Sheathing 54W that the former in- Upper and less excitable surface of former Jeayes and the latter a corresponds with outer or convex sur- ‘