Comparative Electro-Physiology: A Physico-Physiological Study

Fic. 48. Photographic Record former, and the resulting response of Electrical Responses of . : A Potato-tuber is determined by an _ algebraical applied at distance ; 4, Negative yjgorous specimen, whose excit- response to stimulus applied tan : vanometric negativity masks the positivity. The resultant electrical response in general is expressed by the formula N,z—P,, where N, is the galvano- metric negativity due to the true excitatory effect,and P,, the positivity due to the hydro-positive effect, which immediately precedes it. From this, it is clear that, as regards the resultant galvanometric response of vegetable tissues under stimulation, there may occur the two typical cases displayed in the following table :

Conditions . Constituent Factors Resultant Response Excitability great Nez >Pu Galvanometric negativity Excitability diminished Ng <Pxy Galvanometric positivity We thus see how, under the physiological modifications induced by various agents, the normal negative response of a tissue may undergo diminution, or even reversal. We have, for the sake of simplicity, assumed that the two antagonistic effects act on the specimen simultaneously. But, asa matter of fact, under certain circumstances, their time-relations may be subjected to change. Hence various effects of interference may be seen, giving rise to diphasic responses, such as positivity followed by negativity, or the reverse. Or, instead of diphasic, there may even be multi- phasic responses, since it will be shown that very strong stimulus may cause not a single but repeated responses. Some of these effects will be described in detail in a sub- sequent chapter.

I shall here meanwhile describe another method, in which, by means of a selective physiological block, we can unmask the contained hydrostatic effect of positivity, from the ordinary response of galvanometric negativity. The oc- currence of the excitatory contraction, in J/zmosa for example, as shown by the fall of the leaf, depends on a favourable excitatory condition of the tissue. If this ‘motile excitability should be in any way depressed or abolished as, say, by application of ice-cold water on the pulvinus, the true excitatory response could not take place. But if, under

these circumstances, we applied stimulus at the diametrically Opposite point S, (fig. 42), the hydrostatic wave alone would It will thus be seen that the hydrostatic transmission of the indirect effect of stimulus is not to any great extent affected by the loss of excitability of the tissue. Similarly, the transmission of true excitation may be selectively blocked, in a tissue, by the application of various depressing agents, such as anesthetics, without appreciably affecting the passage of the wave of increased hydrostatic tension.

For the present experiment I took a leaf of fern, and made the proximal electrical connection with the petiole, and the distal with an indifferent point on a leaflet. By means of the thermal stimulator (p.38), I now applied successive © uniform stimuli at intervals of one minute, on a point 15 cm. below the proximal .contact. The first. three Fic. 49. Photographic Record of Electrical Response of Petiole of Fern First part of record shows normal negative responses ; second part shows positive response unmasked by selective physiological block of chloro- form ; in the third part is seen the abolition of response when stimulus is applied on anzsthetised area itself.

responses of the series are seen (fig. 49) to be more or less uniform, and negative. Such resultant response is, as has been pointed out, due to the summation of two antagonistic effects, of which the excitatory is predominant. In order therefore to eliminate from this resultant response its excita- tory component, I applied chloroform on a narrow belt inter- mediate between the point of stimulation and the proximal contact. From the next three responses it will be seen that the hydrostatic effect of positivity, represented by down- responses, was thus successfully unmasked. The application of chloroform is thus seen to act here as a selective block,

the wave of increased tension being transmitted across its area, whereas the wave of true excitation is arrested. But the hydrostatic disturbance itself was caused by the excitatory contraction of the distant point. The abolition of the excitability of an intermediate point did not, as we have seen, block the hydrostatic wave. If now the stimulator be brought nearer, and placed over the strongly anzsthetised area, the expulsion of water dependent on true excitation can no longer take place. On doing this, therefore, we find that neither the true excitatory negative, nor its consequent hydrostatic positive effect, is exhibited at the responding point. Response is thus totally abolished.

We thus see that incident stimulus gives rise in the plant to two distinct responsive expressions. In that which we shall consider first—namely, the direct, or true excitatory effect—there is an expenditure of energy, which is indicated by a contraction, negative turgidity-variation, mechanical fall, and galvanometric negativity. In the second, or indirect, of these two effects, we have the opposite of ali these. That is to say, we have here an increase of internal energy, expan- sion, positive turgidity-variation, erection of leaf, and galvano- metric positivity. The galvanometric negativity, due to negative turgidity-variation, is generally speaking of much greater intensity than the galvanometric positivity, due to positive turgidity-variation. Hence, when the two effects act on a tissue in rapid succession, that of electro-negativity masks the positive. The wave of increased hydrostatic tension, by which the indirect effect of stimulus is transmitted, has a greater speed of transmission than that of true excitation. It may be transmitted even across tissues of which the excita- bility has been depressed or abolished, whereas the trans- mission of true excitation is arrested by the intervention of such a physiological block.

The occurrence of these opposed effects of galvanometric negativity and positivity has long been regarded as a pheno- menon of a highly perplexing nature. It was too hastily assumed that wherever these opposed electrical changes took place, they were necessarily to be ascribed to opposite or antagonistic chemical processes—namely, of assimilation and dissimilation. That this, however, cannot be universally true, has here been proved by tracing the two effects to their definite respective origins, in positive and negative turgidity- variations. I shall enter into this subject in greater detail in the subsequent chapters.

Hydraulic transmission of energy in living tissue ~True meaning of tonic con- dition—Opposite expressions of internal energy and external stimulus seen in growth-response— Parallelism between responses of growing and motile organs —Increased internal energy caused by augmentation of temperature finds expression in enhanced rate of growth ; erection of motile leaf; curling move- ment of spiral tendril ; and galvanometric positivity—External stimulus induces opposite effect in all these cases—Sudden variation of temperature, acting as a stimulus, induces transient retardation of growth; depression of motile leaf ; uncurling movement of spiral tendril ; and galvanometric negativity—Laws of mechanical and electrical response.

We have seen in the last chapter that external stimulus, directly applied, induces in an excitable vegetable tissue a contractile response, with concomitant galvanometric nega- tivity. An increase of internal energy, on the other hand, was there seen to give rise to expansion, and galvanometric positivity. These two factors of external stimulus and internal energy are thus seen to be antagonistic in their general expressions. But while stimulus from outside, impinging on an excit- able area, thus caused an expenditure of energy at that area, by inducing excitatory response there, yet it was also found that, by its indirect effect, it brought about an increase of energy in neighbouring tissues. By the sudden contraction, and expulsion of water from the excited area, energy was © transmitted hydraulically, and the consequent positive tur- gidity-variation caused an erectile response of a neighbour- ing motile organ. In the simple case in which the point of receptivity was at a distance from that of response, it was seen that these two effects of stimulus, direct and indirect, were easily discriminated from one another. But as the

stimulator was brought nearer and nearer, the two effects became superposed, and one was masked by the other. Even in this case, however, on careful examination, it is possible to infer the results due to the action of the internal factor. Thus, we may suppose stimulus to be applied directly on the pulvinus of M/zmosa, bringing about a responsive fall of the leaf. The expelled water from the excited pulvinus will — ‘now be forced into the neighbouring tissues, making them over-turgid, and raising their energy above par. On the

cessation of stimulus, the water that has been forced away - will flow back from the region of heightened, to that of lowered tension, and re-establish the normal turgidity of the pulvinus, which had undergone a negative variation, The erectile recovery of the leaf is thus seen to be, not a merely passive process, but an effect dependent on the internal energy of the plant.. This is also shown by the fact that in autumn and winter, when the internal energy is low, the period of recovery is very long, being sometimes as much as eighteen minutes ; whereas in summer, on the other hand, with the increased internal energy of the plant, it takes place nearly three times as quickly. I have elsewhere shown that it is this internal energy which is vaguely referred to as the tonic condition of the tissue, and that it consists of the sum total of energy derived from external stimuli previously absorbed and held latent by the plant. The different forms of stimulus may be very various. We have for instance tonicity, as imparted by light, or phototonus ; by favourable temperature, thermotonus ; by electrical current, edectrotonus; by internal hydrostatic pressure, hydrotonus ; or by the presence of favour- able chemical substances, chemotonus.

We have seen that, as a general rule, external stimulus and internal energy find responsive expressions of opposite sign, the former inducing a negative, and the latter, a positive turgidity-variation. It is, nevertheless, important to demon- strate the extensive applicability of this law, by many different results and different modes of manifestation. And such a demonstration would undoubtedly become still more con- vincing, if we should succeed in discovering some mode of response in which the antagonistic effects of internal energy and external stimulus found opposite expressions.

-- Such an example, of a very striking character, I have in my work on ‘ Plant Response’ shown to be found in growth- response. But the same opposition, between the effects of external stimulus and internal energy, I shall now proceed to demonstrate not only by means of growth-response, but also through mechanical and electrical responses. In the case of srowth, the responsive expression of the growing organ, under increased turgidity, consists of an expansive elongation. If the organ be growing at a uniform rate, an increase of internal energy will enhance that rate. But when external stimulus acts directly on the growing organ, the normal rate of growth is retarded during the action of stimulus. Thus it will be seen that though the mechanical response of a motile organ, and the movement of a growing organ, appear so different, yet these two expressions are not fundamentally distinct. For while in one, the application of direct stimulus, causing nega- tive turgidity-variation and contraction, induces depression of the leaf, in the other, the same negative turgidity-variation and contraction under external stimulus causes a depression of the rate of growth. And, on the other hand, indirect effect of stimulus, er increase of internal energy, inducing positive turgidity-variation, brings about in the one case the erection of the leaf, in the other an enhancement of the normal rate of growth. This parallelism is displayed in detail in the table given below.

It is thus understood that the indication of response to external stimulus is the depression of the motile leaf, or depression of the rate of growth, while the effect of increased internal energy is the erection of the motile leaf, or enhance- ment of the rate of growth. We shall first deal with the responsive expression to that positive turgidity-variation which is due fo the increase of internal energy. One mode of increasing the internal STIMULUS AND INTERNAL ENERGY ON PULVINATED AND GROWING ORGANS

| Mechanical response Growth response Effect of normal turgidity : Effect of normal turgidity : Local action of external stimulus : Local action of external stimulus : and concomitant depression of leaf. and concomitant depression of rate of growth. Action of internal energy exhibited by | Action of internal energy exhibited by (a) Recovery ; (az) Recovery: Re-establishment of turgidity and Re-establishment of turgidity and gradual return of leaf to normal gradual return of organ to normal horizontal position. rate of growth.

(4) Increased hydrostatic pressure : (4) Increased hydrostatic pressure : energy of a plant is by a moderate rise of temperature. And this finds expression in the case of a motile organ, by the erection of the leaf. Thus, when J/zmosa is raised in tem- perature, all its leaves become highly erect. A diminution of energy, on the other hand, by cooling, brings about a depression of the leaves. In the same way, the rate of growth is exalted by rise of temperature. Thus in a growing flower of Crinum lily the normal rate of growth at 30° C. was ‘oo4o mm. per minute, and this was exalted to ‘o113 mm. per minute, or nearly three times, when the temperature was raised*to 35°5° C. Lowering of temperature, on the other hand, greatly de- presses the rate, and may even, if it proceed far enough, cause arrest of growth.

As regards external stimulus, on the contrary, we have seen that its effect on the motile organ is one of depression. The following record (fig. 50) shows that it has a similar influence on the rate of growth. The first part of the curve shows the normal rate of elongation. But after the applica- tion of stimulus of light, growth is not only retarded, but there is an actual shortening of the organ. On the cessation of stimulus, the normal rate of growth is gradually re- established.

I shall next give examples in which the opposite effects of external stimulus and internal energy are exhibited in growth response, motile response, and electrical response. To take first the response of growth: we have seen that a steady rise of temperature brings about an increase of internal energy, while a sudden variation of temperature acts as a stimulus. Thus, if we effect a sudden augmenta- tion of temperature, this will act on the organ, during the period of variation, as a stimulus; but afterwards, when the temperature itself, or its rate of rise, has become

Fic. 50. Longitudinal Contraction and Retardation of Growth under Light in Hypocotyl of Smaps nigra The first part of the curve shows the normal rate of growth. Arrow (1) indicates moment of application of diffuse light, which is seen not only to retard growth, but also to induce a’ marked contraction. The second arrow indicates moment of withdrawal of light, and dotted portion of the curve shows recovery. steady, the condition will act by increasing the internal energy of the organ. These opposite results are seen to be strikingly illustrated by growth response in the case of the following record (fig. 51),

The normal rate of growth at 34° C. was here ‘o15 mm. per two minutes. By a sudden application of heat, raising the temperature of the chamber ultimately by 1° C., a respon- sive contraction was caused, as is seen in the record. But, on the attainment of a steady augmented temperature of 35° C., an increased rate of growth, which now amounted to 024 mm. per two minutes, was observed, owing to the increase of internal energy. Or the same difference may be demonstrated by means of mechanical response. A Mzmosa is placed in a small chamber and subjected to a sudden rise of temperature. In consequence of this there is a preliminary excitatory depression, followed, on the attainment of a steady rise, by gradually increasing erectile re- sponse, which carries the leaf above its original level.

These opposed motile effects can be shown, moreover, even in the case of ordinary plants. We take a_ spiral tendril of Passiflora. In this, the outer or convex surface is more ex- citable than the inner or con- cave, and external stimulus, causing greater contraction of this more excitable outer side, induces a movement of un- curling. This movement corre- Fag, St, gpa cf Groth of Mimosa. The response by and 35° C. increase of internal energy is, The dotted line represents the however, the opposite of this,

change. Note the contractile and consists of a movement of twitch and transient highly ac- : “1. celerated growth which follows. curling. When the tendril is The rate of growth became con- placed in a vessel of water, of uncurling, followed by a movement of curling, when the higher temperature has become steady. The electrical expressions of external stimulus and in- ternal energy are similarly opposed. In fig. 85 (Chapter X.) will be seen a record showing that sudden variation of temperature, acting as an external stimulus, induced a

temperature had the opposite effect—that, namely, of. inducing galvanometric positivity. It is thus seen that while the characteristic effect of external stimulus on an excitable tissue is to cause a nega- tive turgidity-variation, that of increased internal energy is to induce a positive turgidity-variation. The former of these, or negative turgidity-variation, finds electrical expres- sion in galvanometric negativity ; in a motile organ by the fall of the leaf, and in a growing organ by retardation of the rate of growth. The latter, or increase of internal energy, on the other hand, is expressed electrically by galvanometric positivity ; mechanically, by erection of the leaf; and in a growing organ by an acceleration of the normal rate of growth.

We thus arrive at the following laws of response of isotropic organs : 1. Effective stimulation induces contraction and galvano- metric negativity. _ 2. Increase of internal energy induces expansion and galvanometric positivity. Sign of response determined by latent energy of tissue, and by intensity of external stimulus— Sub-tonic, normal and hyper-tonic conditions — The critical level—Outward manifestation of response possible only when critical level is exceeded— Three typical cases: response greater than stimulus; response equal to stimulus ; and response less than stimulus—Investigation by growth-response—Instance of sum of work, internal and external, performed by stimulus constant—Positive response of tissues characterised by feeble protoplasmic activity or sub-tonicity—Enhancement of normal excitability of sub-tonic tissue by absorption of stimulus.

WE shall find, in this and succeeding chapters, that the nature and intensity of response are determined not merely by the intensity of stimulus, but also by the molecular con- dition of the responding substance. The excitatory mani- festation is dependent upon the occurrence of a particular directioned molecular distortion. Hence, if by the action of the stimuli of the environment, an incipient distortion in this direction has already been induced in the tissue, the incidence of even moderate stimulus will then prove sufficient to precipitate visible excitatory manifestation. A tissue in this condition is said to be highly excitable or fully tonic. If, on the cther hand, the tonic condition be less favourable, long- continued stimulation will be necessary to evoke the excita- tory effect. Here, during the first part of application, stimulus will appear to be ineffective. As a matter of fact, however, it is at work to give such a predisposition to the molecules that the action of subsequent stimulus shall be rendered effective.

In the simple case in which the tonic condition is favour- able, a given stimulus will induce a given responsive expres- sion. If further, the tissue, on recovery, return to its original condition, then a second similar stimulus will induce the same responsive expression as the first. These responses will thus be uniform. But if, owing to the after-effect of stimulus, the condition of the tissue itself be changed, the responses will be found to exhibit either staircase increase or fatigue decline, according to the particular molecular con- siderations involved, which will be described in detail in the following chapter.

It will be interesting here, however, to take an extreme case of a tissue in the opposite condition—namely, of great sub-tonicity. It is clear that since the excitability of the tissue is here feeble, there will be little or no outward manifestation of ercztatory response. The incident stimulus will thus be absorbed, and entirely held latent. And the increase of internal energy thus brought about may find expression mechanically by expansion, or electrically by galvanometric positivity.

In the case which we have selected, the excitability of the tissue is too low for the ordinary excitatory expression to occur. Hence the incident stimulus will be entirely absorbed, and there will be a gain of energy without any loss. The whole responsive expression will in this case consist, not of con- traction but of expansion. In other words, it will be exactly opposite to that which is usually consequent upon external stimulus. In order to demonstrate this, I took a seedling of Tamarindus indicus which had been cut off from its supply of external energy, and was consequently sub-tonic. Owing to the insufficiency of internal energy, its growth had, in fact, come to a standstill. On now subjecting this seedling to thermal stimulation, the absorbed stimulus raised the internal energy and found expression in growth expansion. We have seen that the effect of external stimulus on a growing organ in normal tonic condition was the retardation or arrest of growth. But here, in a tissue which is sub-tonic, we find that the effect is exactly opposite. :