Comparative Electro-Physiology: A Physico-Physiological Study

The pulsations to the left were recorded at the ordinary temperature of the room, 29° C. Those to the right, when the temperature had been lowered to 25° C. to cause antagonistic expansion and consequent galvano- metric positivity. This view finds support from the records seen in figs. 129 and 133, given in the next chapter. The first of these (fig. 129) shows the expansion, with consequent physical elongation, of the filament of Passzfora under a rising temperature. In the second (fig. 133) is seen the in- creasing galvanometric positivity of a specimen of Amaranth under similar circumstances.

is below a certain thermotonic minimum, the effect of a rise of temperature will be to enhance the amplitude of response by removing molecular sluggishness. This fact has been illustrated in the gradually heightened mechanical response of the autonomous pulsation of Desmodium gyrans when a plant artificially cooled was allowed to return to the normal temperature of the room (fig. 120). If similarly a plant tissue be first cooled and then allowed to return to the surrounding temperature, its electrical responses to suc- cessive uniform stimuli being recorded throughout, a stair- case increase of response will be observed during the return. When the temperature, however, is raised above a certain optimum, a depression of the amplitude of response begins, not by the depression of excitability, but by the increasing force of recovery due to an augmentation of the internal factor. True depression only takes place when the plant is approaching a condition of heat-rigor.

One very curious effect of temperature-variation which has been touched upon is the marked increase of sensi- tiveness which often makes its appearance as its after-effect. This is seen exemplified in the record given in fig. 124, showing the effect of a cyclic variation of temperature on Eucharzs lily. In another experiment with Scotch kale, the response at the temperature of 30° C. was eleven divisions, and at 50° C. eight divisions, during the thermal ascent. During the descent, however, the amplitude at 50° C. was sixteen, and at 30° C. twenty-three divisions. The sensitiveness was thus doubled. This enhancement may be due in part to the increased molecular mobility consequent on the annealing effect, as it were, of temperature- variation. But it may also be regarded as partly due to the difference of the antagonistic forces which the excitatory response has to overcome during ascent and descent. During the thermal ascent, the opposing expansive force is being rapidly accelerated. During the thermal descent, on the other hand, this is no longer the case, for the force of re- covery is now undergoing a diminution.

When the temperature is raised above a certain critical point, the plant is killed, and its electrical response dis- appears at the same time. This is demonstrated visually in the accompanying photographic record (fig. 127). In this case, normal responses were first obtained at the usual tem- perature of the room. Steam was next introduced into the Fic. 127. Photographic record showing effect of Steam in abolishing Response The two records to the left exhibit normal response at 17° C. Sudden warming by steam induced at first an inorease of response, but five minutes’ exposure to steam killed the plant (carrot) and abolished the response.

Vibrational stimulus of 30° applied at intervals of one minute; vertical line = ‘I volt. plant-chamber, and kept streaming in during the course ot the experiment, electrical responses being recorded mean- while at intervals of one minute. It will be seen that at first a transitory augmentation ot excitability was induced. But this quickly disappeared, and in five minutes the plant was effectively killed, as is shown in the waning and final aboli- tion of response. This experiment affords us a qualitative demonstration of the abolition of response at death under

the influence of high temperature. In the next chapter we shall enter upon an exact determination of this critical point of death. cate It is thus seen that temperature modifies the electrical response of plants. There isa temperature-minimum below which response is abolished. If the plant be kept too long at this temperature it is apt to be killed. In the case of a delicate species like Eucharis, which is highly susceptible of the injurious effect of cold, the electrical response is per- manently abolished by long exposure. But hardier plants, like Holly and Ivy, show revival of electrical response, on a return to a favourable temperature. The electrical response disappears also at a certain maximum temperature con- stituting the death-point.

Different fost-mortem symptoms—Accurate methods for determination of death- point— Determination of death-point by abolition or reversal of normal elec- trical response—Determination of death-point by mechanical death-spasm— From thermo-mechanical inversion—By observation of electrical spasm : (a) in anisotropic organs: (4) in radial organs—Simultaneous record of electrical inversion and reversal of normal electrical response—Remarkable consistency of results obtained by different methods—Tabulation of observations.

IT will be seen from the last chapter that there is in the case of every plant a certain high temperature which is critical, since above it life passes into death. Much difficulty has been experienced in the exact determination of this critical point, because no sure criterion of death was hitherto avail- able, such as would furnish an immediate and reliable indica- tion of its occurrence. The various symptoms of death, such as drooping, withering, discoloration and the escape of coloured cell-sap, do not manifest themselves at the onset of death, but at some time indeterminately later. Even when a plant has been subjected to a temperature in excess of the fatal degree, it continues to appear fresh and living ; and it is not till after some longer or shorter interval that the death symptoms are seen.

To take, for example, the symptom of drooping, it is clear that the loss of turgidity on which this depends cannot at once make itself visible. In a thick tissue, again, death may take place in the superficial layers of the plant, the interior tissues, owing to feeble thermal conductivity, remain- ing comparatively unharmed. Or, if we employ the test of discoloration, which we shall find to occur some time after the initiation of death we find that the exact moment at

which discoloration begins cannot be detected with sufficient precision. When we place the specimen in a thermal bath under a rising temperature, the beginning of discoloration - after death is so slight as to be impossible of detection, and by the time it becomes marked, the temperature has already passed several degrees above the fatal point. I have found, for example, that the colour of the milk-white style of Datura _ .alba has changed to brown by the time that the temperature

of the bath has risen to about 64° C. In the petals of Sesbania coccineum, again, a striking change of colour is detected, under similar conditions. Rich crimson here turns into pale blue at a temperature of about 67° C.. The fila- mentous corona of Passiflora quadrangularis, finally, in which the filaments are barred by purple rings, loses its colour normally at about 68° C. In all these cases, the initiation of the loss of colour must have been imperceptible. Hence, all that can be determined from such experiments is that the death-changes must have commenced at some. temperature ~ lower than 64° to 68° C.

Before proceeding further, it is necessary to obtain a clear idea of what is meant by the death-point. In animals, an early symptom of death consists in the setting in of rigor mortts. But this does not synchronise throughout the body, certain parts of the organism undergoing the death- change earlier than others. Thus the only definition of the death-point which can be made at all precise is that which regards it as the point of initiation of some unmistakable sign of death. I shall next proceed to describe several death-symptoms and the modes by which they may be de- tected with certainty.

With regard to such detection, I Siti pointed out else- where that, theoretically, it should be possible to make such a determination by watching the waning of some effect characteristic of the living condition, the death-point being known by its cessation at a given moment. Such a test, as we shall presently see, is afforded by the electrical responses. The ideally perfect method, however, would be by the detection of some effect which at the moment of death under- went a sudden reversal to its opposite. There would not here be even that minor degree of uncertainty which is in- cidental to the determination of the exact vanishing-point of a waning effect. And such methods are afforded by my discovery of the occurrence of mechanical and electrical spasms at the moment of death.

seen that response to stimulus by galvanometric negativity is _ distinctive of the living condition. When the plant is killed, this normal response disappears. At the moment of death from high temperature, therefore, we may expect to see the abolition of this normal excitatory response of negativity. For this investigation I took a batch of six radishes. The specimens were kept for five minutes previous to each experiment in water at a definite temperature (say of 17° C.), and were then mounted in the vibration-apparatus and their responses observed. Each specimen was next dis- mounted and replaced in the bath at a higher tempera- ture (say of 30° C.) for another five minutes. After this, a second set of responses, to the same stimulus as before, was taken. In this way observations were made with each plant, till the temperature at which response almost or altogether ceased was reached. I give below (p. 195) a table of the results obtained with the six radishes.

From these experiments it would appear that in these cases the responses disappeared at about 55° C. It should be stated here that this investigation was carried out in the winter season in England, and it will be shown later that the incidence of cold has the effect of lowering the normal death-point by about 4° or 5° C. I was next desirous of substituting, for this method of discontinuous observations, one which should be continuous. I, therefore, subjected the specimen—a stem of Amaranth— to a continuous rise of temperature, and took records of responses to uniform stimuli after every few degrees of the ascent. I found here that not only was there a gradual

decrease of response, tending towards its abolition, with rising temperature, but also that, at the death-point, it under- Fic. 128. Record of Electric Responses of Amaranth at various temperatures The response undergoes reversal to positive at the critical temperature of 60° C. tivity to positivity (fig. 128). This was due to the fact that on reaching the death-point, the contained positive com- ponent in response was unmasked by the abolition of the true excitatory effect. But this positive response disappears also after a short time. It will thus be seen that by this method the death-point is capable of determination within very narrow limits, having been, in the present case, near 60° C. When the tissue is thin, this temperature soon proves fatal. But

is necessary, if the interior of the tissue is to be killed effectively. I have also discovered another method of obtaining the death-point with precision, in the symptoms afforded by mechanical responses. For I found thata death-spasm occurs at a certain critical moment in a plant, which is analogous to the death-throe of the animal. The experimental plant— Mimosa, for instance—was placed in a bath of water, whose temperature was being raised gradually, at a uniform rate of, say, 1° C, per one minute and a half, until the death-point was reached. During all this time there was no responsive fall of the leaf, for, we have seen, it isa sudden variation, and not a gradual rise of temperature, which acts as an excitatory stimulus. This gradual rise, on the other hand, increases the internal energy of the plant, by which the turgidity of the pulvinus is continuously augmented. In this process the increase of turgidity is more energetic in the more excitable lower half than in the upper. The greater expansion of the lower side of the pulvinus thus raises the leaf continuously. But immediately on reaching the death-point, there is a reversal of thls movement, and an abrupt fall of the leaf. This spasmodic movement is sudden and well-defined. In a vigorous J/zmosa the death-spasm is found to occur at or very near 60°C. This contraction of death is followed after some time by a fost-mortem relaxation.

That the death-response is an excitatory phenomenon is seen from the fact that any circumstance which lowers physiological activity lowers the death-point also. Thus, after a spell of cold weather, I found that the death-point of Mimosa was lowered from the normal 60° C. to about 53° C. This latter value, it will be remembered, was ap- proximately the same as that obtained with radish, in winter, by the method of electrical response. Again, I find fatigue to induce a lowering of the death-point, the extent of which depends upon the degree of fatigue. When this was moderate, I have found the death-point of MW/zmosa to be lowered to

This spasmodic contraction, indicative of the initiation of death, may express itself in diverse ways. For example, if the tubular peduncle of Ad/zum be filled with water, and raised gradually in temperature, there comes a moment at which a sudden expulsion of the contained water occurs. A spiral tendril of Passzfora, under the same circumstances, exhibits a sudden uncurling. The florets of the ray, in certain Composite, show characteristic movements, either up or down. In all these cases alike, under normal circumstances, the death-point is found to be at or near 60° C.

Turning next to the radial organs of ordinary plants, these also exhibit a sudden longitudinal contraction at the onset of death. I have shown elsewhere how, by means of the Morograph, an instrument which I devised for this purpose, a thermo-mechanical curve is recorded by the specimen, while it is being subjected to the continuous rise of temperature, culminating in the death-point. The ordinate of this curve represents the induced variation of length, and the abscissa the temperature. The expansion described in the case of MWimosa is seen here in the form of a gradual elongation, up to the moment of reaching the death-point. When this point is reached, however, a sudden contraction takes place, giving rise to an inversion of the curve. This turning point is very abrupt. The curve as a whole is thus one of life-and-death, in which the point of inversion separates the two. I give below a photographic record of this thermo-mechanical curve, obtained with the coronal filament of Passtfora. The death-point occurred here at 59°6° C. (fig. 129). The thermo-mechanical curve is very

similar in similar specimens under normal conditions. Fig. 130 gives two records of two different styles of Datura alba, obtained from flowers of the same plant. The death- point is seen to have occurred at 60°C. In recording the thermo-mechanical curve, there is found to be, normally speaking, a continuous expansion up to the death- point. In the case of vigorous specimens, in a good tonic condition, the inversion does not take place till about 59°6° or 60° C. But in less vigorous specimens, a certain hesitation, as it were, is seen to occur in the record at or near 55° C. With vigorous specimens,

Fic. 129. Photographic Record of Thermo-mechanical Curve given by Coronal Filament of Passiflora The first or down part of the so 30° gee oe S* S0° 55° 60° 6S° curve shows expansion, but on reaching death-point, at Fic. 130. Thermo-mechanical Curve of « 59°6° C., there ska sudden Two Different Specimens of Style of inversion, due to spasmodic Datura alba, obtained from Flowers death-contraction. of the same Plant there may be the merest indication of this hesitation ; in other cases, with less favourable tonic condition, the hesitation is prolonged, but the expansion finally proceeds, ahd the death inversion takes place at the usual temperature of about 60° C. When the specimen, however, is enfeebled, or has been subjected to unfavourable circumstances, the point of transient instability becomes fatal, and the inversion takes place there. As an example of what has just been referred to—namely, the influence of unfavourable external circum-

stances in lowering the death-point—it may be mentioned here that the sudden incidence of cold weather will lower it by some 4° or 5° C. Intense fatigue will lower it by as much as 19° C. _ [have already said that this spasm, taking place at the moment of death, is an excitatory response. On this theory it occurred to me that it should also be possible to determine the onset of death by an electrical spasm. It may be well at this point, therefore, to examine some of the conditions under which such a spasm, supposing it to take place, might be displayed most conspicuously. We may suppose a radial organ, with the usual electrical contacts, A and B, to have its temperature raised gradually up to the death-point. The excitatory effect of death may now be expected to cause the galvanometric negativity of a given point. But since these excitatory effects are equal and similar at A and B, they will balance each other, and there will be little or no resultant galvanometric response. In order to obtain a marked resultant effect, then, we must have an organ in which the excitabilities of the two points A and B are different. This difference of excitability, necessary to the exhibition of a resultant response, may be either natural or artificially induced. For the former, we may take a specimen which is not radial, but anisotropic, thus affording us two points of galvanometric contact, possessed of unequal excitabilities.

We have seen that the inner surface of the petiole of Cucurbita maxima was more excitable than the outer. The same is true of the hollow peduncle of Uvzcits lily. There is also a great difference of excitability as between the upper and lower sides of the scale of the bulb of the same lily, in the season of flowering, the concave surface of this scale being more excitable than the convex. Any of these specimens described I find to answer asmizably for the purpose of this investigation.

Taking the petiole of Cucurbita, then, I divided it longi- tudinally, and rejected one-half, ‘thus obtaining a half-tube, of which the inner concave surface was more excitable than the outer convex. Electric contacts were now made through non-polarisable electrodes with equal and opposite areas on the two sides. These adjustments were made in a heating chamber containing electrical arrangements by which the temperature could be raised continuously. This was satisfactorily accomplished by an incandescent electrical lamp which was placed in a second chamber, vertically below | the plant-chamber. There was a wooden partition between the two, by which the light of the lamp was excluded from the specimen (fig. 131). For the radiation itself will be

Electric lamp in the lower compartment raises temperature of the upper. E, E’, electrodes making contacts with the specimen ; T, thermometer. shown to constitute stimulus, and the object in the present case was to eliminate all exciting factors except death itself. By means of side-openings, the heated air was enabled to pass into the plant-chamber, thus raising the temperature. A rheostat included in the lamp-circuit made it possible to adjust the rate of this rise of temperature, its average being about 1° per minute.

The natural current through the petiole is, under normal circumstances, from the less excitable outer to the more excitable inner surface: that is to say, the inner is galvano- metrically positive. This is true when the excitation, due to section made for the purpose of preparation, has subsided. This stimulation, by causing greater excitation of the inner surface, is liable to induce there a temporary negativity. A gradual rise of temperature, as we saw, caused an increased turgidity of the more excitable lower side of the pulvinus of Mimosa, and this increased turgidity was exhibited mechanically by the erection of the leaf. But the electrical sign of increased turgidity is galvanometric positivity. We have also seen that electrical responses occur equally in motile and non-motile tissues. In the petiole of Cucurbita then, on its more excitable inner surface, we obtain, during the gradual rise of temperature, an increasing galvanometric positivity. This is true only, as has been said before, when the rise is continuous, and not marked by fluctuation. For any sudden variation will act as a stimulus, causing galvano- metric negativity of the more excitable inner side. For this reason it is necessary that the rheostatic resistance inter- posed in the lamp-circuit, for the adjustment of the uniform rate of rise of temperature, should be made at the beginning of the experiment, such tissue being very sensitive to this particular stimulatory action. At the commencement of my investigation I experienced much trouble from the erratic movement of the galvanometer spot of light, and the obtaining of a steady electrical curve seemed at that time almost hopeless. Later on, however, I found that these fluctuations were traceable to temperature-variations, unavoid- ably associated with the attempt to regulate the rise of temperature by movement of the rheostatic slide. It is for this reason, then, that the adjustment must be made, once for all, at the beginning.

Carrying out the experiment in this manner, I obtained, with various anisotropic organs, a sudden inversion of the electric curve at the death-point. This death-point was found, in all vigorous specimens, from which traces of injury had been removed by previous rest, to occur accurately at 59°6° or 60° C._ In these electrical curves, the same point of instability, already noticed in the thermo- mechanical curve, was often found to occur at or about 55°C. And if the specimen were not in favourable tonic condition, or had been suffering from injury, the death-point was lowered to this degree. 3 I give below an electrical curve showing the point of inversion at death (fig. 132). It was obtained with the sheathing petiole of Musa. The inner or concave side of this petiole is more excitable, as we have seen, than the outer. These responsive electrical variations were very large, and could not be represented within the limits of the photographic plate. I therefore took a photographic record between the temperature of 54° C. and 67° C. only. The first part of the curve represents the increasing galvanometric posi- tivity of the more excitable inner surface of the specimen. The same process of increasing posi- tivity under the continuous rise of temperature, had been going on Hieltae whctourerkak Reacont previously, it is to be understood, exhibiting Electric Spasmin before arrival at 54° C., at which he Fe Hole oh Ae the photographic record was com- Sudden electric inversion takes ee place at the death-point, menced. This increasing galvano- 59°5°. Record was com- metric positivity corresponds to menced at 54° C., and suc- : . cessive gaps in the record the gradual erection of the leaf in cae C. rise of tem- Yy0sa, and to the expansion of a radial organ, such as a coronal filament of Passzflora, all alike being due.to the positive variation of turgidity. In order to give an indication of the particular temperature at each portion of the curve, the re- cording light was obscured for about 15 seconds after each degree of temperature. The successive gaps, then, are one

degree centigrade of temperature apart. These interruptions, however, were not made after the occurrence of the inversion. As soon as the death-point was reached, in the present case at 59°5° C., there was a sudden inversion of the electrical curve (fig. 132), corresponding with the point of inversion of the thermo-mechanical curve (fig. 129). Each of these curves is seen to bear a striking resemblance to the other. In both cases, the inversion was due to the same fact of sudden excitation, finding expression in the one, in induced galvanometric negativity, and in the other, in mechanical contraction.

In the case of organs which are more or less radial, and in which there is little differential excitability, it is necessary to abolish the excitability of one contact, as, say, by previous scalding. For this experiment I took a leaf of Amaranth, and injured a portion of the lamina by immersion in boiling water. The two contacts were made, one with the petiole, and the other with the injured lamina. On raising the temperature continuously, the more excitable petiole became increasingly positive. The photo- eraphic record in this case was com- menced only on reaching 55° C., and the death-inversion took place at date Pitre, saree 59°5° C. (fig. 133). version -at Death-point,