Researches on Irritability of Plants

Similarly, I have found that the excitatory change of galvanometric negativity is transmitted to a distance through certain plant-organs. Tissues containing fibro- vascular elements, such as stems and petioles, are found to be good conductors of excitation. Indifferent tissues in leaves and tubers possess little power of conduction; in such cases excitation remains more or less localised. The parenchyma in the leaf is thus an indifferent conductor, whereas the midrib and veins are good conductors of exci- tation. Instems also great difference is found, as regards power of conduction, between the fibro-vascular strands and the ground tissue. The results of electrical investigation thus give strong support to the conclusion that plants possess conducting-tissues by means of which the excitatory state may be transmitted to a distance.

The prevailing opinion, however, up to the present has been that in plants like Mimosa there is merely a trans- mission of hydro-mechanical disturbance and no transmission of true excitation comparable with that of animal nerve. That this conclusion is erroneous will be shown from the results of varous inquiries fully described in the next chapter. In all these investigations it is necessary to determine the velocity of transmission with the highest accuracy ; and in

order to eliminate the errors that might be inherent in personal observation, it is desirable that all the data for this determination should be furnished automatically in records made by the plant itself. Successive records, therefore, should enable us to determine with equal accuracy not only the normal velocity but also its variation under given changed conditions. And here the preliminary questions arise: With what degree of accuracy can we determine the normal velocity of transmission? And how far may we depend on the constancy of this velocity, in successive experiments, under normal conditions? As regards these points, some mis- givings might naturally arise. For the factors calculated to interfere with this constancy will in all probability prove to be numerous. First, we may have the variation of excitability at the point of application induced by the stimulus itself. We have, therefore, to find out what is the maximum intensity of stimulus that may be employed without causing fatigue or other deleterious changes in the tissue. Another point to be remembered is the question already discussed in previous chapters of our ability to apply stimuli, in successive experiments, of identical intensity and duration. Unless this can be secured we cannot look for consistent results, inasmuch as the velocity of transmission may to some extent be dependent on the intensity of stimulus. Likewise, if the transmission of excitation should prove to be due to the transmission of a protoplasmic change, it is easy to see that we must allow the tissue a definite time for protoplasmic recovery after each application of stimulus, without which interval consistency of results could hardly be expected.

It was only after a long course of investigations—some of which will be described in the course of the present chapter—that I was able to analyse and provide against these several sources of variation. But even after this, I was by no means prepared for the very great consistency of the results which it has been my good fortune to obtain. For successive determinations, with the same specimen, of the periods required for the transmission of excitation through a given length of conducting-tissue, did not differ from each other by so much as one-twentieth of a second and were often actually identical.

For the purpose of these experiments I used by prefer- ence the petiole of Mimosa, for the reason that in this the conducting-strands situated in the fibro-vascular bundle would be more continuous and evenly distributed than in a branching specimen. In order to determine the velocity of transmission, the stimulus of induction-shock is applied to the petiole at a distance d from the responding pulvinus. Let us suppose # to be the true time taken by the excitation to reach the pulvinus; the initiation of the responsive movement will however be further delayed by the latent period of the pulvinus L. The total time-interval T observed to elapse between the application of stimulus and the initiation of response will therefore be the true time ¢ plus the latent period L. To obtain the true time we have to subtract the latent period L from the observed interval T, thus 7=T—L. The velocity of transmission is then found by dividing the distance by the true time. The necessary data are therefore the distance between the stimulated point and the pulvinus, the time-interval between the application of stimulus and the initiation of response, and the latent period of the individual pulvinus.

In making these determinations the apparatus employed is the same as that for the determination of the latent period. As in these experiments we have to measure time which may be several seconds in duration, the record- ing-plate is made to travel at the relatively slow rate of 2 cm. each second or thereabouts. The vibrating recorder must be selected according to the degree of accuracy that is required. For our present purpose a time-measurement accurate to one-tenth or one-twentieth of a second is ample.

We first obtain a series of records of indirect stimulation. The two electrodes, E and E’, in connection with the exciting secondary coil, are applied on the petiole about 10 mm. apart, the proximal electrode E being at a distance d from the pulvinus. The recording-plate during the course of its descent completes the primary circuit of the induction coil for a definite length of time, which is about one-twentieth of asecond. This gives rise to a definite number of alter- nating shocks to the plant. The stimulus is always applied at a definite instant in the descent of the plate; hence successive records on the same plate always commence on the same level, the vertical line in the record indicating the moment of the application of stimulus. After taking one or more records of the effect of indirect stimulation, an additional record is taken of the effect of direct stimu- lation. This gives the latent period L of the particular specimen.

Before proceeding further I must point out the neces- sity of special precautions for the perfect insulation of the electrodes in connection with the secondary coil. If one of these should happen to touch the table, then, even with connections made for indirect stimulation, a portion of the current would pass through the flower-pot holding the plant and the pulvinus would be directly stimulated by this escaping current or current of leakage. In my own case, it was some time before I discovered that certain anomalous results were to be traced to this particular source of dis- turbance, at first little suspected. To overcome this diffi- culty the flower-pot should be placed on a block of insulating ebonite, the electrodes also being carefully insulated on ebonite rods.

I will now proceed to give the actual records obtained with the arrangements detailed above. In the experiment I am about to describe the specimen of Mimosa was very vigorous. The distance at which the stimulus was applied was 30 mm. from the responding pulvinus, and the inten- sity of stimulus was 3 units. The frequency of the vibrat- ing-recorder was Io per second ; hence the distance between any two successive dots in the record represents a time- interval of one-tenth of a second, and from the record itself it will not be found difficult to estimate intervals of even one-fifth that amount. The lowest of the three records in fig. 82 represents the results of the first experiment. It will be seen that the interval between the stimulus and the

Fic. 82.—Determination of velocity of transmission of excitation in Mimosa. ‘Two lower records are in response to stimulus applied at a distance of 30 mm.; upper record in response to direct stimulation giving the latent period. Recorder to D.V. beginning of response is 16°2 spaces, each of the value of ‘I second. Therefore the total time T is 1°62 second. After a suitable interval necessary for complete recovery of con- ductivity, a second record was taken, under the same con- ditions, onthe same plate. The minimum interval necessary varies from 15 to 20 minutes, depending on the condition of the specimen and the season. It will be seen that the time-interval in this case is the same as before—namely, 162 second. The third record was taken with direct stimu- lation, and from this it will be noted that the latent period

is ‘I2 second. Thus the velocity of transmission as given by both these experiments is identical—namely, In order to put the constancy of these results to a still more rigorous test, I next modified the experiment in the following way, employing the Dvzfferential Method. The stimulus was first applied at a distance d from the respond- ing pulvinus, and the total time T was found from this record. In the next experiment the distance of the point of stimulation was reduced to d,, and its corresponding total time, T,, found in the usual manner. And lastly, a record was taken under direct stimulation. This furnished the value of the latent period L.

It will be seen that we have here three different sets of data for the determination of the absolute value of the velocity of transmission. In two of these we derive the velocity in the usual manner from the distance, the total time, and the latent period. In the third, knowledge of L is not required, as it will be seen that the difference in the times T—T, of transmission observed in the first two cases represents the time taken by the excitation to travel the difference between the two distances d—d,. Hence three separate determinations, Vj, V2, and V3, obtained with the same specimen, are T—L ee TL T—T,

The rigour of this test of constancy will be gauged by the extent to which the different determinations of Vj, Vs, and V; are consistent with one another. In an experiment carried out in this way the intensity of stimulus applied was 3 units, and the vibrating recorder had a vibration-frequency of 10 per second. In the first experiment the point of application of stimulus was at a distance of 30 mm.; the total time was found to be I‘g second. In the next experiment the distance was reduced to half, that is to say, 15 mm., and the total time was found to be 1 second. And lastly, the latent period was determined, under direct stimulation, at ‘o8 second (fig. 83). Thus

Fic. 83.—Determination of velocity by differential method. The records from below to above are in response to stimuli applied at distances of 30mm., 15 mm., and directly. Recorder 10 D.V. The three results thus obtained from independent data are here seen to be extremely consistent. They bear very emphatic testimony not only to the accuracy of the method but also to the constancy of the velocity in a given specimen under unvarying external conditions.

It has been stated that the accuracy of these time- measurements can be pushed to almost any extent. In order to demonstrate this fact, and also to exhibit the high mutual consistency of various determinations, I reproduce another set of records from a different specimen (fig. 84) from which the velocity of transmission is to be determined by the Differential Method. In this case a new recorder was taken, with a vibration-frequency of 20 times per second. Hence the distance between any two successive dots represents a time-interval of one-twentieth of a second. The stimulus intensity was again 3. The lowest record gives us the result obtained when the point of application of stimulus was 30 mm. away from the responding pulvinus. The total time T is here seen to be 2'9 seconds. The next record gives us the result when the point of stimulation was at a distance of 20 mm., and the total time T; is 1°985

Fic. 84.—Determination of velocity by differential method. Uniform stimuli applied at distances of 30 mm., 20 mm., and directly. Recorder second. The third and highest gives the record of direct stimulation, the latent period being shown as ‘085 second. Thus It is thus seen that, taking the precautions described, successive determinations of the velocity of transmission may be arrived at which are of great constancy. It may be said of the velocity of transmission in the petiole of Mimosa that it is constant with a given specimen, but undergoes some variation with different individuals. It is also subject to modifications induced by season. In winter the velocity is much reduced. The highest velocity which I have obtained with summer specimens of the petiole of Mimosa is 30 mm. per second. The lowest, in sluggish specimens, may be as little as 4 mm. per second.

The tabular statement below shows results obtained in twenty-five determinations in different specimens. Maximal stimulus was applied in every case. Having now obtained means for the accurate determina- tion of the velocity of transmission, we next proceed to study the effects of various agencies in inducing changes in the normal rate. And first we must consider the im- portant question of whether or not the velocity is in any degree dependent on the intensity of stimulus. In the corresponding case of conducting animal-nerve there is considerable diversity in the results which have been arrived at. It has been found by some investigators that velocity is independent of the intensity of stimulus. Others have found, on the contrary, that the velocity of trans- mission increases with the intensity of the stimulus, till with a very high intensity it becomes unmeasurable. The results which I shall here describe will probably throw light on this debatable question. It may be said, in anti- cipation, that the effects are to some extent modifiable in a definite way by the condition of the conducting-tissue.

If the specimen happens to be in a sluggish condition, then increasing intensity of stimulus will be found to be attended by increasing velocity of transmission. Again, a moderately strong intensity of stimulus is often found to leave, as an after-effect, increased conducting power. That is to say, to a tissue which has been sluggish, stimulation itself imparts a higher conductivity. In these facts there is a remarkable parallelism to what has already been pointed out in the matter of the amplitude of response of the sub- tonic tissue. In that case we saw that increasing intensity of stimulus gave rise to increased amplitude of response. We saw, further, that following a given stimulus, increased excitability appeared as an after-effect, so that the repeti- tion of an identical stimulus evoked response of enhanced amplitude.

Returning now to the experimental inquiry into the influence of intensity of stimulus on the velocity of trans- mission, I reproduce a set of records (fig. 85) obtained with a tissue which was slightly sluggish. The distance of the point of application of stimulus—namely, 20 mm.—was maintained constant, the intensity being varied in the successive experiments. The vibrating recorder had a frequency of 10 per second. The lowest record is the result of a stimulus-intensity of ‘5. The total time of transmission is seen to have been 2°1 seconds. The true time is obtained by subtracting from this the latent period, the average value of which is found to be about ‘1 second. No appreciable error will be introduced in practice by adopting this average value for the latent period, for its

Fic. 85.—Effect of intensity of stimulus and its after-effect on velocity. Lowest record under stimulus ‘5 ; the next under stimulus 4. Velocity increased under stronger stimulus. Enhancement of conductivity by previous stimulation seen in two upper records under °5 and 4 respec- tively. Velocity high and practically the same in both cases. variations are very slight, being of the order of hundredths of a second. The actual time here taken for transmission is thus 2 seconds, with a stimulus-intensity of ‘5.

The record next above gives the result when the stimulus intensity was 4, that is to say, increased to eight times its original value. The total time is now found to be decreased to 1°6 second, the true time after deducting the latent period being thus 1°5 second. The velocity under this increasing intensity is thus enhanced in the proportion of 2:1°5, or 33 per cent. To find out if there had been any after-effect of stimulation, a record was once more

taken with the original feeble stimulus-intensity of °5. It will be seen from the third record that the time taken for the transmission of excitation was practically the same as that with the previous strong stimulus, showing that this has made the tissue better conducting and that this property has reached a limit of uniformity. In order to test this conclusion further, a fourth record was taken with the second high stimulus-intensity of 4. It will be observed that the time of transmission is now the same as with the feebler intensity.

From these experiments it will be understood that when the tissue is in a somewhat sluggish or sub-tonic con- dition the velocity of transmission is enhanced under increasing intensity of stimulation. This, however, reaches a limit under a maximal stimulus the value of which is about 3 units. The following table gives the results of two sets of experiments on the effect of increased intensity of stimulus on velocity :— The fact that stimulus itself may enhance conductivity in a sub-tonic tissue can be seen in a striking manner in specimens of Mimosa which are in sluggish condition. It will there be found that the application of stimulus on the petiole will at first fail to be conducted. If, however, we apply the same stimulus again after the usual interval of, say, 15 minutes, the excitation which failed in the pre- vious case to be conducted will then reach the pulvinus

and induce the responsive fall. Stimulus had thus imparted conductivity to the tissue. We have seen that when the tissue is in a favourable tonic condition the range between minimal and maximal excitation tends to vanish—that is to say, a moderately feeble stimulus induces the same amplitude of response as the maximal. It appears probable that what was found to be true in the case of motile excitability may be equally applicable to conductivity ; that velocity of trans-

Fic. 86.—Effect of optimum condition on velocity. Plant raised to tem- perature 30°C.; records taken under stimulus -5 (lowest), 2 (highest), ‘5 once more (middle). Velocity practically the same. mission will tend to be constant, even under varying in- tensity of stimulus, when the tissue is in a favourable tonic condition. In order to test this induction I next tried the effect of varying intensity of stimulus on a specimen which had been brought to a favourable tonic condition. We have already noticed how the excitability of the plant is enhanced when the surrounding temperature is raised to 30° (C. OF thereabouts. This was secured by enclosing the experimental plant in a thermal chamber, which was maintained at the uniform temperature of 30° C. The point of stimulation

was at a distance of 30 mm. from the pulvinus, and the frequency of the vibrating-recorder employed was 10 times in a second. In the lowest of the three records shown in fig. 86 a stimulus-intensity of 5 was employed. The next record, the highest in the figure, was taken with stimulus 2, that is to say, four times the former intensity, and the time of transmission was found to be practically the same as with the feeble stimulus. The third, which is the intervening record, was taken when the stimulus had been restored to its original feeble intensity of 5. The results demonstrate that these successive experiments, with varying intensities of stimulus, gave practically uniform results in velocity of transmission of 20 mm. per second.

These experiments confirm the conclusion that when the plant is in an optimum condition its velocity of trans- mission is practically constant, even under varying intensity of stimulus. In ordinary circumstances the velocity in- creases with increasing intensity of stimulus, till a limit is reached under maximal stimulation. On employing the very strong stimulus-intensity of 15 or 20 units, I have sometimes observed a sudden enhance- ment of the normal velocity. In fact, the time elapsing in these circumstances between stimulus and response was tantamount to the duration of the latent period, the velocity of transmission being thus practically infinite. One thing that was noticeable in such experiments was that instead of gradual and continuous enhancement of the velocity, with increasing stimulus, the enhancement which occurred was sudden and abrupt at a certain high intensity. This justifies the conclusion that, in such a case, the stimulation becomes virtually direct by leakage of currents of relatively high tension.

It should be stated here that very strong stimulation has a tendency to induce fatigue, the result of which is seen in the reduction of the rate of transmission in subsequent experiments. The effect of fatigue can also be shown under moderate stimulation by reducing the period allowed for rest between two stimulations. In summer the period of complete recovery of conductivity is about 15 minutes. In winter the same process requires from 20 to 25 minutes. In fig. 87 a pair of records is given showing the reduction of the velocity under fatigue. The upper of the two records ~

Fic. 87.—Effect of fatigue. Upper record when plant fresh ; lower record when fatigued. was taken when the plant was fresh, the distance of the point of stimulation was Io mm., and the intensity of stimulus was 2. The next and lower of the two records was taken after allowing the incomplete resting-interval of only 10 minutes. It will be seen that fatigue has here prolonged the time taken for transmission of excitation. This prolongation is due chiefly to fatigue of conductivity and partly to the prolongation of the latent period. The variation of the latter factor, however, is relatively insigni- ficant, being only, as already stated, of the order of hundredths of a second. Taking the approximate value of

transmission of excitation through a distance of 10 mm. was here 1°32 second when the plant was fresh; when the specimen was fatigued the period of transmiiceie through the same distance was prolonged to 1°7 second. The effect of fatigue is also here depicted in an interesting manner in the records of the responsive movements them- selves. In the upper record of the fresh specimen the movement is seen to have been vigorous, by the comparative erectness of the curve and the distance between the succes- sive dots, representing the amount of the excitatory fall during periods of one-tenth of a second. The lower curve offers a marked contrast in both these respects.