Researches on Irritability of Plants

Effect of intensity of stimulus on velocity and after-effect Effect of cold in the retardation and arrest of transmission . Records of transmitted excitation with the block off ma on . Positive response followed by negative in Biophytum mndee indirect thermal stimulus : : c ; Diphasic response in Biophytum under indirect chemical stimulation Diphasic response in Mimosa due to indirect electric stimu- lation of petiole . : : : - : - Diphasic response in Mimosa due to indirect thermal stimu- lation of stem : - >

Effect of intensity of stimulus in saat fe diphasic response Effect of diminishing distance in transforming positive into diphasic response in Biophytum : : Records of responses to ascending and descending current in Mimosa . : . Records of responses to asdeidine aad Hesvending induction- shock Excitation induced in Biopistum by ttie meee of kattieds snd break of anode . : : : . Effect of anode, opposing systolic contraction in Desmodium . Effect of kathode, opposing diastolic expansion in Desmodium

Arrest at systole by make of kathode, and avastane expansion at break of kathode : : > Arrest at diastole by make of anode, sid ee contraction at break of anode - 5 : Effects of descending and ascending currents at make ae break on nerve-muscle and petiole-pulvinus Record showing abolition of polar excitation at high tet perature : - : - : . Transformation of Type II. to Type I. as atin ett of previous stimulation Polar reactions, Km, Km Am, and Km Am Ab under eradidtin increasing current 5

Multiple response in Averrhoa under a single strong electric shock : : Continuous record of pulsations of Desmodium leaflet ‘on four hours : - - - : : : Record of a single pulsation of Desmodium giving time- réintaaae Effect of application of increased internal hydrostatic pressure on the pulsation of Desmodium : Effect of ligature in inducing arrest of pulsation of Desmodium Response of Desmodium leaflet, originally in a state of and still After-effect of stimulation on pulsation of Desmodium in sak tonic condition see : 5 5 : :

Effect of depletion of energy on pulsation of isolated Desmodium : : Effect of stimulus in renewing pulsation of Desmodium, brought to standstill . 162,163. Extra pulsation induced by induction-shock applied at diastolic phase : é ; Inhibitory effect of transmitted excitation on the paisnten of vigorous leaflet of Desmodium 2 5 Augmentation of pulsation induced by eeaneniitted excitation in less vigorous specimen of Desmodium : Effect of lowering of temperature on the pulsation of heart Effect of lowering of temperature on pulsation of Desmodium

Effect of rise of temperature on amplitude and frequency of pulsation of the heart of frog 171,172. Effect of rise of temperature on the pulsation of Desnaniuet Effect of continuous rise of temperature from 30° C. to 38°5° C. Effect of continuous rise from 30° C. to 42° C. and return to Effect of potassium cyanide - Arrest of pulsation of the heart of frog ai aici Ee the action of dilute lactic acid - : Arrest of pulsation of Desmodium at eiastel by an

of dilute lactic acid : 3 Arrest of pulsation of heart at systole, me the action of dilute sodium hydrate . Arrest of pulsation of etenbdiaen at apstole = the applicaliam of dilute solution of sodium hydrate Action of environment on plant—Revelation of internal condition by character of response—Problems to be solved—Electrical response— Mechanical response—Motile organ in Mimosa pudica—Response in plant and animal—Different phases of the responsive move- ment—Graphic record—Determination of absolute movement of leaf and its time-relations — Characteristic effects of different agencies on the response-curve — Specific difficulties in recording plant-response.

In strong contrast to the energetic animal, with its various reflex movements and pulsating organs, stands the plant in its apparent placidity and immobility. Yet that same environment, which with its changing influences so strikingly affects the animal, is playing upon it also. Storm and sunshine, the warmth of summer and the frost of winter, drought and rain, all these and many more come and go about it. What coercion do they exercise upon it ? What subtle impress do they leave behind ? That they, in their totality, do leave the plant better or worse for their occur- rence, we know. It is evident that internal changes are effected by their agency which are entirely beyond our visual scrutiny. Would it be possible to trace this general action of the environment into some detail, and then follow out the question of its particular effects upon the vegetal

whether a given influence has contributed to the plant’s well-being or the reverse, whether it has left it more or less excitable, whether it has rendered it more or less energetic ? It is conceivable that internal changes which eluded our direct vision might nevertheless be brought within the range of our observation if we could obtain any sort of answer from the plant itself to a questioning shock. In such a case, the feebleness or vigour of the reply would in itself doubtless constitute a measure of the vitality of the organism. It appears obvious that if any given influence had rendered the plant more excitable, this fact would be mani- fested by the greater intensity of its response. In a very excitable condition we may suppose the slightest shock of stimulus would evoke a very large responsive expression ; in a state of depression, on the other hand, the strongest stimulus would induce only a feeble reply. The relation between the stimulus and the response would thus form a gauge of the physiological condition of the organism. The invisible fluctuating changes taking place in the plant, under the changing conditions of the environment, might in this way be made to reveal themselves.

All this presupposes, however, that the plant will answer in some tangible way to the impinging testing stimulus, and that it may be possible to obtain some record of this answer. The possibility of this will be further discussed presently. There are many important problems which wait for their solution till some such means of inquiry is found. What, for instance, are the various forms of stimulus which evoke an answering reaction in the plant? Again, has a given plant-tissue, like animal muscle, any definite percep- tion-period capable of exact measurement ? Is the respond- ing tissue susceptible of fatigue? Is the intensity of its answer dependent on the intensity of the blow? Is the excitation that may be caused at one point transmissible to a distance, as along animal nerve ? Is such transmission, supposing it to occur, fundamentally of the same nature

tissue that might twitch persistently, like the cardiac tissue of the animal? Ifso, are these rhythmic pulsations charac- teristically similar? Is there, again, any general resem- blance between responsive actions in plant and animal? Going deeper, since the same protoplasmic basis underlies them both, are these reactions to be regarded as essentially the same, though different in degree? If this last were true, then since the simpler explains the more complex, might not the physiological reactions of the plant be expected to elucidate many of the obscurities in the similar reactions of animal tissues ?

We return, then, to the question, Is the plant capable of furnishing any such responsive indications as we have supposed ? I have shown elsewhere that all plants give response to impinging stimulus by a definite electrical change,’ which can be recorded by means of suitable apparatus. For the purpose of the present work, however, it will be convenient to employ the more conspicuous motile indications afforded by certain plants, pre-eminent amongst which is Mimosa pudica.

The most prominent motile organ in Mimosa consists of a mass of tissue known as the pulvinus, at the joint or arti- culation of the primary leaf-stalk. The swollen mass on the lower side of this organ is very conspicuous. Under excita- tion the parenchyma, in this more effective lower half, undergoes ‘contraction,’ in consequence of which there is a fall of the leaf. This sudden movement constitutes the mechanical response of the leaf to the impinging stimulus, just as the contractile movement of a muscle in similar circumstances forms its characteristic mechanical response.

Digressing for a moment to consider the phenomenon of excitatory contractions in general, it may be said that our present knowledge is not complete as to the minutia 1 Bose: Friday Evening Discourse—Royal Institution, May 1901; of the process by which these are brought about. In muscle it is supposed that during the act of contraction there is a transfer and redistribution of fluid material.! In the case of Mimosa there is known to be an escape of fluid from the excited cells; there is a diminution of turgor. It is sup- posed that this may in some unknown way be connected with a diminution of pressure within the cell.?

In the case of the stamens of Cyneree, Pfeffer 3 observed a contraction under excitation of as much as 30 per cent. of the original length. There is an escape of water from the cells into intercellular spaces. The mode in which the fall of turgor takes place is uncertain, and various supposi- tions have been made to account forit. It has been thought that the escape of fluid is brought about by the elastic cell wall which forces liquid out of the cell, when the protoplasm lining it has become permeable under excitation. There may in addition be an active contraction of protoplasm which might force the liquid out of the cell-vacuole. This latter supposition is regarded by many as improbable, though the observations of Schutt and Benecke indicate that under stimulation the protoplasm of a diatom contracts away from the cell wall. Similar withdrawal of protoplasm

1 “Schafer, working on the highly differentiated wing-muscle of the wasp, concludes that each sarcomere contains a darker substance near the centre, divided into two parts by Hensen’s disc. At each end of the sarcomere the contents are clear and hyaline. In the act of contraction the clear material flows, according to Schafer, into tubular pores, in the central dark material.’—Starling: Elements of Human Physiology, 8th edition, p. 91. 2 “When the pressure in the cell decreases, we naturally assume this to be due to decreasing osmotic pressure, a decrease which may well amount to 2} to 5 atmospheres, and may be due either to the transformation of osmotically active substances into bodies with larger molecules, or to alterations in the permeability of the plasma, and an excretion of materials from the cell. As evidence of excretion of material we may quote the fact that Pfeffer observed crystals of unknown nature appearing on evaporation of the liquid expressed from the intercellular spaces. Still there are several reasons for doubting this conclusion. It is a remarkable fact that plasmo- lytic research affords no evidence of any decrease in osmotic pressure,’’— Jost: Plant Physiology, English edition, 1907, p. 515.

5 Cf. Pfeffer: Physiology of Plants, vol. ili., English edition, p. 75. has been observed in Spirogyra by Nageli and in Nitella by Hofmeister. Whatever theory may be held, the undoubted fact in these two cases, of plant and animal alike, is the occurrence of afundamental excitatory protoplasmic change which finds external expression in alteration of form. If we now record the responsive movement, we shall be recording what is an effect of excitatory change, either in plant or animal. Whether or not this fundamental change is similar in the two cases can only be decided by comparing the records due to excitation, in plant and animal tissues, under all possible variations of external conditions.

The fall of the leaf of Mimosa is brought about in consequence of the contraction of cells in the lower half of the pulvinus. I shall for convenience describe the fall as the contractile movement, in contradistinction to the erectile movement brought about by the recovery of cells into normal turgid and expanded condition. In studying the excitatory reactions of the plant, under external stimulus, we have to determine, first, what time elapses between the incidence of the shock and the initia- tion of a perceptive responsive movement. This constitutes the determination of the Latent Period. We have next to find out at what rate this responsive movement of the leaf takes place, and after what time the contractile phase of the movement is exhausted. After a short pause the plant gradually recovers from the effect of the shock, and the leaf is re-erected to its former position. We there- fore want to know the various rates at which recovery gradually takes place. In order to secure these data, it will be necessary to make a graphic record of the entire responsive movement of the plant organ. This record, further, must furnish us not only with the amount, but also with the time-relations, of this movement. This would involve the construction of a writing-lever which, deflected by the pull of the falling leaf, would be capable of tracing on a writing-surface, moving at a known uniform rate. the

concomitant curve. For this there must be an axis, sup- ported on frictionless jewelled bearings, and carrying two arms of a horizontal lever and a thin vertical wire with a bent tip, to serve as the writer. The different parts, as far as possible, should be made of aluminium, to secure the utmost lightness. A point of the petiole of the responding leaf would be attached by a silk thread to one arm of the lever, the other having on it a small weight, to act as counter- poise. On the fall of the leaf, under ex- citation, it would pull down with it the attached arm of the lever. The vertical writer would then also move, say, to the left. If the finely pointed bent end of the writer were to press lightly against the smoked surface of a glass plate, which was allowed to fall, at a uniform rate, by means of clockwork, a curve would then be traced which would not only record the responsive movement and recovery but also give their time-relations (fig. 1). To obtain the latter, it would be necessary to know the rate of movement of the plate on which successive vertical lines might be traced by a time-marker at Fic. 1—Diagrammatic intervals of, say, one minute. ee ee RE In order to find out the absolute movement of the leaf we must know the degree of magnification or reduction that has been effected by the recording arrangement. This will depend upon the relative lengths of the writer and the lever, and the distance of the point of attachment on the leaf from the pulvinus. When the lengths of the lever-arm and the writer are equal, then the writer will describe a movement which is equal to that of the point of leaf-attachment. By shortening the arm of the lever to half the length of the writer, we should obtain the magnification of two. This

shortening might be accomplished by attaching the thread nearer to the fulcrum. Proceeding in the manner indicated, any magnification, however high, can be obtained. Reduction, again, may be effected with equal ease. When the point of attachment is exactly midway between the pulvinus and the tip of the leaf, then the movement executed by it will be half that described by the extremity of the leaf. By bringing the point of attachment nearer Fic. 2.—Response-curve of primary leaf of Mimosa; the vertical lines below the record indicate intervals of one minute each.

to the pulvinus, we can obtain whatever reduction may be required. The resultant magnification or reduction of the record will thus depend, in any given case, on two factors—namely, the relation between the length of the writer and the length of the lever-arm, on the one hand; and, on the other, the relation between the distance of the point of attachment from the pulvinus and the entire length of the leaf. Thus if the lever produce a magnification of four, and the point of attachment cause a reduction to half, the resulting magnification will be 4 x $=2. As the movement of the primary petiole of Mimosa is considerable, the records taken are normally either equal or reduced to two-thirds. A record taken in this manner is given in fig. 2. The height

of this curve gives the amplitude of the movement, and the horizontal distance measures the corresponding time. The up-line here, a 0, indicates the responsive fall, and the descending line, 0 c, the gradual erection, due to recovery. The responsive movement was initiated within an exceedingly short time after the application of stimulus—in this case an electrical shock—and the fall was completed also within a relatively short period. In a record which will be given later, taken on a faster-moving plate, these characteristics will be seen better. The recovery, however, is a slow process, the earlier part being comparatively quick and becoming slower towards the end. The entire recovery is here seen to require I2 minutes.

If we were able to apply a stimulus of exactly identical intensity at regular and suitable intervals, and if the physio- logical condition of the responding tissue remained constant, then we should obtain a series of responsive twitches which would be practically identical. But if the physiological condition were to undergo any change, under environmental conditions, then the record would give us indications of that internal change, otherwise entirely beyond our power of scrutiny.. Thus if the plant were to become depressed, the amplitude of the pulse would undergo a diminution. If on the other hand its excitability should be enhanced, that fact would be indicated by an increase in the amplitude of the response.

The mere amplitude of the twitch, however, affords only a broad indication of the physiological condition of the tissue. There are many factors the effects of which find expression in subtler changes of the response-curve. One agency, for instance, will make the plant more alert. This is at once reflected at that part of the curve which corre- sponds to the Latent Period. This becomes shorter. The ascent of the curve will also be more abrupt. Another agency will induce, let us say, a contrary change. Different agencies, similarly, will bring about definite changes in the contracting and relaxing portions of the curve. The

varying effects of freshness and fatigue, of stimulating or depressing drugs, of heat or cold, of the environment, are in this way clearly revealed by the characteristic flexures of the curve. Thus, by means of testing-blows, we are able to make the plant itself describe those obscure internal changes which would otherwise have entirely escaped us. In fact, the phytographic records would, in the case of plants, supply us with all the information that myo- grams afford in the case of animal tissues. The experi- mental difficulties which the plant offers are, however, very great. In the case of muscle-contraction, the pull exerted is considerable and the friction offered by the recording-surface constitutes no essential difficulty, though even here the time-relations of the curve are, I have reason. to think, rendered somewhat unreliable by this friction. In the case of plants the contractile movement is relatively feeble, and in the movement of the leaflet of Desmodium, for instance, a weight so small as four-hundredths of a gram is enough to arrest the pulsating leaflets. When employing the very lightest lever, the extremely minute friction offered by the smoked-glass surface of the recording-plate is sufficient in this case to cause complete cessation of the record. Even in the leaf of Mimosa, the friction offered is enough to distort the curve to such a serious extent that errors are introduced into the amplitude and time-relations amounting to more than 50 per cent. These difficulties have been overcome by the successful devising of my Resonant Recorder, an account of which will form the subject of the next chapter.

Obscure modifications of internal condition of plant, resulting from changing factors of environment, may be revealed by records of plant’s response to testing-blows. intensity of reply measures the physiological efficiency of the plant tissue for the time being. Automatic records of mechanical and electrical responses of plant can be obtained with suitable apparatus. For recording the mechanical responses of Mimosa and other sensitive plants, a writing-lever is employed. Distor- tion of record by friction is apt to introduce error, which has to be eliminated.

Advantages of intermittent contact in record—Two types of apparatus : the Oscillating Recorder and the Resonant Recorder—Coercer and Vibrator—Perfect tuning—Recorders with standardised frequencies— Slide and clockwork—The record its own chronogram—Smoked surface and its fixation—Adjustments of the writer—Records with continuous and intermittent contacts. Ir was stated in the last chapter that however light the contact may be, and however smooth the glass recording- surface, the record was still apt to be either arrested, or seriously distorted, on account of friction. As long as I employed the ordinary method of continuous contact of the writing-point with the glass surface, it was impossible to overcome this particular difficulty. It occurred to me at last that the problem might find a solution if I could succeed in making an intermittent instead of a continuous writing- contact. I have solved this problem by devising two different types of apparatus, which I have called respectively the Oscillating Recorder and the Resonant Recorder. In the former, the Trecording-surface itself is made by an electro-magnetic device, to vibrate to and fro, thus bring- ing it into periodic contact with the writing-point.1. This apparatus is extremely convenient for the general pur- pose of recording responses in which the measurement of excessively short intervals of time is not essential.

But there are many important problems, such as the determination of the latent period, and accurate determina- tion of the velocity of transmission of excitation, in which we require time-measurements of the order of one-hundredth of a second. It will be shown that these determinations can be carried out with great precision, by means of the intermittent dots themselves, when the periodicity of their recurrence is rendered perfectly constant. For such pure poses, then, we require a frequency of intermittent contacts amounting to something like a hundred times per second.

It was clearly impossible to make the heavy plate- carrier oscillate with such a high frequency. There remained only the theoretical alternative of causing the writing- point to vibrate to and fro, at the required frequency, so as to make the necessary intermittent contacts with the surface of the recording-plate. The advantage of this intermittence may be understood from a concrete example. It will be remembered that the writing-point, under the action of the responsive fall, moves parallel to the surface of the recording-plate. If now, by means of some mechanism, the writing-point be made to vibrate to and fro, say, ten times each second, at right angles to the plate, this will in no way affect the record beyond the fact that instead of a continuous line a dotted line will be traced. The record will not now labour under the defects inseparable from the friction of continuous contact. Instead of this, we shall have the vibrating writer tapping a record which is practically free from friction. For it will be understood that, as in our concrete example, a recording-point which is vibrating ten times each second will execute one entire to-and-fro movement in one-tenth of a second. The duration of contact, at the extreme forward end of the swing, will represent only a small fraction, say one-fifth, of the entire period of one vibration. Hence after each contact, lasting only one-fiftieth of a second, the recording-point is absolutely free to take up the movement impressed upon it by the moving leaf. In a record lasting for one second the sum of the intermittent contacts will then amount to one-fifth, and the period of entire freedom to four-fifths of a second. We can thus see the theoretical

advantages of an intermittent over a continuous contact. What has been said of the writer vibrating ten times in a second holds good equally in those cases where the vibration-frequency is much higher. One great difficulty we encounter in carrying out this idea lies in giving the recording-point an impulse exactly perpendicular to the direction of its recording movement. If the recording-writer be made of fine steel- wire, and if we place behind it a small electro-magnet— the pole consisting of a rectangular piece of soft iron, at right angles to the wire—then, by sending a momentary strong current through this electro-magnet, a pull will be exerted on the wire which will make its recording-tip strike for an instant against the glass recording-surface. As the steel wire has to be made extremely fine, in order to reduce to a minimum the inertia of the recorder, the resulting pull exerted on it is very slight, unless an exces- sively strong current be sent round the electro-magnet. Again, unless the intermittent closures of the electric circuit be properly timed, the writing-index may be subjected to attraction in the course of its journey, now to the recording- surface and again away from it. In the latter of these cases its vibration, on which the intermittent contact depends, is totally destroyed.