#Fruit.#--Egg-plant (_Solanum Melongena_).

#Negative variation.#--Taking the leaf-stalk of turnip we kill an area on its surface, say B, by the application of a few drops of strong potash, the area at A being left uninjured. A current is now observed to flow, in the stalk, from the injured B to the uninjured A, as was found to be the case in the animal tissue. The potential difference depends on the condition of the plant, and the season in which it may have been gathered. In the experiment here described (fig. 6, _a_) its value was 13 volt.

[Ill.u.s.tration: FIG. 6.--(_a_) EXPERIMENT FOR EXHIBITING ELECTRIC RESPONSE IN PLANTS BY METHOD OF NEGATIVE VARIATION. (_b_) RESPONSES IN LEAF-STALK OF TURNIP TO STIMULI OF TWO SUCCESSIVE TAPS, THE SECOND BEING STRONGER.

A and B contacts are about 2 cm. apart, B being injured. Plant is stimulated by a tap between A and B. Stimulus acts on both A and B, but owing to injury of B, effect at A is stronger and a negative variation due to differential action occurs.]

A sharp tap was now given to the stalk, and a sudden diminution, or negative variation, of current occurred, the resting potential difference being decreased by 026 volt. A second and stronger tap produced a second response, causing a greater diminution of P.D. by 047 volt (fig. 6, _b_). The accompanying figure is a photographic record of another set of response-curves (fig. 7). The first three responses are for a given intensity of stimulus, and the next six in response to stimulus nearly twice as strong. It will be noticed that fatigue is exhibited in these responses. Other experiments will be described in the next chapter which show conclusively that the response was not due to any accidental circ.u.mstance but was a direct result of stimulation. But I shall first discuss the experimental arrangements and method of obtaining these graphic records.

[Ill.u.s.tration: FIG. 7.--RECORD OF RESPONSES IN PLANT (LEAF-STALK OF CAULIFLOWER) BY METHOD OF NEGATIVE VARIATION The first three records are for stimulus intensity 1; the next six are for intensity twice as strong; the successive responses exhibit fatigue. The vertical line to the left represents 1 volt. The record is to be read from right to left.]

#Response recorder.#--The galvanometer used is a sensitive dead-beat D'Arsonval. The period of complete swing of the coil under experimental conditions is about 11 seconds. A current of 10^{-9} ampere produces a deflection of 1 mm. at a distance of 1 metre. For a quick and accurate method of obtaining the records, I devised the following form of response recorder. The curves are obtained directly, by tracing the excursion of the galvanometer spot of light on a revolving drum (fig. 8). The drum, on which is wrapped the paper for receiving the record, is driven by clockwork. Different speeds of revolution can be given to it by adjustment of the clock-governor, or by changing the size of the driving-wheel. The galvanometer spot is thrown down on the drum by the inclined mirror M. The galvanometer deflection takes place at right angles to the motion of the paper. A stylographic pen attached to a carrier rests on the writing surface. The carrier slides over a rod parallel to the drum. As has been said before, the galvanometer deflection takes place parallel to the drum, and as long as the plant rests unstimulated, the pen, remaining coincident with the stationary galvanometer spot on the revolving paper, describes a straight line. If, on stimulation, we trace the resulting excursion of the spot of light, by moving the carrier which holds the pen, the rising portion of the response-curve will be obtained. The galvanometer spot will then return more or less gradually to its original position, and that part of the curve which is traced during the process const.i.tutes the recovery. The ordinate in these curves represents the E.M. variation, and the abscissa the time.

[Ill.u.s.tration: FIG. 8.--RESPONSE RECORDER]

We can calibrate the value of the deflection by applying a known E.M.F.

to the circuit from a compensator, and noting the deflection which results. The speed of the clock is previously adjusted so that the recording surface moves exactly through, say, one inch a minute. Of course this speed can be increased to suit the particular experiment, and in some it is as high as six inches a minute. In this simple manner very accurate records may be made. It has the additional advantage that one is able at once to see whether the specimen is suitable for the purpose of investigation. A large number of records might be taken by this means in a comparatively short time.

#Photographic recorder.#--Or the records may be made photographically. A clockwork arrangement moves a photographic plate at a known uniform rate, and a curve is traced on the plate by the moving spot of light.

All the records that will be given are accurate reproductions of those obtained by one of these two methods. Photographic records are reproduced in white against a black background.

#Compensator.#--As the responses are on _variation_ of current of injury, and as the current of injury may be strong, and throw the spot of light beyond the recording surface, a potentiometer balancing arrangement may be used (fig. 9), by which the P.D. due to injury is exactly compensated; E.M. variations produced by stimulus are then taken in the usual manner. This compensating arrangement is also helpful, as has been said before, for calibrating the E.M. value of the deflection.

[Ill.u.s.tration: FIG. 9.--THE COMPENSATOR A B is a stretched wire with added resistances R and R'. S is a storage cell. When the key K is turned to the right one scale division = 001 volt, when turned to the left one scale division = 01 volt. P is the plant.]

#Means of graduating the intensity of stimulus.#--One of the necessities in connection with quant.i.tative measurements is to be certain that the intensity of successive stimuli is (1) constant, or (2) capable of gradual increase by known amounts. No two taps given by the hand can be made exactly alike. I have therefore devised the two following methods of stimulation, which have been found to act satisfactorily.

[Ill.u.s.tration: FIG. 10.--THE SPRING-TAPPER]

#The spring-tapper.#--This consists (fig. 10) of the spring proper (S), the attached rod (R) carrying at its end the tapping-head (T). A projecting rod--the lifter (L)--pa.s.ses through S R. It is provided with a screw-thread, by means of which its length, projecting downwards, is regulated. This fact, as we shall see, is made to determine the height of the stroke. (C) is a cogwheel. As one of the spokes of the cogwheel is rotated past (L), the spring is lifted and released, and (T) delivers a sharp tap. The height of the lift, and therefore the intensity of the stroke, is measured by means of a graduated scale. We can increase the intensity of the stroke through a wide range (1) by increasing the projecting length of the lifter, and (2) by shortening the length of spring by a sliding catch. We may give isolated single taps or superpose a series in rapid succession according as the wheel is rotated slow or fast. The only disadvantage of the tapping method of stimulation is that in long-continued experiment the point struck is liable to be injured.

The vibrational mode of stimulation to be presently described labours under no such disadvantage.

#The electric tapper.#--Instead of the simple mechanical tapper, an electromagnetic tapper may be used.

[Ill.u.s.tration: FIG. 11.--THE TORSIONAL VIBRATOR Plant P is securely held by a vice V. The two ends are clamped by holders C C'. By means of handles H H', torsional vibration may be imparted to either the end A or end B of the plant. The end view (_b_) shows how the amplitude of vibration is predetermined by means of movable stops S S'.]

#Vibrational stimulus.#--I find that torsional vibration affords another very effective method of stimulation (fig. 11). The plant-stalk may be fixed in a vice (V), the free ends being held in tubes (C C'), provided with three clamping jaws. A rapid torsional vibration[9] may now be imparted to the stalk by means of the handle (H). The amplitude of vibration, which determines the intensity of stimulus, can be accurately measured by the graduated circle. The amplitude of vibration may be predetermined by means of the sliding stops (S S').

#Intensity of stimulus dependent on amplitude of vibration.#--I shall now describe an experiment which shows that torsional vibration is as effective as stimulation by taps, and that its stimulating intensity increases, length of stalk being constant, with amplitude of vibration.

It is of course obvious that if the length of the specimen be doubled, the vibration, in order to produce the same effect, must be through twice the angle. I took a leaf-stalk of turnip and fixed it in the torsional vibrator. I then took record of responses to two successive taps, the intensity of one being nearly double that of the other. Having done this, I applied to the same stalk two successive torsional vibrations of 45 and 67 respectively. These successive responses to taps and torsional vibrations are given in fig. 12, and from them it will be seen that these two modes of stimulation may be used indifferently, with equal effect. The vibrational method has the advantage over tapping, that, while with the latter the stimulus is somewhat localised, with vibration the tissue subjected to stimulus is uniformly stimulated throughout its length.

[Ill.u.s.tration: FIG. 12.--RESPONSE IN PLANT TO MECHANICAL TAP OR VIBRATION The end B is injured. A tap was given between A and B and this gave the response-curve _a_. A stronger tap gave the response _b_. By means of the handle H, a torsional vibration of 45 was now imparted, this gave the response _c_. Vibration through 67 gave _d_.]

#Effectiveness of stimulus dependent on rapidity also.# In order that successive stimuli may be equally effective another point has to be borne in mind. In all cases of stimulation of living tissue it is found that the effectiveness of a stimulus to arouse response depends on the rapidity of the onset of the disturbance. It is thus found that the stimulus of the 'break' induction shock, on a muscle for example, is more effective, by reason of its greater rapidity, than the 'make'

shock. So also with the torsional vibrations of plants, I find response depending on the quickness with which the vibration is effected. I give below records of successive stimuli, given by vibrations through the same amplitude, but delivered with increasing rapidity (fig. 13).

[Ill.u.s.tration: FIG. 13.--INFLUENCE OF SUDDENNESS ON THE EFFICIENCY OF STIMULUS The curves _a_, _b_, _c_, _d_, are responses to vibrations of the same amplitude, 30. In _a_ the vibration was very slow; in _b_ it was less slow; it was rapid in _c_, and very rapid in _d_.]

Thus if we wish to maintain the effective intensity of stimulus constant we must meet two conditions: (1) The amplitude of vibration must be kept the same. This is done by means of the graduated circle. (2) The vibration period must be kept the same. With a little practice, this requirement is easily fulfilled.

The uniformity of stimulation which is thus attained solves the great difficulty of obtaining reliable quant.i.tative values, by whose means alone can rigorous demonstration of the phenomena we are studying become possible.

FOOTNOTES:

[8] A preliminary account of Electric Response in Plants was given at the end of my paper on 'Electric Response of Inorganic Substances' read before the Royal Society on June 6, 1901; also at the Friday Evening Discourse, Royal Inst.i.tution, May 10, 1901. A more complete account is given in my paper on 'Electric Response in Ordinary Plants under Mechanical Stimulus' read before the Linnean Society March 20, 1902.

I thank the Royal Society and the Linnean Society for permission to reproduce some of my diagrams published in their _Proceedings_.--J. C. B.

[9] By this is meant a rapid to-and-fro or complete vibration. In order that successive responses should be uniform it is essential that there should be no resultant twist, i.e. the plant at the end of vibration should be in exactly the same condition as at the beginning.

CHAPTER IV

ELECTRIC RESPONSE IN PLANTS--BLOCK METHOD

Method of block--Advantages of block method--Plant response a physiological phenomenon--Abolition of response by anaesthetics and poisons--Abolition of response when plant is killed by hot water.

I shall now proceed to describe another and independent method which I devised for obtaining plant response. It has the advantage of offering us a complementary means of verifying the results found by the method of negative variation. As it is also, in itself, for reasons which will be shown later, a more perfect mode of inquiry, it enables us to investigate problems which would otherwise have been difficult to attempt.

When electrolytic contacts are made on the uninjured surfaces of the stalk at A and B, the two points, being practically similar in every way, are iso-electric, and little or no current will flow in the galvanometer. If now the whole stalk be uniformly stimulated, and if both ends A and B be equally excited at the same moment, it is clear that there will still be no responsive current, owing to balancing action at the two ends. This difficulty as regards the obtaining of response was overcome in the method of negative variation, where the excitability of one end was depressed by chemical reagents or injury, or abolished by excessive temperature. On stimulating the stalk there was produced a greater excitation at A than at B, and a current of action was then observed to flow in the stalk from the more excited A to the less excited B (fig. 6).

But we can cause this differential action to become evident by another means. For example, if we produce a block, by clamping at C between A and B (fig. 14, _a_), so that the disturbance made at A by tapping or vibration is prevented from reaching B, we shall then have A thrown into a relatively greater excitatory condition than B. It will now be found that a current of action flows in the stalk from A to B, that is to say, from the excited to the less excited. When the B end is stimulated, there will be a reverse current (fig. 14, _b_).

[Ill.u.s.tration: FIG. 14.--THE METHOD OF BLOCK (_a_) The plant is clamped at C, between A and B.

(_b_) Responses obtained by alternately stimulating the two ends.

Stimulation of A produces upward response; of B gives downward response.]

We have in this method a great advantage over that of negative variation, for we can always verify any set of results by making corroborative reversal experiments.

By the method of injury again, one end is made initially abnormal, i.e.

different from the condition which it maintains when intact. Further, inevitable changes will proceed unequally at the injured and uninjured ends, and the conditions of the experiment may thus undergo unknown variations. But by the block method which has just been described, there is no injury, the plant is normal throughout, and any physiological change (which in plants will be exceedingly small during the time of the experiment) will affect it as a whole.

[Ill.u.s.tration: FIG. 15.--RESPONSE IN PLANT (FROM THE STIMULATED A TO UNSTIMULATED B) COMPLETELY IMMERSED UNDER WATER The leaf-stalk is clamped securely in the middle with the cork C, inside the tube T, which is filled with water, the plant being completely immersed. Moistened threads in connection with the two non-polarisable electrodes are led to the side tubes _t t'_. One end of the stalk is held in ebonite forceps and vibrated. A current of response is found to flow in the stalk from the excited A to the unexcited B, and outside, through the liquid, from B to A. A portion of this current, flowing through the side tubes _t t'_, produces deflection in the galvanometer.]

#Plant response a physiological or vital response.#--I now proceed to a demonstration of the fact that whatever be the mechanism by which they are brought about, these plant responses are physiological in their character. As the investigations described in the next few chapters will show, they furnish an accurate index of physiological activity. For it will be found that, other things being equal, whatever tends to exalt or depress the vitality of the plant tends also to increase or diminish its electric response. These E.M. effects are well marked, and attain considerable value, rising sometimes, as has been said before, to as much as 1 volt or more. They are proportional to the intensity of stimulus.

It need hardly be added that special precautions are taken to avoid shifting of contacts. Variation of contact, however, could not in any case account for repeated transient responses to repeated stimuli, when contact is made on iso-electric surfaces. Nor could it in any way explain the reversible nature of these responses, when A and B are stimulated alternately. These responses are obtained in the plants even when completely immersed in water, as in the experimental arrangement (fig. 15). It will be seen that in this case, where there could be no possibility of shifting of contact, or variation of surface, there is still the usual current of response.

I shall describe here a few crucial experiments only, in proof of the physiological character of electric response. The test applied by physiologists, in order to discriminate as to the physiological nature of response, consists in finding out whether the response is diminished or abolished by the action of anaesthetics, poisons, and excessively high temperature, which are known to depress or destroy vitality.

I shall therefore apply these same tests to plant responses.

#Effect of anaesthetics and poisons.#--Ordinary anaesthetics, like chloroform, and poisons, like mercuric chloride, are known to produce a profound depression or abolish all signs of response in the living tissue. For the purpose of experiment, I took two groups of stalks, with leaves attached, exactly similar to each other in every respect. In order that the leaf-stalks might absorb chloroform I dipped their cut ends in chloroform-water, a certain amount of which they absorbed, the process being helped by the transpiration from the leaves. The second group of stalks was placed simply in water, in order to serve for control experiment. The narcotic action of chloroform, finally culminating in death, soon became visually evident. The leaves began to droop, a peculiar death-discolouration began to spread from the mid rib along the venation of the leaves. Another peculiarity was also observed.

The aphides feeding on the leaves died even before the appearance of the discoloured patches, whereas on the leaves of the stalks placed in water these little creatures maintained their accustomed activity, nor did any discolouration occur. In order to study the effect of poison, another set was placed in water containing a small quant.i.ty of mercuric chloride. The leaves here underwent the same change of appearance, and the aphides met with the same untimely fate, as in the case of those subjected to the action of chloroform. There was hardly any visible change in the appearance of the stalks themselves, which were to all outer seeming as living as ever, indications of death being apparent only on the leaf surfaces. I give below the results of several sets of experiments, from which it would appear that whereas there was strong normal response in the group of stalks kept in water, there was practically a total abolition of all response in those anaesthetised or poisoned.

#Experiments on the effect of anaesthetics and poisons.# A batch of ten leaf-stalks of plane-tree was placed with the cut ends in water, and leaves in air; an equal number was immersed in chloroform-water; a third batch was placed in 5 per cent. solution of mercuric chloride.