Researches on Irritability of Plants/Chapter 3
CHAPTER III
METHODS OF STIMULATION
In the case of contractile animal muscle, various stimuli give rise to excitation, and it is a very remarkable fact that the same stimuli exercise a similar excitatory influence on the pulvinus of Mimosa. Classifying these stimuli, we find that they are:—
- 1. Mechanical.—A blow will excite animal muscle and cause mechanical response. A similar effect is induced by a mechanical blow in the pulvinus of Mimosa. A prick or cut also will cause contraction in either.
- 2. Chemical.—Various chemical agents are found to induce excitation in both animal and vegetal contractile tissues. Thus dilute hydrochloric acid or ammonia causes excitation of both muscle and pulvinus.
- 3. Thermal.—The application of a hot wire will induce responsive contraction in both cases.
- 4. Electrical.—The muscle may be excited by an induction-shock. The pulvinus of Mimosa is also excited by such shocks. Other modes of electrical stimulation, such as that of condenser-discharge and that of the application of a constant electrical current, are found effective in causing excitation of animal tissues. It will be seen in the course of the present chapter that plant tissues also may be excited by similar methods.
In all these cases excitation may be either direct or indirect. In the case of muscle, with its attached nerve, we may cause excitation directly by applying the various forms of stimulus on the muscle itself, or indirectly by applying them on the nerve. In the latter case excitation is transmitted by the conducting-tract—the nerve—and reaching the muscle after a brief and definite interval, induces there the usual contraction.
Taking the case of Mimosa, we may similarly have either direct or indirect excitation. Excitation is direct when it is applied, say, on the contractile pulvinus itself. It is indirect when it is applied on the petiole, at a distance from the pulvinus. Certain tissues in the petiole conduct this excitation, which, reaching the pulvinus after a definite interval, induces a responsive contraction.
It is usually maintained that in the case of Mimosa there is no true conduction of excitation, but that this contention is not justified will be fully demonstrated in a subsequent chapter. We have, then, in correspondence to the nerve and muscle preparations of the animal, plant-specimens, consisting of petiole and pulvinus. Indirect excitation for specific experiments is effected in the animal through the nerve, and in the plant through conducting-strands embedded in a tissue, as in the petiole.
Although we thus have various forms of stimulus at our disposal for inducing individual and isolated responsive contractions in Mimosa, yet we are confronted with very great difficulties when we wish to obtain a series of uniform excitations for quantitative investigation. It is obvious that chemical forms of stimulus would be impossible for successive excitations. The objection to a mechanical blow, as the stimulus to be employed, lies in its liability to cause a mechanical jar and thus to disturb the record.
The ideal form of stimulation would be one the intensity of which might be maintained uniform in successive experiments, or varied in a definite and known manner. Another great obstacle to be overcome in practice is the avoidance of injury which is caused by the stimulus itself. The application of stimulus above a critical intensity induces a depression or abolition of excitability of the tissue.
As the result of long investigation for the purpose of securing various forms of quantitative stimulus, I find that one mode of thermal and three modes of electrical stimulation may be rendered practicable for our purpose. These four different methods will be described in some detail below.
Electro-thermic Stimulation
The thermal mode of stimulation can, however, be rendered practicable by the electrical mode of the generation of heat. A loop of fine platinum-wire is made to clasp round the petiole which is to be excited, and is connected with an electrical circuit by means of fine flexible silver-wire (fig. 6). The circuit can be completed by a metronome interrupter, the current from the battery flowing for a definite length of time during, say, a single or definite number of beats of the metronome. This produces a sudden thermal shock, enough to cause excitation. Successive uniform stimuli can thus be applied. By means of a variable resistance included in the circuit, the intensity of the stimulus can be increased or diminished. Care must, however, be taken that the heat produced in the platinum loop shall not be such as to scorch or otherwise injure the tissue.
Another mode of stimulation—namely, that of thermal radiation—can also be employed, though not so conveniently as the former. A certain area may be rendered radiant by the passage of an electrical current. A Nernst electrical lamp can be conveniently utilised for the purpose. This, when rendered incandescent, gives out radiation of constant intensity. This radiation consists not only of light rays but also of a large proportion of obscure heat-rays. The excitatory value of the latter is more efficient than the luminous rays. The radiant surface of the Nernst lamp is suitably placed in front of a concave metal mirror, by means of which the rays can be focused upon any point that is desired.
Stimulation by Constant Current
I have found that Mimosa and other sensitive plants show certain very remarkable excitatory effects under the action of a constant current. The characteristic feature of these is that excitation is not induced during the passage Of the current but only at its initiation or cessation. The excitatory effect in this case is further conditioned by the point
Fig. 8.—Responses to stimulation by constant electric current.
of entry, or anode, and that of exit, or kathode. The specific characteristics of this mode of stimulation will be found fully described in the chapter on the Polar Effects of Currents in Excitation. It need only be mentioned here that, in the matter of all these peculiar effects, the plant tissue behaves in a manner exactly similar to the animal tissue. A series of records obtained from Mimosa by the stimulus of a constant current are shown in fig. 8.
Stimulation by Condenser Discharge
Another practical method of stimulation is that of condenser discharge. The condenser consists essentially of two conducting-plates—which may be two sheets of tin-foil—separated by a sheet of non-conducting material, such as mica or paraffined paper. The capacity of the condenser is increased by enlarging the effective area of the plates. The diagram in fig. 9 illustrates this mode of excitation. By increasing the number of cells, the charging e.m.f. may be increased until a suitable value is obtained which is efficient for excitation. This will depend on the excitability of the plant-specimen. About 2 volts charging ·5 microfarad will in general be found sufficient. K is a special spring-key by which the condenser may be charged
Fig. 9.—Direct stimulation by condenser discharge; C, condenser, K, key. (a) intra-electrodal and (b) indirect extra-electrodal mode of stimulation.
or discharged. The plant to be excited is included in the electrical circuit. When K is pressed down, the condenser is charged, the instantaneous charging current passing in one direction. The upper arrow in the diagram shows the direction of this charging current. When the key is released, it springs back and discharges the condenser. The instantaneous discharge current now flows in a reverse direction (fig. 9).
The electrical connections with the plant are diagramatically shown, in this and other figures, by two lines. In practice the connections have to be made by means of thread moistened in dilute saline solution. In certain experiments it is necessary to avoid complications arising from electrolytic polarisation. In these it is advisable to use non-polarisable electrodes for making the connections with the plant tissue. Such non-polarisable electrodes may be of the usual U-tube type. The shorter limb of the glass U-tube is filled with kaolin paste in normal saline. A cotton thread moistened in saline protrudes from this and makes the connections with the tissue. The longer limb is filled with zinc sulphate solution, into which dips a zinc rod. For ordinary purposes, however, a much simpler contrivance is found effective. A narrow cork is partially hollowed out and paraffined. The well thus formed is filled with dilute saline solution. The bottom is pierced for the entry of a cotton thread into the saline. A thick silver-wire, whose surface has been covered electrolytically with a film of chloride, pierces the side of the cork and dips into the saline solution. The silver wire forms one of the two electrodes, and the cotton thread makes the necessary electrical connection with the tissue.
For the purpose of excitation we may make the two electrical connections, one at or near the pulvinus itself, and the other on the petiole at a short distance. It will be shown later that when the electrical current leaves the tissue by the pulvinus, that point becomes the seat of excitation. Thus by making the pulvinus the point of exit of current, or kathode, we may cause direct excitation. Or we may have the pulvinus included between the two electrodes, so that the electrical current passes through it (fig. 9, a). This connection we may designate the intra-electrodal. Here, in certain circumstances, the excitation throughout the tract becomes diffuse and practically instantaneous. And lastly, the two electrical connections may be made side by side, say about 1 cm. apart on the petiole, at a moderate distance from the pulvinus. The excitation thus caused in the petiole reaches the pulvinus, as I have already said, by conduction. This connection we may call extra-electrodal (fig. 9, b).
I give below a series of records (fig. 10) of response to stimulation by condenser discharge. The plant was highly excitable, and excitation was caused by the discharge of ·1 microfarad condenser charged to ·3 volt.
Stimulation by Induction-shock
Excitation may be induced by means of a single or repeated shock from an induction coil. In my own experience I was at first under the impression that this mode of stimulation was not suitable for repeated quantitative experiments, as I found that the plant was liable to become insensitive owing to the fatigue or injury caused by the shock. Later, however, I was able to trace this difficulty to the
Fig. 10.—Record of responses to stimulation by condenser discharge.
employment of an intensity of shock which was in excess of a certain critical value. I had in fact been misled by the prevailing belief that the excitability of the plant was considerably lower than that of the animal. Hence I employed an intensity of current which was unnecessarily high. This induced fatigue and consequent insensitiveness. Afterwards I discovered that in so far as its sensitiveness to electrical stimulation was concerned, Mimosa was in no way inferior to the animal. Quantitative results will be given later, in justification of this statement. Avoiding, then, an intensity of stimulus which was too great, and allowing proper resting-intervals, I found that the efficiency of induction-shock as a mode of stimulation was all that could be desired. It has also the great advantage of allowing successive stimuli to be maintained constant, or to be increased in a known manner.
The induction coil consists of a primary made of a few turns of thick wire enclosing a bundle of soft iron wire—thus forming an electro-magnet—and of a secondary consisting of a larger number of turns of thin wire. The secondary coil can be made to approach or recede from the primary, by means of a slide. When a current is suddenly started in the primary coil, by pressing a key an instantaneous make-induction-current is induced in the secondary. When, by releasing the key, this current is broken, an instantaneous break-induction-current, whose direction is opposite to that of make, is induced in the secondary. Owing to the greater suddenness with which the break is effected, the intensity of the break-shock is greater and of shorter duration than that of the make-shock. The intensity of either make- or break-shock may be increased by bringing the secondary nearer the primary. We can obtain successive uniform shocks, of either make or break, by maintaining the distance between the two coils constant.
We may subject the tissue to shock of either make or break at will by employing an additional short-circuiting key. When this key is down, the shock from the secondary coil is practically diverted across the better conducting-path provided by the key, so that for practical purposes none passes through the plant tissue. If it is desired to cause successive excitations by make-shock only, then the short-circuit key is raised when the primary circuit is made, and pressed down when this is broken. For exciting by break-shock, the short-circuit key is pressed when the primary is made and raised when the primary is broken (fig. 11).
Effects of Make- and Break-shock
In the response of animal tissue it is well known that while single induction-shocks are effective in the case of quickly reacting skeletal muscles, they induce hardly any contractile effect in the more sluggish smooth muscles. Vegetal protoplasm also is commonly regarded as little capable of excitation by these shocks, behaving in this respect like the sluggish smooth muscles amongst animal tissues. Again, while the break-induction-shock of higher intensity and shorter duration is more effective in exciting the quickly reacting skeletal muscle, in the case of the sluggish smooth muscle it is the make-shock of low intensity and long duration that proves more efficacious. That
Fig. 11.—Arrangement for applying single make- or break-shock; K, key in the primary circuit. The secondary circuit may be short-circuited by the second key.
the inference commonly made about the reaction of vegetal protoplasm to single induction-shocks is not of universal application, is strikingly seen in the response of pulvinus of Mimosa. Here, so far at least as single induction-shocks are concerned, its reaction appears more analogous to that of skeletal than of smooth muscle; as a position of the secondary in relation to the primary can be found in which, while a single make-shock is ineffective, a single break-shock is quite efficient. In order to render the make-shock effective, the secondary has here to be pushed in nearer to the primary, thus increasing the intensity of the shock. A pair of records will be given in a later chapter (figs. 20, 21), showing the relative ineffectiveness of the make-shock compared with the break-shock.
Excitation by Tetanising Shocks
It will be shown later that shocks individually ineffective become effective by repetition. It is thus possible to excite a plant by subjecting it to a number of relatively feeble make- and break-shocks. The advantage of this mode of stimulation is that, owing to the low intensity of these shocks, the liability of the tissue to injury is very much reduced. Such alternating tetanising shocks can be produced by means of an automatic spring-interrupter, included in the primary circuits. This interrupter consists of a steel spring carrying at its free end a soft-iron armature which faces one pole of the electro-magnet of the primary. An adjustable contact-rod touches the spring and completes the primary circuit. But the completion of the circuit magnetises the electro-magnet, which, pulling the armature, breaks the contact, thereby interrupting the primary current. The electro-magnet is thus demagnetised, the armature is released, and the spring returns suddenly, re-establishing the circuit. By this automatic make-and-break we obtain alternating induction-currents in the secondary.
When we wish to subject the experimental tissue to the additive effects of these shocks, of a given short duration, this may be accomplished by including in the primary circuit a metronome, which in the course of a single beat closes the circuit for an approximately definite duration. If the duration of closure be, say, one-fifth of a second, and if the frequency of the spring-interrupter be 50 times per second, the number of alternating double-shocks given to the tissue would be ten.
A method of securing still greater accuracy in the duration of the tetanising shock will be described in another chapter, where also may be seen records obtained by that mode of excitation. In subsequent chapters we shall also study in detail the various characteristics of response and its time-relations.
Summary
The ideal method of stimulating a plant is one in which the intensity might be maintained uniform or varied in a definite and known manner.
If the stimulus exceeds a certain critical value, the tissue is injured with concomitant diminution or abolition of excitability.
One practical method of quantitative stimulation is by electro-thermic stimulus, where the plant-tissue is subjected to a sudden and definite thermal variation.
The plant may also be excited by the action of a constant current.
Another method of excitation is by the discharge of a condenser.
And lastly, excitation may be induced in the plant by a single induction-shock or by a series. As in the skeletal muscle of animals, so in the pulvinus of Mimosa, the break-shock is more effective than the make-shock.