Researches on Irritability of Plants

It has thus been shown that when a feeble or moderate stimulus is applied at a relatively great distance there occurs only the positive response. As the distance is reduced the antagonistic negative response makes its appearance, there being at first a considerable time-interval between the posi- tive and the negative. On reducing the distance still further the interval becomes reduced. The positive will then, as we shall see later, become masked by the predominant negative.

I shall now pass under review the various characteristics which distinguish the positive from the negative impulse. (1) The positive impulse travels much faster than the negative. (2) The conditions for the exhibition of the positive impulse are that the stimulus should be moderate or feeble and applied at a distance. A semi-conducting tissue again is more suitable for its exhibition than a_ highly conducting tissue. (3) While an increase of intensity of stimulus gives rise to enhanced velocity of negative impulse, it- produces little or no change in the velocity of positive impulse. Thus Table IV shows that while raising the intensity of stimulus from 5 to 8 units shortened the transmission-

period of the negative from 9°4 seconds to 4°6 seconds, it induced no variation in the transmission-period of the positive impulse which remained unchanged at *6 second. (4) When the distance to be traversed is reduced say to half, the period of transmission of the negative impulse is also reduced to half. But with the positive impulse the diminution is very slight. Thus in fig. 103 we find that with a distance of 20 mm. the period for the propa- gation of the positive impulse was I second; when the distance was reduced to half, the transmission-period was reduced not to half a second but to eight-tenths of a second.

It is difficult from these data to arrive at any definite conclusion as regards the nature of the positive impulse. Is this impulse physical or physiological? If the former, could it be hydro-mechanical? In consequence of a feeble stimulus applied at a distance, we have at the responding pulvinus, a positive turgidity variation, expan- . sion and erection of the leaf, evidently brought about by the forcing in of water. This presupposes a forcing out of water somewhere else, probably at the point of application of stimulus. It may be supposed that an active contraction occurred in the plant-cells under excitation, in consequence of which the sap was forced out giving rise to a hydraulic wave. On this supposition the positive impulse is to be regarded as hydro-mechanical. There is, however, no definite experimental proof in support of this theory. I shall presently describe the experiment I devised to settle the question. Unfortunately the results obtained were not as decisive as I hoped they would be.

It should be clearly understood that even if the positive impulse were found to be hydro-mechanical, it has nothing whatever to do with the propagation of the excitatory or negative impulse. For if the negative response took place in consequence of the impact of the positive impulse, then the negative should take place at a definite interval after the positive, the interval being equal to the latent period of the responding pulvinus. I have shown that the average latent period of the primary pulvinus of Mimosa is ‘1 second. In unfavourable circumstances even, I have never found it to exceed *3 second. The latent period of the leaflet of Biophytum or Averrhoa is about ‘4 second. The latent period of the responding organs of the various sensitive plants mentioned may therefore be taken as less than 1 second. If between the positive and negative response there is any relation of cause and effect, then the negative should occur within a second of the positive. But this is by no means the case. Referring to fig. ror we find that the positive impulse reached the pulvinus 4 seconds after the application of the stimulus, and instead of the excitatory response taking place within a second, we find that it did not occur till 37 seconds after! I give below a table which shows the very lengthened periods which are often found to elapse between the arrival of the positive impulse and the initiation of the excitatory response :—

Transmission- Transmission- ; 7 : i Difference Specimen. period of period of of sane + impulse. —impulse. ‘ seconds seconds seconds Mimosa ie 4 41 37 Biophytum .. 2 29 27 Averrhoa v3 22 65 43 The results just described, together with the fact that we may have a positive response without the subsequent negative (cf. lower record fig. 103), show conclusively that the excitatory response is independent of the positive impulse. The physiological character of the negative impulse has, moreover, been fully demonstrated by various crucial experiments described in the last chapter.

For deciding the question as to the physical or physio- logical character of the positive impulse, I employed the test of local application of cold on the velocity of trans- mission. It was found that while moderate application of cold delayed the transmission of the negative impulse, it had little effect in retarding the transmission of the positive. Further test by the physiological block induced by excessive cold was, however, less satisfactory. This block not only arrested the negative impulse, but sometimes brought about an apparent abolition of the positive also. This latter result cannot, however, be taken as decisive. The amplitude of positive response is naturally small ; under unfavourable circumstances it may have become so diminished as to pass unnoticed. In these circumstances the question of the character of the positive impulse may for the present be left an open one.

It has been shown that as the distance of the point of application of stimulus is reduced, the transmission period of the negative impulse is reduced at a greater rate than that of the positive. When the point of application of stimulus is at some distance, the negative impulse lags con- siderably behind the positive ; but as the distance is reduced, the lag tends to disappear. It thus happens that when the distance of application is sufficiently reduced, the positive effect is masked by the predominant negative.

In an experiment with Biophytum, electric stimulus was applied at a distance of 30 mm. from the responding leaflet. The positive response was initiated 2 seconds after the application of the stimulus and persisted for a further period of 12 seconds; the negative impulse then reached the leaflet, giving rise to the reversed negative response (fig. 104). The transmission period of the negative impulse is thus 14 seconds for a distance of 30 mm. The distance was next reduced to half, or 15 mm., and on repeating the stimulus

Fic. 104. — Effect of dimi- nishing the distance of application of stimulus. The lower record shows Ae ; diphasic response in Bio- Fic. 105.—Positive (3), diphasic phytum, positive followed (2), and neeahive FESspanee by negative. By applying (t) of Mimosa brought stimulus nearer, the posi- aleu By, radu Sete tive is masked by the tion of distance of appli- predominant negative cation of stimulus. the positive impulse was found masked, the negative occurring alone after an interval of 6°5 seconds.

I will next describe another expetiment carried out with Mimosa. Stimulus was applied on the stem at a distance sufficiently great to give only the positive response. This is shown in the lowest record in fig. 105. The point of application of stimulus was then brought somewhat nearer, with the result that a diphasic response took place, positive followed by negative—as depicted in the middle record. And finally, on applying stimulus still nearer, the positive was completely masked by the predominant negative.

We have hitherto dealt with the question of the suppres- sion of the positive by the negative. It is sometimes possible to unmask this suppressed positive by separation of the two impulses brought about by the differential effect of cold. Stimulus is at first applied on the stem at a distance near enough to give rise to the resultant negative response. A strip of cloth is then wound round the intermediate conduct- ing portion of the stem, and the region gradually cooled. The result of this cooling is to reduce the velocity of trans- mission of true excitation.

I have in this manner often succeeded, by careful and gradual cooling of the intermediate region, in converting a resultant negative into a diphasic—positive followed by negative ; further cooling gave rise to the positive alone. In the case of the diphasic response, the result was brought about by the lowering of the velocity of transmission of excitation by cold, in consequence of which the negative lagged behind the positive. In the case of the conversion into positive, the effect was due to the arrest of the excitatory negative impulse.

The effect induced by a given stimulus is modified, as we have seen, by its point of application. A stimulus thay be applied directly on the responding organ, or it may be applied at a distance from it. Following the usual terminology we shall designate the former as Direct Stimu- lation, and the latter as Indirect Stimulation. When stimulus is applied directly on the responding organ, there is induced an excitatory fall of the leaf concomitant with contraction and negative turgidity variation. This parti- cular reaction we shall designate as the DirEcT EFFECT of stimulus. When, on the other hand, a feeble stimulus is applied at a distance, there occurs only a positive response

of erection of leaf with concomitant expansion and positive turgidity variation. For simplicity we shall designate this particular reaction as the INDIRECT EFFECT of stimulus. If the intervening tissue be highly conducting and the stimulus sufficiently strong, then the excitatory negative effect masks the positive. In such a case the effect of indirect stimulation is the same as that caused by direct stimulation. But if the intervening tissue be semi-conduct- ing, or if the stimulus be feeble or applied at too great a distance, then we obtain the positive or INDIRECT EFFECT.

Two diametrically opposite effects may thus be induced by an identical stimulus, depending on direct or indirect application. The existence of the positive or indirect effect of stimulus has hitherto been unsuspected. It must be taken into full account in unravelling the com- plexities of reaction in a responding organ. The laws of Drrect and INDIREcT EFFEcTs of stimulus may thus be enunciated :— The effect at the responding region of a strong excitation transmitted through a short distance, or through a good con- ducting channel, is negative, being the same as the effect under direct stimulation. The response is by negative turgidity vari- ation, contraction, fall of leaf, and electrical change of galvano- metric negativity. This is the Direct Effect of Stimulus.

The effect of feeble stimulus transmitted through a great distance, or through a semi-conducting channel, is_ positive. The responsive reaction is by positive turgidity variation, expan- sion, erection of leaf, and electrical change of galvanometric positivity. This is the Indirect Effect of Stimulus. A single stimulus gives rise in the plant to two impulses, positive and negative. The positive travels at a faster rate and induces positive or erectile response of the responding leaf. The negative or excitatory impulse travels at a slower rate and induces in the responding leaf a contractile fall. The laws of Drrect and INDIREcT EFFEctTs of stimulus are—

The effect at the responding region of a strong excitation transmitted through a short distance, or through a good con- ducting channel, is negative ; the response is by negative tur- gidity variation, contraction, fall of leaf, and electric change of galvanometric negativity. The effect of feeble stimulus transmitted through a great distance or through a semi-conducting channel is positive. The response is by positive turgidity variation, expansion, erection of leaf, and electric change of galvanometric positivity.

Polar excitation in animal tissues—Anomalous reactions in Protozoa— Mono-polar method—Current Reverser—Excitatory polar action in plant—Method of record—Effects of ascending and descending currents in animal and plant—Records giving time-relations— Potential slide—Measurement of E.M.F. and current—Potential key- board. Towarps the end of a previous chapter I made brief refer- ence to certain remarkable excitatory effects which I have observed in the plant tissue at the initiation or cessation of a constant current. Electrical currents are well known to have certain characteristic effects in the case of animal tissues. On applying a current of feeble intensity to a muscle-preparation by means of two suitable electrodes, it is found that, at the moment of its sudden starting, an excitatory contraction is initiated at the point of kathode— that is to say, where the current leaves the tissue ; no such effect takes place at the anode, the point at which it enters. With current of feeble intensity there is, again, no excitatory effect either at the anode or the kathode on the cessation or break of current. It should be remembered that these marked exhibitions of excitation are caused by sudden variation of the current. They do not take place if the current variation is made very gradual.

These excitatory effects occur not only when the appli- cation of the current is direct but also when it is indirect. That is to say, when the two electrodes are applied on excitation with a feeble current is initiated only at the kathode at ‘make,’ this excitation being then conducted along the nerve to cause the contraction of the terminal muscle. It has been ascertained that this particular type of effect is by no means universal. In working on the polar effects of currents on Protozoa, it was found by Kiihne, Verworn, and others that the phenomena there displayed are more or less the opposite. Hence it has been assumed by some that the law of polar reaction in unfibrillated protoplasm is entirely different from that which holds good in animal tissues in general.

I shall, however, be able to show that the reactions in the undifferentiated protoplasm of the plant body are identical with those of highly differentiated animal tissues. Experi- ments with plants, moreover, led me to the discovery that what is known as Pfliiger’s Law is not a complete statement of the polar reactions that take place in living tissues. For it will be shown that under high electromotive forces a new class of phenomena comes into prominence, culminating in a more or less complete reversal of the normal reactions. From a consideration of these it would appear as if the effects on Protozoa were perhaps to be regarded as less anomalous than has hitherto been supposed.

In studying the polar effects of an electrical current on the plant, I shall first take a simple case and employ the pulvinus of Mimosa as the contractile organ by which the excitatory action is to be detected. A straight form non- polarising electrode is employed for making the necessary electrical connections. This consists of a glass tube the lower half of which is filled with kaolin paste moistened with normal saline. A wet thread hangs down from the kaolin and makes contact with the plant. The upper half of the tube, above the kaolin paste, is filled with a saturated solution of zinc sulphate ; an amalgamated thin rod of zinc dips into the zinc sulphate and serves as the electrode. In the Mono-polar method of experiment one contact is now

made at or near the pulvinus, and the other, with a distant indifferent point, lower down on the main stem. For suddenly starting or stopping a current, or for making an electrode anodic or kathodic, we may use Pohl’s Com- mutator or Reverser (fig. 106). When the commutator is in the middle position, the current is interrupted; when tilted to the left, the current enters the plant through the stem and leaves it by the pulvinus, which thus becomes the kathode. On partially tilting the commutator to the right, the current is broken. On tilting further to the right, the

Fic. 106.—Pohl’s commutator, for causing make and break of kathode or anode. current is suddenly reversed and the pulvinus becomes the anode. .For experiment I took a leaf of Mimosa which was in a fairly sensitive condition ; the electrical connections were made in the manner already described. The electrical resistance offered by the plant tissue between the two points of contact was found to be half a million ohms. An E.M.F. of 4 volts was found effective in inducing excitation at make, when the pulvinus was the kathode. The intensity of the exciting current was feeble, being 8 micro-amperes. A micro-ampere, it should be noted, is one-millionth part of an ampere. When the commutator was turned to the

left, the pulvinus was suddenly made the kathode, and this induced an excitatory fall of the leaf. It has been said that the exciting action is in general induced by sudden variation, and not during continuation of current. Hence, if after the excitatory fall the current is continued, the leaf is found to re-erect itself and to have its excitability restored. The current is now broken; this induces no excitation. The commutator is then tilted to the right, the pulvinus being made the anode; this again induces no excitation, nor is there any further excitation at the break of the anode. It is thus seen that with a feeble current the polar effects in Mimosa are precisely the same as in animal tissues— namely, excitation only at the make of the kathode. Having thus described the fundamental reaction of plant tissues under feeble currents, we have still to study the effects of stronger currents and the modifications that may be induced in the normal reactions under changing conditions, both internal and external. Indisputable evidence on such points can only be obtained if it is possible to have automatic records made by the plant itself. This I have been able to secure by the method of record already described, with the addition of a contrivance by which definite signals are marked at the base of the record, at the moment of application of either kathode or anode, its continuation, break, renewal, and so forth. For this a rotating reversing key has a disc attached to the axis. Semi-rotation of this axis to the left or to the right, or its position half-way between, renders the pulvinus kathode or anode, or suffices to break the current. The disc has a thread wound round its circumference, so that when the commutator-axis is rotated in one direction the thread is wound, and in the opposite direction unwound.

This thread is ultimately led to, and wound about, a second wheel with a long index, which is to serve as the marker. This second wheel is suitably fixed above the recording-plate, with the tip of the index resting on it at the same vertical line but a little lower than the response-recorder. When the commutator- key is turned to the left, the current is suddenly established, the pulvinus being made kathode. The marker, which has up to this moment been tracing a horizontal line, will simultaneously mark an up-line, as shown in the record. During the continuation of the current, the recorded signal will appear as horizontal but at a higher level. The break of kathode will be indicated by a down-line, stopping at the original level. Anode-make is produced by turning the key to the right, and this is indicated in the record as a down-line passing below the horizontal level. A horizontal line below the general level indicates the continuation of the anode. An up-line reach- ing the general level indicates the break of anode.

Two records are reproduced (fig. 107) exhibiting this polar Fic. 107.—Record of polar exci. Yeaction under feeble current Response seen to take place censitive specimen. The only at make of kathode. A 2 : : ode or anode. Up-lineabove will be seen that response normal denotes kathode- took place only at) kaGwemes either at kathode-break or anode-make or break. The next response was taken under the increased E.M.F. of 4 volts. The results depicted are similar to those in the last case, the only difference being the exhibition of an increased excitatory effect at kathode- make due to the higher voltage.

In the mono-polar method above described we have the effect of one particular electrode, quite distinct and isolated from the other. But when both the electrodes are placed on a conducting tissue, then there is no such distinctive isolation and the excitatory effect initiated at either of the electrodes may be transmitted to the terminal motile organ, there to induce contractile response. We have an example of this in the nerve-and-muscle preparation of a frog, where under feeble current excitatory contraction of the muscle is seen to take place at make of both ascending and descend- ing currents in the nerve. The terms ‘ascending’ and “descending ’ may be found somewhat confusing, especially in plant-experiments. The meaning will be clear if we remember that the current is said to be descending when it flows towards the responding motile organ. In the case of descending current in a nerve-and-muscle preparation, the kathode is proximal as regards the responding muscle ; with the ascending current, the kathode is distal.

As already stated, excitation takes place in nerve-and- muscle preparation at make of both ascending and descend- ing currents. This is explained as due to make of the proximal kathode in the case of descending, and make of the distal kathode in the case of ascending, currents. It is true that in the latter case the excitation has to traverse the anodic area before it can reach the motile organ. The anode induces, in general, a depression of conductivity which may even block the passage of excitation. In the present case, however, this blocking action is ineffective on account of the feebleness of the current.

The fact that it is the proximal kathode of the descending, and the distal kathode of the ascending, current which form respectively the seats of excitation, may be proved by taking time-measurements of the interval between ‘ make ’ of the current and the response of the muscle. With the descending current the interval should be shorter, since in such a case the kathode is proximal. This has been found to be the case. Characteristics in every way similar are found in experi- ments with the petiole and pulvinus of Mzmosa. In this case the petiole contains the conducting tissue and the