Plant Response as a Means of Physiological Investigation

In the case of Biophytum, when the temperature is maintained at a uniform favourable degree, the periods of the Fig. 124. Induction of Autonomous Response in Biophytum, at Moderately High Temperature of 350 G. Note the diminution of amplitude of response with the gradual loss of latent energy, consequent on falling temperature. The pulsations came to a stop below 290 C. autonomous pulsations become very regular. It has been said that these pulsatory movements are maintained by means of energy absorbed, and with Biophytum I found an added opportunity of demonstrating this fact. A particular plant had been kept at a uniform temperature of 350 C, under which the young leaflets gave autonomous responses. The heating current, by which this temperature was maintained uniform, was now stopped, and the chamber gradually

cooled down. One of the young leaflets had been attached to the Optical Lever, and records were now taken continuously. It was very interesting to observe how, with the gradual loss of absorbed energy, the pulsating movements of the plant became diminished in amplitude (fig. 124) till they came to a stop. We shall find exactly the same thing in the case of Desmodium. I referred in the last chapter to certain observations of my own, in which the leaflets of Biophytum were found, under favourable circumstances of light and temperature, to give rise to apparently spontaneous movements, which could not be traced to any definite varying external cause. From the experiment just described, however, we see that it was the continuous stimulus of favourable temperature and light combined that caused this rhythmic movement, which appeared as automatic.

(b) Resumption of automatic movements in Desmodium.— Similarly in Desmodium, if the isolated leaflet in which all movement has been brought to a stop, but not to a condition of permanent rigor, be raised in temperature, and if the tissue be maintained uniformly at this higher point, the pulsatory movement will be found to commence and persist for a long time. The initiation and maintenance of the responses here, again, is undoubtedly due to the renewed supply of energy.

A new difficulty now arises, however, from our habit of regarding stimulation as dependent upon some sudden variation of external conditions. For we are here confronted with a case in which uniform and continuous application of heat produces stimulation. It may be urged that there has been some variation of environmental conditions, in the fact of the rise of temperature before the constant point was reached. But it has to be remembered, that this rise was purposely made very gradual, and even if a single stimulation had been caused by this preliminary thermal variation, it would have evoked only a single response, or at most a few multiple responses But how are we to account for these

long-continued periodic pulsations, which are kept up during the whole time in which the leaflets are maintained at an unvarying given temperature ? We are thus led to see that stimulation is not in all cases dependent upon the occurrence of a sudden change in the exciting cause. On the contrary, excitation may be produced, as we have seen, under a constant and uniform exciting condition. If we analyse the multiple responses produced in Biophytum as the after-effect of the application of a strong thermal stimulus, we find, as said before, that some part of this thermal energy remains latent, and afterwards gives rise to recurring pulsations.

5. Of internal hydrostatic pressure. — The isolated animal heart, when in a state of standstill, is found to renew its excitatory pulsation under an increase of internal hydrostatic pressure. I shall show later (p. 349) that a Desmodium leaflet, similarly, when in a state of standstill, can be made to resume its excitatory rhythmic activity, by increasing the internal hydrostatic pressure. The true meaning of ' tonic ' condition. — We have thus seen that under the continuous action of a constant source of external stimulus, multiple responses are produced. We have also seen that the excess of energy absorbed remains latent in the tissue, in consequence of which, even on the cessation of external stimulation, the pulsatory movements are maintained for a longer or shorter time. It is thus the excess of stimulus absorbed which renders the tissue excitable, or 'tonic' Hence we may have tonicity imparted by light, photo-tonus ; by favourable temperature, thermo-tonus ; by electric current, electro-tonus ; by internal hydrostatic pressure, hydrotonus ; or by the presence of favourable chemical substances, chemo-tonus. WTe have seen that each one of these, by itself, was competent to give rise to multiple responses. It has been shown further that there is no hard- and-fast line between such multiple and automatic responses, the one passing imperceptibly into the other.

determined by the sum total of the latent energy derived from all the above-named factors. This internal factor, of latent energy, will be shown to play a very important part in all response-phenomena, and as it thus becomes necessary to have some convenient means of referring to it, we shall henceforth designate it as the Internal Energy of the plant. Automatic movements in plants are thus only exhibited under favourable tonic conditions. It has been shown that the plant displays rhythmic activity when subjected to different forms of constant stimulus, and we have now investigated separately the effects of such constant stimuli — chemical, electrical, thermal, photic, and hydrostatic. As has been explained before, if a given stimulus be not of sufficient intensity to evoke visible response, yet the absorbed energy may render the tissue capable of responding to another subsequent stimulus, which by itself would have been ineffectual ; in other words, the tissue is made excitable by the presence of a stimulus which has not of itself been adequate to cause response. When this latent excitability exceeds a certain amount, any further increase may be expressed in a visible manner by mechanical pulsation. Now, taking the various forms of stimulus to which the plant is constantly exposed — namely, warmth, light, moisture, and the action of the various chemical reagents, organic and inorganic, present in it or absorbed by it — it is clear that each of these exerts its stimulating effect independently ; any one by itself may then make the tissue excitable to the verge of response ; in this condition, though there is no outward sign of the fact, there is a considerable amount of latent energy ; and the incidence of any second and additional form of stimulation is now sufficient to precipitate the excitation in visible form. These considerations will show how, by the cumulative and additive effects of all the forms of constant stimulation mentioned above, the plant may become so highly excitable as to manifest the fact by giving rise to responses which appear as automatic. The tonic condition is thus the latent excitatory

condition, which is determined by the sum total of all these exciting factors. In connection with these facts, it is well to bear in mind that the excitability imparted by a stimulus does not always increase continuously with the intensity. On the contrary, there may be an optimum intensity beyond which excitability may be diminished. We have now seen that the energy which expresses itself in pulsatory movements may be derived by the plant, either directly from immediate external sources ; or from the excess of such energy, already accumulated and held latent in the tissue, aided by the incidence of external stimulation ; or from an excessive accumulation of such latent energy alone. In the last case, however, if the plant were kept isolated from all external supply of energy, it is clear that its reserve would become exhausted, and its automatic movements would cease. I have already described an experiment in which this arrest of pulsation took place in the case of a specimen of Desmodium kept in a dark room. We then saw that revival of response was only brought about when fresh stimulus was applied. And we have also seen the converse, namely, the ordinarily responding Biophytum, when supplied with excess of energy, become automatically responding.

Cause of rhythmicity. — Having, then, seen that it is a constant source of energy, external or internal — the latter being really derived from a previous absorption of external stimulus — which maintains the so-called automatic movements in plants, we have still to determine how it is that a latent or constantly acting external stimulus can find only periodic expression ? In connection with this we have seen how, when the minimal factor of stimulating intensity is exceeded, there is a manifestation of the fact by visible response. I have also shown (p. 245) that after each excitatory discharge there is a marked diminution of both conductivity and excitability ; and a new stimulus, or existing excess of stimulus, owing to the loss of these properties, is retarded for a time from producing any effect on the motile tissue. It

is only after the lapse of an interval that the protoplasm regains its original properties. There is thus an oscillatory variation of conductivity and excitability. It will therefore be seen how, under the circumstances, a constant stimulus, or a stimulus which is latent in the tissue, can find an excitatory expression only in a pulsating manner. Perhaps a physical model will enable us to visualise this process. Imagine a reservoir into which flows a constant supply of water. An elastic conducting pipe is led from the reservoir, and this pipe is constricted by a compressing spring. On the far open end of this pipe abuts the flattened end of an indicating lever. When water has been supplied for some time, its level is gradually raised, producing an increasing pressure. At a certain point, when the pressure becomes sufficiently great, the spring which keeps the elastic tube constricted gives way, and there is an impulsive discharge of water, which, impinging against the lever, gives rise to a visible response. But the yielding spring again closes, the tube is once more constricted, and thus by the oscillation of the spring which regulates the conduction of water, pulsating hydraulic impulses are kept up. On account of the oscillating mechanism, the outflow, and consequent mechanical response, are periodic, though the supply is constant. In this model the first period, before the pressure of water becomes sufficient to force open the spring, corresponds to the latent period in plant response ; the oscillation of the flow of water corresponds to the oscillation of conductivity ; and the responding lever corresponds to the motile leaflet. Similarly, an ill-fitting spring tap is thus often thrown into a pulsating movement, by the constant pressure of water from the main, and there is then seen a rhythmic play of the water-jet.

1 After-effect ' and its relative persistence. — In the experiment on Biophytum under the continued action of light, we saw that for a period of one minute, during which the plant was absorbing light, there was no response. Then after this latent period of one minute, the energy absorbed reached the verge of response. The excitatory discharge was followed by a retardation of conductivity and excitability, which was, however, gradually recovered, and there were produced a second and then successive periodic responsive movements.

In the hydraulic model, if the capacity of the reservoir be small, the cessation of the water-supply will cause an immediate cessation of the rhythmic movement of the indicating lever. In Biophytum, similarly, we found that the periodic response continued as long as energy was supplied, and that the movement soon stopped on the stoppage of the supply of external energy. But if the capacity of the reservoir be great, the accumulation may be sufficient to maintain the oscillation for a considerable length of time, even after the main supply is cut off. And we see that in Desmodium the responsive movements continue in a persistent manner, though the immediate source of stimulation be interrupted. A similar difference in the persistence of after-effects, depending on relative capacities for storage of energy, is seen in the two classes of inorganic substances, which are distinguished as fluorescent and phosphorescent. In the first, the responsive emission of light caused by a preceding excitation is extremely short-lived ; but in the latter it continues long after the light stimulus has ceased to act. Thus Desmodium may continue to exhibit rhythmic movements, though not at the moment exposed to the marked action of any special source of stimulus. But for the display of long-continued rhythmic movements it should previously have absorbed a considerable amount of energy from an external. source — in other words, it must have been exposed to those circumstances which produce a favourable tonic condition.

We have thus obtained some insight into that very obscure phenomenon which is known as the after-effect. By the inertia of the organism there is a certain loss of time before response begins to take place, and this determines the latent period. But when the stimulus has already initiated movement, the responding organ will, through the same inertia, continue to show this movement even when the stimulus has ceased to act. There is another factor, however, which

determines the persistence of this after-effect, namely, the larger or smaller capacity of the tissue itself to hold energy latent within it. In the attempt to investigate the cause of automatic movements in Desmodium, there were three points of inquiry which had to be determined. First, there was the question of the seat of excitation ; second, that of the nature of the stimulus which maintained the rhythmic pulsation ; and third, the determination of the process by which constant stimulus found periodic expression.

I have shown that the seat of excitation in Desmodium is neither central nor peripheral, but is in, or in immediate contiguity with, the motile tissue itself, which resembles in this respect the animal heart, the seat of excitation there also being in the cardiac tissue itself (Chapter XXVI.). I have also shown that as the cause of excitation it is not necessary to have any sudden variation. A constant stimulus, of whatever nature, is found efficient to produce excitation.

I have also demonstrated that periodic pulsation is produced in Desmodium at standstill, by a constant thermal stimulus ; also that periodic pulsations are produced in Biophytum and in Desmodium by the constant action of a chemical stimulant. I have shown further that rhythmic excitation was produced in Averrhoa carambola by the passage of a constant electric current. And, finally, I have shown that, just as in the retina, under constant stimulus of light we have periodic visual excitations, so also in Biophytum, under constant stimulus of light we obtain periodic excitations which give rise to rhythmic movements. In the animal tissue, similar multiple rhythmic responses are met with under constant stimulus.

Since we have found it to be a fundamental characteristic of the tissue of a rhythmically-responding plant like Biophytum or Desmodium to give a response which cannot be increased by any excess in the stimulus-intensity (maximal response or none), we should expect that the excess over the minimally effective stimulus must remain latent in the tissue. This is evidenced by the fact that on applying a very strong stimulus we obtain, not a single but multiple responses. Thus all the various forms of constant stimulus to which it is exposed — warmth, light, moisture, and the different chemical reagents, organic and inorganic, present in or absorbed by it — become latent, and the sum total of all these stimulating factors determines its ' tonic ' condition.

When this accumulated latent energy exceeds a certain value, it is visibly manifested in the form of the so-called 1 automatic ' movements. The periodicity of the excitatory discharges which give rise to rhythmic movements in a plant that is under constant latent stimulus, is brought about by the peculiarity which has been demonstrated, that after each discharge the conductivity and excitability of the tissue are diminished, and are only gradually regained. This oscillation in the conductivity regulates the outflow of energy, and causes the rhythmicity of the responsive movements.

In Desmodium, the seat of excitation of the lateral leaflets lies in the motile organ itself. Multiple response is produced in the plant, as in the animal, under constant thermal, chemical, or electrical stimulus. The retina, under constant stimulus of light, exhibits periodic visual pulsations. Similarly, the leaflet of Biophytum, and that of Desmodium at standstill, under continuous stimulation of light, exhibit rhythmic mechanical pulsations.

Biophytum, when raised to a temperature of about 290 C, becomes automatically responding. Under these circumstances, the only difference between the so-called automatism of Biophytum and Desmodium is that, in the latter case, the critical thermo-tonic condition is arrived at about 120 C. earlier. The energy which expresses itself in pulsatory movements may be derived by the plant, either directly from immediate external sources, or from the excess of such energy, already accumulated and held latent in the tissue, aided by the incidence of external stimulation, or from an excessive accumulation of such latent energy alone.

By i tonic ' condition is meant the latent excitatory condition of the plant, as determined by the sum total of the stimulating factors which are, or have been, derived from its environment. In other words, the tonic condition depends on the internal energy of the plant. In rhythmic tissues, a constant stimulus, external or internal, finds pulsatory expression in consequence of the oscillatory variation of conductivity and excitability. The duration of rhythmic movements, in the absence of any external exciting cause, depends on the amount of energy previously absorbed and held latent in the plant. The persistence of this after-effect, therefore, depends also on the greater or less capacity of the tissue for storage of energy. These rhythmic movements thus appear to be automatic, but when the reserve is exhausted, the so-called automatic movements come to a stop. Renewal of pulsatory movements can then take place only on the supply of fresh energy from without.

The recorder and experimental chamber — Absolute measurement of period and amplitude of Desmod/2/m-osciWa.tion — Responsive significance of up and down movements deduced from (a) analogy with response of Mimosa ; (b) test of increased internal hydrostatic pressure — • Systolic' contraction and ' diastolic' expansion of Desmodhtm pulvinus — Mode of application of chemical reagents — Action of chemical reagents modified by : tonic condition of plants ; strength of solution ; and duration of application — Effect of anaesthetics — Effect of alcohol — Effect of carbonic acid — Effects of ammonia and of carbon disulphide — Effect of copper sulphate solution, either when applied externally, direct on the pulvinus, or internally — Spark-record of Desmodiumpulsation.

Having thus, in the last chapter, traced the causes of autonomous movements in plants, I shall now, taking Desmodium gyrans as the type, describe the effect of various agencies on the so-called ' spontaneous ' responses of its lateral leaflets. With regard to the experimental arrangements for making the record, I have already in Chapter I. described how records may be obtained, using the intact plant for experiment. The automatic movements of the leaflets, however, persist, even after the petiole bearing them is detached, the cut end being kept in water. Under proper conditions, the rhythmic pulsations of the detached specimen will continue for a couple of days. In order, therefore, to subject the motile organ to various modifying conditions, it is much more convenient to use such a specimen than the whole plant.

The recorder and experimental chamber. — As the extent of the movement of the tip of the leaflet is considerable, no magnification is necessary for the record. A single cocoon thread is attached to the middle of the leaflet by a drop of shellac varnish, the other end of the thread being tied to the longer arm of the Optic Lever, which is 30 cm. long. The distance of the recording drum from the mirror is also 30 cm. The length of the lever arm being thus equal to the distance of the drum, and the extent of the angular movement being by reflection doubled, it will be seen that the movement is magnified twice. The thread is attached, however, to the middle of the leaflet, and the record there-

Experimental Apparatus for Making Records of Pulsation of Desmodium Leaflet, P, mounted in u-tube in plant chamber, and attached to long arm of Optical Lever, L ; M, mirror attached to fulcrum-rod ; D, recording drum ; 1, O, inlet and outlet pipes for gases and vapours introduced into plant chamber ; c, electric heating coil. fore gives the movement of the tip unmagnified. The records given in some of the figures are thus without magnification. In others, again, the records are on a reduced scale.

It will be understood here that the extent of movement will vary with the length of the leaflet, some of these being very small, and others relatively large. In the apparatus shown in fig. 125, the smaller chamber contains the motile leaflets mounted in a tube filled with water. By means of an inlet-pipe, different gases may be introduced into the chamber for any desired length of time, after which fresh air may be re-introduced. In order to produce variations of temperature there is a coil of wire for electric heating.

The automatic movements of the leaflet, both up and down, take place in some cases as a number of jerks, which may pass gradually into continuous movement. In others they are continuous from the beginning. From the normal or highest position, the leaflet, generally speaking, sinks somewhat rapidly. Having reached its maximum depressed position, there is a pause, after which there is rather a slow return to its original position. The up and down motion is in some cases approximately straight. In other cases, the sub-petiole of the leaflet is slightly twisted after its descent, and the curve described becomes more circular. For the purpose of the present investigation, specimens were selected in which the movement of the leaflet took place gradually, without jerl?s, the up and down movements being approximately in a straight line.

Absolute measurement of period and amplitude of Desmodium - oscillation. — The period of a single oscillation varies with the temperature and the tonic condition of the plant. In winter it may be as long as five, in summer as short as two minutes, and when the temperature is artificially raised, the period may be even further reduced to one minute. As a concrete example, affording a clear idea of the general characteristics of the pulsatory movement of Desmodiiun, I shall give the following results, obtained from a photographic record which gives the extent of the absolute movement (fig. 126).

The up movement of the record means down movement of the leaflet in this and all subsequent records. The period of the complete vibration was in this case 35 minutes, of which the down movement was accomplished in the course of IMJ minute. The up movement was relatively slower, and was accomplished in two minutes. The mean amplitude of pulsation — that is to say, the vertical distance travelled by the tip of the leaf — was 25 mm. The fact that the down movement is, generally speaking, relatively the quicker is seen visually demonstrated in the photographic record of uniform pulsations obtained

with another specimen (fig. 127). In connection with this, certain peculiarities of photographic action should be borne in mind. It is found that a short exposure gives an image which in the case of a line is very thin and sometimes consists of only the faintest impression; but when the exposure is prolonged the line is much thickened. Hence, in the records, the faint or more sharply outlined portions indicate responsive down movements which were relatively rapid. In fig. 127, therefore, these differences of line afford us a graphic representation of the various rates of movement, and durations of pause, in the different parts of the curve. In all the photographic records given here and elsewhere, we are thus able to distinguish the down movements by the relative thinness of the recording line. In fig. 134, at the end of this chapter, will be found a spark-record of the pulsation of Desmodium in which the different rates of movement at different stages of the response can be distinguished at a glance.