Plant Response as a Means of Physiological Investigation

Growth-response and excitatory response.— If we compare these multiple growth-responses with the multiple mechanical responses of Biophytum (fig. 116), their similarity is at once evident. As in that case, so here also, recovery is not complete, and the series of effects produced is therefore additive in both cases. With Biophytum^ however, when the leaflet is depressed to the utmost, a limit is reached ; but in growth there is no such limit, and the summation of effects may go on indefinitely, until senility and death supervene.

Yet there is a certain difference between these responses. In the case of Biophytum, the response is due to a sudden diminution, but in growth, to a sudden increase, of turgidity. This might at first sight appear anomalous, but I shall presently show that both are expressions of an excitatory reaction ; for we have seen that when the responsive organ of Biophytum is directly excited there is an expulsion of water, and the response is brought about by negative turgidityvariation. This variation we shall for the sake of convenience distinguish as the direct effect of stimulation. We saw in the last chapter, however, that an increase of the internal energy of a plant gives rise to an opposite response, that is to say, one characteristic of positive turgidityvariation. The two responsive turgidity-variations, then, both negative and positive, are alike in being expressions of the excitatory reaction, though the negative variation is the effect of external stimulus applied directly to the responding organ, while the positive variation is to be regarded as the effect of the internal energy of the plant. This internal energy may itself have been derived previously by the plant from external sources of stimulation, or the internal energy of a given point may result from the application of a stimulus at a distance. The pumping-in of water by the stimulated root is an example of the latter case ; the cells are thus made tense, and the potential energy of the tissue is raised above par. Again, we may conceive of another interesting instance as follows. When stimulus is applied at a distance the excitatory expulsion of water gives rise to a wave of increased turgidity, which produces an abnormal positive response, and this positive turgidity-variation we shall designate as the indirect effect of stimulation.

The wave of true excitation may in this case reach the organ after that of positive turgidity-variation, and give rise to the direct effect of stimulation, that is to say, depression of the leaflet. We may next imagine that the seat of stimulus is at so great a distance from the responding organ, that the transmitted excitation becomes too much enfeebled, by the long tract which has to be traversed, to produce the excitatory negative response. In this case, nevertheless, the sudden expulsion of water at a distance will give rise to a wave of increased turgidity, which will reach the responding organ, and produce there only that response which is characteristic of positive turgidityvariation.

Positive response as indirect effect of excitation. — Numerous experiments have been described exhibiting the negative turgidity-variation as the direct effect of stimulation. I have already described the production of positive turgidity- Fig. 170. Responses of Leaf cf Artocarpiis to Thermal Stimulat Thick dots show points of application of stimulation. In a stimulus was applied near the responsive pulvinoid and gave rise to normal response of fall — here represented as up movement— preceded by preliminary erectile twitch. In b stimulus was applied at a greater distance. The true excitatory effect did not reach the organ, and we obtain positive erectile response of positive turgidity-variation, here represented as down.

variation as the indirect effect of stimulation (p. 400). A fuller demonstration of this, by the electrical method, will be found in Chapter XXXVII. I shall here give an experiment which establishes the fact by means of mechanical response. In fig. 32 (reproduced in fig. 170, a) was shown a series of normal mechanical responses of negative turgidity-variation obtained from the leaf of Artocarpus. The conductivity of the petiole in this case is relatively feeble, and these normal responses, preceded by preliminary positive twitches, were obtained when stimulus was applied at a distance of

3 mm. from the responsive pulvinoid. I then repeated the experiment with the same leaf, but applying stimulus at the greater distance of 5 mm. The responses now consisted of a series of up movements of the leaf, indicative of positive turgidity-variations (fig. 170, b), the direct effect of stimulus not reaching the organ. Now, turning our attention to the growing organ, we find that the fibro-vascular element, which possesses the power of conduction to a high degree, is not yet fully established in the zone of growth. If, then, contiguous to the growing zone there be a mass of active tissue thrown into a state of rhythmic excitation, it is to be expected that the indirect effect of such stimulation will alone act, and give rise in the region of growth to pulsations of increased turgidity. It will be remembered from the last chapter'that water is conducted by preference along the fibro-vascular elements ; and since these strands end below the zone of growth, it is clear that there must be in this region an accumulation of water, and consequent over-turgidity of the tissue ; a condition which is, as we know, sufficient to initiate rhythmic excitation.1 This region, then, acts like the actively excitable tissue of Colocasta, which, as we saw, gives rise to spasmodic expulsions of water. In the latter case there is, however, a channel by which the water escapes, thus relieving the pressure on the tissue ; but the growing organ offers only a cul de sac, and the constant repetition of hydrostatic blows thus effects those positive turgidity-variations that are to result in the responsive elongations and incomplete recoveries of the tissue, bringing about growth-movements.

* Inner stimuli.' — It is thus seen that growth represents the indirect effect of stimulus ; its motive power residing in the rhythmic activity of the internal tissues of the plant. This rhythmic activity has been shown to be, in its turn, the result of the tonic condition of the plant, that is to say, of the sum total of energy previously absorbed, and held 1 Or the over-turgidity of the growing region may be sufficient of itself to initiate rhythmic activity.

latent in the tissue (p. 314), which we have designated as the internal energy. We have thus succeeded in defining the actual nature of those ' inner stimuli ' to which the phenomenon of growth is usually vaguely ascribed. From what has been said, it is clear that the responsive peculiarities of the growing region are not per se in any way different from those of any other excitable tissue, the apparent contrast between negative and positive turgidityvariations — that is to say, between responsive contractions and responsive expansions — having been shown to depend upon the fact that in one case we see the direct, and in the other the indirect, effects of excitation. If this be so, it follows that the direct application of external stimulus to a growing tissue ought to have the normal effect of excitatory contraction. In other words, while the action of the so-called ' inner stimuli,' or internal energy, gives rise, as explained above, to responsive expansions, the direct effect of external local stimulation must be the production of responsive contractions. That this is the case, will be shown in the next chapter.

Each of these multiple growth-responses consists of a sudden elongation, due to a pulse of increased turgidity, followed by an incomplete recovery. The difference between elongation and recovery is the irreversible growth-effect. In ordinary excitatory response there is a pulse of diminished, and in growth-response a pulse of increased, turgidity. Both are, however, effects of excitatory reaction. When a tissue is locally excited, it gives a response of negative turgidity-variation, that is to say, of contraction. This is the direct effect of stimulus.

When the source of stimulation is behind, and the intervening tissue does not conduct excitation, then the excitatory expulsion of water finds expression in a positive turgidityvariation, which produces an expansion of the responding zone of growth. The growth-response is thus the indirect effect of stimulation. The rhythmic activity of internal tissue supplies its motive power to the zone of growth. This activity, depending on the tonic condition of the plant, constitutes the ' inner stimuli ' to which growth is to be ascribed.

Characteristics common to growth and to other forms of rhythmic response : (i) Periodic groupings— (2) Effect of external stimulus in renewal of growth when at temporary standstill — (3) Renewal of growth-pulsation by positive turgidity-variation — (4) Effect of increased internal hydrostatic pressure — (5) Effect of ascent of sap on growth —Effect of temperature on growth — Comparison of various types of multiple response — Effect of external tension on growth — Effect of direct application of stimulus on the growing region — Similarities between motile and growth responses — Direct and indirect effects of stimulus, and laws of growth.

Having shown in the last chapter that growth is a form of multiple or rhythmic response, I shall now proceed to demonstrate in detail the fact that in it also are found various phenomena which are characteristic of rhythmic response in general. (1) Periodic groupings.- -In the multiple response of Biophytum, and in the autonomous response of Desmodium, we have noticed the occurrence of various periodic groupings, the simplest of which consisted of an alternate waxing and waning of the pulses. In multiple growth-responses, similarly, we are able to detect such groupings, of which the simplest was shown in fig. 169 (c). When a continuous series of records is taken, extending over some time, these groupings undergo various changes, as is illustrated in that figure, the three series (a), (b), and (c) having been taken with the same plant at different intervals. It will there be seen that the pulse-records in (c) represent an alternate waxing and waning of amplitude ; that in those of (&) the responses are small at the beginning and large at the end ; and, finally, that in the

responses of (a) this order is reversed, the amplitude being at first great, and undergoing a steady decrease to the end. (2) Effect of external stimulus on growth when at standstill. — It will be remembered that the rhythmic excitation of Desmodium comes to a standstill under unfavourable circumstances — that is to say, when the sum total of internal energy has fallen below par ; and when this has happened, the application of fresh external stimulus is found to renew the activity. Growth-response, similarly, comes to a stop when the plant is in an unfavourable /

condition with regard to light, temperature, or moisture. I shall now show that under such circumstances the application of external stimulus is found to be competent to renew growth. Taking a specimen of the hypocotyl of Tamarindus indzcus, in which growth had come to standstill, I stimulated the plant by thermal means, and this was found to renew the multiple response of growth, as is clearly seen in fig. 171. The pulses are here characterised by interesting periodic variations. The period of each is relatively long, the average value being about six minutes. In some other cases the renewed pulsations were so rapid as almost to appear continuous. When the tonic condition of the plant was very low, the energy supplied by brief stimulation was only sufficient to maintain the rhythmic growth-activity for a short time, and after this the plant would again return to the state of standstill. It will thus be seen that when the tonic condition of the plant is below par, the applied external stimulus is absorbed, and, becoming latent, serves as internal energy for the production of growth-response (cf. p. 462). I shall adduce other instances of this in the course of the present chapter.

Fig. 171. Renewal of Growth-pulsation by Thermal Stimulus in Tamarindus indicus originally at Standstill Fig. 172. Initiation of Erectile Response in leaf by Supply of Water to partially Drought-rigored Mimosa (3) Renewal of growth by positive turgidity-variation. — We have seen that in Desmodium in a state of standstill, increased internal hydrostatic pressure renewed the rhythmic activity. It was also stated in the last chapter, that growth is a responsive expression of the positive turgidityvariation. We have seen further that the mechanical expression of the positive turgidity-variation in a dorsiventral organ takes the form of erectile response. Thus this erectile response and growth-elongation are to be regarded as two different forms of expression of the same internal activity.

If we take for example a plant of Mimosa which is underturgid, for want of sufficient supply of water, but not to the extent of drought-rigor, the leaves are found to assume a certain horizontal position, corresponding to the degree of turgidity. If we now supply the plant with water, poured on at the roots, the consequent sudden increase of suctional pumping activity is seen in the positiveerectile response of the leaf (fig. 172). Similarly, when an ordinary plant, under the same circumstances, has its growth brought to a standstill, the growth-elongation is found to be renewed on the application of a fresh supply of water. This experiment was carried out on a seedling of Cucurbita 12 cm. in height, growing in a small pot, which had come to growth-standstill for reasons described. Two cc. of water was supplied to the dry soil about the roots, and

Fig. 173. Initiation of Growthpulsation by Small Supply of Water to Drought-rigored Seedling of Ctuurbita The first thick dot represents application of water, which induces growth for three minutes only. A second application at the second dot renews it for the second time. growth-response was initiated after a latent period of eleven seconds (fig. 173). It will be remembered that the positive turgidity-variation, on which growth depends, is hydrostatically transmitted at a much quicker rate than the state of excitation itself. In the present case, the responsive elongation in the growing region at a distance of about 12 cm. took place eleven seconds after the application of water at the root. There was a certain loss of time before the cells in the growing region became of sufficient turgidity to initiate growth. The small supply of water which had been given was enough to maintain growth for three minutes only, after which the plant came again to a standstill. Another 2 cc. of water was now applied, and the latent period was found reduced, as we should have expected, being now three seconds only. This was due to the fact that the cells had not now to absorb water before they could be sufficiently turgid. This renewed growth-activity was again, however, exhausted after about three minutes ; and it was very interesting to observe how the response of growth followed, for a little while, after each such doling out of water.

(4) Effect of increased internal hydrostatic pressure. — We found, in the case of Desmodium, when the cut petiole, carrying the motile leaflets, was subjected to increased hydrostatic pressure, applied by means of the U-tube, that the rhythmic activity of the leaflet, as shown by its quickening, was thereby increased (p. 320). We saw also in that case that this increased frequency, due to increased pressure, reached an optimum, and that beyond this, under excessive pressure, the pulsations became irregular, or even came to a stop.

In order, then, to study the effect of hydrostatic pressure on growth, I mounted the specimens — in this case an entire seedling of Balsam and a cut flower of Crinum—m U -tubes supplied with water, and proceeded to take records, first under normal conditions— z>. the level of water in the two limbs being the same— and then under a gradually increasing hydrostatic pressure. These records are made, it should be said, only when the rate of growth under the changed

conditions has become uniform. This occurs, generally speaking, after a period of variation which does not exceed two or three minutes. I give here a table embodying the results of the experiments on Balsam, and on Crinum, which show how increase of internal hydrostatic pressure increases the rate of growth up to an optimum, after which there is a diminution of growth. Table showing Effect on Growth of Increased Internal Hydrostatic Pressure The curve seen in fig. 174 exhibits graphically the relation between these internal pressures and corresponding growths in the case of Balsam seedling. It will be seen that after a certain moderate rate of growth has been attained by increase of pressure, the curve becomes a straight line ; that is to say, after this point, equal variation of pressure produces equal variation in the rate of growth. But in the first part of the curve, where the rate of growth is feeble, an equal increase of pressure causes a disproportionately large increase in the rate of growth. This is still more strikingly shown when the growth, to begin with, is zero — that is to say, at standstill ; in such a case, by gradually increasing the internal pressure, we arrive at a point where growth begins abruptly, after which increasing pressure causes an increasing rate of growth. But if the pressure be now brought back to a point just below that at which growth was initiated, it is found not to be arrested, but to persist. Thus the curve does not here return upon itself.

Since the various growth-curvatures are brought about by the variations of internal hydrostatic pressure caused by the action of external stimulus, this quantitative demonstration of the effect of internal pressure on growth, is of much theoretical importance. I have already shown the connection between mechanical response and growth-response, and demonstrated the fact that the erectile mechanical response and growth-elongation are but different expressions of increased internal activity. I shall now show how the suctional and growth responses are related to each other, and in what manner the action of the former affects the latter.

(5) Effect of ascent of sap on growth. — We have already seen that the positive turgidity-variation on which growth depends is brought about, under normal conditions, by the ascent of sap. As regards the latter, we have seen that when the root is subjected to the stimulating action of warm water there is a sudden augmentation induced in the rate of suction. The application of cold water, on the other hand, induces the converse effect. The mechanical response of the plant to warm or cold water, applied at the base, was shown to be manifested in the erection or depression of the leaves of Mimosa or Biophytum (p. 400).

These agents are seen in the following experiments to produce parallel effects on growth. A growing Crinum Lily was taken, and its normal rate of growth ascertained to be •005 mm. per minute. Ice-cold water was now applied at its base, and this was found to cause an almost immediate arrest of growth. As the temperature, however, was gradually restored to that of the surroundings, the rate of growth was also slowly recovered. Five minutes after, the rate was only

Pig. 174. Curve showing Relation between Internal Hydrostatic Pressure and Rate of Growth {Crinum Lily) •ooi mm. per minute, or one-fifth of the original rate. It was only after about half an hour that the original rate of growth was once more attained. I next applied warm water at the base, with the result that the rate of growth was almost instantaneously enhanced to '125 mm. per minute, or twenty-five times the normal! That this effect was not due to the rise of temperature as such, is shown by the fact that it was almost instantaneous, and that, moreover, as will be shown in the next chapter, the maximum rate of growth of Crinum at the optimum temperature is only about three or four times as great as the normal.

From these experiments we see that the energy applied at the root is transmitted hydraulically to the growing region by the ascent of sap, where a certain amount of work is performed in causing an increase of turgidity, and thus producing in the cells a state of tension. Growth is now caused not simply by the presence of water, but rather by the energy conveyed by that water. It is well to bear in mind, at this point, that the mobility or plasticity of the responding growing region is also an important factor in the production of growth ; for if the molecular mobility of the zone of growth be in any way reduced, the transmitted pressure, which was formerly effective, will now become ineffective to bring about growth.

(6) Effect of temperature on growth. — We have seen, in studying the pulsatory movements of Desmodium, that the rise of temperature, within certain moderate limits, increased the rhythmic activity of the plant, as shown in the increased frequency of pulsation. At a maximum temperature, again, above 400 C. these movements almost disappeared, there being now produced very rapid oscillations, of so small an amplitude as to be almost incapable of detection. This will be seen in fig. 175, where the normal pulsations, of a period of 2*5' at 300 C, are seen reduced to a period of only 10" at 420 C. With this, the amplitude also is so far reduced as to be visible only on very careful inspection. We meet with

corresponding phenomena in growth-response. It will be shown in the next chapter that the rate of growth increases with the rise of the temperature up to a certain optimum. At a determinate maximum, however, which is about 440 C, growth is arrested, but this arrest does not, as we have just seen in the corresponding instance of Desrnodium, imply a total cessation of internal activity. In making experiments on a seedling of Balsam, I obtained the record shown in the upper part of fig. 176, the temperature being 340 C, which is below the optimum. The average period of a single pulse

Fig. 175. Photographic Record showing the Slow Pulsations of Large Amplitude of Desmodium Leaflet at 300 C. to become very much Quickened and Reduced in Amplitude at 420 C. was in this case 125 seconds, and owing to the considerable amplitude of each pulsation, combined with its incomplete recovery, the average of the resultant rate of growth was as much as '074 mm. per minute. On now raising the temperature to 440 C. I obtained the lower of the two records in fig. 176, showing no resultant growth. It is interesting to observe the process by which the cessation of growth comes about in this case. For it is clearly seen from the record that there is no cessation of activity. On the contrary, we find that the frequency of oscillation has become increased from four pulsations to ten,

in the course of 50". The average period has thus fallen from 12-5" at 340 C. to 5" at 440 C. The amplitude of pulsation at the same time is found to be decreased, and this, with the fact that recovery is now complete, accounts for the resultant cessation of growth. Comparison of various types of multiple response. — At this point, it is worth while to compare two or three types of multiple response. In Bioftkytum we have seen that, by reason of incomplete recoveries from negative turgidityvariation, the multiply-responding leaflet gradually becomes depressed below its original level. In contrast to this we have in growthresponse those incomplete recoveries from positive turgidity-variations which have the effect of gradual elongations In the pulsation of Desmodium again we have an intermediate instance where, response and recovery being equal, the responding organ is ultimately neither raised nor depressed. It is interesting to note, therefore, that in raising the temperature of a growing organ to the maximum, and thus abolishing the resultant elongation, we bring on a condition of equality of response and recovery which in so far resembles the pulsation of Desmodium.

Effect of external tension on growth. — Great advantages are afforded by the method of magnified record, which enables us to detect instantly the immediate and after effects on the specimen of any changes of external conditions. It is Upper record shows slow pulsations with incomplete recoveries at 340 C. Lower record shows quickened pulsations with complete recoveries at 440 C. The magnification employed is 500 times. thus easy to obtain exact records of the effect of tension on growth. The normal record is first taken, with the very slight tension exerted by the recording lever itself. The short arm of the lever, -5 cm. in length, is, it should be remembered, attached to the growing organ. A rider, half a gramme in weight, can be placed on the longer arm of the lever, at distances of '5, 1, 1*5, 2, 2*5, or 3 cm. from the fulcrum. The effective tension may thus be gradually increased, and the corresponding effects on growth recorded. It may be stated here that, generally speaking, any sudden change of external conditions, such as sudden cooling, sudden warming, or sudden variation of tension, acts on the organ as an external stimulus ; and I shall presently show that an external stimulus always induces a contraction or retardation of growth. When the organ is subjected to sudden increase of tension, the preliminary effect of contraction occurs therefore, as we should expect. But after this temporary disturbance has disappeared we are able to observe the permanent effect of increase of tension on growth. For these experiments I took different specimens of Crinum Lily, and the results obtained show that increase of tension enhances the rate of growth. This increase, however, appears to reach a limit at a certain optimum point, beyond which increase of tension would seem to retard growth. The following table exhibits the results of two experiments on different specimens of Crinum.