The Nervous Mechanism of Plants

The intensity of current initiating excitatory impulse IS often extremely feeble ; in Biophytum it may be as little as 0-5 micro-ampere, an intensity of current which cannot be detected even by the highly sensitive tip of the human tongue. Experiment T^. — Two electric connections M'Cie mneie on the left side of an erect stem of Mimosa fudica, Fig. 6. Excitation at kathode-make Km and at anode-break Ab. at points intermediate between the leaves 4 and 6 {tig. 6). A fairly strong current of 6 volts was maintained. On starting the current an excitatory impulse was initiated at the kathode which was above, and leaves 6 and 8 fell

in serial succession. Owing to electrotonic block, to be presently explained, the excitation could not be transmitted downwards. When the current was broken, excitation was induced at the anode below, and the impulse was transmitted downwards, the leaves 4 and 2 falling one after another. Experiment 16. Effect of a stronger current. — ^The E.M.F. was increased to 10 volts. This gave rise to a stronger impulse at kathode-make, which ascended on the left side and caused the leaves to fall in sequence from below upwards. After the impulse had reached the apex, it crossed over to the right side, the direction of propagation becoming reversed from an ascending on the left side to a descending impulse on the right. The effect of moderately strong unilateral stimulation by the polar action of current is thus the same as those by scratch-stimulus, and by induction-shock (Experiment 3).

x\nother method is available for determining the true nature of the transmitted impulse in plants. When a polarising current from a battery is maintained through a length of animal nerve, a transmitted nervous impulse is arrested ; the physiological block persists during the passage of the current, conduction being restored on the cessation of the blocking current. It is of special interest to have thus at our disposal a physiological block which can be put ‘ on ’ or ‘ off ’ in succession. It is obvious that such a block could arrest neither the movement of sap nor a hydro-mechanical disturbance caused by a sudden variation of pressure.

Experiment 17. — In this investigation it is desirable to employ a testing stimulus which can be either maintained constant or increased in a quantitative manner. With the help of a sliding induction-coil it is easy to obtain an intensity of stimulus which is always effective. The proximal of the two exciting electrodes was placed on the main petiole, at a distance of 30 mm. from the primary pulvinus. Halfway between the point of excitation and the pulvinus were placed the two polarising electrodes 5 mm. apart, through which a constant current could be maintained, producing the electrotonic block.

The record of the response of the leaf was obtained by means of the Resonant Recorder described in Chapter VI. was applied at intervals of twenty minutes, by which time the leaf had re-erected itself. The electrotonic block was put ‘ off ’ and ‘ on ’ alternately. The signal below indicates the application of the test-stimulus ; the block was put on at B and B. Fig. 7. Records of transmitted excitation with Xhe record (fig. 7) the block off and on. Arrest of transmitted 4.1 1 4.1

tation was invariably arrested whenever the blocking current was applied, and was restored on the cessation of the current. Inasmuch as the transmission of impulse in the petiole of Mimosa is arrested by an electrotonic block, as is the transmission in an animal nerve, the obvious conclusion is that transmission is essentially the same process in both ; if it be called * nervous ' in the case of the animal, there is equal reason for applying to it the same term in the case of the plant. The excitatory impulse in plants will therefore be designated henceforth as the nervous impulse.

Protoplasmic excitation is induced in plants by the polar action of a constant current. The impulse generated is purely physiological. The following Laws of Polar Excitation in Plants have been established : The polar reactions of the undifferentiated protoplasm of the plant-body are thus identical with those of highly differentiated animal tissues. Excitation in certain sensitive plants is found to occur under a current so feeble that it cannot even be detected by the very sensitive tip of the human tongue.

The normal impulse in the conducting tissue of plants is therefore due not to any hydro-mechanical disturbance but to the propagation of protoplasmic excitation as in the nerve of animals. Weak unilateral polar excitation is conducted only on the stimulated side of the stem; under stimulation with a stronger current, the impulse ascends on the stimulated side, and, after crossing at the apex, descends on the opposite side. The effect produced is similar to that under unilateral scratch-stimulation and stimulation by induction-shock.

The nervous impulse in an animal is arrested by the interposition of an electrotonic block, the conduction being restored on the cessation of the electrotonic current. Similarly, transmission of impulse in the plant can be repeatedly arrested and restored by the alternate application and removal of the electrotonic block. These results conclusively prove that the transmission of excitatory impulse in plants is essentially similar to that of the nervous impulse in animals.

The nervous impulse is itself invisible, and its propagation along the nerve can only be detected by the electric change which accompanies it. The nervous tissue is imbedded in the interior of the plant, and the problem of its definite localisation is surrounded by numerous difficulties. I have, however, been successful in solving it by my Electric Probe. When this is gradually introduced transversely into, say, the petiole, a sensitive galvanometer in circuit with the Probe remains practically quiescent until the tip of the Probe comes in contact with the nerve through which protoplasmic excitation is being transmitted ; the passage of the impulse is then detected by an electric response of galvanometric negativity.

The petiole contains four main vascular bundles shown in the micro-photograph of the transverse section (fig. 8) : E is the epidermis ; C is the cortical tissue ; S is the hollow cylinder of sclerenchyma for ensuring mechanical strength and protection of the conducting tissue in the interior ; F is one of the four vascular bimdles ; O is the central pith. The four vascular bundles converge at the pulvinus, and appear to coalesce into an almost continuous ring.

The following is a tabular statement of micrometric measurements of the distance of the different tissues from the surface and of their thickness. Table III.— Showing Distance from Surface and Thickness of THE Different Tissues in the Petiole Fig. 8. Transverse Section of Petiole of Mimosa showing the four main bundles enclosed in protecting cylinder of sclerenchyraa (two stray bundles at upper corners not shown). E, epidermis; c, cortex; s, sclerenchyma ; ^ f, one of the four main vascular bundles ; o, pith.

The results of the method of the removal of the various tissues are always brought forward in support of the view that the xylem is the conductor of impulse. The tabular statement given above shows how minute are the thicknesses of the different layers ; there is no distance between them, for they are continuous. It is only under microscopic examination that it is at all possible to discriminate where one tissue ends and the other begins. It is therefore practically impossible to remove by hand (without the microscope to guide it) any particular layer, leaving the others uninjured. Again, if conduction persists after removal of the outer phloem, it does not at all follow that the xylem is the transmitting tissue, for the inner phloem would still be left for conduction. The method of removal of various tissues for the determination of the transmitting tissue is not only crude and beset with numerous sources of error, but the inferences drawn from the results contradict each other. Thus while some observers came to the conclusion that the xylem was the transmitting tissue, Haberlandt arrived at tbe definite opinion that the phloem, and not the xylem, was the conducting tissue. The only certain method for localising the conducting tissue is the electrical, and the results of this point definitely to the phloem as being the conductor of excitation.

Objection has been raised to my assertion of the existence of an inner phloem ; it has been said that, unlike the external phloem, the inner tissue consists only of parenchyma-ceUs, and that it contains no tube-ceUs. The above objection is entirely groimdless, as will be seen from what follows, where I show how by means of selective staining it is possible to distinguish two neighbouring systems of tissue having different functions, or to establish the similar functional activities of two tissues which happen to be separated from each other.

The first step is to distinguish the two phloems from the other tissues. It is now recognised that the outer phloem is a conductor of excitation. In order to prove that there is an internal conducting phloem, it is necessary to establish the following points : reactions as does the external; moreover, the reaction of the phloems, both external and internal, should distinguish them from the interposed xylem, and also from other adjacent tissues such as parenchyma of cortex and of pith.

is associated with the presence of tubular cells, if the internal phloem is conducting, it must be characterised by similar tubular cells. the distinction in a striking manner. In order to secure a strong contrast, it is necessary to regulate the duration of application, for too prolonged action causes diffuse staining. By exact timing of the application, hsematoxylin and safranin produce a very marked contrast between the xylem and the phloem. While the interposed xylem is stained red, the two phloems, external and internal, are stained deep violet. The other tissues are but faintly coloured.

In another method the section is first subjected for a short time to the action of haematoxylin, and is then treated for a longer period with Bismark brown. Examination of the transverse section shows that while the xylem, the parenchyma of the cortex, and the pith are coloured brown, the external phloem p and a tissue on the inner side of the xylem, which is in fact the internal phloem p\ are stained a deep violet (c/ fig. g). Thus the staining reaction of the external and internal phloem is not only the

same but is characteristically different from those of the neighbouring tissues. Microscopic examination of the longitudinal section.— In order to determine the anatomical characteristics of the two phloems, a longitudinal section of one of the bundles Fig. 9, Transverse and Longitudinal Sections of a Single Vascular Bundle. Left iigure : Transverse section. The dotted vertical line indicates the passage of the Electric Probe. c, cortex ; s, sclerenchyma ; p, external phloem ; x, xylem ; internal phloem ; o, pith.

Figure to right : Longitudinal section of the bundle, Kole elongated tubular ceils, both in the external and in the internal phloem. (The section passed through one side of tiie bundle and not through the middle.) was made. This is shown under high microscopic magnilication (fig. 9), in which E is the epidermis ; C, parenchyma of the cortex ; S, sheath of protective sclerenchyma ; P, elongated tube-cells of the external phloem ; X, the vessels of the xylem ; P', tube-cells of inner phloem ; and 0, the pith. The tubular cells in the inner phloem are as definite and distinct as those in the outer phloem. Relatively few of

the tubular cells of the outer phloem have perforated septa, le. are sieve-tubes/ the other cells being imperforate, as are also the septa of the tubular cells of the inner phloem. But it must not be assumed that protoplasmic continuity is necessary for conduction of excitation, for I shall explain in the next chapter that excitation is transmitted across synaptic membranes in the animal nerve, and that similar transmission of excitation takes place in the plant across the septa of the conducting cells.

The fact that the inner phloem does actually conduct excitation is proved by my investigation on the localisation of the conducting nerve in the petiole by means of the Electric Probe, previously referred to, which will be fully described in Chapter X. The line of passage of the Probe is indicated by the dotted vertical line in Tig. 9. Galvanometric negativity, due to transmitted excitation, was found to occur only when the Probe came in contact with the phloem ; the cortex, the xylem, and the pith did not show this characteristic reaction. The electric excitation was found to exhibit two maxima, one inside and the other outside the xylem. It was in fact this which led to the unexpected discovery of the two conducting phloem-strands, the bundle being thus bi-collateral.

The second physiological method of demonstrating the two conducting phloems is the, detection of two definite excitatory impulses generated by separate stimulation of the external and internal phloems, fully described in Chapter XVL I may digress, for a moment, to point out that the presence of two conducting phloems in the bundles would appear to afford a satisfactory explanation of certain anomalous results obtained by Pfeffer in his experiments on the effect of an anesthetic, chloroform, on conduction in the petiole. This method is not only crude but also

^ Sieve-tubes would appear to Lave a physiological function other than tiiat of conduction. very uncertain in its action. It has already been pointed out that it is extremely doubtful if external application of chloroform would effectively paralyse the nerve in the interior of the petiole. It is, however, conceivable that after prolonged application a small quantity of the narcotic might, by absorption, get across to the outer phloem ; the Fig. 10. ::Tramverse Section of the Stem. (One half sliown in the fignre.)

X5dem would, however, interpose an additional barrier to the passage of the narcotic to the internal phloem. In such a case there would be paralysis of conduction in the outer phloem, while the inner phloem would still function in conducting excitation. Now it is obvious that, while a superficial mechanical stimulus will initiate an excitatory impulse in the outer phloem, a deep wound or cut is necessary to cause excitation of the inner phloem. The above considerations wiU probably explain Pfeffer’s observation that while the excitation induced by a deep wound-stimulus was always conducted across the superficially narcotised

area, the transmission of the excitation induced by mechanical stimulus applied on the surface was often blocked during transit through the chloroformed region. The anaesthetic reached and paralysed the external phloem, but failed to reach the internal phloem. The supposition that the conducting mechanism in the stem is different from that of the petiole is altogether groundless. Fig. lo shows the circular arrangement of a number of bundles in the stem, each bundle being laterally contiguous to the next. A magnified micro-photograph of one of the main bundles is given in fig. ii. The different tissues are essentially the same as those in the petiole : C is the cortex, S the sclerenchyma, P the external phloem, P' the

internal phloem, and 0 the pith. In the petiole, growth in diameter is completed within a short time, and little trace of a cambium is seen in the section; but in the stem the cambium-layers c are conspicuous. There are two opposite main bundles in the stem which give off lateral branches to the odd and even series of leaves. A vertical section of a young stem in the plane of the leafinsertions (fig. 12) shows this in a very clear manner. After treatment of the section with hsematoxylin and safranin, the inner and outer phloems of each bundle appeared separately as two violet strands. The phloemstrands give off lateral branches to the leaves, thereby assuring conducting continuity between the stem and leaves. An impulse initiated by stimulation of the stem can thus be propagated in a centrifugal direction to the leaves ; and an impulse generated in the leaves can, on the other hand, travel centripetally to the stem, and be then conducted up and down so as to cause the fall of the leaves. It will also be noted that the two main strands of

Longitudinal section of stem bearing leaves on each side. The two ascending bundles ff' give lateral branches to leaves, and meet at apex. The double phloem stained violet stands out against the background. Pulvinus of leaves shaded. conducting phloem converge and meet at the apex of the stem. The establishment of this continuity explains how it is that, under moderately strong unilateral stimulus, the ascending impulse crosses over at the apex and becomes reversed into a descending impulse on the opposite side. (Experiment 3.)

It has been pointed out that the continuity of the conducting phloem explains the unilateral propagation of excitatory impulse along the two main conducting strands. Inspection of fig. 10 makes it also clear that besides these main strands there are intermediate ones. The conducting phloems of the latter are not continuous with, hut contiguous to those of the two main strands. There is thus a ' block ’ or resistance to the spread of excitation in a lateral direction ; hence unilateral excitatory impulse is conducted lengthwise along the line of least resistance and not laterally across the stem.

Experiment 18. Diffuse excitation under stronger stimulus, — ^The resistance to the spread of excitation can however be overcome, as in the animal conducting tissue, by the application of a stronger stimulus. I repeated the experiment with all the specimens in which unilateral propagation of excitation occurred iinder diverse modes of moderate stimulation — by polar action of constant current, by scratch-stimulus and by induction-shocks. In the present series of experiments the stimuli were appropriately increased in intensity, with the result that the excitation, which had hitherto remained unilateral, now became outspread, causing responsive fall of all the leaves on hath sides of the stem.

Nervous impulse induces no visible change in the conducting tissue. The only means of its detection is by the concomitant electrical change of galvanometric negativity. The tissue transmitting excitation in the plant has been definitely localised by means of the Electric Probe. The conducting tissue is the phloem. Microscopic examination shows that the four main bundles of the petiole are separate from each other. They converge at the pulvinus and form an almost continuous ring.