Comparative Electro-Physiology: A Physico-Physiological Study

Returning now to our original question, we have first to determine whether excitation causes any motile effect in nerve. Under observation, it is easily seen that when the nerve is excited by tetanic electrical shocks it increases in thickness and at the same time shortens in length. We have here a phenomenon in every way analogous to the thickening and shortening of muscle under excitation. The contraction which occurs in nerve, moreover, is of an order by no means microscopic. I give here a record (fig. 313) of the contractile response of nerve under continuous stimulation by fairly strong tetanising electric shocks. This record was obtained by means of the ordinary lever-recor@er, the magnification employed being only three times. The induced contraction in this particular case was about 14 per cent. It will also be seen that this contraction reached a limit, at which state of maximum contraction the nerve remained for a considerable time. After this we observe a tendency

to decline, owing to fatigue. In some other cases, moreover, I have obtained a contraction of as much as 20 per cent. If we wish to obtain a series of successive responses, however, it is desirable to avoid over-stimulation of the tissue. In order, then, to obtain a response-record under moderate stimulation, we have to employ a higher magni- fication.. This magnification, if made about 200 times, is more than sufficient for all practical purposes, and the photo- graphic records given in the course of the present chapter are of this order. With long ‘specimens of nerve, however, a magnification of fifty times would be enough, and in the

Fic. 313. Record of Contractile Response in Frog’s Nerve under Continuous Electric Tetanisation. course of the next chapters, I shall give certain records on this scale, obtained directly on a smoked glass surface. The apparatus used for the purpose was the Kunchangraph (Sanskrit, Aunchan, contraction), which I had already devised afd employed in recording the contractile responses of plant- tissues. This apparatus, as adapted for the purpose of recording mechanical response in nerves, consists of, first, a nerve-chamber, N; secondly, a modified Optical Lever, 0 ; and thirdly, a photographic recorder, D (fig. 314).

_ Of these, the nerve-chamber consists of a small rectangular ebonite box, the front of which is closed by a semi-cylindrical N, nerve chamber containing nerve with electrical connections, E E’, FIG, 314. Optical Kunchangraph for Record of Mechanical Response of Nerve Thread tied to lower end of nerve, and attached to short arm of optic lever, O. Beam of light from L reflected from mirror of optical lever, oO, falls on recording-drum, D. Adjustment of reflected spot of light made by micrometer screw, Ss. Periodic electric stimulation at intervals of one minute is automatically made by means of key regulated by clock-work. Air bubbles through water at w, and is led on by india-rubber tubing, T, to nerve-chamber, thus kept humid. By proper manipulation of stop-cock any vapour—as chl oroform—con- tained in vessel V, may be passed through nerve-chamb er, subsequent responses showing effect.

glass cover. The nerve is placed vertically within this, and held, at its upper end, by a clamp. The lower end of the nerve is connected with the short arm of the Optical Lever by means of a thread, which passes through a hole in the floor of the chamber. A second thread of cotton moistened with saline solution hangs loosely from the end of the nerve, and is connected with the electrode E’.. When the electrodes E and E’ are put in connection with the secondary of an induction coil R, the entire length of the nerve is subjected to. direct excitation. When, on the other hand, we wish to study the effect of transmitted excitation, the nerve is lightly clamped at B (fig. 315). Excitation is then induced in the portion of the nerve A a, and after transmission through the inter- vening tract, causes the motile effect in the responding portion of the nerve B C, :

One precaution which I find to be very necessary is the maintenance of the properly humid condition in the i ._,_nerve-chamber. This is_ specially Sai keeping the chamber moist, by a large quantity of blotting-paper soaked in water, is not sufficient to bring about the maintenance of the normal excitability of the nerve for any length of time. This need was met by keeping moist vapour in uniform circulation through the nerve-chamber. An air-bag is kept under suitable pressure, and the air, bubbling through water in the vessel W, is made to enter the nerve-chamber through an entrance-pipe, and to escape by an exit-pipe. In warm weather it is well to keep fragments of ice in the water-vessel. By proper mani- pulation of the stop-cock of the air-bag, a gentle stream of cooled and humid air is kept in constant circulation through

the chamber. Observing these precautions, I have been able to obtain responses from a given nerve for as much as three hours continuously, whereas, without this care, they would have come to a stop in a very short time. By a modification of this arrangement, we are also enabled to study the effect on the excitability of the nerve of various gases and vapours contained in a second vessel, v. A series of responses is first taken, under normal conditions—that is to say, when the nerve is surrounded simply by a moist atmosphere. On now turning a three-way tap in a given direction, the water-vapour can be made to: pass through the vessel V, filled with the given gas or vapour, before reaching the nerve-chamber. The series of responses then obtained will show either the immediate or the after-effect of the reagent at will. For it is easy, by means of the three-way cock, to shut off the gas and re-establish the first or normal condition, after which the responses will afford an indication of the nature of the after-effect.

_ The lower end of the nerve, as has been said, is attached to the arm of the lever which passes through the fulcrum-rod. A light .mirror is fixed on the fulcrum-rod, its face being downwards. The pull caused by the excitatory contraction of the nerve causes rotation of the fulcrum-rod, and this in turn gives rise to a deflection of the spot of light reflected from the mirror. A responsive relaxation of the nerve would give rise, on the other hand, to a deflection of the spot of light in the opposite direction. The long arm of the lever, it will be noticed, is here the ray of light. The responsive movement of the spot of light is recorded on a moving photographic plate vertically below the mirror, and whose movement, regulated by clockwork, is in a direction at right angles to that of the spot of light. The photographic plate, or the film wrapped round the drum, moves under a fixed wooden cover, not shown in the figure, which is provided with a narrow incised slit. The length of this is parallel to the direction of the movement of light, and at right angles to that of the plate or film. The advantage of having the plate

vertically below the mirror lies in the fact that a lighted candle may be placed in the dark room without spoiling the record by diffuse illumination. The only way in which such diffuse light could now find access to the plate would be by reflection from the ceiling. But if the ceiling of the experimental room is blackened, or a black cover placed over the nerve-chamber at a certain height, even this possibility is eliminated. The advantage which the observer enjoys, when, instead of groping in semi-darkness, he can work in a fairly well-lighted room is obvious. By making the arm of the lever to which the nerve is attached sufficiently short, and by placing the recording plate sufficiently far away, a wide range of magnification, from several hundreds to several thousands, may be obtained. It may sometimes be desirable to subject the nerve to a certain amount of tension, and this is secured by placing a small weight on the arm of the lever. With high magnification, due adjustment, which is very troublesome, lies in bringing the spot of light con- veniently over the recording plate. This difficulty is obviated, however, by means of a fine micrometer screw S which moves the whole nerve-chamber up or down, in relation ‘to the Optical Lever. The adjustment of this screw in a right- handed manner then moves the spot of light in one direction, say to the left, while its left-handed rotation moves it to the right. This movement can be made very fine, and the spot adjusted to any part of the photographic field.

It remains to deal with the possible disturbances inci- dental to the high magnification employed. Apprehension, in this matter, is often more fanciful than real. Disturbances might no doubt occur, however, when proper conditions are not secured for the experiment. If the nerve-chamber, for example, be supported on a different stand from that of the Optical Lever, then the slightest tremor of the common pedestal would result in relative movements of the two supports, causing constant disturbance of the spot of light. Under these conditions, heavy stone pedestals, erected on steady foundations, afford no security against the ground-

vibrations of a busy city. But when both the nerve- chamber and the Optical Lever are fixed to the same supporting-rod, relative movements, due to external disturb- ance, are practically eliminated. This common supporting- rod may be screwed securely to a wall. With these precautions, I have been able to take records, without the least dis- turbance from the adjacent electric tram line. Asa matter of fact, when the magnification required is only of a few hundred times, nothing but gross carelessness could allow any source of disturbance to remain. It is only when the magnification has to be pushed to the order of a hundred thousand that unusual care is necessary to avoid errors of disturbance. One precaution which should, however, be taken, is that arising from disturbance of the mirror by convection currents of air. The remedy for this is obvious, namely; a suitable glass cover.

This is the order of magnification which is necessary for the recording of response under a degree of stimulation - usual in making observations of excitatory electrical variation with a very sensitive galvanometer. But while the sensitive- ness of the galvanometric method of detecting response is here nearing its limit, that of the mechanical method is in its first stage only, and how greatly the sensitiveness of the latter may be exalted when required will be shown in the next chapter.

I shall: ‘flow “explain how easy it is to study the aiisin: logical variations induced in the animal nerve under various agencies by means of the mechanical response. The following experiments were performed on specimens of the sciatic nerve of frog. A well-known reagent for abolition of ex- citability of the nerve is ammonia. Its effect on mechanical response is seen in fig. 316. In all the following experiments, the stimulus applied was by fairly strong tetanising electrical shocks, which were usually of two seconds’ duration. Two series of records were taken, successive responses being recorded at intervals of one minute, before and after the

application of the chemical reagent. In fig. 316, the normal responses seen in the first series are found to be abolished when the nerve has been subjected to strong vapour of ammonia for some time. It should be mentioned here that this abolition takes place under the action of a strong dose. When highly diluted with air, the vapour ey cause a temporary exaltation. In the next figure (fig. 317) is shown the effect of szorphia. After the application of this solution for a certain length of time, the response is seen to ‘be abolished. The strength of application which brings about this abolition I find to vary according to the condi-

Fic. 316. Photographic Record of Effect of Ammonia. on Mechanical Response of Frog’s Nerve FiG.. “317: Photographic First series of responses are nor- Record showing Abolition mal. Second series show effect of Mechanical Response of ammonia in practical aboli- of Frog’s Nerve by Action tion of response. of Solution of Morphia tion of the nerve. Another agent by which the mechanical response of the nerve is found to be abolished is aconite. And it is of special interest to note that I have often found this to act as an antidote, for the revival of response previously almost completely abolished by morphia. ‘The condition of the nerve here also appears to be a determining factor in the mutually antidotal action of these two poisons. A strong application of alcohol after long-continued action

abolishes the response of nerve. But its preliminary effect is often one of exaltation, as seen in fig. 318. I shall next describe the effect of chloroform, which dis- plays many interesting features. We have seen that when a tissue is excited by impinging stimulus, two opposite effects are induced : one of these is the increase of energy, by the absorption of stimulus, and the other is the expenditure of energy by excitatory response. The former, as we have seen, finds expression in galvanometric positivity and expansion. The latter, on the other hand, is exhibited: as galvanometric negativity and contrac- tion. In the record of excitatory response, the former of these elements is generally masked by the predominant negative or contractile effect. We have also seen that this hidden positive may be unmasked. in either of two: ways: first, by retarding the expression of one effect in relation to the other ; or, second, by abolishing the excitatory negative altogether. In the first of these cases, the negative response is converted into diphasic, say positive followed by Fic. 318. Photographic

: Record showing Pre- negative. In the latter, the response limitiary Bimaltation it becomes positive, by the suppression Mechanical Response : . of Frog’s Nerve after of the negative. An example of this Application of Alcohol unmasking of the positive element, by suppression of the negative, we have already seen to occur under the action of chloroform (cf. fig. 49). This demon- stration was made on a vegetable tissue, the test employed being electrical.

The experiment which I am now about to describe is interesting from the fact that effects parallel to those there seen in a vegetable tissue are in it shown to occur also in the highly specialised animal nerve. The unmasked ‘electro-positive effect, moreover, is here seen to correspond with an expansive response of the tissue. A record of the various phases in the effect of chloroform on the mechanical response of nerve is found in fig. 319. It will be seen here that the first effect of chloroform was to cause a great enhancement of ex- citability, which in this case lasted for about a quarter of an hour. I have given only two responses of this series. After this, the responses began to decline, and another very

Fic. 319. Photographic Record showing Effect of Chloroform on Mechanical Response of Frog’s Nerve First pair of responses, normal; second pair, preliminary exaltation on application of chloroform; last series exhibit subsequent effect of chloroform in unmasking the positive component as diphasic response. Expansion is here followed by contraction. Note regular waning of both components with growing aneesthetisation. interesting reaction made its appearance. The impinging stimulus had hitherto induced only an immediate contractile response. But by the action of the chloroform the excitatory effect was delayed, and the positive, or mechanically ex- pansive response was unmasked in the form of a preliminary downward twitch. Response was now, therefore, diphasic— positive followed by negative. Immediately on the applica-

tion of stimulus, as may be seen from the record, there is a sudden expansive movement downwards, followed by an equally rapid reversed movement of contraction upwards, and this followed again by a slow recovery. Each of the successive stimuli evokes the same diphasic responsive sequence. It must be noted that the downward twitches are the preliminary, and not the after-affect. It is also interesting to note, as the tissue approaches death, under the continued action of chloroform, how regularly in both negative and positive directions the responses decrease in amplitude.

We shall next undertake an independent investigation into the causes which bring about the three types of response —abnormal positive, diphasic, and normal negative—known to be exhibited in the electrical response of the animal, and already demonstrated as occurring also in that of the vege- tal nerve. While discussing these three types of electrical response and their variations in Chapter XXXI. it was stated that the differences of effect involved were due to changes in conductivity and excitability, brought. about by varying tonic conditions. It was also explained, in the same place, that the continued isolation of so highly excitable a tissue as nerve, from its accustomed supply of energy, would be sufficient of itself to depress its tonic condition below par, with concomitant depression of its conductivity and excitability. The result of this depression of excitability will be to render inefficient a stimulus which was formerly efficient, to evoke the true excitatory reaction of galvano- metric negativity. The absorbed stimulus will now induce only a responsive positivity. The depression of conductivity also would cause the transmission: of the hydro-positive, instead of the excitatory negative, wave. Owing, then, to the joint action of these two factors, stimulus induces a positive response—the so-called ‘abnormal ’—at a distant responding point, when the tonic condition of the tissue has become depressed. Absorption of stimulus, however, by supplying the requisite energy, raises the tonic condition, with con- sequent restoration of conductivity and excitability. As

a result of this, the abnormal positive will pass into normal negative response, through an intermediate diphasic, after the impact of a series of stimuli, or after tetanisation. The enhancement of conductivity and excitability thus conferred on the tissue by the absorbed stimulus will now act by still further tetanisation, to bring about the enhance- ment of the normal negative response. Starting thus, with the most depressed condition of the tissue, and sub-. jecting it to continuous action of stimulus, we obtain four typical stages : (1) the abnormal, passing after short tetanisa- tion into (2) the diphasic ; this in its turn giving place to (3) the normal negative alone ; which finally becomes (4) the enhanced negative. |

In studying electrical response, both of animal and vege- tal nerves, under appropriate experimental conditions, we have already seen various examples of these different types of response and their transformations. But under such modes of experiment as have been described, the effects were, as already stated, due to joint changes in excitability and con- ductivity. I shall now, however, describe a still simpler experimental arrangement, in which the stimulus is applied directly on the tissue, and the responsive variations are, therefore, due to variations in the excitability alone. These changes, moreover, will be recorded by means of their direct mechanical expression, namely, contraction, or its opposite expansion.

With regard to abnormal response, I have already stated that this is brought about, not by ‘staleness,’ or the moribund condition, with its concomitant chemical changes, but by the run-down of the energy of the tissue in isolation. On investigating this subject, by means of mechanical response, with its superior sensitiveness, this conclusion finds inde- pendent support of the strongest character. On taking even the freshest specimen, I generally find that its responses at first are the abnormal positive. These gradually pass into moderate negative through diphasic. This is due to the raising of the tonic condition by the absorption of

the stimulus, and after a series of stimulations, the isolated tissue, which was originally depressed, has its tonic condition so much heightened, that the responses are enhanced to an unprecedented magnitude. A specimen, in fact, which was at first almost irresponsive, may generally be brought to any state of exalted excitability desired, with concomitant increase in amplitude of response, by merely subjecting it for a certain length of time to the action of impinging stimulus.

Fic. 320. Photographic Record showing Abnormal Positive converted into Negative Response after Tetanisation First series, abnormal positive ; second series, persistence of this positive after very brief tetanisation ; third series, conversion to negative, after a tetanisation of longer duration. I shall now describe in detail some of the principal experiments, Selecting a specimen of frog’s nerve, I took a series of responses to electrical shocks, of three seconds’ duration, at intervals, in each case, of one minute. The testing stimulus was kept always the same throughout the experiment, except for certain intervening periods of tetani- sation. The variations seen in the responses thus give a visual demonstration of the variations in excitability. The record of these is given in fig. 320. The responses in

the first series are by the abnormal positive variation; that is to say, by expansion. The tissue here being sub-tonic, the impinging stimulus could not induce the true excitatory effect. The tissue. was now subjected to short-lived tetanisation. But the absorbed stimulus was not yet sufficient to induce the normal responsiveness. The next series of. records, = therefore, still exhibited the abnormal positive response. Tetanic shocks of longer duration were next applied. This gave rise to a short- lived positive twitch down- wards, succeeded by large contractile response upwards, After the cessation of the second tetanisation, the absorbed energy is seen to have brought the tissue to a condition of more or less normal responsiveness. This is seen in the third series, where the first responses are diphasic, but the positive component (the downward twitch) becomes perceptibly

Fic. 321. Photographic Records show- smaller and the negative ing Gradual Disappearance of Positive larger, in each of the succeed- Element in Diphasic Mechanical . Tt. wheal Responses of Frog’s Nerve and INS responses. shou Note also the staircase increase. recovery from positive ts much quicker than trom negative response. This fact is important, in connection with certain psycho-physiological phenomena to be described in a later chapter. The effect of successive stimuli, in enhancing normal response, when the nerve is not yet in maximum. tonic condition is illustrated in a still more striking manner in the record given in fig. 321, obtained with a different specimen

of frog’s nerve. In this, also, the first series of responses was purely positive. But the record shown here begins at the point where, in consequence of previous tetanisation, response has become diphasic. Here it will be noticed that the true excitatory effect of contraction is undergoing a con- tinuous increase, while the abnormal positive is decreasing. The excitatory response, indeed, becomes so great as to be incapable of record within the plate. _

I have already shown how similar in every respec are the responsive characteristics of the vegetal nerve to those of the animal. This fact finds an interesting illus- tration in the various phases of its mechanical response. That is to say, plant nerve in a sub-tonic condition gives positive, passing into diphasic and normal negative response, under tetanisation. On arriving at this second stage of diphasic response, successive responses undergo enhance- ment in a manner precisely the same as holds good in the corresponding cases with frog’s nerve. This is sufficiently illustrated in the two records given side by side in fig. 321, the first of which, as already said, is of frog’s nerve, and the second, of nerve of fern. We see here again, as already in numerous cases before, how the responsive pecu- liarities and their modifications in the one are in every respect paralleled by those of the other. The only differ-— ence between them lies in the degree of their excitability, that is to say, two stimuli of equal intensity will in general induce a more intense excitatory effect in the nerve of frog than in that of fern; or in order to obtain from both an equal intensity of response, we must, in the case of fern, employ. a stronger stimulus. We have seen that in con- sequence of the absorption of stimulus, not only does the abnormal positive phase disappear, giving place to the normal negative, but the subsequent negative responses themselves also show an enhancement in a staircase manner. I give here (fig. 322) another record showing the mechanical response of frog’s nerve to undergo this staircase enhance- ment. From this effect then it is easy to understand that an