Comparative Electro-Physiology: A Physico-Physiological Study

We thus see that by the employment of this method the only change induced at the two electrodes will be the excitatory change, the physical polarisation-factor being eliminated. Thus, on subjecting two points, A and B, to equal stimulation, the induced galvanometric negativity, at both the points, will be equal, if the natural excitabilities of . the two have been the same. But if the tissue be anisotropic, and the natural excitability of one point, say b, greater than that of A, then we shail obtain a resultant responsive current, which will flow in the tissue from the more excited B to the less excited A, the induced galvanometric negativity of B being relatively the greater. We have here what is merely a repetition, by electrical stimulation, of the results which

were described in Chapter IX., as obtained by thermal and mechanical stimulus. How perfect and how consistent, due precautions being taken, these results may be rendered, will be seen from the numerous records in this and the following chapters. I have postponed till now the consideration of the mode of application of these alternating shocks. The usual alter- nating current from a Ruhmkorff’s coil would be entirely unsuitable for delicate and crucial experiments ; first, because the excitatory values of the slow make- and quick break-shocks are unequal ; and, secondly, because such currents leave their residual polarisation effects.

These defects I have been able, as stated in the last chapter to avoid, by quick reversals of the primary current which actuates an induction-coil. When the primary current is reversed from the js/us to the mznus direction, we obtain an induction-current due to magnetic varia- tions of lines of force, from, say, plus n to minus n. When the primary current is re-reversed, from mznus to plus, we obtain an opposite induction-current, due to magnetic variation, from mznus n to plus n. It will be seen that if these reversals of the primary current are made with equal rapidity, the alternating induced currents will be equal and opposite. The reversals are accomplished by means of a Pohl’s commutator, worked up and down by a crank, in connection with an electric revolving motor (fig. 170). The intensity of the induction-shock may be varied by sliding the secondary nearer to, or further away from, the primary. Having now described the general means of producing equi- alternating electric shocks, it still remains to explain two distinct methods of applying them, for the determination of the differential excitability of the tissue. These may be distinguished as: (1) the Method of the AFTER-EFFECT ; and (2) the Method of DirRECT-EFFECT. According to the first of these—the method of the after-effect—the tissue is excited for a definite length of time, and the excitatory effect observed, by connecting it immediately afterwards with the

galvanometer-circuit. The manner in which this is done will be understood from fig. 170. We have a highly insulating electrical key, K, of ebonite. P and Q are connected with two points A and B of the tissue, whose relative excitabilities are to be determined. A spiral spring keeps the key down, con- necting the two points in the specimen with the galvanometer. Any existing difference of potential, as between the two points Fic. 170. Experimental Arrangement for Determination of Excitatory After-effect of Equi-alternating Electrical Shocks

M, electrical moter working Pohl’s commutator for alternate reversal of current in primary, P. Note that the connecting-rod, A, works simply up and down, causing reversals of current ; $s, secondary coil; k, ebonite key kept down by elastic spring, the two surfaces of the specimen being thus in circuit with galvanometer, G. When key is pressed, these are put in circuit with exciting coil, s. When key is released, after-effect of excitation on specimen exhibited by galvano- meter deflection. C, the compensator.

is balanced by the compensating potentiometer c. Under these circumstances, the galvanometer spot of light would remain steady, whether the key was up ordown. By pressing the key K, the galvanometer-circuit is broken, and the tissue is put in series with the exciting circuit S, which forms the secondary of the induction-coil. When the key is again released, the galvanometer-circuit is rapidly made, at a definite short interval after the cessation of the exciting shock, and the resulting deflection of the galvanometer indicates the differ-

ential excitation as between A and B. In this way records may be taken af a series of the after-effects of brief excitations, at intervals of, say,a minute. The direction of this responsive current, which in the tissue is from the more to the less excitable, enables us to determine the relative excitabilities of the two points, A and B. Much more delicate is the second method, that, namely, which depends on the record of the Direct-Effect of equi- alternating shocks; but for its perfect working, certain difficulties have to be overcome. One of the first conditions to be fulfilled lies in the perfect equality of the alternating shocks. The importance of this will be understood on observing the effect of the alternating shocks given by a Ruhmkorff’s coil, when actuated by a vibrating hammer. Here, the make- and break-shocks are of unequal intensity and duration, and the following sources of disturbance come into play: (1) a galvanometric drift in one direction or the other ; (2) a resultant inequality of polarisation-effects ; and (3) the inequality of the excitatory values of the two shocks.

The galvanometer-drift, owing to the inequality of the induction shocks, becomes very troublesome, when we have to employ, as is necessary, an instrument of high sensibility. If the differential excitability of the specimen be very great, this drift may be masked by the predominant excitatory effect. In other cases, however, the excitatory effect itself may be overpowered by the drift. The difference of intensity as between the make- and break-shocks in the Ruhmkorff’s coil described, thus becomes a strongly dis- turbing element. The necessity to make the two shocks absolutely equal will be understood when we find that alternating telephonic currents, which are generally re- garded as equal and opposite, induce a drift of the galvanometer in one direction or the other, on account of a slight difference of intensity between the two alternating currents. 4

The difficulty arising from inequality of polarisation- effects is tao obvious to require further elucidation. Still more important, moreover, is the disturbing factor last mentioned, that of the inequality of excjtatory values as between the make- and break-shocks themselves. This element of uncertainty is very clearly seen in the experi- ments of Von Fleischl on the response of nerve. The resulting deflection he found to be in the direction of the - break-shock. The explanation of this phenomenon has hitherto been regarded as a matter of great difficulty, authorities being much divided on the subject. In these experiments, alternating currents from a Ruhmkorff’s coil are sent through the galvanometer and the nerve, in series. The two points on the nerve, A and B, are presumably of equal excitability. A make-shock of relatively lower E.M.F. passes, say, from A to B, followed by a break-shock of higher E.M.F..in the opposite direction, from B to A. Confining our attention to the excitatory effects of these shocks, we have at A, during make and break, the following four effects : (a) feeble anode-make; (4) feeble anode-break ; (c) strong kathode-make ; and (d@) strong kathode-break. At B, on the other hand, at the same time, we shall have: (a’) feeble kathode-make ; (6’) feeble kathode-break ; (c’) strong anode- make ; and (@’) strong anode-break. Now, since the excitatory value of anode-break is probably the stronger and more persistent, and since the intensity of this effect will depend, within certain limits, on the intensity of the anodic shock, it follows that the (@’) or strong anode-break effect at B will, as a general rule, be the most conspicuous of these. That is to say, as a result of all these excitatory effects in combination, greater galvanometric negativity will be induced at B than at A, the responsive current being thus from B->A, in the same direction as the break shock, which was the actual result.

In any case, whatever may have been the cause of this, it is clear that the employment of such unequally exciting shocks of make and break would be fatal to any attempt to determine accurately the natural difference of excitability as between the two points. I may state here, that when I have employed absolutely equal alternating shocks on a specimen of nerve, I have obtained no resultant deflection whatever, showing that such shocks induce exactly equal excitations in an isotropic tissue. But if the excitability of one of the two points be first abolished by killing, then a definite resultant responsive current is obtained, from the excitable living to the inexcitable dead. So perfect were in fact the results secured by means of these equi-alternating electric shocks, that I was desirous not only to detect, but also to record photographically, the responses thus obtained. In this a certain difficulty is experienced, inasmuch as the alternating shocks are apt to render the recording spot of light tremulous, and thus to spoil the photographic im- pression. This may, however, be overcome by making the alternation frequency so high, in reference to the period of the needle or suspended coil of the galvanometer, that the unsteadiness of the deflection ceases.

I shall now describe the practical means employed to obtain equi-alternating shocks of any frequency that may be desired. This I have been able to do in several ways, and, among others, by using a Rotating Reverser. This consists of an ebonite disc, on the periphery of which there are strips of metal of equal breadth, and separated from each other by equal distances. The odd strips (1, 3, 5, and so on) are connected together and led to a metallic ring on the left of the disc. The same is done with the even strips, which are led to the right. The two electrodes of a battery are led through a key, K, to these two metallic rings and are con- nected with them by means of brushes. Thus one ring, with all the odd strips, is connected, say, with the positive, and the other, with all the even strips, with the negative pole of the battery. The current is led off by a second pair of brushes, placed diametrically opposite to each other on the disc, in the primary circuit, P, of an induction coil (fig. 171). Let us suppose the upper brush to be connected with an odd strip, the lower will then be connected with an even. The current in the primary coil now flows in one direction. When the disc is rotated, so as to bring the next pair of strips in

contact with the brushes, the upper will then be connected with the even strip and the lower with the odd. Thus the direction of the current will be reversed, and rapid rotation of the disc will give rise to equi-alternating currents in the primary of the induction coil. This will in turn induce equi-alternating induction currents in the secondary, the intensity of which can, as already said, be varied within wide limits by appropriate changes of distance between the primary and the secondary. The number of strips in the apparatus used is fifty, and when the disc is rotated, by

Fic. 171. Method of Direct Effect of Excitation by Equi- alternating Shocks R, rotating reverser, in circuit with primary coil, P. Duration of stimula- tion determined by metronome, M._ 8, secondary coil in series with ' specimen and galvanometer. means of an electrical motor, at a rate of one revolution per second, there will be fifty alternations of current in a second. The duration of the application of the stimulating shock to the tissue is regulated by a metronome, which completes the primary circuit for a definite short length of time. When the metronome, M, is so adjusted as to complete the circuit for *5 second, then a stimulus of that duration will be imparted at each stroke. A second interrupting key, not shown in the figure, is included in the circuit. When this key is closed, a single beat of the metronome gives a stimu- lating shock of *5 second’s duration. ‘The key is now opened

for one minute for recovery. In this way, records of response and recovery are obtained, at intervals, say, of one minute. Another very effective means of producing equi-alter- nating shocks is by the employment of an alternating- current dynamo, driven by an electrical motor, M (fig. 172). The alternating current is led to the primary of a Ruhm- korffs coil in the usual manner. The motor is driven by an electrical supply from the street mains, its speed being adjusted by a regulation of the current, which is effected by

M, motor, rotating armature of alternating-current dynamo, D; R, liquid rheostat, in circuit with street-mains, for regulating speed of rotation of motor; Pp’, idle coil; P, primary coil; I, resonating index; s, secondary coil, in series with specimen and galvanometer. Duration or excitation determined by pressure of key, K. an electrolytic rheostat, R. As the dynamo is provided with a permanent horse-shoe magnet, the intensity of the alter- nating current is determined by the speed of rotation of its armature. If the speed be kept always constant, the number of alternations will also be constant, and the ex- citing value of the electric shocks will depend simply upon the distance of the primary from the secondary. It is thus possible day after day to use the same intensity of stimu- lation, and thus to compare the relative excitabilities of

different tissues. The constancy of the speed of rotation of the alternating-current dynamo is secured by means of the resonating index, I. This consists of a short steel spring with a long index. When the frequency of alternation is the same as the natural period of vibration of the spring, the resonator is thrown into strong sympathetic vibration. At first the rheostatic resistance, which determines the speed of the motor, is made slightly too large. The movable plate is now gradually brought nearer, till the proper speed has been arrived at, and this point is at once indicated by the induced vibration of the resonator. | .

A further difficulty has to be overcome in the main- tenance of the uniformity of speed. When the open circuit of the alternating dynamo is closed, by the interposition of the primary of the Ruhmkorff’s coil, the speed undergoes a sudden diminution, owing to the work which the dynamo has now to perform. In order to avoid this fluctuation, then, the dynamo circuit is kept closed by means of an idle primary coil, P’, which is a duplicate of the primary, P, of the Ruhmkorff’s coil. When the key, K, is pressed, the alter- nating current is transferred from P’ to P. There is thus no fluctuation in the speed of the dynamo, and the duration of the closure determines that of the stimulus. I may mention here, that instead of employing a separate motor to drive the alternating-current dynamo, I have sometimes used, with equal success, a motor transformer, giving rise to alter- nating currents. It is easy to construct a very compact and portable form of this latter apparatus.

In this manner we may apply uniform stimuli of equi- alternating shocks at regular intervals of time, say of one minute. The usual preliminary test of the successful elimination of all sources of disturbance may here be made in the following way. The kaolin ends of the non-polaris- able electrodes are connected with each other, without the interposition of a specimen, and alternating shocks from the secondary are passed through the circuit. These should give rise to no deflection in the galvanometer. It may be

said here that I use a D’Arsonval type of galvanometer, in which, instead of a suspended needle, we have a suspended coil. There is thus here not even the remote contingency of disturbance which might arise from the demagnetisation of the magnetic needle. Having thus tested, by null action, the symmetry of the electrodes and the galvanometer, the differentially excitable tissue, say the sheathing petiole of Musa, is interposed, with its concave and more excitable surface upwards. On now: applying excitation by equi- alternating shocks, the responsive current will be found to flow downwards, from concave to convex, giving a deflection of the galvanometer, say to the right. And this deflection will continue to be to the right, even if the battery current (fig. 171) be reversed by means of key K. The direction of the excitatory current, moreover, depending solely, as it does, on the relative excitabilities of the two surfaces of the specimen, will remain constant, even if the connections with the secondary coil, S, be reversed. The zinc rod, N, of the non-polarisable electrode in connection with the concave surface (fig. 172) has thus, up to the present, shown induced galvanometric negativity, the galvanometric deflection being to the right. But if we exchange the zinc rods of the non- polarisable electrodes, it will then be N’ which will be con- nected with the more excitable concave surface, and it will now be this electrode N’ which will show galvanometric nega- tivity. This reversal of the galvanometer deflection with the reversal of the electrodes affords additional confirmation of the greater excitability of the concave surface of the specimen of Musa. |

In these experiments the existing current of rest may be balanced previously by a potentiometer. But this is not absolutely necessary. I give below a series of records obtained with a specimen of the sheathing petiole of Musa (fig. 173), in which we know the inner or concave surface to be more excitable than the outer or convex. The responsive current is seen under this form of electrical stimulus, as we found to be the case under mechanical and thermal stimulation, to flow from the more excitable concave to the less excitable convex. In order next to demonstrate the physiological character of these responses, I subjected the tissue to the action of chloroform, and the record in the second part of the figure shows the consequent Bepresion of the response.

The great delicacy and pliability of this mode of applica- tion of stimulus enable us to attack many difficult problems, on the difference of excit- ability between two points in a tissue, with perfect ease. To how many distinct in- vestigations it can be suc- cessfully applied will be set forth in detail in succeeding chapters. As there is nothing to prevent the two exploring . electrodes from being applied on any two points, however distant, of the same organism,

‘Fic. 173. Photographic Record of Re- sponse of Petiole of usa to Equi- alternating Electric Shocks, before and after Application of Chloroform. it is seen that we have here a means of determining, not only the differential excit- ability of any two points of the same organ, but also that of any two organs of the same specimen. For the present I shall, however, content myself with giving a few instances only in illustration of the ex- treme delicacy of this method in detecting physiological differences as between two points.

We shall first turn our. attention to those physiological modifications which are due to the a-symmetrical action of the environment on the organism, and here we shall select the case of the plagiotropic stem of Cucurbita. We have seen that in the recumbent stem of this plant the tissue of the upper side is rendered relatively fatigued by the con- tinuous action of sunlight, and thus becomes permanently less excitable than the lower side. We have also found that

while the natural resting-current was from the less excitable upper to the more excitable lower side, the responsive cur- rent under mechanical stimulation was in the opposite direc- tion—namely, from the lower to the upper (p. 112). Using now the electrical form of stimulus, we obtain results which are identical. Fig. 174 gives a series of such responses under equi-alternating electrical shocks. Curiously enough, as pointed out in the last chapter, I have detected a similar plagiotropy in the case of the eel. The head of the fish was cut off, and voluntary action thus eliminated ; electrical connections were then made, after a period of rest, with the dark dorsal upper surface, and the colourless skin of the ventral or lower. A natural current was now found to flow from the upper

of Responses of Plagiotropic Fic. 175. Electrical Responses of Stem of Cucurbita to Equi- Eel to Equi-alternating Electrical alternating Electric Shocks Shocks Direction of responsive current from Current of response from ventral ventral to dorsal surface. surface to dorsal. surface to the lower, as in the case of the plagiotropic stem of Cucurbita. Electrical excitation was now applied, and the result was a responsive current from the more excitable lower to the less excitable upper surface again, as in the case of Cucurbita. In fig. 175 is seen a series of records in illustration of this.

Another investigation which I thought might be interest- ing had reference to the variegated colouring of certain foliage leaves, A striking example of this is found in the tropical elephant creeper (Pothos), the rich green of which is barred by longitudinal streaks of milk-white. This dis- tribution of colour is found even in the youngest and most vigorous leaves. The question whether such colouring was accidental or associated with physiological differences could, I thought, be determined by the delicate mode of investiga- tion which was now at my disposal. On making electrical . connections, then, with the green and white portions of a leaf, I found that the natural current of rest was from white to green through the tissue, and, on further testing the differ- ential excitability in the usual manner, the responsive current was observed to flow from the green to the white. This showed that pallidity was here associated with a depressed physiological condition, |

Currents of rest and action—Currents in animal skin—Theories regarding these —Response of vegetal skin—Stimulation by Rotary Mechanical Stimula- tor—Response of intact human skin—Isolated responses of upper and lower surfaces of specimens—Resultant response brought about by differ- ential excitability of the two surfaces—Differences of excitability between two surfaces accounted for— Response of animal and vegetal skins not essentially different—General formula for all types of response of skin— Response of skin to different forms of stimulation gives similar results— Response to equi-alternating electric shocks : (1) Method of the After Effect ; (2) Method of Direct Effect—Response of grape skin—Similar response of frog’s skin—Phasic variation of current of rest induced as result of successive stimulation in (a) grape skin ; (4) frog’s skin ; (c) pulvinus of Mimosa—Phasic variation in autonomous mechanical response of Des- modium gyrans—Autonomous variation of current of rest—True current of rest in skin from outer to inner—This may be reversed as an excitatory after effect of preparation—Electrical response of skin of neck of tortoise — Electrical response of skin of tomato —Normal response and positive after- effect — Response of skin of gecko—Explanation of abnormal response,

IN this and the next few chapters it is my intention to make an inquiry into the responsive peculiarities of the skin, epithelium, and glandular tissues, alike in plant and animal. By the study of such simple cases as are found in plants, it should be possible to obtain’a clear insight into the various factors which go to make the corresponding phenomena in animal tissues so complicated and obscure as to be difficult of reconciliation with each other.

It is not possible in a short space to give any but the briefest summary of the work hitherto done on this extended subject in animal physiology. All that can be attempted is to indicate some of the leading theories and_results, at the same time drawing attention to those outstanding questions which still remain open. Some of the methods which I have employed in the investigation of plant phenomena, moreover, have proved so highly satisfactory that records will be given of the results obtained by their means in the case of animal tissues also. And this will, I hope, show the great reliability and simplicity which it is thus possible to intro- duce into the investigation as a whole.

With regard to the electrical effects in animal skin, epithelium, and glands, the inquiry resolves itself into the determination of, (1) the direction of the current of rest ; (2) that of the excitatory current ; and, lastly, (3) a consideration of theories regarding these. The first of these, the current of rest, was found by Du Bois-Reymond and Engelmann in the skin of frog to be ‘ingoing ’—that is to say, passing from the outer surface to the inner. Hermann also found a similar current in the skin of eel. He regarded the source of electro-motive action as lying in the partial mucin-metamor- phosis of single cells. From the fact that in the toad, where the ingoing current is specially strong, the skin glands are vigorously developed, and from the discovery by Rosenthal that in the mucous glands of the stomach the current is also ingoing, it was assumed that the observed electro-motive forces were due to the glandular nature of the tissues. The skin current of the frog and of the fish, and the glandular current of the stomach, are thus usually regarded as due to the same cause.