Comparative Electro-Physiology: A Physico-Physiological Study

consequence of repeated responses there is also a relatively greater contraction and loss of turgidity in that lower half. The leaflets are thus closed by the additive effect of multiple normal negative responses or downward movements. When a Szophytum plant is kept in the dark the leaflets undergo a closure which is outwardly similar to that induced by light, but actually arises from a cause precisely opposite. Under strong stimulus of light a general contractile effect drives the water inwards to the interior of the plant, and, the loss of turgor in the lower halves of the pulvini being, as we have just seen, greater than in the upper brings about the closure of the leaflets. But in complete darkness the reverse process is set up. The water returns to the pulvini, making them over turgid. Under this condition of excessive turgor, however, the upper half of the pulvinus becomes more turgid than the lower, and, being thus rendered more convex, closure of the leaflets is brought about. Ex- posure to light in this condition : Fic. 241. Multiple Mechanical induces a greater loss of turgor by Response of Leaflet of the upper than by the lower half, Biophyium under the Con.

outspread or erected position of the leaflets by a series of small positive movements. Thus, as the result of internal changes, the response of an organ may undergo reversal from the ordinary negative to positive. The change from light to darkness, by which we have here seen the character of response to be so greatly modified, occurs in the diurnal periodic alternation. And we can see that, in consequence of such an imparted periodicity, an alternation of phase is impressed upon the organism, which will carry with it a periodic variation of excitability. If this diurnal periodicity acted alone its after-effect would be comparatively simple,

but it is complicated by other periodicities, such as that of temperature variation, which do not always coincide in maxima and minima with these variations of light. We have seen from mechanical indications that multiple excitations are induced by light. We have, therefore, to determine whether similar multiple excitations can be detected electrically. Fig. 242 is a photographic record of a series of such electrical responses ob- tained from the lamina of a leaf of Bryophyllum sub- jected to the continuous action of light. We see here an alternation of phase in the response, negative being followed by positive in each case throughout the series. This constitutes a parallel

case to that of the mechanical Fic. 242. Photographic Record of ; ; Multiple Electrical Response in FeSponse in fig. 241, inasmuch covery from each negative phase, the base line is gradually tilted upwards. But this need not always occur, for the two phases may be equal, and in that case the base line remains horizontal. We have seen, in the chapter on Multiple and Autono- mous Response, that these effects are due to the absorption of an excess of energy. When this absorption is great the energy may find an outward expression, even after the cessation of the stimulus. An example of this was seen in the mechanical response of a Desmodium \eaflet (fig. 141): The plant was at first in a sub-tonic condition, and the auto- nomous pulsation of its lateral leaflets had come to a stand- still. One of these was now exposed to the continuous action of light and its record taken. It will be noticed that under this stimulus of light multiple responses were initiated, which persisted for a time as an after-effect, even on the ces- sation of light. In taking electrical records of the after-effect

of stimulation by light, I have obtained similar multiple after-effects. And in this connection I have discovered certain characteristic peculiarities of the electrical after-effect which would appear to throw much light on the obscure subject of after-effects in the retina. The electrical after-effect of stimulus of light varies greatly in different conditions of the tissue, but is capable of classifica- tion under three different types. ‘ / The first of these types refers - to those multiply responding tis- sues which give the usual single response of galvanometric nega-

light. When such specimens are a ‘a at subjected to the continuous action \ of this stimulus they give multiple \ responses whose phases alternate e\ our attention to the first two pairs of such phasic alternations under continuous stimulation of light (fig. 243), we observe that, in the first period, excitatory . negativity attains its maximum at J, after which the phase becomes reversed to positive, in which process the curve may arrive at the original base line, or stop short of this or go beyond it. The maximum of

sentation of Phasic Alter- nations, and After-effect in Type I. phasic. alternations are a 4, ba',a' b’,o'a' (—+—+); 6 is here the maximum negative, and a’ the maxi- mum positive. Withdrawa of stimulus at point of re- versal a’ causes unmasking of positive component, which is exhibited by overshooting of the curve in the positive this phase, a’, we shall designate as maximum positivity. Under the still-continued action of the stimulus the phase now changes once more to negative, and so on.

It has already been stated (p. 100) that these periodic alternations in phase were brought about by antagonistic as was there pointed out, in those cases which normally give negative response, the internal energy may be so increased by the continuous absorption of impinging stimulus as to induce a continuously increasing antagonistic reaction of positivity. Hence, in such cases, negativity is gradually diminished, and ultimately reversed. After the attainment of maximum positivity, however, the negative element once more becomes predominant.

With a fresh specimen, exhibiting multi-phasic responses under the continuous action of light, I find it easy to obtain a convincing demonstration of the presence of the positive factor by a sudden cessation of stimulus at an appropriate moment. If the stimulus be stopped at 4, the increasing internal energy and natural tendency to recovery will con- spire with each other, and the after-effect will, generally speaking, be 6a’. That is to say, this effect is the same as that of natural recovery.

When the second or positive phase has reached its maxi- mum a’, the response under the continued action of stimu- lation is once more reversed to negative a’ J’, as we have seen. At this point of reversal a’, the positive, is balanced by the negative. If then at this point the impinging stimu- lus be suddenly withdrawn, the positive element, finding itself unopposed, will overshoot the line of balance, and the curve will proceed in the positive direction towards c (fig. 243). This phasic sequence (Type I.) then, during stimulation, and as an after-effect of it, should thus be (— + -:-), the dotted sign representing the after-effect. That this is actually so, is seen in the following series of photographic records (fig. 244), where the dotted portion of the record exhibits the after-effect. The intensity of this positive after-effect varies with the freshness and vigour of the specimen. The influence of fatigue, in the gradual diminution of the effect, is seen in the two subsequent series (4) and (c) obtained from the same specimen. This diminution culminates in the gradual diminution of the positive after-effect, and its conversion into a negative after-effect, as seen in (d).

plified by specimens which are not very fresh. The sequence ishere(— + ...). In some instances, if the stimulus be stopped at the end of the first negative phase, we obtain a small increment of negativity as the after-effect. The sequence here, then, is (— ...). Fic. 244. Photographic Record of Phasic Alternations, showing Direct and After Effects of Light in Type I., represented by Bryophyllum a: First record of the series. Positive after-effect represented by. dotted curve is here strong. 4andc: Less strong positive after-effects due to fatigue. Inc light was stopped slightly beyond the second phase of maximum positivity; d, owing to fatigue, after-effect converted into negative. Dotted portions of curve represent, as usual, the after-effect on cessation of light.

Turning to Type III., we take specimens whose charac- teristic response to short-lived stimulus of light is by galvanometric positivity, the multiple phases in which, under its continuous action, may be expected to_be in the se- quence represented in figure 232, that is tosay(+ — + —) As the responses here are the exact opposite of those in Type I., we may expect to obtain the unmasking of the nega- tive element in the response, as the after-effect, on the with- drawal of stimulus. Here there should be cessation of stimulus at the end of the second phase, in this case, maximum mznus. The negative element, thus freed from its opposing positive, will demonstrate its presence by over-shooting. in the negative direction (fig. 245). This is seen in the next two figures (246 and 247). The specimen ; taken was the petiole of cauli- flower, in that particular condition which gives the positive as its immediate response. The _alter- nation of plus-minus-plus-minus, under continuous stimulation, is seen in the first part of fig. 246. Stimulus was now stopped ata point short of the maximum nega- tivity, and we see the consequent overshooting in the negative direction. In the next figure (fig. 247), is seen a single alter- nation of p/us-minus during stimu- lus, in a different specimen, with

FIG. 245. Diagrammatic Repre- sentation of Phasic Alter- nations and After Effect in Type III. During action of light the phasic alternations are a 0, b a’, a’ b', b’ a" (+—+-—);3 is here the maximum positive, and a’ the maximum nega- tive. Withdrawal of stimulus at point of reversal a’ causes unmasking of negative element, which is exhibited by the overshooting of the curve in the negative direc- tion, a’ ¢. its after-effect and recovery from that effect. The impinging stimu- lus was in this second case stopped at the exact point of maximum negativity, and the overshooting

curve shows, by its abrupt steep- ness, its sudden freedom from the restraint imposed by the opposite element of positivity. Thus, in this instance of Type III., we arrive at the typical formula of (+ — ...). It may then be said that under this head, of alternation of phase, we have examined representative cases of the two ex- treme Types I. and III., between which lie many variations, classified as intermediate, or Type II. plants to light is not essentially different from their response

to other forms of stimulus. The various effects observed are the electrical concomitants of those excitatory effects which we had already seen to be exhibited in mechanical response. In a Fic. 246. Photographic Record of Phasic Alternation, showing Direct and After Effects in Type III., represented by with a Second Specimen of Petiole of Cauliflower Cauliflower, representative of Continuous line represents effect during appli- Type III. cation of light. Stimulus was withdrawn slightly before the attainment of the second maximum negativity, resulting in over- shooting of curve in negative direction. Dotted portion of record represents after- effect on cessation of light,

The stimulus of light was in these cases withdrawn exactly on the attainment of the negative maximum. Dotted portions of the record ex- hibit the after-effect. highly excitable tissue the direct effect gives rise to galvano- metric negativity. The indirect effect of stimulus, however, is one of galvanometric positivity. Positive response may also be obtained from a tissue which is not highly ex- citable or in a sub-tonic condition or fatigued. The pre- sence or absence of chlorophyll is not a determining factor in electrical response. As in the case of mechanical and growth responses, so also in the electrical, continuous stimu- lation gives rise to multiple responses, as its direct or after- effect. ‘The sequence of phases, under the direct action of continuous stimulation, may be (— + — +) or(+ — + —).

Taking into account both the direct and after-effect, the observed results may be classified under three types. In. the first type, that of fresh and highly excitable tissues, the formula is(— + -i-). In the second or intermediate type, it is (— + ...) or (— ...). And in the third type we have the formula (+ -— ...). Response of retina—Determination of true current of rest—Determination o: differential excitabilities of optic nerve and cornea, and optic nerve and retina —The so-called positive variation of previous observers indicates the true excitatory negative—Retino-motor effects—Motile responses in nerve— Varying responsive effects under different conditions—Reversal of the normal ° response of light due to (1) depression of excitability below par ; (2) fatigue— The sequence of responsive phases during and after application of light— Demonstration of multiple responses in retina under light, as analogous to those in vegetable tissues—Three types of after-effect—Multiple after- excitations in human retina—Binocular Alternation of Vision—Demonstra- tion of pulsatory response in human retina during exposure to light.

THE nature of the electrical reaction of the retina under stimulus of light constitutes a problem which has attracted many investigators, chief and earliest among these being Holmgren, Kiihne and Steiner, Dewar and McKendric. The results obtained by these workers have been confirmed by the later researches of others; but while their general observa- tions are fairly concordant, the way in which the phenomena they have described are related to the excitatory reaction is a question which has hitherto remained undetermined. Observed results, in fact, would seem to show that the electrical reactions of the retina are of a nature quite different from those exhibited by other animal tissues. For whereas nerve or muscle, for example, responds on excitation by negative variation, the retina, under the same normal con- ditions, is said to exhibit a positive variation. The subject is very. much complicated, moreover, by the confusion which has unfortunately arisen as to the meaning of the terms positive and negative. These positive and negative varia- tions are so named in reference to the existing current ot

rest, which, as we have already seen, does not constitute a very definite or unvarying standard. A still further source of complication is introduced again when, under certain internal changes in the retina itself, the normal response is found to be reversed. The present inquiry, then, resolves itself into the follow- ing questions :—(1) What is the true current of rest, and do the various currents which have been observed really fall under this head or not? (2) What is the galvanometric character, positive or negative, of the excitatory reaction of the retina—that is to say, is the excitatory phenomenon in the eye similar to, or different from, that of other tissues? ' (3) May we not discover, in the case of the retina, those mul- tiple excitatory reactions which we have already found to be induced by light in vegetable tissues? (4) And, lastly, what are the after-effects of this particular stimulus in the retina?

Of these we shall deal first with the question of the direction of the current of rest. In an eyeball which is isolated entire it has been found that the nerve is negative to the cornea, and the current of rest is therefore believed to flow through the eye from nerve to cornea. In an isolated nerve-retina preparation, on the other hand, the rod-surface of the retina is negative to the nerve, the current of rest being thus from retina to nerve. These observed currents, however, may not be true currents of rest, but rather excitatory after-effects, due to injury in preparation. In such cases we have seen that the naturally more excitable becomes persistently nega- tive. We have also found that the true current of rest flows from the less to the more excitable, the latter being thus galvanometrically positive. In order, therefore, to determine the true nature of the resting-current, we have first to deter- mine which of each two surfaces, nerve and cornea, and nerve and retina, is the more excitable.

I have already described how, by means of equi-alter- nating electrical shocks, the differential excitability of a preparation can be determined. We saw also that under such conditions the resulting responsive current flows from the more to the less excitable. The experimental arrange- ment used in this investigation is shown in fig. 248. In such an experiment, equi-alternating shocks are given to the’ pre- paration by means of a secondary coil included in the circuit. The existing current, from nerve to cornea, may be balanced previously by a potentiometer. Whether the record be taken under balanced or under ordinary conditions, it is found that in anormal eyeball the responsive current is from the nerveto the cornea, showing that the nerve is relatively the more excitable of the two. Fig. 249 gives a series of such responses. It will also be noticed that the excitatory current is in the same direction

Fic. 248. Experimental '. Arrangement for Deter- mination of Differential © _. Excitability of Optic — ; Nerve and Cornea Fic. 249. Series of Photographic Records of ‘Excitatory Responses in Frog’s Eye to Equi-alternating Electric Shocks at Inter- _ vals of One Minute C, injury current, or so-called current of rest; R, re- sponsive current. as the existing current, and thus constitutes a positive varia- tion of it. Thzs positive variation of the current in the eyeball thus indicates a true excitatory reaction of the optic nerve. From the fact that the nerve is more excitable than the cornea, it is clear that the sectioning of it for isolation of the eyeball, acting as an intense stimulus, will result in its excita- tory negativity, which will persist for a time and slowly dis- appear. Owing to this fact, a current flows from the more excited nerve to the less excited cornea. That this current

is not the true resting-current, but rather an excitatory effect consequent on preparation, is supported by the known fact that it undergoes a decline more or less rapid. On the other hand, from the fact just demonstrated that the nerve is the more excitable, we should expect that, under natural or primary conditions—that is to say, inthe absence of excitatory disturbance—the resting-current would be from the less to the more excitable, or in other words, from cornea to nerve, This | conclusion I was able to verify by carefully dissecting away the socket of the frog’s eye, and making connections with the longitudinal surface of the un- detached nerve and the cornea. Under these ideal conditions the true resting-current was detected, and was, as expected, from the cornea to the nerve.

In order, next, to determine whether the current, observed to flow, in a nerve-retina prepara- < tion from retina to nerve, is a ‘ee true: resting-current, or merely Fic. 250, Experimental Arrange- an excitatory after-effect, I pro- ment for Demonstration of ‘ i Differential Excitability as bee ceeded to determine, in the tween Retina and Optic Nerve — manner already described, which excitable. Under the excita- tory effect of equi-alternating shocks, the responsive current was found to flow from the retina to the nerve, thus proving that the retina was the more excitable (fig. 250). The first series of responses in fig. 251 gives a record of these effects with a moderate intensity of stimulation, while the second series shows the responses under an intensity nearly twice as great. The so-called current of rest observed in the nerve-retina preparation is thus to be taken as due to that after-effect of preparation which is inseparable from the isolation of such a highly excitable tissue. /¢ well also be

noticed that the true excitatory effect on the retina ts tn the same direction as the existing injury-current, and thus constitutes a positive variation of tt. . Thus, as regards forms of stimulus other than light, such as, for example, the electrical, the responsive reaction of the retina is by induced galvanometric negativity. This at once disposes of the doubt that the general reaction of the retina might be of a different sign from that of any other tissue. But we have still to determine whether or not the stimulus of light, in particular, induces the same normal negative change, The next question to be taken up, then, is that of the true nature of the responsive variation induced in the

Fic. 251. Series of Photographic Records of Excitatory Responses in Frog’s Retina to Equi-alternating Electric Shocks at Intervals of One Minute Responsive current from retina to nerve. First series show response to stimulus of moderate intensity ; the second series to stimulus of intensity nearly twice as great. retina by light. We have seen that the effect recorded as normal by numerous observers, whether in the eyeball or in the isolated retina (Kiihne and Steiner) was a positive varia- tion. But since we now know that retinal response to stimu- lation in general is not unlike that of other tissues, and since, with regard to the stimulus of light'in particular, we have seen it induce true excitation in vegetable tissues, we might expect the responsive reaction of the retina to light to take place by galvanometric negativity. We must then accept one of two conclusions. Either the positive change is a misnomer for the phenomenon observed in the eye, or the inference which we have drawn from the analogy of vegetable tissues is not justified.

As, in reference to this latter point, however, it may be urged that the retina is exceptionally sensitive to light, while the reactions of vegetable tissues are generally slug- gish, it may be worth while to point out here that vegetable tissues are not so insensitive as is generally supposed, but are often, on the contrary, highly susceptible to the: action of light. It. was, for instance, found by Darwin that the coty- ledons of Phalaris canariensts were, in the course of some _ hours’ exposure, curved towards a small lamp, placed ata distance from them of 12 feet.. The intensity of the stimulus of light was in this case extremely feeble. I have myself observed, again, the remarkable sensitiveness to light of the terminal leaflet of Desmodztum,.which, on the mere striking of a match in its vicinity, was thrown into a state of pulsatory movement.

I shall, in the course of the present chapter, describe certain definite effects on the retina, which will prove, in an - unmistakable manner, that, when exposed to light, it under- goes a change of galvanometric negativity. It is now there- fore necessary to show clearly that what has been described as the positive variation is really due to the excitatory nega- tive change of the retina. What has to be demonstrated, then, is the way in which this simple underlying reaction of negativity comes to appear as a positive variation of the opposite-directioned currents of rest in the a and isolated retina respectively.

As regards experiments on the eyeball, with contacts at nerve and cornea, we have seen that. the existing injury- current is from nerve to cornea, and that this undergoes a positive variation when the nerve is stimulated in any way. This is because the added. negativity induced in the nerve gives rise to an increase, or positive variation of, the existing current (fig. 248). Now, when light falls on the eye it -acts on both the cornea andthe retina. The former, however, is relatively inexcitable, especially to so moderate a stimulus as that of light. But the retina is excited, and its excitatory condition is rapidly transmitted to the optic nerve, which

thus becomes galvanometrically negative. The so-called positive» variation observed under such an experimental arrangement, then, is really the result of the true excitation of the retina conducted to the optic nerve. In experiments- on nerve-and-retina preparations, with contacts on the nerve and the retina, the existing injury- current is from retina to nerve.- When do¢/ nerve and retina are excited simultaneously, by equi-alternating electrical shocks, we have seen that, on account of the greater ex- citability of the retina, the resultant responsive current, here differential. is from retina to nerve, constituting a positive variation of the existing injury-current (p. 419). In the case of stimulation by light, however, it is the retina which is directly excited. The added negativity thus induced in the retina gives rise to an increase or positive variation of the existing injury-current (fig. 250). Hence, in all these cases, the so- called positive variation really indicates the normal excitatory negative effect. The apparent anomaly involved in the supposition that the response of the retina to light was positive, and thus of opposite character to that of other ex- citable tissues, is thus seen to be due to a misinterpretation of observed results. It is unfortunate that, as a consequence of this misinterpretation, the effects described by different observers, of the response of the eye as ‘ positive,’ are really to. be understood as ‘negative, and vice versa. When quoting these results, therefore, I shall always give the actual effect indicated in italics and in parentheses,