On the anatomy and physiology of the retina

1249

5th.H LeysBruxs

1250

ARCHIV

ANATOMIE UNDPHYSIOLOGIE. FORTSETZ~JNG DESVONREIJ,, R,EIJ, u. AUTENR,IETJT, J. F. MECKET,, JOIT. MiiJIT~ER, REICIHER’J’ u. l)U BOIS-REYMOND

HERAIJSGEGEBENEN ARCI~IVES.

HERAlJSGE:G:ERE:N VON DR.

WILH. HIS PROFESBOREN

DER

UND I)R.

ANATOWIE

AN

WILH. BRAlJNE, DER

IJNIVERSIT~T

I,EIPZIC3,

UND I.h

EMIL

J’ROFESSOR

DER

IX-J BOlS-REYMOND, PHYSIOLOQIE

AN

DER

UNIVEIIRIT;iT

~ER,,,s.

JAHRGANG1877. PHYSIOLOGISCHE ABTHEILUNG.

LEIPZIG, VERLAG

VON

VEIT

1877.

& COMP.

1251

ARCHIV

PHYSIOLOGIE. PHYSIOT,OGISC!HE ABTHEILUNG DES ARCHIVES FOR ANATOMIE UNll PHY SIWOGIE.

JAHRGANG1877. MIT Jl~J,ZSCHNITTEN IM TEXT UND 11 TAFET,N.

LEIPZIG,’ VEHIrA(:

VON

VEIT &

1877.

COML’.

1252

Zur Xnntotnie und Physiologic der Ketina. VOU

Prof. hus dem Laboratorium

Franz

Boll.

ftir vergleiohende dnatomie Achte Nittheiluqq. 1

uud Physiolqie

zn Rom.

(Hierzn ‘hf. I.)

Seitdem zuerst durch He i n ri c h M ii 11 er die in der sogenannten musivischen Schicht der Retina vereinigten Elemente der Stab&en uud Zapfen als die Endorgane des Nervus opticus in Anspruch genommen wurden, sind die ~~ikroskopil~er t~nausgesetzt bestrebt gewescn fiir diese Behauptung den a~atomischen Beweis zu fiihren. Dennoch ist es trotz der jahrelangen Bemtihungen der ausgezeichnetsten Anatomen nicht gelungen, diesen Nachweis zu fiihren und die Nervenbalm nnfzuspiireu, durch welche die Substanz der Stabchen nnd Zapfen mit den Fasern des Sehnerven in Verbind~~ngsteht. Die Erfolglosigkeit aller dieser Bestreb~~ng~nhat einzelne Forscher sogar vermocht, den von H. Xii I ler eingeschlagenen Weg wieder zu verlassen und die Endigungen des N. opticus in anderen Schiohten der Retina zu suchen. So hat man die Elemente der Membrana fenestrata als die Endze~len des Sehnerven in Anspruch nehmen wolten; und ich selbst bewahre noch einen im Jahre 1871 niederges~hriebenen Auf&z auf, in welchem ich ausffihre, dass mit grosserer Wahrscheinlichkeit als die Stibchen und Zapfen die sechseckigen Epithelzellen des Retinalpigmentes als die percipirenden Endorgane des N. opticus anzusehen seieu. Han ist, giaube ich, bisher bei der ~n~rsuch~~ng der ~ervenendigung in der Netzhaut anatomischerseits von einer zu engen Vorstellungsweise ausgegangen, indem man in der St%bchen- und Zapfensehicht beharrlich nach demselben einfachen Schema der Endigung suchte, welches in den anderen einfacher gebauten Neuroepithelien z. B. fur die Geruchs- und 1 ~~~an~~ung~

1876-l 877.

i&w 2% AccaAmia

dei Gncei.

Dritte

Serie.

Enter

The&

Vision Rrr. Vo!. Il. pp. 1249 lo 1265. Pergamon Press 1977.Pnnd

m Great Bnlmn.

ON THE ANATOMY AND PHYSIOLOGY THE RETINA*

OF

PROFESSORFRANZ BOLL From the Laboratory for Comparative Anatomy and Physiology in Rome.’

Ever since Heinrich Miiller first suggested that the portions of the rods and cones that constitute the retinal mosaic are the end organs of the optic nerve, microscopists consistently have tried to find anatomic evidence for this assertion. But despite years of effort by the best anatomists, it has been impossible to demonstrate this and to discover the neural path by which the rods and cones are connected to the fibers of the optic nerve. The futility of these attempts has even induced a few experimenters to abandon the line of investigation initiated by Miiller and to look in other retinal layers for the terminations of the optic nerve. Thus some have tried to invoke constituents of the outer granular layefi as terminal cells of the optic nerve; and I still have the draft of a manuscript written in 1871 in which I suggest that it is more plausible to regard the hexagonal cells of the pigment epithelium as the perceptive end organs of the optic nerve than the rods and cones I believe that heretofore the viewpoint from which we have approached the anatomic investigation of the nerve endings in the retina has been too narrow in that we have always tried to fmd in the layer of rods and cones the same simple arrangement of endings that has been shown to exist in other, simpler neural epithelia, such as for example those of the olfactory and gustatory nerves We are taught that in these, each of the fine, primitive nerve fibrils terminates upon a single epithelial cell; and there has been a determined attempt to establish this same relationship in the retina, though these efforts are known to have met with no success so far. At present only the greatest of optimists would hope that we will one day discover the nerve fibrils which connect at one end with the substance of the rods and cones and at the other with the optic nerve fibers Though I am not one of them, I am also not * (Translated by Ruth Hubbard with the help of Helene Hoffmann). 7 Boll uses the term, membrcrwjenestrata. which according to Polyak (S. L Polyak, The Retina, Chicago: University of Chicago Press 1941, p. 194) is the outer granular or internuclear layer in which the bipolar and horizontal cells make synaptic contact with the rods and cones. (R.H.) ’ Previous communication: Trans. R. Acad. dei Lincei, 3d Series, Part 1, 1876-77. pp. 4-35. * [On the Anatomy and Physiology o/ the Retina], Monotsberichte der Berliner Akudemie, 12. November, 1876. [On the Physiology of Vision and Co&r Sensution,] Ibid. 1I January, 1877, with addenda of 15. February 1877. The contents of both essays are in general agreement with two short communications which I-presented to the R. Academia dei Lincei in Rome on 3. December. 1876 and 7. January. 1877.

among those pessimists who “flee into the desert because not all their dreams of spring came true” and who deny to the rods and cones their significance as the visual end organs of the optic nerve because we have not been and will not be able to demonstrate this anatomically. It is my conviction as an anatomist that the physiological units which perceive light and color are very complicated structures that are formed by a combination of elements in the rod layer and the pigment epithelium. They should therefore be regarded histologically as double or twin cells, completely analogous to those that have recently been shown in the terminal cells of the auditory nerve in the cochlea. I assume that for each of the anatomical and physiological units that result from the combination of these two element% the anatomical connection with the fibres of the optic nerve is established by means of delicate processes of the pigment epithelium which continue into the rod layer and whose unpigmented endings I have been able to follow through the external limiting membrane. In addition it does not seem at all improbable that such complicated composites, as I imagine the end organs of the optic nerve to be, also make other connections with the nervous system, perhaps by means of the rod and cone fibrils. In this paper I do not intend to develop in detail the anatomical reasons on which I base these assump tions regarding the terminations of the optic nerve. This will perhaps be done elsewhere. However, I believe that the evidence I am about to report will suffice, even in the absence of anatomical proof, to allay any possible remaining doubts and to confirm Heinrich Miiller’s original postulate that the perceiving end organs of the optic nerve can only be located in the retinal mosaic. The following experiments and observations, some parts of which have been communicated previously in two reports to the Berlin Academy,’ are primarily physiological, although they originate in an anatomical discovery: they concern a hitherto almost completely unacknowledged peculiarity of the substance that composes the outer segments of vertebrate rods and of the physiologically and perhaps also phylogenetically equivalent organs of invertebrates (e.g., the rods of higher molluscs and the rhabdomes of anthropods), whose characteristic platelet structure has been the subject of numerous investigations since its discovery by Hannover (1840). In 1842 Krohn first called attention to the fact that in cephalopods this substance is red. This observation was extended by numerous reports of other authors regarding many different classes of invertebrates. Among vertebrates, Heinrich Miiller in 1852 first described the red color exhibited by the rods of the

1253

12%

FRANZ Bou

frog

retina. However it was left to me to discover that this red coior is an inseparable charac~~stic that it is part of the nature of the substance that forms the red platelets and is the same in all animals in whose eyes it occurs. The frog is the most suitable animal for demonstrating this red color. If one bisects an eyeball and with fine tweezers pulls the retina away from the dark background of the pigment epithelium and choroid, the retina looks intensely red in the first moment, so that one could imagaine than the tweezers had picked up a blood-clot. During the next ten or at most twenty seconds (stage I) this color fades gradually until it disappears completely except for a slightly yellowish gloss. For the next 30-60 see and sometimes longer (stage II) the retina exhibits a satiny sheen. Gradually this also disappears and the retina becomes completely transparent and stays this way a quarter hour or longer (stage III). After that it eventually becomes cloudy and opaque (stage IV) Microscopic examination reveals that the red color of the first stage and the satiny sheen of the second reside exclus ively in the material that constitutes the platelets of the outer limbs. Toward the end of the second stage this substance swells and changes and its rdractive index approaches that of the other retinal layers, so that in the third stage the retina becomes completely transparent The opalescence of the retina in the fourth stage is due not to changes in the rod layer but to the ~oag~tion of proteins in other retinal layers. But how has it been possible that such striking phenomena which take the same form in the eyes of almost all animals have heretofore escaped the attention of scientists? My initial explanation was that one is dealing with an extremely ephemeral property of the living retina, that can be seen only during the first short moments after death, and which escaped earlier investigators only because they never examined a retina sufficiently quickly for it to be fresh enough and they always lost that first precious moment after the animal’s death, those critical C&20 see within which I have ahnost always seen the red color disappear completely. But I soon found that this could not be the whole explanation and could at best be part of the story. As I continued with my experiments I soon noticed that I could often not see the red color of the retina at all, although I had prepared it as quickly as ever and the critical 10-20 set from the animal’s death to the preparation of the retina had definitely not elapsed. In spite of this I often could no longer detect a trace of the red color. Eventually I could find no way out of all the conflicting observations except to assume that some other 3 It was not unusual for frogs which had been exposed to the sun for more than an hour in the cylindrical glass jars to be found dead with all their muscbs in heat rigor! 4 At the beginning of my experiments madequate obaervations led me to imagine that the intensity of visual red should be thought of as increasing steadily if the animals spent a very long time (several weeks) in darkness. More recent experiments taught me something different and I must now assume that visual red attains its maximal intensity after a relatively short time (twelve hours; that is a night’s rest) and that longer periods af darkness produce no further increase.

physiological condition affects the disappearance 01 the red color besides the animal’s death and the cessation of the normal supply of nutrients. Thus 1 soon arrived at the notion that the red color is not a constant property of the living retina but must be subject to physiological change, and that the retina bleaches not only when the animal dies and when it is removed from the eyeball, but probably under certain conditions even in life. Once I had gone this far, it was no longer difficult to guess the effective physiological agent and I do not think that I deserve special credit to have soon hit upon light as the significant factor that determines the presence or absence of the red color of the retina. It was easy to raise this guess to a high degree of certainty: animals which had been exposed for long intervals to the sun or even to diffuse, bright daylight never exhibited the red color in their retinas: but it was always there as soon as the animals had spent long periods of time in darkness. From this I concluded that in life the red color of the retina is constantly destroyed by light entering the eye and is just as constantly restored by the physiological supply of nutrients, so that the red color can be observed only after the eye has been in darkness long enough to allow time for it to accumulate. One of the first series of experiments that I undertook was designed to determine the time within which “visual red”, as I shall henceforth call the red color of the retina, is consumed by light. I simul~~usly placed a dozen frogs which had spent a very long time in complete darkness into sun-lit glass vessels and examined a pair of eyes every five minutes. During the first such experiments, which I performed last November, I was unlucky in that the weather and sunlight varied, so that I obtained times which were much too long, judging by a later series of experiments. This second investigation, which took place during the second half of January, 1877 under a completely cloudless sky and very intense sunlight3, revealed a considerable fading of visual red even after the initial ftve min. After IOmin one could detect only a slight red shimmer; and this was present only very rarely after 15 mm when the retina usually was already completely colorless. At the end of half an hour there was never a trace of the initial pigmentation and the satiny sheen of the dying retina was never yellowish, but only clear white. The same experiments were performed simultaneously in front of a northern window of the laboratory, where bright diffuse daylight, but not direct sunlight could reach the frog eyes, with the result that complete bleaching of the retina took two-three times longer in diffuse daylight than in direct sunlight: after two hours visual red had been consumed completely in all the eyes. To decide the second question, within what time bleached visual red is reconstituted, I took the opposite course. A dozen frogs which had been exposed to direct sunlight for more than an hour were returned to complete darkness and examined successively. In these frogs the first traces of redness never tetumed sooner than an hour and were usually still very faint after an hour and a half. However, after two hours the colour usually was very intense and scarcely increased further after longer periods of darkness4. After these temporal investigations only one,

On the anatomy and physiology of the retina further experiment was needed to convert my hypoth-

esis that light destroys visual red into a certainty: the ~on~mtion that in the eye partial i~u~ation of the retina destroys visual red only in the illuminated areas and nowhere else. That this must be so was a near certainty a priori; in addition, I had already noticed that those parts of the retina which were more shielded from light, such as those near the ora serrutu, frequently still looked red even when visual red had disappeared completely from the center. But I did not want to omit such an experiment: I partially closed the shutter% so that only a relatively narrow stripe of sunlight entered the room. Into this I placed the eye of a curarized, dark adapted frog.’ Examination of the retina after 1Omin showed it to be divided by a clearly delineated colorless stripe into two red halves 0nly after this experiment, which surely could be mod&d in many ways but which I did not pursue further, I felt justified in stating the conclusion of my first communication, which is as follows: “The intrinsic color of the retina in uiw is constantly destroyed by light entering the eye. Diffuse daylight causes the red color of the retina to fade. Prolonged exposure to direct sunlight (a high degree of light adaptation) bleaches it completely. In darkness, the intense color soon recovers. “This objective change of the outer limbs by light surely must play a part in the act of seeing.” Ona my initial experiments had led me to recognize the ~s~b~~ of visual red when exposed to light, I Italy should have begun to m-examine more carefully my original assumption regarding its great physiological instability. I had taken this so completely for granted when I began my investigations that I felt it scarcely needed further proof. But, initially, this simple and logical idea did not occur to me! Even after I had begun to understand and appreciate the full extent of the effect of light on visual red, I continued to believe the red color of the retina to be a very transient characteristic of the live tissue that was extinguished almost immediately after the animal’s death or upon cessation of the normal life processes This assumption, which alone enabled me to ~d~s~d how all my predecessors who had worked with fresh retinas had failed to discover visual red was initially quite unaffected by my new knowiedge: I was still as inclined as ever to attribute the rapid loss of color of the isolated retina almost more ’ This is how, for brevity’s sake, I plan to refer to frogs that have remained in complete darkness for long periods of time. 6 I had just performed these experiments with dead frogs the day my first communication to the Berlin Academy was returned to me for proof-reading Throughout the article the great physiotogical labifity of visual red was assumed and was stated cxplicitiy at the end. Since I did not think it right to make substantive changes in the paper, once it had been presented to the Academy, I Iimited my revision to inserting into the text of the thesis, “The basic, lifetime color is present only in live animals and outlasts their death by only a few moments”, the words. “at least in warm-blooded animals”, thus expressing by implication my newest realization that the fading of the retina in amphibians is elicited not by death but by illumination. ‘The reason why such artificial lights do not destroy visual red is discussed below.

1255

life processes than to a direct effect of light I continued to believe that for the demonstration of visual red the greatest speed in preparing the retinas was essential, and my first communication was still written entirely under the ir&uence of this interpretation. However a chance observation soon showed me the correct way and demonstrated the true significance of the two relemnt factors: the direct effect of light and the cessation of normal life processes. Following the bright, clear weather that had prevailed until then, by midNovember the days became dark and cloudy and forced me, if I did not want to discontinue my work completely, to undertake the microscopic examination of the retina in -poor light”. This immediately resulted in‘ a very considerable prolongation of the time course of the “first stage- and the red color of the retina now lasted not just 2Osec as before, but up to 5 min and even longer. This constantly recurring fact was clear evidence that I had previously underestimated the effect of direct illumination on the isolated retina and had assigned too much importance to the cessation of the normal life processes that accompanies the preparation of the retina. Therefore I decided to investigate systematically the relative significance of these two factors for the sudden fading of the isolated retina-l-he experimental method was very simpIe: I simuitaneously decapitated a dozen dark adapted frogs and kept their heads dark in order to examine their eyes consecutively at stated intervals. But I was not con& dent that this experiment would yield positive results and I therefore cautiously examined the first eye after only 5 min. To my great surprise I found its retina as beautifully red as if it had been prepared right after the animal’s death. My astonishment grew as the same result was repeated after longer and longer intervals: I found visual red intact in frogs (and also in bony and cartilagenous fishes) that had been dead up to 24 hours; thereafter, it seems to disappear rather suddenIy.6 The visual red of mammals that had been dark adapted and killed in darkness was quite similar in its stability, which astonished me ail the more as I had thought that in earlier, incomplete ophthalmoscopic experiments (of which I will say more later), I had been able to see visual red vanish at the moment of death or at least soon thereafter. Also in mammals I have seen several cases in which visual red lasted up to 12 hours or more after death. Consequently these experiments proved that the red color is definitely not very labile, as I had originally assumed but is rather a fairly stable characteristic of the retina. This made it possible to improve and perfect the method of investigation which until then had remained very inadequate. I then began to prepare retinas in semi-darkness with the shutters almost closed, or in gas or ~~e~~t with complete exclusion of daylight’, and to admit daylight only later, when the specimen was already under the microscope. Unfortunately this method did not prove useful for microscopic examinations, because my own eye was always too blinded by the sudden transition from darkness into the light, or from reddish-yellow light into daylight to allow me to make clear and precise observations Several seconds always elapsed before I could satisfactorily register the microscopic to the cessation of physiolog&al

1256

F~UNZBOLL

image of the retina. But during this interval the retina always lost most of its color, so that (all in all) the new method of preparing the retina in darkness and examining it in the light offered hardly any advantage over the old procedure in which the preparation and examination were undertaken at the same level of illumination. Therefore I now once again almost always use the old method and turn to the new procedure, of working in darkness, only in the special cases for which a particularly precise and time consuming preparation is required, e.g. when comparing the central and peripheral portions of the same retina. The exclusion of daylight has many more advantages for the chemical and physical investigations of visual red than for the microscopic observations. In these ex~~rn~~ I am guided exclusively by the one point of view that has dominated my thinking since the moment I discovered visual red. For I asked myself the following question: Is visual red 8n intrinsic color that is inherent in the substance composing the platelets of the outer segments? Or does it owe its existence to the optical effect produced by a stack of intrinsically colorless platelets? The first alternative would imply that the outer segments contain a specific pigment which might be called “erythropsin” and which bears a similar relationship to the substance of the rods as hemoglobin does to the stroma of red blood cells; like the stroma, the ground substance of the outer segments could be thought of as colorless. Like the blood pigment this erythropsin would have to have a definite chemical composition and just as various gases transform hemoglobin, it would be transformed by light (and presumably in specific ways by different colors) and converted into other physiological compounds In this case the nature of the light sense and of the different color sensations would have to be sought in the production and transformations of these various chemical compounds under the influence of light, hence in a photochemical process As against this “photochemical theory” of the nature of visual red and of the light sense, another ro~u~tio~ is possible which by contrast might be called the ~pho~phy~ theory”. This theory assumes that there is no such special pigment as erythropsin that permeates the colorless rod stroma, but attributes the red color of the rods to a purely physical phenomenon, the optical effect of the stacking of intrinsically colorless platelets Accordingly visual red would belong to the category of interference phenomena and more specifically to the narrower class of the so-called colors of thin films. The colors of thin lamellae, where they occur in nature, are almost always known to fluoresce and to iridesce and are almost never constant. Yet thin layers can also give rise to completely constant and homogenous colors; according to the theory this must in fact always occur when there is not a single thin ’ Already W. Zenker tried in his [“Attempts at a Theory of Color Vision’J (Max Schultz& Archio, mikrosk. Anar. 111, 248, 1867) to establish a relationship between the platelet structure of the outer segments and the wavelengths of light. g [On the Rods and Cones of the Retina] Archio. mikrosk. Anut. III, 215, 1867.

lamella but an orderly, plane parallel, layered system. a set of very many thin lamellae all of which have the same refractive index and diameter and are separated by equal interspaces. Such a system must extinguish all light rays that enter it, with the single exception of those rays whose phase difference is always a whole wavelength or zero and to which the constants of the system are, so to speak, tuned Such a system, therefore, will and always must appear in a quite spe&c uniform hue. The photophysical theory of visual red therefore only requires the assumption that each outer segment represents such a system of thin lame&e that is tuned to the wavelength that corresponds to the red color. According to this theory vision and the perception of color would arise from material changes which the incident light waves induce in the system One would have to assume that the light waves can provoke changes perhaps in the thickness of the individual lamellae or in the distances between them, and one could very well imagine that a specific change in the constants of the system would correspond to each individual wavelength. In that case the specific kinds of changes themselves would evoke the particular qualities of the light sense that correspond to the sensations of the individual w1ors.s I am aware that there is disagreement among microscopists over the physiological significance of the platelet structure of the outer segments, which is the basis of the photoph~~ theory of visual red and of the sensations of light and color, and that some regard this as a sign of post mortem decomposition and as a kind of coagulation. To the extent that this opposition is directed especially against the findings of Max Schultze9 and F. W. Zenker, who estimate that the outer mgments of frogs consist of approximately 30 identical platelets of 0.0005 mm thickness, it is in fact not entirely unjustified: for I, too, cannot accept these formations as physiological constituents of the outer segments These platelets, as described by the above authors, which are more or less distinct and regular in the presence of different reagents, and especially after treatment with 10% of sodium chioride, first of all never exhibit the uniform thickness that Max Schultze and Zenlcer attribute to them but are really discs of quite variable thickness. These platelets are nothing other than packets of variable thickness into which different numbers of the real lamellae are stuck together. The real lamellae are probably much more numerous and delicate than those described by Max Schultze and Zenker which are O.OOOS mm thick. Their existence can be deduced only from the pattern of extremely fine cross striations of the rod stroma which is sometimes seen when absolutely fresh and still clearly red rods are examined under oil immersion and particularly favorable illumi~tion and which alway run parallel to the direction in which the rods fragment. This is true because during the preparation of fresh material, the rods, which seem to consist of a very brittle substance, break very readily into several fragments whose fracture faces arc alway precisely perpendicular to the rod axis Although there is therefore a sufficient anatomical basis for the photophysical theory of visual red, the central question was which of the two theories, that

On the anatomy and physiology of the retina

were’equally- justified from an anatomical point of view. was preferable for other reasons. Of course I was aware that my knowledge of physics and chemistry was not sufficient for me to be able to resolve this question completely; but I wanted at least to undertake some sort of preliminary investigation, if only to be able to form my own opinion in this matter. My starting point in this preliminary investigation was the following dilemma: if visual red is a chemical compound and if erythropsin really exists and is not merely a pretty name, then it must be possible to separate it from the substance of the rods and perhaps to dissolve it or somehow isolate it. On the other hand if visual red is not a chemical substance, not a pigment, but only an optical effect of the lamellae, then it will never be separable from them and always will exist or disappear only as part of the rods. In that case of course it would be impossible to isolate visual red On the other hand it might be possible to destroy or to alter visual red by chemicals and procedures which could change the physical but not the chemical form of the structures, e.g., mechanical compression. All relevant experiments of course have to be performed with complete exclusion of daylight. In order to isolate erythropsin from the matrix of the outer segments it seemed appropriate to use the same procedures as are used for the separation of hemoglobin from the stroma of red blood corpuscles: that is, freezing the retina and treating it with ether, alcohol and chloroform. All these experiments yielded negative results in that none of them succeeded in separating the red color from the rods and dissolving it. One can freeze and thaw the retina two or three times in a drop of aqueous humor without its losing color; only in time does it become paler and linally colorless. However, this loss of color occurs within the outer limbs themselves; it never can be seen first to leave the rods The process is the same if the retina is treated with ether, chloroform or alcohol. With these reagents visual red is destroyed, but never extracted from the rods. Moreover, ether and chloroform take much longer (up to several hours and more) to bleach the retina than does alcohol, which causes the retina to lose all its color within a few minutes. One thing that struck me upon the addition of ether and chloroform was that the color of the rod layer first changes from red to lemon yellow, which then becomes paler and paler and finally disappears completely. Since I do not intend to anticipate investigators, who have more experience than I in physiological chemistry, and to undertake a systematic examination of visual red with different chemical reagents, my investigation of visual red has been limited to the three compounds I have mentioned, and only to certain other solutions whose effect, on myelinated nerve fibers I had specifically investigated before, namely 0.75% physiological saline, distilled water, 10% sodium chloride, glycerol, caustic potassium hydroxide, and acetic acid To test just these reagents whose “This color is so exactly the same as the beautiful golden color of the oil droplets that are contained in the pigment epithelial cells of the frog, that it is very probable that these represent material stored for the regeneration of erythropsin.

1257

effects on myelinated nerve fibers I had already examined, seemed advantageous because a number of known facts suggest that considerable chemical similarities exist between the matrix of the outer segments and that of the myelin sheath, such as their reaction with osmium, which is very significant and common to both. An examination of these reagents showed that most of them are able to preserve visual red for a relatively long time: thus physiological and 10% saline do so up to 48 hours and glycerol almost as long; distilled water is less satisfactory and makes visual red disap pear almost completely after about 24 hours. By contrast, concentrated potassium hydroxide destroys visual red almost instantly. The behavior of acetic acid is very peculiar: it immediately transforms the red color of the rods to an intense golden yel10w’~ which fades only gradually upon exposure to light and does not disappear until much later. (The substance that is formed when the rods are bleached by light does not undergo this reaction.) The separation of visual red from the substance of the rods was not achieved with any of these reagents. As opposed to these chemical experiments, which were directed towards the chemical isolation of visual red from the rod substance, I also tried to make visual red disappear from the rods by the purely mechanical means of compression. The idea of this experiment arose from the frequent observation that the retina ten&d to bleach quite suddenly the very moment I covered it with a cover slip in order to examine it under the microscope. I particularly noticed this with retinas that have more delicate rods (those of mammals and bony and cartilagenous fishes), less so in frogs, whose considerably larger rods are presumably more resistant to pressure. Therefore I performed this experiment first with the dog retina, which I compressed in the dark between two plane-parallel slides. When I brought it into the light I found that it had lost every trace of color and had assumed a completely white, satiny sheen. I have repeated this experiment often and always with the same result, also by candle light. In this case, I could always observe very clearly that at the moment of compression the retina first assumed a very intense green sheen and only later became completely colorless, a fact which tends strongly to support the view regarding the nature of visual red that was characterized above as the photophysical theory. This is as far as I have gone in my preliminary investigations into the nature of visual red As regards the question posed above, the experiments so far favor the photophysical theory in that they have been unable to fulfill the basic postulate of the photochemical theory, to isolate visual red from the substance of the platelets. Moreover, they have discovered a chemically ineffective method, which successfully destroys visual red within the platelets by mechanical means. On the other hand, it is difficult to explain as an entirely physical phenomenon the striking color change of visual red in acetic acid which looks much more like a purely chemical reaction. Under the circumstances, I am of course far from assuming that the few experiments I have done so far contain the proper solution to this difficult problem and enable one to decide between the photo-

1258

FRANZBOLL

chemical and the photophysical theories. Rather, 1 expect this to come only from investigators who are more at home in these matters than I; they also will have to determine to what extent the two alternatives I have set up concerning the nature of visual red are justified in theory and in fact and whether it would not perhaps be more correct to assume a simultaneous and dual photochemical and photophysical action of the light rays on the substance of the platelets. I must also leave a second question to the more informed opinion and experience of others and wait for the specialists in ophthalmology to determine pre-

cisely in what way visual red contributes to the red color of the illuminated fundus of the eye. Of course I asked myself this question as soon as I discovered the red color of the retina and undertook a series of ophthalmoscopic investigations on mammals which led me to the conviction I expressed in my first communication that “the red color of the fundus. when viewed with the ophthalmoscope. does not come from the illuminated blood vessels of the choroid but depends mainly on the intrinsic red color of the retina.” Indeed I had noted during ophthalmoscopic examinations of dark adapted mammals that the spaces between the larger blood vessels that can be seen with the naked eye are a much darker red than the spaces in eyes which have been previously illuminated, in fact they are almost the same red as that of the larger blood vessels themselves. Furthermore, I believed at that time that the sudden fading of the red fundus which I observed in chloroformed animals at the moment of death was due to the sudden disappearance of visual red which could survive for only a few moments But further investigations soon revealed that at least this second assump

I1 By contrast visual red in the frog eye is clearly visible against the pigment epithelium if one removes all the refractile layers and looks sideways at the retina, which then looks like deep, dark-red velvet. I* With these it is not necessary to cut a window into the sclera since it admits enough light as it is. I3 I have not been able to discover the reason for this color, which is peculiar to the frog; I assume that it must be sought in the particularly fine distribution of the pigment processes within the rod layer of amphibia; this would confer on the rod layer the characteristics of a scattering medium and hence. it would appear bluish in reflected light. 141n mammalian eyes visual red can be se&n with the ophthalmoscope up to twelve hours after death. After that the fundus always looks white in the ophthalmoscope, a fact that may prove useful for forensics (for verification of death). IS In this connection I want to call attention to an interesting experiment regarding the subjective perception of visual red: if on awakening in the morning (preferably in a previously completely darkened room, which is suddenly illuminated by sunlight) one opens the eyes and immediately shuts them again, the entire visual field appears intensely red. (In this red visual field there appear simultaneously, as others have already reported, both Purkinje’s cobweb and the rust-colored mrrcula Mea.) If one now keeps the eyes open for a while and then shuts them again. the same phenomenon appears, but much fainter, and sirnilarly a third and fourth time, but always fainter until it finally merges into the completely normal situation.

tion must be completely wrong: I found even in mammals that visual red usually lasts for a considerable time after death and it therefore seemed more reasonable to attribute the paling of the eye ground to the cessation of the circulation of blood. This also cast doubt on my first thesis, that the red color of the fundus is not due to the blood vessels but to visual red and I was almost ready to give it up completely when I found that the ophthalmoscopic examination of frogs that had been in sunlight or in the dark showed no difference in the color of their eyegrounds. Both looked like the same bluish-grey ink. Therefore it seemed clear that the red color of the illuminated fundus was entirely independent of visual red and that the latter must for some reason be inaccessible to ophthalmoscopic examination. But this conclusion was premature and I soon was able to convince myself that I was probably dealing only with a specific peculiarity of the frog eye and not with a general property of visual red. When I prepared an isolated frog eye in such a way that daylight could fall directly onto the retina (by cutting a small window in the lateral wall of the bulbus) and I then looked through the cornea, pupil and the lens into the fundus of the eye, by this method of illumination it also had a very bright bluish-grey hue, irrespective of whether the frogs had been exposed to sunlight or had been kept in darkness. Their visual red could never be detected by these methods of illumination and observation” even when further examination demonstrated that it was present in quantity. But when I performed the same experiment with the excised eye of a dark adapted mammal, e.g., a guinea pig’ 2, the fundus did not look bluish-grey, as in frog? 3. but quite distinctly red Moreover, this red color was definitely due to the presence of visual red and did not arise from the blood vessels. which are always completely collapsed and anemic in excised eyes. The same red color can also be identified with a head-mirror in the excised eyes of dark adapted mammals; whereas the fundus of excised eyes, which had been previously exposed to light, looked not red but pale. whether it was viewed directly through the cornea, pupil and lens or examined with an ophthalmoscope.14 Therefore it is obvious that the red color of the fundus which can be seen with the head-mirror in living mammals or humans is a mixed phenomenon produced by the cooperation of two factors, the blood vessels and visual red, to which a third usually is added, namely the red hue of the light source. The latter is readily eliminated by using white light or monochromatic light that is not red, so that in each experiment one need only decide how much of the red color of the fundus is due to visual red and how much to the blood vessels. Simple reflection and direct observation suggest that there must be great fluctuations in this regard. In eyes whose visual red has been completely or almost completely depleted by illumination, the red color of the fundus must arise solely from the blood vessels, whereas in rested eyes the optical effect of visual red must be added to that of red blood. In fact I have been able to see clearly in humans that in the morning, immediately after waking in a dark room’ ‘, “eye red” (as I shall call the red cola!- of the ophthalmoscopic view) is much

On the anatomy and physiology of the retina

more intense

than during the day, after visual red has been used steadily in the light With this decisive observation I was satisfied and

did not perform further ophthalmoscopic examinations, partly bemuse, having once established this principle, I thought I could well leave the further investigation to ophthalmologists with more experience in this area and partly because I lacked a suitable instrument with which to tackle the individual questions with sufhcient scientific rigor. One such instrument is the “ophthalmospectroscope” which I constructed a spectroscope that has a drilled concave mirror fastened in front of its slit I constructed such an instrument, of necessity, by attaching the concave mirror of an ordinary Liebreich ophthalmoscope to a small hand spectroscope. With this inst~m~t I could clearly demonstrate the absorption bands of hemoglobin in the light that was reflected back from the eye of an albino rabbit. I did not succeed in making further observations with my insufficient instrument, apparently bemuse it was very poorly centered With a correctly centered ophthalmospectroscope, in which the focal point of the attached concave mirror coincides precisely with the axis of the spectroscope, it should be easy to answer all questions concerning the nature of visual red by establishing in each case the characteristics of the light leaving the eye and its positive and negative deviations from the spectrum of the light source used to illuminate it. I was prevented from performing such a series of experiments, first of a11because I lacked a well made instrument, but also because I was busy with experiments concerning another problem relating to visual red; namely an examination of the way in which it is changed by.llght of different colors. I performed this series of experiments no less than three times, each with approximately 50 frogs. The first time I undertook them (in December, 1876) I was led by flawed experiments to the mistaken notion that the changes of visual red, its destruction by light and its recovery in the dark, generally require much more time than the few minutes and hours which later, more exact experiments showed to be the extreme time limits Starting from this ~~on~ption I had If, In connection with the experiments with the sun’s spectrum in the darkroom, I made a most peculiar observation which perhaps may have some use for practical ophthalmology (in the diagnosis of color blindness and the likcj On a white wall at a distance 4-5 m from the prism I produee as bright and big a sun spectrum as possible and place my eye into it by looking at the prism while I accommodate for infinity. I then see a strongly luminous center. surrounded by an area of more weakly luminescent points, like a mosaic. I think that it is legitimate to interpret this image as a reproduction of the mosaic of the yellow spot because the diameterof the entire apparition is different in the various spectral colors into which I bring the eye. The luminous surround is small in red light, increases in yellow and becomes largest in green light, then diminishes again in blue and even more in violet light. These facts agree so well with the results obtained by a different and much more laborious method regarding the differences in sensitivity of the retinal periphery to different colors that I have used this as a lecture demonstration in order to demonstrate to everyone at a stroke the entire theory of the localization of the color sensations in the retina.

1’59

to assume that it is possible to elicit chronic changes in the visual red of animals which were exposed for long periods, e.g. weeks, to only one kind of light. Therefore I kept frogs in glass containers of different colors and examined their eyes only after 8-14 days. convinced that the changes which I couid demonstrate in them were due to the long times they had spent in single-colored light and hence should be considered chronic. Of course I had to abandon this idea as soon as more exact experiments showed me the appropriate time relationships. I became convinced that I could not expect chronic alterations from this sort of experimental sequence but that, as with Penelope’s labors, each night’s darkness would undo any retinal changes that might have occurred during the day. Naturally from this new viewpoint I could no longer regard the observed changes as chronic and having been begun a week or more before; I had rather to assume that each observation was conditioned by the few hours of exposure to monochromatic illumination that had occurred on the day the eye was examined Since during the entire time in which these experiments were done sunshine was rarely continuous and almost all the days received the average illumination of a white cloudy sky, the results of these experiments and the retinal changes must be regarded as due to several hours of monochromatic illu~nation at a moderate light intensity. In the second series of experiments in which I used the same containers and color filters, I was favored by constant and very intense sunshine and therefore had access to very bright single-colored lights, whose effects I timed precisely in order to determine the difference between the effects of short and more prolonged monochromatic illumination. In this way the first two series of experiments already yielded all the data I needed to determine the effect of every color at a moderate light intensity and long duration, and at high light intensity and short and long durations. The third series of experiments, therefore, was not designed to discover new facts, but to -confirm and secure the results of the first two experiments by means of a better method. The colored filters I had used for the first experiments were in part defective and it was therefore desirable to repeat them with really pure colors I finally did this (in February of this year) by exposing the atropinized eyes of frogs that had been immobilized with curare for a longer or shorter time to the effects of a specific portion of the sun’s spectrum, which I had cast into a darkroom by means of a heavy flint-glass Men: prism.i6 Concerning the objective changes in the rod layer that correspond to the different physiological states of the retina, I ascertained the following facts: 1. Compiete darkness The color of a retina that has been kept in complete darkness is “red” and not purple, as I called it in my first report: for it corresponds not to the color that results from an overlap of the two extreme ends of the spectrum but rather precisely to the middle of the spectral red I call this color “visual red” or the “basic color of the retina”. If one views the mosaic of the rod layer with the microscope, the vast majority of rods exhibit the same red color that is characteristic of the entire retina. Among these red rods

1260

FRANZBOLL

there are occasional ones that have a very pale green color. If one follows the fading of the retina under the microscope, one sees that as the red rods lose their color, they become distinctly yellowish red and fmally almost completely yellow. Usually the color of the green rods disappears even sooner than that of the red ones. When the whole, unmagnified retina fades, its color is the same as that of the fading red rods. II. White sunlighr After prolonged action of direct sunlight or bright diffuse daylight the retina looks completely colorless; as it dies it assumes not a yellowish but a pure white satiny sheen. No microscopic difference can be detected among the rods; all appear uniformly colorless and transparent. III. Colored light (1) Red fight. In red light the red color of the retina is intensified and turns into a hue that is deeper and darker than the basic color of the retina: one could designate this tint, which is exactly like a certain shade of so-called Pompei red, as “red brown”, indeed almost as brown. This change is more intense the stronger the red light and the longer it has acted When fading, the retina first assumes a yellow-red and finally an almost a brown-yellow hue, which differs from that of the fading basic color by its greater saturation and by the lack of any hint of red Under the microscope the red rods show the same red-brown color that is peculiar to the whole retina: the green rods that are distributed among them are of a much livelier color than the green rods in a retina that has been kept in the dark. (2) Yeflow light. Even at the highest intensity and after prolonged exposure, yellow light changes the basic color of the retina only slightly. Whereas red tight deepens the basic retinal color, yellow lightens it, so that normal visual red can be regarded as almost exactly half-way between the changes produced by red and by yellow lights. Therefore the color elicited by the action of yellow light is best designated as a more transparent visual red. It also changes to yellow-red and yellow as the retina fades. The green rods look exactly the same after exposure to yellow or red light.” (3) Green light. When the retina is exposed to green light there is a distinct dilIcrence depending on whether the light has been more or less intense or has acted for a longer or shorter time. The first effect I’ The fact that red and yellow light alter the basic color of the retina so very slightly has an immediate and very important practical consequence to prepare the retinas and experiment with visual red under total exclusion of daylight and only under artificial illumination with reddish yellow candle- or gaslight. I* It would be very desirable if these experiments were repeated soon with a-quartz prism; until now I have been unable to obtain one here in Rome. I9 When I finished my first series of experiments, all of which had been done with colored lights of moderate intensity, I believed that each primary color elicits a specific corresponding color change in the retina and that the complete loss of retinal color cannot be achieved by any single -continued

on page 1261.

of a very bright green light (or its equivalent-a longer exposure to green light of medium intensity) is to change the basic color of the retina to “purplered”, which fades to a pretty, in the end very pale. rose color, but never to yellow. After prolonged action of very intense green light, the retina does not remain purple-red but becomes a dull violet; this tic+ let becomes paler and paler until the retina finally looks almost completely colorless. Under the microscope the red rods display at different times the hues that correspond to these color changes. The green rods look peculiarly turbid dark green: in this prep aration, they often are still colored intensely green when the surrounding red rods are already extremely pale. It has seemed to me as if their number had increased considerably in comparison with a retina that has remained in darkness or in red and yellow light. (4) Blue and uiolet light. Just as with green light. when using blue and violet light one must consider its intensity and duration in addition to the quality of the exposure. Under dim blue and violet lights or very bright but only brief ones, the basic color of the retina is changed to dirty “violet” in the microscope. But with prolonged exposure to intense blue and violet, this retinal “violet” becomes paler and paler until finally the retina appears completely colorless Microscopic examination gave me the same impression regarding the proportion of green rods as with retinas exposed to green light: their number seemed approximately doubled as compared with retinas that had remained in darkness or in red or yellow light Their color is the same as after exposure to green light. The majority of the other rods under the microscope has a completely clear, bluish-red hue which fades to a distinctly light violet. While they fade, green rods usually retain their color for a considerably longer time than red rods and they still look distinctly green when the others are already nearly colorless This is especially prominent in those retinas which faded in response to in uivo illumination with only blue and violet light; it is less pronounced when blue and violet light acted only moderately on the living retina. Complete fading took place only on the slide in daylight, not in life. (5) Ulrmoiolet rays. According to my experiments, ultraviolet rays have no physiological effect on the living retina and cannot change the basic color in any way even after prolonged irradiation. r ’ These observations regarding the objective changes of the rod layer under the influence of various colors of light can be summarized as follows: The basic color of the retina is changed in different ways by lights of various wavelengths. All long wavelength rays shift the basic color towards the less refrangible parts of the spectrum while intensifying it. All shorter wavelengths change it toward the more refrangible parts of the spectrum and at the same time weaken it. Probably the wavelength as well as the light intensity are significant in both kinds of changes. This can be. proved with certainty at least in the shifts toward the more refrangible parts of the spectrum: the same degree of change can be effected with a more intense (and longer) exposure to less short wavelengths (green) as with shorter wavelength lights (blue and violet) at a decreased, intensity and duration.”

On the anatomy and physiology of the retina

it is not possible to correlate the fact that the physiological destruction of visual redZo increases steadily as one moves to shorter wavelengths, with the continuously increasing chemical effectiveness of sunlight in this part of the spectrum; for the physiological destruction of visual red stops at the limit of the visible spectrum, whereas the chemical effect is known to extend far beyond it.” However,

Besides these chemical changes in the rod layer, light, also evokes in it a second series of changes, which may be no less remarkable than the first: I

have found that in the frog the epitheliil pigment that is dist~buted within the rod layer is not stationary but moves and behaves quite differently in response to different conditions of retinal illumination. In the courSe of my investigations I could not help but notice the fact that the preparation of the retina proceeded quite differently under different physiological conditions. In eyes that had been kept in darkness the retina, inclu~ng the rod layer, always came ioose from the pigment epithelium easily and as a continuous membrane and microscopic examination showed it to be almost completely free of epithelial pigment. This was even more true if the retina had been in red light; less so, after yellow light. By contrast the preparation was much less successful with retinas that had been bleached by daylight or by green, blue, or violet light: the retina usually tore into several pieces which had variable amounts of pigment epithelium stuck tightly lo them. From these observations I concluded that light changes the consistency of the retina and the pigment epithelium in the following way: in white, green, blue and violet light there is a softening of the rod layer as well as of the substance of the pigment epithelium, so that both continuedfrom page 1260. color but requires the concerted action of all colors, i.e.

white tight. Neither assumption was sustained by the results of the hter experiments at more intense illuminations, which showed that the same kind of effect on the basic color of the retina must be attributed to all short wavelength rays, albeit to different extents. Not only white daylight, but also violet and blue, as well as to a lesser extent green, are able to effect the complete bleaching of the retina. Under these circumstances a characteristic change in the basic retinal color can in fact be attributed only to ted (and perhaps also to yellow), whereas the shorter wavelength colors, such as green, blue and violet. are unable to elicit speci& and definite changes in the basic color. The changes produced by the latter colors are characteristic only in a relative sense (at the same intensity and duration of the colored light in questionh never absolute. 2oThe physiological benefit of the yellow pigment of the nmcrda Iurea that is located in the anterior retina1 layers apparently must be sought in the fact that it shields visual red against blue and violet lights ‘i The alterations of the green rods seem to obey similar rules as those of the red ones Their pale green basic color (which becomes apparent after prolonged periods of darkness) is also changed in two ways: namely by the long wavelength rays (red) to an intense tight-green and by the

shorter wavelengths @mea, blue, and violet) to a dull darkgreen. Therefore I must correct the statement in my second report, in which I ascribed to red and green light the same effect on the basic color of the green rods and in which I assumed that a different change took place only in blue and violet light.

become more fragile and tend to stick together

1261 when

one tries to separate them. I felt forced to attribute the opposite effect to darkness, as well as to red and to a lesser degree to yellow light, a hardening of the rod layer and the pigment epithehum; and I assumed that during the detachment of the former. the epithelial processes are drawn out from among the rod layer. However along with this explanation I could also imagine another, namely the assumption that the pigment granules change their position in the light. Ihis latter, bolder hypothesis was more correct: for from a considerable number of eyes that had been fixed in alcohol, taken from frogs that had been kept, in &rkness or red light, and others from frogs that had been kept in white, blue, or violet light, the fact emerged quite unequivocally that depending on the different physiological states of the rod layer, the dis-

ifibution of epithelial pigment was also different: in eyes from the first group the spaces between the rods were always completely free of pigment, whereas in the latter, dense brown strands of pigment extended to the bases of the rods and to the external limiting membrane. This important observation, which makes it highly probable that the epithelial cells participate directly in the visual act, was followed immediately by others that were no less interesting. Like the pigment granules of the hexagonal epithelial cells, one can also show that the oil droplets that are contained in the same cells have quite specific relationships to the physiological processes that take place in the rod layer. These structures, which in frogs are known to have a magnifment golden yellow cobr, have long been the subject of my special attention and I had often indulged in the most daring physiological speculations about their function. The above mentioned reaction with acetic acid by which visual red is changed to an intense golden yellow that looks entirely the same as the color of the oil droplets finally showed me the correct way to understand the physiological signi&ance Of these pu22hng structures: namely, that their pigment is the substance from which visual red is constantly regenerated as it is consumed by light. If this was the correct hypothesis, anatomical examination would have to provide quite defmite evidence for it: this material must be more

~~tif~y visible in rested, red retinas, and sparser in those that had lost their coior in the light, and finally scatcest in eyes which had been put back into the dark to regenerate visual red after many hours of intense illumination and which were examined when the regeneration of visual red had only just been completed (that is, after about two hours). In fact, microscopic examination showed di&rences that

agreed with these expectations. It was impossible to show, as I had initially expected, a striking di3&nce in the number of oil droplets, e.g. a numerical decrease, in retinas that *had been active: the individual variations seem to be too great for the physiological decreases and increases to show thenAves clearly. Occasionally I found ~~~~y more oil droplets in a few retinas that had been strongly ilhuninated than in those that had long been resting. But another

change appeared that is much more substantive than a merely quantitative difference; in dark adapted frogs all oil droplets have the same homogeneous

1262

FRANZBOLL

color, whereas in frogs which one might expect to have used this material the epithelial cells contained pale yellow and even many completely colorless droplets from which all the yellow pigment had been extracted, in addition to the intense goldyellow droplets. This observation makes it extraordinarily likely that there is a direct relationship between visual red and the gold-yellow pigment in the oil droplets. This makes the chemical reality of erythropsin and the photochemical theory of the light sense extremely probable.22 All these observations as well as those on the outer limbs and the components of the pigment epithelium, e.g., the pigment and the oil droplets, show for the first time that in the sense organs various physiological states occur in parallel with changes in their material composition. Thus they lill exceedingly well a serious gap in theoretical physiology by proving that just as in the muscles and the electric and luminous organs which are innervated by centrifugal deeply gold-yellow

‘* Further experiments have made it very likely that the gold-yellow pigment of the oil droplets of the pigment epithelial cells is also photosensitive and approximately to the same, slight extent as is the modified form of visual red that is produced by acetic acid This fact has led me to a very simple picture regarding the significance of the various forms which the light sensitive structures assume within the different classes of vertebrates. In those classes in which both the substance that is layered in the platelets and visual red are either slightly developed or absent (birds and reptiles), the colored oil droplets are located in the visual cell layer itself and presumably serve directly in the sensation of light and color. A higher stage of development of the organ of vision is represented by those classes of vertebrates (mammals, amphibia, fishes) in which the pigmmt of the oil droplets is not located within the visual cell layer itself and bears no direct relationship to the visual act, but is stored within the epithelial cells and represents the material from which the more photosensitive visual red is formed, 23 It is perhaps not an improbable assumption that simple light sensitivity, the differentiation of light and darkness, is mediated exclusively or primarily by states of excitation of the pigmmt epithebal cells, whereas the various qualities of the tight sensation, i.e., the colors, are signalled by alterations of visual red; in what way the behavior of the rods and cones diners in this latter process I cannot guess. ‘a Recently Schwalbe has described two anatomically completely different kinds of rods in the frog retina (Grtije und Siimish, Handbuch der gesamtnten Augenh+lkunde, I., p. 406, 1874). ” I am obliged to be so hesitant concerning the occurrence of green rods in other classes of vertebrates because I have so far been able to examine only those with very thin rods. Among mammals I have had available only the mouse, rat, rabbit, guinea pig, bat, dog and cat, and in the rods of all these animals visual red disappears so extraordinarily fast that, though I never could discover even a trace of grem rods, I still have to leave open the question of their existence. With the extraordinarily rapid disap pearance of the color in the examined rods it remains possible to assume that the green rods only became unrecognizable, but not that they were never present in the examined retina. With somewhat greater certainty than for mammals, I believe I can assert the lack of green rods in cartilaginous fishes. Their rods are sufiicimtly strong to maintain their color for a short time both under the microscope and under the pressure of a cover slip, and always, without exception, they look pale reddish.

nerve fibers so also in the sense organs that are supby centripetal fibers, quite specific material, physical, chemical and anatomical changes correspond to the physiological states of rest and activity. That such changes must take place, of course, could be deduced (I priori from the law of conservation of energy: for it was inconceivable that the conversion of the different physical agents (e.g., light and sound waves) into nervous activity that occurs in the end organs of the sensory nerves could take place as though they were immaterial and without simultaneous objective changes in the end organs. But until now, such changes had not real!y been shown to take place. These changes in the rod and epithelial layers of the retina which I have described above form the material basis upon which it will be possible to construct a complete physiological theory of vision and color sensations. But too many difficulties still lie in the way of establishing a direct relationship between the physiological processes and the material changes to conceive of elaborating such a theory of vision at this time. Two of the most difficult questions that arise in this connection and that will be soluble only after very extensive and prolonged investigations, concern (1) the different significance of the rods and cones23 and (2) the function of the green rods which one always observes in the retinas of amphibians. Should one, quite aside from the cones, distinguish two morphologically24 and functionally different kinds even among rods, the majority red and the minority green? Or should one not rather assume a fundamental identity of all retinal rods and consider the red and green ones only as different manifestations of identical structures that are evoked by different physiological conditions and perhaps also by the regenerative processes? This latter alternative is supported by the fact that no difference among the rods can be observed in a retina that has been exposed to white sunlight, so that it therefore contains only a single class of these elements. This same view is supported by the previous observations concerning the increase in the number of green rods produced by green and blue light But unfortunately it must be said that just these latter observations cannot yet be considered absolutely certain, because, for many reasons, it is very likely that the proportion of green and red rods in each retina is not uniform, but is different in the different regions, in the center and the periphery. But if this is so, it at once become ‘precarious to compare two retinas with respect to their relative abundance of green rods, and I therefore dare speak only with great reservations for the objectivity of my observations regarding the increase in the number of green rods in green and blue light. However as long as the significance of the green rods is not &r&d, indeed as long as one does not even know whether they occur only in amphibians or also in the higher and highest vertebrates, i.e., in mmmals and particularly humans,25 it will be very difficult to exploit these results for a theory of color sensation. The next job in this field must be to carry out the same series of investigations as have been done with frogs, with an animal whose retina is as close as possible to that of humans, hence on monplied

On the anatomy and physiology of the retina

keys. Perhaps it will be possible there to obtain results that bear a simple relationship to the facts about the color sensations of the human retina that have been derived from subjective observations. Given this agreement, it might be possible to deduce a securely based theory of color sensation. At the moment it is unfortunately hardly more than an idle occupation if one were to assume a priori that the facts observed in frogs are essentailly like those in humans26 and wished from this point of view to examine how to relate them to the most important data of the contemporary physiological science of color, namely tbe phenomena of color contrast and cf after-images, as well as the Young-Hehnholtz theory. In some respects these subjective data signi& cantly contradict the results of objective observations and it does not seem easy to devise a theory by which both can be fitted together in a really satisfactory way and brought into agreement. Only in one instance does there appear to be obvious agreement: in reference to color blindness The fact that green rays evoke only a slight alteration in the basic color of the retina, but blue and violet a much stronger one, could contain the direct explanation of why the great majority of colorblinds cannot specifically distinguish red and green, whereas only very few colorblinds confuse red and blue. Accordingly one would have to conclude that this latter abnormality rep resents the highest degree of color blindness which always includes red-green blindness as a lesser component However it would be premature to venture any further in this field and perhaps try to explain some other facts of physiological optics on the basis of the new discoveries about the processes that occur in the rod layer. Therefore I shall abstain from any further arguments regarding this subject and will conclude by calling attention to two more viewpoints which forced themselves upon me repeatedly during my investigations, and with increasing probability, and whose discussion seems to me to be of some use for the general physiology of sensations. The first of these considerations concerns the locus of sense perception. Modem sensory physiology is dominated by the idea that to the terminal extensions of the sensory nerves within the sense organs, such

1263

as the retinal mosaic and the sensory cells in the cocblea, there correspond central end organs within the center, which to a degree reproduce anatomically the pattern of the sensitive points in the periphery; and one imagines that the mind abstracts its impressions and perceptions only from the physiological excitation of these representatives of the sensitive terminals which are located within the central organ. One therefore assumes for each sensory perception a double process, e.g. in vision a specific excitation in the end organs of the optic nerve, which is conducted by the optic nerve fibers to the brain where it reproduces itself once more in the “central end organ”, and one allows the mind to construct its sensations only from this second, duplicated excitation process, so to say from the copy of the first excitation that occurred in the periphery. No one will deny that ihis entire formulation is arbitrary. I want to go further and suggest that it is also useless and complicates rather than simplifies the question regarding tbe nature of sensations. The great puzzle regarding the nature of perception remains unresolved whether the sensory impression is produced in the periphery or in the center, since moving it into the center only pushes the problem into a different place but does not solve it. One must still determine how the mind grasps the picture that is reproduced in the central organ. Therefore I find it simpler to assume that the quality of the sensations is already established in the retina itself and that the mind reads the different states of the sensory nerve endings directly in the periphery, so that they do not need to be registered centrally in a specific receiving device from which they must then be transmitted to the mind to be perceived2’ From this point of view one would have to assume that the changes that occur in the end organs of the sensory nerves are transformed directly into consciousness. With reference to this conversion into consciousness one can imagine two routes Gn the one hand one can assume that the mind treats the changes which occur in the sense organs during physiological activity as raw material which it arranges by itself and from which it produces sensations by interpreting these changes in the way that it is able. In this case there need be no detkrite relationship between the nature of the material changes that take place in the end organ and the nature and quality of the sensory ” Supplementary Note. That visual red also occurs in process; just as there need be no relation between the human retina was ascertained objectively, in addition the form of a printed word and the nature of the to the results of ophthalmoscopic examinations, by Schenk thing it signifies. Indeed in this “interpretation and Zuckerkandl on the occasion of an execution that took theory” one could even imagine a fundamental differplace in Vienna on 5 March, 1877. (Wiener med. Wochenence and even a complete contradiction between the schrgt, No. 11. 13 March, 1877.) objective nature of the signal and the manner in ” If it were correct to assume the existence of special which the mind interprets it From this viewpoint it central end organs for the sensory nerves, one would expect that the origins of the optic and acoustic nerves would not be unthinkable that the mind interprets in the brain would show a degree of complication and the C-vibrations of an auditory hair as the tone, A, a richness of structure that correspond to the large variety and conversely the A-vibrations of the hair as the and quantity of the sensations that would need to be reprotone, C; cooling the nerve endings that serve temperaduced. But this is not at all the case: on the contrary, ture sensations could be interpreted as an increase the origins of these nerves are not at all different anatomiin temperature, warming them as a decrease. And in cally from the origins of ordinary sensory nerves the field of color sensations one wodd have to SupplementcvyNote. Only after 1 had written this footassume that the yellowing and blueing of the retina note did I become aware of the nice paper by W. Miiller need not necessarily signify yellow and blue to the in which the same view is expressed, though for different reasons (Die Stommesentwicklungdes Sehorgones innerhalb mind, but quite arbitrarily, for instance, red and green des Typw der Wirbelthime, Leipzig, i875. p. 52) or even the reverse, blue and yellow.

1264

FRANZ BOLL

So much for the first possibility, which I would like to call the “interpretation theory” and which has until now exclusively dominated sensory physiology. Against this. the “identity theory” emphasizes the ides of a detinite and necessary connection between the material process in the sense organ that accompanies the sensation and the image it evokes in the mind. Many of the details reported in this communication are of significance for such a theory; also from the domain of the other senses, especially for the sensa‘a This discussion regarding the interpretation and identity theories does not refer so much to the differentiated sense organs of humans as to the development of sensory mechanisms and their specific energies among animals in

general.

tions of hearing, taste, and temperature. many facts can be related within this viewpoint: “That by the influence of different agents (light and colors. sound waves. heat, savory substances) objective changes are evoked in the end organs of the sensory nerves which are identical with the content of the sensations and subjective perceptions that they evoke.” If it became possible really to establish this concept completely for the individual sense organs, it would immediately provide an entirely new solution to the ancient question regarding the reality of the content of our sense perceptions2s Rome. 6. March 1877.

The series of colored fields designated la, 2a, 3a, and 4a represents at the original intensity the color of the retina and the digerent physiological changes that are inflicted on it by colored light. Fig. la corresponds to the basic color of the retina or visual red. Above it, Fig 2a represents the hue that results from the prolonged e&t of red light Both lower fields. Fig. 3a and 3b. represent the different degrees of the changes evoked by short wavelength light; Fig. 3a corresponds to the hue produced by a short or dim irradiation with green light; Fig. 4a represents the further color change that develops after a longer or more intense exposure to green light or after a shorter and less intense irradiation with blue and violet light. The adjacent series of colored fields denoted by 1b, 2b. 3b and 4b reproduces the colors which visual red and its various physiological modifications assume as they fade. Each faded color corresponds to the adjacent darker field Thus Fig. lb represents the hue assumed by visual red as it fades; Fig. 2b corresponds to the color which is characteristic of the fading of the modified form of visual red that is produced by red light; Fig. 3b represents the faded color of Fig. 3a, and Fig 4b, the faded color of Fig. 4a. This latter field, 4b, also represents the highest degree of the physiological alteration of visual red by short wavelength light and therefore could readily be placed into the first series of colored fields as the final number, 5a. The figs numbered 5 to 9 reproduce the mosaic of the rod layer of the frog in its different physiological states. Fig. 5 corresponds to a retina that has been in the dark for a long time. The color of the red rods corresponds to visual red; the few green rods are very pale. Fig. 6 represents a retina that was exposed to red light for a relatively long time. The red rods appear a darker reddish brown; the color of the green rods seems considerably more intense. The illustrations in Figs. 7-9 represent three sequential stages of the changes that occur in the retina following exposure to short wavelength tight. In all three the number of green rods seems greater and their color a dull dark-green. The red rods appear purplish red in Fig. 7, red-violet in Fig. 8, and pale violet, indeed almost colorless, in Fig. 9. (Color plate by the courtesy of Prof. Ch. Baumann.)