Quantum Relations of the Rat Electroretinogram

The rat retina is uniform and contains almost exclusively rods. Therefore the rat eye, when uniformly illuminated, produces a gross electroretinogram (ERG) which is simply related to the activity of the individual retinal sources of the ERG. Characteristics of ERG's are shown on an intensity scale of the average number of quanta absorbed per rod per stimulus flash obtained by direct accurate measurement of all quantities involved. An independent check on the accuracy of these measurements is applied to pigment-bleaching data reported by Dowling (1963). When ERG characteristics are placed on this scale it is found that: (a) The b-wave can usually be observed when fewer than one out of two hundred rods absorbs a quantum, the threshold being determined by the noise of the preparation. (b) Near threshold the b-wave amplitude is proportional to intensity. (c) The a-wave appears when there are more than two to four absorptions per rod per flash. (d) The b-wave latency decreases with intensity, and the amplitude becomes proportional to the logarithm of intensity when fewer than one out of ten rods absorbs a quantum. This implies that the b-wave sources must combine excitation from more than one rod (probably more than seven). Therefore the b-wave cannot arise from independent rods or rod-bipolar synapses, but probably reflects activity of entire inner nuclear layer cells.     

The Rat Electroretinogram : I. Contrasting effects of adaptation on the amplitude and latency of the b-wave

Characteristics of the electroretinogram (ERG) produced by the essentially all rod eye of the rat are presented as functions of the number of quanta absorbed by each rod per stimulus flash. The ERG's were obtained with 1.5 msec. stimulus flashes and uniform illumination of the entire retina. Under these conditions, distortions in the ERG due to stray light are minimized, and the ERG more accurately reflects the activity of its retinal sources. The effects of background light and two forms of dark adaptation were studied and compared. The results, especially for the b-wave, permit an interpretation in terms of two distinct processes. One process appears to determine the b-wave latency. This process is almost independent of the state of adaptation of the retina. The other process does not affect the latency, but determines the b-wave threshold and amplitude. This process strongly depends upon the state of adaptation. Moreover, the effects of dark adaptation on this amplitude-determining process are almost identical with the effects of background light.     

A Common Fluence Threshold for First Positive and Second Positive Phototropism in Arabidopsis thaliana

The relationship between the amount of light and the amount of response for any photobiological process can be based on the number of incident quanta per unit time (fluence rate-response) or on the number of incident quanta during a given period of irradiation (fluence-response). Fluence-response and fluence rate-response relationships have been measured for second positive phototropism by seedlings of Arabidopsis thaliana. The fluence-response relationships exhibit a single limiting threshold at about 0.01 micromole per square meter when measured at fluence rates from 2.4 × 10⁻⁵ to 6.5 × 10⁻³ micromoles per square meter per second. The threshold values in the fluence rateresponse curves decrease with increasing time of irradiation, but show a common fluence threshold at about 0.01 micromole per square meter. These thresholds are the same as the threshold of about 0.01 micromole per square meter measured for first positive phototropism. Based on these data, it is suggested that second positive curvature has a threshold in time of about 10 minutes. Moreover, if the times of irradiation exceed the time threshold, there is a single limiting fluence threshold at about 0.01 micromole per square meter. Thus, the limiting fluence threshold for second positive phototropism is the same as the fluence threshold for first positive phototropism. Based on these data, we suggest that this common fluence threshold for first positive and second positive phototropism is set by a single photoreceptor pigment system.     

Internal conversion in the photosynthetic mechanism of blue-green algae

1. In Chroococcus a quantum of light absorbed by phycocyanin has 90 per cent the chance of doing photosynthesis that a quantum absorbed by chlorophyll has. 2. By a process analogous to internal conversion in radioactivity (but with the linear dimensions and the wave length 10⁴ times larger) there will be transferred from phycocyanin to chlorophyll See PDF for Equation (a number of the order of 100) quanta for every one emitted as fluorescent light by the phycocyanin in the Chroococcus cell. 3. The yield of fluorescent light in Chroococcus is between 1 and 2 per cent. 4. The transfer of energy by internal conversion can account for the photosynthesis by phycocyanin observed by Emerson and Lewis.     

RESPIRATORY EFFECTS UPON THE VISUAL THRESHOLD

Measurements are reported of the effects of respiratory stresses upon the absolute threshold of peripheral (rod) vision. Since subjects were kept wholly dark adapted and the photochemical system of the rods therefore stationary, the changes recorded may be assumed to have originated more centrally. To this degree the measurements provide a quantitative index of central nervous imbalance. Breathing room air or 32 to 36 per cent oxygen at about double the normal rate causes the visual threshold to fall to approximately half the normal value within 5 to 10 minutes. This change is due primarily to alkalosis induced by the hyperventilation, and can be abolished or reversed by adding carbon dioxide to the inspired mixtures. Normal or rapid breathing of 2 per cent carbon dioxide causes no change in threshold; with 5 per cent carbon dioxide the threshold is approximately doubled. Breathing 10 per cent oxygen at the normal rate also approximately doubles the threshold. This effect is compensated in part by rapid breathing. When 10 per cent oxygen is breathed at twice the normal rate the threshold usually falls at first, then slowly rises to supernormal levels. Due primarily to variations in their breathing patterns subjects yield characteristically different responses on sudden exposure to low oxygen tensions with breathing uncontrolled. The threshold may either rise or fall; and on release from anoxia it may rise, or fall to normal or subnormal levels. The threshold adjusts to anoxia rapidly; exposures lasting 5 to 6 hours do not produce greater or more persistent changes than those of much shorter duration.     

THE TOTAL LUMINOUS EFFICIENCY OF LUMINOUS BACTERIA

Methods are described for measuring the light emitted by an emulsion of luminous bacteria of given thickness, and calculating the light emitted by a single bacterium, measuring 1.1 x 2.2 micra, provided there is no absorption of light in the emulsion. At the same time, the oxygen consumed by a single bacterium was measured by recording the time for the bacteria to use up .9 of the oxygen dissolved in sea water from air (20 per cent oxygen). The luminescence intensity does not diminish until the oxygen concentration falls below 2 per cent, when the luminescence diminishes rapidly. Above 2 per cent oxygen (when the oxygen dissolving in sea water from pure oxygen at 760 mm. Hg pressure = 100 per cent) the bacteria use equal amounts of oxygen in equal times, while below 2 per cent oxygen it seems very likely that rate of oxygen absorption is proportional to oxygen concentration. By measuring the time for a tube of luminous bacteria of known concentration saturated with air (20 per cent oxygen) to begin to darken (2 per cent oxygen) we can calculate the oxygen absorbed by one bacterium per second. The bacteria per cc. are counted on a blood counting slide or by a centrifugal method, after measuring the volume of a single bacterium (1.695 x 10^–12 cc.). Both methods gave results in good agreement with each other. The maximum value for the light from a single bacterium was 24 x 10^–14 lumens or 1.9 x 10^–14 candles. The maximum value for lumen-seconds per mg. of oxygen absorbed was 14. The average value for lumen-seconds per mg. O₂ was 9.25. The maximum values were selected in calculating the efficiency of light production, since some of the bacteria counted may not be producing light, although they may still be using oxygen. The "diet" of the bacteria was 60 per cent glycerol and 40 per cent peptone. To oxidize this mixture each mg. of oxygen would yield 3.38 gm. calories or 14.1 watts per second. 1 lumen per watt is therefore produced by a normal bacterium which emits 14 lumen-seconds per mg. O₂ absorbed. Since the maximum lumens per watt are 640, representing 100 per cent efficiency, the total luminous efficiency if .00156. As some of the oxygen is used in respiratory oxidation which may have nothing to do with luminescence, the luminescence efficiency must be higher than 1 lumen per watt. Experiments with KCN show that this substance may reduce the oxygen consumption to 1/20 of its former value while reducing the luminescence intensity only ¼. A partial separation of respiratory from luminescence oxidations is therefore effected by KCN, and our efficiency becomes 5 lumens per watt, or .0078. This is an over-all efficiency, based on the energy value of the "fuel" of the bacteria, regarded as a power plant for producing light. It compares very favorably with the 1.6 lumens per watt of a tungsten vacuum lamp or the 3.9 lumens per watt of a tungsten nitrogen lamp, if we correct the usual values for these illuminants, based on watts at the lamp terminals, for a 20 per cent efficiency of the power plant converting the energy of coal fuel into electric current. The specific luminous emission of the bacteria is 3.14 x 10^–6 lumens per cm². One bacterium absorbs 215,000 molecules of oxygen per second and emits 1,280 quanta of light at λ_max = 510µµ. If we suppose that a molecule of oxygen uniting with luminous material gives rise to the emission of 1 quantum of light energy, only 1/168 of the oxygen absorbed is used in luminescence. On this basis the efficiency becomes 168 lumens per watt or 26.2 per cent.     

Uridine 5'-phosphate photohydrate formation in irradiated Escherichia coli 30S ribosomes: photochemistry and relation to ribonucleic acid-protein cross-linking

Irradiation of aqueous buffered solutions of Escherichia coli 30S ribosomes with doses of 254-nm radiation greater than 10(19) quanta causes formation of uridine 5'-phosphate (UMP) photohydrates in ribosomal 16S RNA (rRNA). The number of molecules of UMP photohydrate formed at doses less than 2 x 10(20) quanta is linearly dependent on dose of absorbed 254-nm radiation. Maximum UMP photohydrate formation is dependent on initial ribosome concentration. When solutions containing 1 A260 unit of 30S ribosomes/mL were irradiated with greater than 2 x 10(20) quanta of 254-nm radiation, maximum photohydrate formation was equal to 47 residues/ribosome. Irradiation of solutions containing 2 A260 units/mL with greater than 7 x 10(20) quanta caused formation of 102 UMP photohydrates/ribosome. These values correspond to conversion of either 15 or 33%, respectively, of the total UMP content of 30S ribosome 16S rRNA to photohydrates. Target theory analysis of UMP photohydration in 30S ribosomes showed that UMP photohydrates are formed by single-hit kinetics from two photochemically distinct precursors. Of the total 16S rRNA UMP residues, 10% was included in the most rapidly (low dose) reacting fraction. The respective photohydration cross sections are 0.014 (low dose) and 0.0095 cm2/muEinstein (high dose) for ribosome solutions containing 2 A260 units/mL. UMP photohydrate content of irradiated 30S ribosomes was compared with that of previous data for the extent of RNA-protein cross-linking at equivalent doses of absorbed 254-nm radiation. This comparison showed that at least two UMP photohydrates form per RNA-protein cross-linking event in 30S ribosomes irradiated with a dose of 254-nm radiation (1.5 x 10(19) quanta), which causes cross-linking of only three ribosomal proteins to 16S rRNA.     

QUANTITATIVE STUDIES ON THE PHOTOLETHAL EFFECTS OF QUARTZ ULTRA-VIOLET RADIATION UPON PARAMECIUM

Paramecia grown under controlled conditions were irradiated at known intensities of light of wave-lengths 2537, 2654, 2804, 3025, and 3130 A. The approximate absorption of the light by the Parmecia was found to be greatest and of the same order of magnitude at the three shortest wave-lengths, considerably less at 3025, and least at 3130 A. Paramecia did not die when irradiated with high dosages of intense light of wave-length 3130 A. At the other wave-lengths 50 per cent vesiculation occurred when between 10¹² and 10¹³ quanta had been absorbed by a Paramecium. This would indicate that a very large number of molecules in a Paramecium are affected before vesiculation occurs.     

The quantic and statistical bases of visual excitation

1. The photochemical theories of vision cannot provide a valid interpretation of the facts over the whole range of brightness. The fact that liminal excitation is increased by the absorption of a very small number of quanta, each absorbing rod receiving a single quantum, excludes the intervention of the mass action law which is the basis of all photochemical theories. 2. Owing to the quantic structure of light and to the random distribution of quanta in a faint light pencil, there must exist numerical relations between the threshold energy on the one hand and the size of the retinal area stimulated and the stimulation time on the other, whatever may be the inner mechanism of liminal excitation. When taking as a basis Van der Velden's experimental results, viz. that two quanta absorbed during a certain interval of time are sufficient to raise threshold excitation, the probability calculus enables us to compute the course of threshold energy in relation to the stimulation time and to the stimulated retinal area. No arbitrary parameter is needed to do so; the only constant to be used is found by experiment. 3. The quantic and statistical theory of visual excitation that we put forward in the present paper enables us to predict the validity of Ricco's law within what we call a "quasi-independent unit" and the validity of Piper's law within a test area made up of a certain number of such units. This theory does not correspond exactly with Piéron's law for foveal threshold in relation to the size of the stimulated area, but the deviation is probably due to an artefact; viz., the action of the micronystagmus. 4. Experiment proves that in region IV of the retina, 15 degrees temporally from the fovea of the right eye of two observers, Ricco's law applies strictly in rod vision from 2'12' to 31'36' and, perhaps, further on. 5. In the same region, from 12'30' to 31'36', Piper's law applies strictly in cone vision of extremely red light. 6. In peripheral vision with extremely red light the photochromatic interval has been found to be null. 7. Our theoretical interpretation of the term "quasi-independent unit" fits well with the histological data of the retina. 8. Numerical deviations of the theoretic time law of threshold intensity from the empirical course may be due to the existence of a relative refractory period of the ganglion (or bipolar) cells. This mechanism would be a sort of instantaneous adaptation of nervous elements and would explain the fact that the sensation level increases very much slower than the brightness level, in a range of the brightness scale where the photochemical adaptation cannot account for this phenomenon.     

The interplay o light and heat in bleaching rhodopsin

Rhodopsin, the pigment of the retinal rods, can be bleached either by light or by high temperature. Earlier work had shown that when white light is used the bleaching rate does not depend on temperature, and so must be independent of the internal energy of the molecule. On the other hand thermal bleaching in the dark has a high temperature dependence from which one can calculate that the reaction has an apparent activation energy of 44 kg. cal. per mole. It has now been shown that the bleaching rate of rhodopsin becomes temperature-dependent in red light, indicating that light and heat cooperate in activating the molecule. Apparently thermal energy is needed for bleaching at long wave lengths where the quanta are not sufficiently energy-rich to bring about bleaching by themselves. The temperature dependence appears at 590 mµ. This is the longest wave length at which bleaching by light proceeds without thermal activation, and corresponds to a quantum energy of 48.5 kg. cal. per mole. This value of the minimum energy to bleach rhodopsin by light alone is in agreement with the activation energy of thermal bleaching in the dark. At wave lengths between 590 and 750 mµ, the longest wave length at which the bleaching rate was fast enough to study, the sum of the quantum energy and of the activation energy calculated from the temperature coefficients remains between 44 and 48.5 kg. cal. This result shows that in red light the energy deficit of the quanta can be made up by a contribution of thermal energy from the internal degrees of freedom of the rhodopsin molecule. The absorption spectrum of rhodopsin, which is not markedly temperature-dependent at shorter wave lengths, also becomes temperature-dependent in red light of wave lengths longer than about 570 to 590 mµ. The temperature dependence of the bleaching rate is at least partly accounted for by the temperature coefficient of absorption. There is some evidence that the temperature coefficient of bleaching is somewhat greater than the temperature coefficient of absorption at wave lengths longer than 590 mmicro;. This means that the thermal energy of the molecule is a more critical factor in bleaching than in absorption. It shows that some of the molecules which absorb energy-deficient quanta of red light are unable to supply the thermal component of the activation energy needed for bleaching, so bringing about a fall in the quantum efficiency. The experiments show that there is a gradual transition between the activation of rhodopsin by light and the activation by internal energy. It is suggested that energy can move freely between the prosthetic group and the protein moiety of the molecule. In this way a part of the large amount of energy in the internal degrees of freedom of rhodopsin could become available to assist in thermal activation. Assuming that the minimum energy required for bleaching is 48.5 kg. cal., an equation familiar in the study of unimolecular reaction has been used to estimate the number of internal degrees of freedom, n, involved in supplying the thermal component of the activation energy when rhodopsin is bleached in red light. It was found that n increases from 2 at 590 mµ to a minimum value of 15 at 750 mµ. One wonders what value n has at 1050 mµ, where vision still persists, and where rhodopsin molecules may supply some 16 kg. cal. of thermal energy per mole in order to make up for the energy deficit of the quanta.     

An apparatus for measuring photosynthetic quantum yields and quanta absorption spectra of intact plants

Abstract An apparatus is described which consists of an assimilation chamber mounted in the centre of a light-integrating Ulbricht sphere, irradiated through two fibreoptic light pipes. This arrangement provides totally diffuse radiation in the sphere. The quantum flux density in die sphere is measured by an integrating quantaspectrometer connected to the sphere by eight fibreoptic light pipes. The quantaspectrometer gives the spectral and the total quantum flux density in the sphere. By measuring the quantum flux density in the sphere before and after the insertion of a plant the number of absorbed quanta per unit time and plant area can be determined. In addition, the spectral distribution of the absorbed quantum flux density is obtained. CO₂-assimilation per unit time and plant area is determined in the same apparatus. The totally diffuse radiation within the sphere minimizes mutual shading between branches and leaves. Hence, well-defined light-responsecurves of photosynthesis can be obtained by plotting the flux density of C0₂-assimilation as a function of the absorbed quantum flux density, and the quantum yields of intact plants can be calculated. Examples of photosynthetic quantum yield determinations and quanta absorption spectra are given for some plant species.     

INTERMITTENT STIMULATION BY LIGHT : III. THE RELATION BETWEEN INTENSITY AND CRITICAL FUSION FREQUENCY FOR DIFFERENT RETINAL LOCATIONS

When measurements of the critical fusion frequency for white light over a large range of intensities are made with the rod-free area of the fovea, the relation between critical frequency and log I is given by a single sigmoid curve, the middle portion of which approximates a straight line whose slope is 11.0. This single relation must be a function of the foveal cones. When the measurements are made with a retinal area placed 5° from the fovea, and therefore containing both rods and cones, the relation between critical frequency and log I shows two clearly separated sections. At the lower intensities the relation is sigmoid and reaches an upper level at about 10 cycles per second, which is maintained for 1.25 log units, and is followed by another sigmoid relationship at the higher intensities similar to the one given by the rod-free area alone. These two parts of the data are obviously separate functions of the rods at low intensities and of the cones at high intensities. This is further borne out by similar measurements made with retinal areas 15° and 20° from the fovea where the ratio of rods to cones is anatomically greater than at 5°. The two sections of the data come out farther apart on the intensity scale, the rod portion being at lower intensities and the cone portion at higher intensities than at 5°. The general form of the relation between critical frequency and intensity is therefore determined by the relative predominance of the cones and the rods in the retinal area used for the measurements.     

Optomotorische Reaktionen der Fliege Musca Domestica

Summary The torque exerted by the housefly Musca domestica during fixed flight was used as a measure of the optomotor reaction of the insect elicited by the rotation of cylindrical patterns with periodic distributions of surface brightness. Measurements were made of the dependence of the reaction on the wave length, speed of rotation, contrast, and mean brightness of the stimulus patterns. The effect on the reaction of modulation of the light illuminating the stimulus pattern was examined. Further experiments indicated that stimulation of only one of the two complex eyes is sufficient to elicit an optomotor reaction, and that there is overlap between the visual fields of neighboring photoreceptor units in the complex eye. Estimates of the rates of absorption of light quanta by individual ommatidia in the complex eye indicated that these rates are low enough that the Poisson statistics of the light quanta results in a significant level of noise in the light signals received by the photoreceptors, when the brightness of the stimulus pattern is low but still sufficient to elicit a measurable reaction. The contrast that is required of a rotating stimulus pattern in order to elicit a just-measurable reaction was found to depend upon the mean brightness of the pattern in a manner that is consistent with the hypothesis that the noise due to the statistics of the light quanta absorbed by the photoreceptors in the complex eye is a principle cause of the breakdown of the optomotor reaction at low values of the contrast and mean brightness of the stimulus pattern.

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Mit Unterstützung durch ein Fellowship des National Institute of Neurological Diseases and Blindness, US Public Health Service und ein Fellowship der National Science Foundation, USA.

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Herrn Freiberg danken wir für das Anfertigen der Abbildungen.     

Auslösung von Elementarprozessen durch einzelne Lichtquanten im Fliegenauge

Summary Light quanta impinging upon the photopigments located in the rhabdomeric receptor structures of the fly's compound eyes trigger photochemical reactions which in turn elicit miniature receptor potentials (bumps). The paper mainly deals with the problem whether a single quantum of light is sufficient, or whether a coincidence of quanta and/or elementary photochemical events is necessary to trigger a miniature receptor potential.The experiments were based on tests of the optomotor responses of fixed flying flies suspended in a rotating patterned cylinder with periodic distributions of inner surface brightness. The tests were made under two different light programs: 1) Illumination constant in time 2) Illumination by periodic light pulse sequences with various frequencies. Average light fluxes absorbed by the receptors were equal in both programs. Theoretical considerations lead to the following conclusions: The strength of the optomotor responses to the light programs 1 and 2 should not differ from each other in the case of single quantum processes. However for multiquantum processes light program 2 should be more effective than light program 1 as it favours the coincidence of quantum absorptions per unit time. But these theoretical conclusions are valid only if two conditions are fulfilled in the experiments: a) The pulse frequency of light program 2 has to be kept below a certain limit which is determined by the kinetics of the photochemical systems. Otherwise light program 2 gets averaged in time and in principle can be not more effective than light program 1. b) The rates of quanta absorbed by the receptors have to be kept low enough to guarantee that the concentration of unbleached pigment molecules remains practically unchanged as compared with the concentration in darkness. Accordingly the test experiments were carried out with light pulse frequencies ranging from 500 to 1/120 cycles per second. Intensities were used which corresponded to an average quantum flux effective for one rhabdomeric structure ranging between 10 and 250 quanta per second.The interpretation of the experimental results is in accordance with the hypothesis that one single quantum of light is sufficient to trigger an elementary photochemical reaction and that in turn one single photochemical event can elicit a miniature receptor potential. At present time the experiments do not allow conclusions about the possible occurrence of coincidence-functions of synapses at the level of the first optical ganglion which receive their information via fibers leading off from the receptors.In one of the appendices of the paper, the transinformation flux into a receptor is calculated, taking into consideration the Poisson noise of the quanta disrupting the signal at extremely low quantum rates.

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Ein Teil der in dieser Arbeit abgedruckten Ergebnisse wurde bereits in zwei vorläufigen Mitteilungen publiziert, Reichardt (1965, 1966).     

THEORY AND MEASUREMENT OF VISUAL MECHANISMS : I. A VISUAL DISCRIMINOMETER. II. THRESHOLD STIMULUS INTENSITY AND RETINAL POSITION

Monocular threshold stimulus intensities (ΔI_o, photons) were measured along the 0–180° meridian of human retinae for three observers. The test image was small (= 0.08°) and of short duration (= 0.20 second). ΔI_o was found to decrease as the angular distance from the fovea was increased. Actual counts of the number of retinal elements per mm.² along the 0–180° meridian (Østerberg) were compared with the obtained results. No direct correlation was found to exist between visual sensitivity and the number of retinal elements. Binocular threshold stimuli were also measured along the same meridian. The form of the function relating binocular visual sensitivity and retinal position was discovered to be essentially similar to that for monocular sensitivity, but is more symmetrical about the center of the fovea. The magnitude of the binocular measurement is in each case smaller than that of the monocular threshold stimulus intensity for the more sensitive eye. The ratio is statistically equal to 1.4 (a fact which suggests Piper''s rule). These results are shown to be consistent with the hypothesis that the process critical for the eventuation of the threshold response is localized in the central nervous system. They are not consistent with the view that the quantitative properties of visual data are directly determined by properties of the peripheral retina.     

Glucose Absorption, Carbohydrate Accumulation, Presence of Starch, and Rate of Photosynthesis in Barley Leaf Segments

Fresh and dry weight changes in barley leaf segments were measuredafter the segments had been floated on a 5 per cent glucosesolution at the light compensation point (3.I W m^-2) or on distilledwater at a higher illuminance. The accumulation of dry matterhad a small inhibiting effect on photosynthesis when it hadresulted from previous photosynthesis, but when a similar accumulationresulted from a previous absorption of glucose, marked inhibitionof photosynthesis was found which increased with the weightof glucose absorbed. Both the quantitative determination ofstarch in the segments and an examination of the distributionof radioactivity among water-soluble, alcohol-soluble, and insolublefractions after segments had either absorbed ¹⁴C-glucose orused ¹⁴CO₂ in photosynthesis, showed that in the case of photosynthesisa greater proportion of the dry weight accumulation was in theform of starch. However, this starch fraction constituted onlyabout 10 per cent of the total increase in dry weight so thatan explanation of the differential inhibition is unlikely tobe in these terms.     

Performance of cat retinal ganglion cells at low light levels

Responses of brisk-sustained cat retinal ganglion cells were examined using receiver operating characteristic (ROC) analysis. Stimuli were brief luminance changes superimposed upon a weak steady pedestal ranging from 27 to 47,000 quanta (507 nm) per second at the cornea. Overall quantum efficiencies of cells ranged up to approximately 13% and were compatible with previous estimates at absolute threshold. The main work was done on on-center cells, but a small sample of off-center units behaved similarly. Experimental ROC curves verified a set of qualitative predictions based on a theoretical treatment of performance, assuming that response variability resulted solely from quantum fluctuations. However, quantitative predictions were not fulfilled. The discrepancy could be resolved by postulating a source of added internal variance, R, the value of which could then be deduced from the experimental measurements. A ganglion cell model limited by a fixed amount of added variance from physiological sources and having access to a fixed fraction of incident quanta can account quantitatively for (a) slopes of ROC curves, (b) variation of detectability with magnitude of both increments and decrements, and (c) performance over a range of pedestal intensities. Estimates of the proportion of incident quanta used ranged up to 29% under some conditions, a figure approximately matching estimates of the fraction of corneal quanta that isomerize rhodopsin in the cat.     

Kinetics of the Photocurrent of Retinal Rods

The shapes of the photocurrent responses of rat rods, recorded with microelectrodes from the receptor layer of small pieces of isolated retinas, have been investigated as a function of temperature and of stimulus energy. Between 27 and 37°C the responses to short flashes can be described formally as the output of a chain of at least four linear low-pass filters with time constants in the range 50-100 msec. The output of the filter chain is then distorted by a nonlinear amplitude-limiting process with a hyperbolic saturation characteristic. Flashes producing ~30 photons absorbed per rod yield responses of half-maximal size independently of temperature. The maximum response amplitude is that just sufficient to cancel the dark current. The rate of rise of a response is proportional to flash energy up to the level of 10⁵ photons absorbed per rod, where hyperbolic rate saturation ensues. The responses continue to increase in duration with even more intense flashes until, at the level of 10⁷ photons absorbed per rod, they last longer than 50 min. The time-courses of the photocurrent and of the excitatory disturbance in the rod system are very similar. The stimulus intensity at which amplitude saturation of the photocurrent responses begins is near that where psychophysical “rod saturation” is seen. An analysis of these properties leads to the following conclusions about the mechanism of rod excitation. (a) The kinetics of the photocurrent bear no simple relation to the formation or decay of any of the spectroscopic intermediates so far detected during the photolysis of rhodopsin. (b) The forms of both the amplitude- and rate-limiting processes are not compatible with organization of rhodopsin into “photoreceptive units” containing more than 300 chromophores. Even at high stimulus intensities most rhodopsin chromophores remain connected to the excitatory apparatus of rods. (c) The maximum rate of rise of the photocurrent is too fast to be consistent with the infolded disks of a rod outer segment being attached to the overlying plasma membrane. Most of the disks behave electrically as if isolated within the cell. (d) Control of the photocurrent at the outer segment membrane is not achieved by segregation of the charge carriers of the current within the rod disks. Instead, it is likely to depend on control of the plasma membrane permeability by an agent released from the disks.     

Electron Microscopy of the Tapetum Lucidum of the Cat

The fine structure of the tapetum of the cat eye has been investigated by electron microscopy. The tapetum is made up of modified choroidal cells, seen as polygonal plates grouped around penetrating blood vessels which terminate in the anastomosing capillary network of the choriocapillaris. The tapetal cells are rectangular in cross-section, set in regular brick-like rows, and attain a depth of some thirty-five cell layers in the central region. This number is gradually reduced peripherally, and is replaced at the margin of the tapetum by normal choroidal tissue. The individual cells are packed with long slender rods 0.1 µ by 4 to 5 µ. The rods are packed in groups and with their long axes oriented roughly parallel to the plane of the retinal surface. Each cell contains several such groups. Cells at the periphery or in the outer layers of the tapetum are frequently seen to contain both tapetal rods and melanin granules, the latter typical of the choroidal melanocytes. Also melanocyte granules may have intermediate shapes. These observations plus the similar density of the two inclusions lead to the belief that the tapetal rods may be melanin derivatives. A fibrous connective tissue layer lies between the tapetum and the retina. The subretinal capillary network, the choriocapillaris, rests on this layer and is covered by the basement membrane of the retinal epithelium. The cytoplasm of the retinal epithelium exhibits marked absorptive modifications where it comes in contact with the vessels of the choriocapillaris. This fibrous layer and the basement membrane of the retinal epithelium apparently comprise the structural elements of Bruch's membrane.     

Divergent photic thresholds in the non-image-forming visual system: entrainment, masking and pupillary light reflex

Light is the principal cue that entrains the circadian timing system, but the threshold of entrainment and the relative contributions of the retinal photoreceptors—rods, cones and intrinsically photosensitive retinal ganglion cells—are not known. We measured thresholds of entrainment of wheel-running rhythms at three wavelengths, and compared these to thresholds of two other non-image-forming visual system functions: masking and the pupillary light reflex (PLR). At the entrainment threshold, the relative spectral sensitivity and absolute photon flux suggest that this threshold is determined by rods. Dim light that entrained mice failed to elicit either masking or PLR; in general, circadian entrainment is more sensitive by 1–2 log units than other measures of the non-image-forming visual system. Importantly, the results indicate that dim light can entrain circadian rhythms even when it fails to produce more easily measurable acute responses to light such as phase shifting and melatonin suppression. Photosensitivity to one response, therefore, cannot be generalized to other non-image-forming functions. These results also impact practical problems in selecting appropriate lighting in laboratory animal husbandry.