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.     

Detection of red and green flashes: evidence for cancellation and facilitation.

Green and red flashes of light will differentially stimulate the middle- and long-wavelength sensitive cones. Interaction of cone signals was studied by measuring increment thresholds for combinations of green and red flashes on a yellow adapting field. When the yellow adapting field was at 10.000 trolands (td), green and red incremental flashes (1 degree, 200-msec duration) produced cancellation when presented simultaneously and facilitation when presented sequentially. A green incremental flash (1.15 degrees, 200 msec, 5000-td adaptation field) and red decremental flash, or vice versa, produced facilitation when presented simultaneously. The results can be explained by color-differencing, opponent-mechanisms. The cancellation effect for the simultaneous incremental flashes largely disappeared when the flashes were exposed briefly (10 msec) or reduced in size (0.04 degrees). It is unlikely that the stimuli were exclusively detected by achromatic, luminance channels, as suggested by previous work, since observers could partially distinguish the hue of threshold flashes of 570- and 590-nm light (0.04 degrees, 10 msec) on a bright yellow field.     

Neural response mechanisms in the photoreceptive pineal organ of goldfish

In order to classify the different cell types involved in signal transmission of the photoreceptive pineal organ of the goldfish, Carassius auratus, intra- and extracellular electrical responses were recorded from photoreceptors and second-order neurons. Photoreceptor responses to light consisted of hyperpolarizing potentials up to 30 mV. The responses were graded with intensity and their voltage-intensity relation followed the hyperbolic function V/Vmax = In/In + sigma n. Latencies varied between 500 msec for responses near threshold and 60 msec for supersaturating flashes. The response duration increased up to 60 sec for flashes 2 log units above the saturation level. Action spectra of individual photoreceptors peaked at lambda max = 530 nm and corresponded to measurements of extracellular slow mass potentials or spike potentials. Slow mass potentials exhibited similar characteristics as intracellular recorded photoreceptor potentials with respect to latency, voltage-intensity curves and spectral sensitivity. Ganglion cells showed maintained discharges under conditions of steady illumination. The discharge rate changed inversely with the logarithm of steady illumination over a range of 8 log units. The response to light flashes was purely achromatic and consisted of inhibition of the maintained discharge. The physiological properties demonstrate that the pineal organ of the goldfish is an effective functional photoreceptor organ operating both in dim and in bright light. The light-induced hyperpolarization of photoreceptors lead to an inhibition of the nervous discharge of ganglion cells. The direct flow of information from photoreceptors to ganglion cells is the basic channel of data processing in the goldfish pineal.     

The Dependence of the Photopupil Response on Flash Duration and Intensity

The changes in pupil size were recorded by infrared pupillographic methods in response to light flashes of different durations and intensities for a 13 degree 34 minute centrally fixated circular field. For such stimuli, the threshold intensities for (rod) vision and for the pupil response were found to be about the same. The response amplitudes were related to the logarithm of the flash energy, the reciprocity law remaining valid up to about one-half second. The curve relating flash energy and pupil response was clearly divisible into two parts commensurate with the duplex character of the human retina. A similar dichotomy appears in curves relating response amplitude to response latency. Since the pupil response is determined by total flash energy, intense long flashes produce larger pupil responses than shorter (and perceptually brighter) ones of the same intensity.     

A duplex retina and the electroretinogram in the nocturnal Perodicticus potto.

The presence of cones in potto's retina has been proved beyond doubt although they are very restricted in number (1 cone for 300 rods). Morphologically, speaking there is no point in calling these cones "rudimentary" except for their slender outer segment. There are red sensitive elements in that retina at wavelengths beyond the spectral sensitivity of visual purple and it is tempting to assume that these elements are cones. The ERG evoked from these elements by red light differs from that in response to white and blue light. They dark-adapt faster than the receptors sensitive to blue and white flashes. However in some of their properties, for example fusion frequency, these cones behave like rods in other species. As these few cones seem to activate the bipolar cells nearly as effectively as the numerous rods, it is suggested that these cones may be responsible for day vision in the potto.     

The Rat Electroretinogram : II. Bloch's law and the latency mechanism of the b-wave

Electroretinograms were obtained from the all-rod eye of the rat with uniform illumination of the entire retina and stimulus flashes of less than 3 msec. duration. Bloch's law of temporal summation was verified for the b-wave latency by varying the time between two equal intensity flashes and observing that no change occurred in the latency when measured from the midpoint of the two flashes. The results of this and other experiments are described in terms of a simple but general model of the latency-determining mechanism. It is shown that this latency mechanism acts as if it depends on a linear additive process; and also that a hypothetical excitatory substance which triggers activity in the sources of the b-wave must accumulate rapidly in time after the flash, approximately as t⁸. The rate at which this substance accumulates is accurately represented by the diffusion equation for more than 4 to 6 log units in the flash intensity. This suggests that the rate-determining step in the latency mechanism may be diffusion-limited.     

蛙离体视网膜中杆系统对锥系统的抑制作用

在蛙离体视网膜标本上研究了由条件刺激(F_c)引起的快速暗适应(RDA)过程中b 波的变化。当F_c超过一定强度时,以501nm 闪光为测试刺激(F_(t(501)))时b波阈值单调下降,先为锥相后为杆相,两者过渡处呈现杆-锥转折;但以654nm闪光为测试刺激时(F_(t(654))),阈值先略有下降,继而上升,之后再下降。F_(t(654))应阈值在RDA 过程中的这种升高是反映575锤系统的活动受到502杆系统抑制的结果,433杆系统对此没有明显的贡献,主要依据是:(1)F_(t(654))反应阈值的上升在时间上与暗适应曲线的杆相的出现一致;(2)在RDA 过程的锥相,反应的光谱敏感性与锥色素吸收光谱一致,而在F_(t(654))反应阈值上升到最大值时,反应的光谱敏感性与502杆色素的吸收光谱非常接近;(3)对502杆系统作用等效的F_(c(439))、F_(c(501))和F_(c(616))作用后,F_(t(654))的敏感度变化在整个RDA 过程中基本相同。     

The statistical detection of threshold signals in the retina

1. Photographic records of impulses from single ganglion cells in the cat's retina were made while the retina was stimulated by flashes occurring once a second. Ten flashes at each of several intensities near threshold were used. 2. For the purpose of statistical analysis, the number of impulses (x) falling within a critical period following each flash was used as an index of the response. Histograms of x were plotted and used to calculate rates of transfer of information by the ganglion cell for the case of an ideal experiment, the yes-no choice, in which flashes of intensity I and blanks are to be distinguished. 3. The information rate increased (a) with increasing stimulus intensity and (b) with the number of identical flashes or blanks presented successively in a block. The intensity chosen as threshold by the experimenter, who observed the impulses visually and aurally, corresponded to an average information rate for single flashes of 0.7 bit/flash, compared to the maximum possible rate of 1 bit/flash. A threshold intensity giving 0.4 or more bit/flash, if presented in blocks of six identical flashes, corresponded to 0.95 or more bit/block, or near certainty. Thus the calculation of information rates using the index x provides an estimate of threshold at least as sensitive as those obtained during an experiment, which were made only after observing the responses to five to ten flashes of the same intensity. 4. The index x has statistical properties similar to those of the "index of neural activity" used by Tanner and Swets (1954) in their statistical model of human vision, and represents a possible physical interpretation of their index. However, x gave values (0.5 to 1.5) of the parameter called the slope which were consistently smaller than their values (2.1 to 3.1).     

Response entrainment of movement fibers in the optic tract of crayfish

The response properties of jittery movement fibers (JMF) in the crayfish optic tract reacting to a non-moving temporally patterned light were analyzed. The JMFs usually show no response during the regular flickering of stationary light with a flash duration of less than 50 msec when the stimulus frequency is between 4 and 20 per second; however they do respond when the flickering stops if a certain number of flashes have been given. The response appears about 50 msec after the first missing flash, i.e., the latency of the response after the last flash of the train changed from 100 to 300 msec. Thus, the “off” response at the end of the flicker is entrained to the stimulus repetition interval and locked onto the time of the first missing flash. The response of a sustaining fiber to an identical stimulus has quite different features as illustrated in Fig. 2. Some of the fibers show responses to the beginning part of the flicker but not necessarily to each flash, and habituate after several flashes. When a single flash longer than 250 msec is given, the fiber shows an “off” response with about 50 msec latency, as it does to sustained light. Some fibers show a double burst of “off” discharge to single flashes; the first at 50 msec is followed after 120 msec by the second one. However, when the flash duration is between 250 and 50 msec, a single flash elicits little or no response. The latency of the “off” response is as much as 300 msec for short single flashes less than 50 msec. An “on” response to flashes of light is observed when the inter-stimulus interval is more than 5 sec. The responses to the beginning part of flicker train are not simply locked to the just preceding flash except the “on” response to the very first one, but they can be the long latency responses to the flash before that. This response is modified in latency by the succeeding flashes in flicker trains and becomes entrained to the missing flash. Four types of entrainment are classified on the basis of the change in latency from the missing flash with regard to the number of flashes in a train. In most cases, 10 flashes are sufficient to entrain the response to the first missing flash. Non-resposiveness, i.e., habituation, during a regular flicker, may be due to an active inhibitory process, initiated by each succeeding light pulse. The response to the missing flash, therefore results from a disinhibited modified response to the last flash. Some JMFs continue to respond to the flicker even after a considerable number of flashes but only when the repetition interval is about 120 msec corresponding well to the interval of the double burst “off” discharge, thus the JMF has a resonant frequency of about 8 Hz. The JMFs appear to be acting as an irregularity detector in temporal sequence.     

Neural and Photochemical Mechanisms of Visual Adaptation in the Rat

The effects of light adaptation on the increment threshold, rhodopsin content, and dark adaptation have been studied in the rat eye over a wide range of intensities. The electroretinogram threshold was used as a measure of eye sensitivity. With adapting intensities greater than 1.5 log units above the absolute ERG threshold, the increment threshold rises linearly with increasing adapting intensity. With 5 minutes of light adaptation, the rhodopsin content of the eye is not measurably reduced until the adapting intensity is greater than 5 log units above the ERG threshold. Dark adaptation is rapid (i.e., completed in 5 to 10 minutes) until the eye is adapted to lights strong enough to bleach a measurable fraction of the rhodopsin. After brighter light adaptations, dark adaptation consists of two parts, an initial rapid phase followed by a slow component. The extent of slow adaptation depends on the fraction of rhodopsin bleached. If all the rhodopsin in the eye is bleached, the slow fall of threshold extends over 5 log units and takes 2 to 3 hours to complete. The fall of ERG threshold during the slow phase of adaptation occurs in parallel with the regeneration of rhodopsin. The slow component of dark adaptation is related to the bleaching and resynthesis of rhodopsin; the fast component of adaptation is considered to be neural adaptation.     

The induction kinetics of bacterial photophosphorylation. Threshold effects by the phosphate potential and correlation with the amplitude of the carotenoid absorption band shift

1. ATP synthesis (monitored by luciferin-luciferase) can be elicited by a single turnover flash of saturating intensity in chromatophores from Rhodopseudomonas capsulata, Kb1. The ATP yield from the first to the fourth turnover is strongly influenced by the phosphate potential: at high phosphate potential (?11.5 kcal/mol) no ATP is formed in the first three turnovers while at lower phosphate potential (?8.2 kcal/mol) the yield in the first flash is already one half of the maximum, which is reached after 2–3 turnovers.2. The response to ionophores indicates that the driving force for ATP synthesis in the first 20 turnovers is mainly given by a membrane potential. The amplitude of the carotenoid band shift shows that during a train of flashes an increasing ΔΨ is built up, which reaches a stationary level after a few turnovers; at high phosphate potential, therefore, more turnovers of the same photosynthetic unit are required to overcome an energetic threshold.3. After several (six to seven) flashes the ATP yield becomes constant, independently from the phosphate potential; the yield varies, however, as a function of dark time (t_d) between flashes, with an optimum for t_d = 160–320 ms.4. The decay kinetics of the high energy state generated by a long (125 ms) flash have been studied directly measuring the ATP yield produced in post-illumination by one single turnover flash, under conditions of phosphate potential (?10 kcal/mol), which will not allow ATP formation by one single turnover. The high energy state decays within 20 s after the illumination. The decay rate is strongly accelerated by 10^?8 M valinomycin.5. Under all the experimental conditions described, the amplitude of the carotenoid signal correlates univocally with the ATP yield per flash, demonstrating that this signal monitores accurately an energetic state of the membrane directly involved in ATP synthesis.6. Although values of the carotenoid signal much larger than the minimal threshold are present, relax slowly, and contribute to the energy input for phosphorylation, no ATP is formed unless electron flow is induced by a single turnover flash.7. The conclusions drawn are independent from the assumption that a ΔΨ between bulk phases is evaluable from the carotenoid signal.     

Multifocal Electroretinograms

A limitation of traditional full-field electroretinograms (ERG) for the diagnosis of retinopathy is lack of sensitivity. Generally, ERG results are normal unless more than approximately 20% of the retina is affected. In practical terms, a patient might be legally blind as a result of macular degeneration or other scotomas and still appear normal, according to traditional full field ERG. An important development in ERGs is the multifocal ERG (mfERG). Erich Sutter adapted the mathematical sequences called binary m-sequences enabling the isolation from a single electrical signal an electroretinogram representing less than each square millimeter of retina in response to a visual stimulus¹.Results that are generated by mfERG appear similar to those generated by flash ERG. In contrast to flash ERG, which best generates data appropriate for whole-eye disorders. The basic mfERG result is based on the calculated mathematical average of an approximation of the positive deflection component of traditional ERG response, known as the b-wave¹. Multifocal ERG programs measure electrical activity from more than a hundred retinal areas per eye, in a few minutes. The enhanced spatial resolution enables scotomas and retinal dysfunction to be mapped and quantified.In the protocol below, we describe the recording of mfERGs using a bipolar speculum contact lens.Components of mfERG systems vary between manufacturers. For the presentation of visible stimulus, some suitable CRT monitors are available but most systems have adopted the use of flat-panel liquid crystal displays (LCD). The visual stimuli depicted here, were produced by a LCD microdisplay subtending 35 - 40 degrees horizontally and 30 - 35 degrees vertically of visual field, and calibrated to produce multifocal flash intensities of 2.7 cd s m^-2. Amplification was 50K. Lower and upper bandpass limits were 10 and 300 Hz. The software packages used were VERIS versions 5 and 6.     

Visual Adaptation in the Retina of the Skate

The electroretinogram (ERG) and single-unit ganglion cell activity were recorded from the eyecup of the skate (Raja erinacea and R. oscellata), and the adaptation properties of both types of response compared with in situ rhodopsin measurements obtained by fundus reflectometry. Under all conditions tested, the b-wave of the ERG and the ganglion cell discharge showed identical adaptation properties. For example, after flash adaptation that bleached 80% of the rhodopsin, neither ganglion cell nor b-wave activity could be elicited for 10–15 min. Following this unresponsive period, thresholds fell rapidly; by 20 min after the flash, sensitivity was within 3 log units of the dark-adapted level. Further recovery of threshold was slow, requiring an additional 70–90 min to reach absolute threshold. Measurements of rhodopsin levels showed a close correlation with the slow recovery of threshold that occurred between 20 and 120 min of dark adaptation; there is a linear relation between rhodopsin concentration and log threshold. Other experiments dealt with the initial unresponsive period induced by light adaptation. The duration of this unresponsive period depended on the brightness of the adapting field; with bright backgrounds, suppression of retinal activity lasted 20–25 min, but sensitivity subsequently returned and thresholds fell to a steady-state value. At all background levels tested, increment thresholds were linearly related to background luminance.     

The scotopic electroretinogram of the sugar glider related to histological features of its retina

The flash electroretinogram (ERG) was used to characterize the scotopic retinal function in a marsupial. Key parameter values of the a- and b-waves of adult male sugar gliders, Petaurus breviceps breviceps, elicited with ganzfeld flashes were determined under dark- and light-adapted conditions. Using standard histological methods, the thicknesses of the major layers of the retina were assessed to provide insight into the nature of the ERG responses. The ERG and histological results were compared to corresponding data for placental C57Bl/6 mice to establish whether the functional retinal specialization that underlies scotopic visual function in a marsupial parallels that of a placental mouse. The sensitivity of the a-wave assessed with the Lamb and Pugh (Invest Ophthalmol Vis Sci 47:5138–5152, 2006) “model” and that of the b-wave assessed with standard methods were lower in the sugar glider compared to the mouse. The thickness of the sugar glider retina was two-third of that of the mouse. The high-intensity flash ERG of the sugar glider substantially differed in shape from that of the mouse reflecting perhaps structural and functional differences between the two species at the level of the inner retina.     

THE FLICKER RESPONSE CONTOUR FOR PHRYNOSOMA (HORNED LIZARD; CONE RETINA)

The lizard Phrynosoma, with purely cone retina, provides a simplex flicker response contour (log critical flash intensity as a function of flash frequency). It is well described as a normal probability integral (F - log I). The Phrynosoma curve differs markedly, in higher slope and in higher median intensity level, from that obtained under the same conditions for the turtle Pseudemys, also with entirely cone retina. Other comparisons having a bearing on the duplexity doctrine are discussed.     

Flash photolysis of rhodopsin in the cat retina

The bleaching of rhodopsin by short-duration flashes of a xenon discharge lamp was studied in vivo in the cat retina with the aid of a rapid, spectral-scan fundus reflectometer. Difference spectra recorded over a broad range of intensities showed that the bleaching efficacy of high-intensity flashes was less than that of longer duration, steady lights delivering the same amount of energy. Both the empirical results and those derived from a theoretical analysis of flash photolysis indicate that, under the conditions of these experiments, the upper limit of the flash bleaching of rhodopsin in cat is approximately 90%. Although the fact that a full bleach could not be attained is attributable to photoreversal, i.e., the photic regeneration of rhodopsin from its light-sensitive intermediates, the 90% limit is considerably higher than the 50% (or lower) value obtained under other experimental circumstances. Thus, it appears that the duration (approximately 1 ms) and spectral composition of the flash, coupled with the kinetic parameters of the thermal and photic reactions in the cat retina, reduce the light-induced regeneration of rhodopsin to approximately 10%.     

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.     

Slow PIII component of the carp electroretinogram

The slow PIII component of the electroretinogram (ERG) was studied in the isolated, aspartate-treated carp retina. Although the latter is richly populated with cones, slow PIII appeared to reflect almost exclusively the activity of rods; e.g. the spectral sensitivity of the potential paralleled closely the rod pigment curve, its operating range (i.e. the V-log I curve) was limited to 3 log units above absolute threshold, and raising background intensities to photopic levels produced saturation of the increment threshold function without evidence of a cone-mediated segment. Only after bleaching away a significant fraction of the porphyropsin was it possible to unmask a small photopic contribution to slow PIII, as evidenced by a displacement in the action spectrum to longer wavelengths. The spatial distribution of the slow PIII voltage within the retina (Faber, D.S. 1969. Ph.D. Thesis. State University of New York. Buffalo, N.Y.; Witkovsky, P.J. Nelson, and H. Ripps. 1973. J. Gen Physiol. 61:401) and its ability to survive aspartate treatment indicate that this potential arises in the Muller (glial) fiber. Additional support for this conclusion is provided by the slow rise time (several seconds) and long temporal integration (up to 40s) of the response. In many respects the properties of slow PIII resemble those of the c-wave, a pigment epithelial response also subserved by rod activity. On the other hand, the receptoral (fast PIII) and the b-wave components of the ERG behave quite differently. Unlike slow PIII, response saturation could not be induced, since both potentials are subserved by cones when the stimulus conditions exceed the limits of the scotopic range. Receptors appear to govern light adaptation at photopic background levels; both fast PIII and b-wave manifest identical incremental threshold values over this range of intensities. However, under scotopic conditions, the sensitivity of the b-wave is affected by luminous backgrounds too weak to alter fast PIII threshold, indicating a postreceptoral stage of adaptation.     

Energy and predation costs of firefly courtship signals

Animal courtship signals include many highly conspicuous traits and behaviors, and it is generally assumed that such signals must balance the benefits of attracting mates against some fitness costs. However, few studies have assessed the multiple costs potentially incurred by any one courtship signal, so we have limited understanding of the relative importance of different costs. This study provides the first comprehensive assessment of signal costs for Photinus fireflies (Coleoptera: Lampyridae), using controlled experiments to measure both the energy and predation costs associated with their bioluminescent courtship signals. We measured energy required to generate bioluminescent flashes, using differential open-flow respirometry, and found that flash signaling results in only a nominal increase in energy expenditure above resting levels. These results suggest that the energy required to generate bioluminescent flashes represents a minor component of the total cost of firefly courtship. However, controlled field experiments revealed that visually oriented predators imposed major costs on firefly courtship signals, with higher signaling rates significantly increasing the likelihood of predation. Together with previous results demonstrating that female fireflies prefer more conspicuous courtship signals, these results support the importance of multiple-receiver communication networks in driving signal evolution.     

Control of Retinal Sensitivity : III. Lateral Interactions at the Inner Plexiform Layer

Both the "on" and the "on-off" ganglion cells in the mudpuppy retina generate graded responses over a narrow range of log test intensities. Sustained full field or surround backgrounds change the range of center log test intensities that elicits the graded response for both cell types. The on-off, but not the on ganglion cells are further affected by moving or flashing surround backgrounds. These cells are hyperpolarized, threshold is elevated, and the entire graded range of response is elicited by a higher range of log center test intensities. Depolarizing activity is elicited in amacrine cells by moving backgrounds that affect the on-off ganglion cells, but bipolar activity is unaffected. These results suggest that the amacrine cells at the inner plexiform layer mediate a third stage of sensitivity control in the retina, increasing threshold for response to change specifically in the on-off ganglion cells.