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Many retinal degenerative diseases result in vision impairment or permanent blindness due to photoreceptor loss or dysfunction. It has been observed that Pde6b rd1 mice (rd1), which carry a spontaneous nonsense mutation in the pde6b gene, have a strong phenotypic similarity to patients suffering from autosomal recessive retinitis pigmentosa. In this study, we present a novel mouse model of retinitis pigmentosa generated through pde6b gene knockout using CRISPR/Cas9 technology. We compare this Pde6b-KO mouse model to the rd1 mouse model to gain insights into the progression of retinal degeneration. The functional assessment of the mouse retina and the tracking of degeneration dynamics were performed using electrophysiological methods, while retinal morphology was analyzed through histology techniques. Interestingly, the Pde6b-KO mouse model demonstrated a higher amplitude of photoresponse than the rd1 model of the same age. At postnatal day 12, the thickness of the photoreceptor layer in both mouse models did not significantly differ from that of control animals; however, by day 15, a substantial reduction was observed. Notably, the decline in the number of photoreceptors in the rd1 model occurred at a significantly faster rate. These findings suggest that the C3H background may play a significant role in the early stages of retinal degeneration.

The avian magnetic compass allows orientation during migration and is shown to function properly under short-wavelength but not long-wavelength visible light. Therefore, the magnetoreceptive system is assumed to be light- and wavelength-dependent and localized in the retina of the eye. Putative candidates for the role of primary magnetosensory molecules are the cryptochromes that are known to be expressed in the avian retina and must be able to interact with phototransduction proteins. Previously, we reported that in migratory birds change in magnetic field direction induces significant effects on electroretinogram amplitude in response to blue flashes, and such an effect was observed only in the nasal quadrant of the retina. Here, we report new electroretinographic, microscopic and microspectrophotometric data on European robins, confirming the magnetosensitivity of the retinal nasal quadrant after applying the background illumination. We hypothesized that magnetoreceptive distinction of this region may be related to its morphology and analyzed the retinal distribution and optical properties of oil droplets, the filtering structures within cones. We found that the nasal quadrant contains double cones with the most intensely colorized oil droplets compared to the rest of the retina, which may be related to its magnetosensory function.

Green rods (GRs) represent a unique type of photoreceptor to be found in the retinas of anuran amphibians. These cells harbor a cone-specific blue-sensitive visual pigment but exhibit morphology of the outer segment typical for classic red rods (RRs), which makes them a perspective model object for studying cone-rod transmutation. In the present study, we performed detailed electrophysiological examination of the light sensitivity, response kinetics and parameters of discrete and continuous dark noise in GRs of the two anuran species: cane toad and marsh frog. Our results confirm that anuran GRs are highly specialized nocturnal vision receptors. Moreover, their rate of phototransduction quenching appeared to be about two-times slower than in RRs, which makes them even more efficient single photon detectors. The operating intensity ranges for two rod types widely overlap supposedly allowing amphibians to discriminate colors in the scotopic region. Unexpectedly for typical cone pigments but in line with some previous reports, the spontaneous isomerization rate of the GR visual pigment was found to be the same as for rhodopsin of RRs. Thus, our results expand the knowledge on anuran GRs and show that these are even more specialized single photon catchers than RRs, which allows us to assign them a status of "super-rods".

Migratory birds are known to be sensitive to external magnetic field (MF). Much indirect evidence suggests that the avian magnetic compass is localized in the retina. Previously, we showed that changes in the MF direction could modulate retinal responses in pigeons. In the present study, we performed similar experiments using the traditional model animal to study the magnetic compass, European robins. The photoresponses of isolated retina were recorded using ex vivo electroretinography (ERG). Blue- and red-light stimuli were applied under an MF with the natural intensity and two MF directions, when the angle between the plane of the retina and the field lines was 0° and 90°, respectively. The results were separately analysed for four quadrants of the retina. A comparison of the amplitudes of the a- and b-waves of the ERG responses to blue stimuli under the two MF directions revealed a small but significant difference in a- but not b-waves, and in only one (nasal) quadrant of the retina. The amplitudes of both the a- and b-waves of the ERG responses to red stimuli did not show significant effects of the MF direction. Thus, changes in the external MF modulate the European robin retinal responses to blue flashes, but not to red flashes. This result is in a good agreement with behavioural data showing the successful orientation of birds in an MF under blue, but not under red illumination.

Migratory birds can detect the direction of the Earth's magnetic field using the magnetic compass sense. However, the sensory basis of the magnetic compass still remains a puzzle. A large body of indirect evidence suggests that magnetic compass in birds is localized in the retina. To confirm this point, an evidence of visual signals modulation by magnetic field (MF) should be obtained. In a previous study we showed that MF inclination impacts the amplitude of ex vivo electroretinogram (ERG) recorded from isolated pigeon retina. Here we present the results of an analysis of putative MF effect on one component of ERG, the photoreceptor's response, isolated from the total ERG by adding sodium aspartate and barium chloride to the perfusion solution. Photoresponses were recorded from isolated retinae of domestic pigeons Columba livia. The retinal samples were placed in MF that was modulated by three pairs of orthogonal Helmholtz coils. Light stimuli (blue and red) were applied under two inclinations of MF, 0° and 90°. In all the experiments, preparations from two parts of retina were used, red field (with dominant red-sensitive cones) and yellow field (with relatively uniform distribution of cone color types). In contrast to the whole retinal ERG, we did not observe any effect of MF inclination on either amplitude or kinetics of pharmacologically isolated photoreceptor responses to blue or red half-saturating flashes. A possible explanations of these results could be that magnetic compass sense is localized in retinal cells other than photoreceptors, or that photoreceptors do participate in magnetoreception, but require some processing of compass information in other retinal layers, so that only whole retina signal can reflect the response to changing MF., Competing Interests: The authors have declared that no competing interests exist.

Purpose: Accumulating evidence suggests that dopamine, the major catecholamine in the vertebrate retina, may modulate cAMP-mediated signaling in photoreceptors to optimize vision in the light/dark cycle. The main putative mechanism of dopamine-induced adaptation changes in photoreceptors is activation of D 2 -like receptors (D 2 R), which leads to a decrease of the intracellular cAMP level and reduction of protein kinase A (PKA) activity. However, the mechanisms by which dopamine exerts its regulating effect on the phototransduction cascade remain largely unknown. The aim of the present study was to investigate the effects of dopamine and dopamine receptor agonists on rod photoresponses., Methods: The experiments were performed on solitary rods of the Rana ridibunda frog. Photoreceptor currents were recorded using a suction pipette technique. The effects of dopamine (0.1-50 µM) and selective dopamine receptor agonists-D 1 R agonist SKF-38393 (0.1-50 µM), D 2 R agonist quinpirole (2.5-50 µM), and D 1 -D 2 receptor heterodimer agonist SKF-83959 (50 µM)-were examined., Results: We found that, when applied to the rod inner segments (RISs), dopamine and dopamine receptor agonists had no effect on photoresponses. In contrast, the rods responded to dopamine and all agonists applied to their outer segments by decreasing sensitivity to light. At the highest tested concentration (50 µM), the most prominent effect on light sensitivity was induced by D 1 R agonist SKF-38393, while dopamine, D 2 R agonist quinpirole, and D 1 -D 2 receptor heterodimer agonist SKF-83959 produced somewhat lower and approximately equal effects. Moreover, SKF-38393 reduced sensitivity at all tested concentrations starting from the smallest one (0.1 µM), whereas dopamine and quinpirole started their action from the higher concentrations of 2.5 µM and 50 µM, respectively. In addition, dopamine, SKF-38393, and quinpirole, on average, did not change the intracellular calcium level as judged from the "exchange current", while SKF-83959 increased it by ~1.3 times., Conclusions: Dopamine induces a decrease in rod sensitivity, mostly by reducing the activation rate of the cascade, and to a much lesser extent, speeding up the turning off of the cascade. The sign of the reaction to all tested drugs, lack of selectivity of dopamine and dopamine receptor agonist action, and analysis of factors that determine sensitivity of photoreceptors suggest that, in rod outer segments (ROSs), dopamine action is mediated by D 1 -D 2 receptor heterodimers but not D 1 R or D 2 R alone. This work supports the assumption made earlier by other authors that dopamine exercises its regulatory effect via at least two independent mechanisms, which are cAMP and Ca 2+ mediated.

Rod photoreceptors of the vertebrate retina produce, in darkness, spontaneous discrete current waves virtually identical to responses to single photons. The waves comprise an irreducible source of noise (discrete dark noise) that may limit the threshold sensitivity of vision. The waves obviously originate from acts of random activation of single rhodopsin molecules. Until recently, it was generally accepted that the activation occurs due to the rhodopsin thermal motion. Yet, a few years ago it was proposed that rhodopsin molecules are activated not by heat but rather by real photons generated within the retina by chemiluminescence. Using a high-sensitive photomultiplier, we measured intensities of biophoton emission from isolated retinas and eyecups of frogs ( Rana ridibunda ) and fish (sterlet, Acipenser ruthenus ). Retinal samples were placed in a perfusion chamber and emitted photons collected by a high-aperture quartz lens. The collected light was sent to the photomultiplier cathode through a rotating chopper so that a long-lasting synchronous accumulation of the light signal was possible. The absolute intensity of bio-emission was estimated by the response of the measuring system to a calibrated light source. The intensity of the source, in turn, was quantified by measuring rhodopsin bleaching with single-rod microspectrophotometry. We also measured the frequency of discrete dark waves in rods of the two species with suction pipette recordings. Expressed as the rate constant of rhodopsin activation, it was 1.2 × 10 -11 /s in frogs and 7.6 × 10 -11 /s in sterlets. Approximately two thirds of retinal samples of each species produced reliably measurable biophoton emissions. However, its intensity was ≥100 times lower than necessary to produce the discrete dark noise. We argue that this is just a lower estimate of the discrepancy between the hypothesis and experiment. We conclude that the biophoton hypothesis on the origin of discrete dark noise in photoreceptors must be rejected., (© 2019 Govardovskii et al.)

Application of molecular photochromic ion channel blockers to recover the visual function of a degenerated retina is one of the promising trends in photopharmacology. To this day, several photochromic azobenzene-based compounds have been proposed and their functionality has been demonstrated on cell lines and knockout mouse models. Further advance necessitates testing of the physiological activity of a great number of new compounds. The goal of this study is to propose animal models of photoreceptor degeneration that are easier to obtain than knockout mouse models but include the main features required for testing the physiological activity of molecular photoswitches. Two amphibian-based models were proposed. The first model was obtained by mechanical deletion of the photoreceptor outer segments. The second model was obtained by intraocular injection of tunicamycin to induce the degeneration of rods and cones. To test our models, we used 2-[(4-{(E)-[4-(acryloylaminophenyl]diazenyl}phenyl)amino]-N,N,N-triethyl-2-oxoethanammonium chloride (AAQ), one of the compounds that have been studied in other physiological models. The electroretinograms recorded from our models before and after AAQ treatment are in agreement with the results obtained on knockout mouse models and reported in other studies. Hence, the proposed models can be used for primary screening of molecular photochromic ion channel blockers.

Purpose: To identify steps of the phototransduction cascade responsible for the delay of the photoresponse., Methods: Electrical responses of fish ( Carassius ) cones and Rana ridibunda frog rods and cones were recorded with a suction pipette technique and as an aspartate-isolated mass receptor potential from isolated perfused retinas. Special attention was paid to sufficiently high temporal resolution (1-ms flash, 700 Hz amplification bandpass). Stochastic simulation of the activation steps from photon absorption to the formation of catalytically active phosphodiesterase (PDE) was performed. In addition, a deterministic mathematical model was fit to the experimental responses. The model included a detailed description of the activation steps of the cascade that enabled identification of the role of individual transduction stages in shaping the initial part of the response., Results: We found that the apparent delay of the photoresponse gets shorter with increasing stimulus intensity and reaches an asymptotic value of approximately 3 ms in cones and greater than or equal to 10 ms in rods. The result seems paradoxical since it is suggested that the delay occurs in the chain of steps from photon absorption to the formation of active transducin (T*) which in cones is, on average, slower than in rods. Stochastic simulation shows that actually the steps from photon absorption to T* may not contribute perceptibly to the delay. Instead, the delay occurs at the stage that couples the cycle of repetitive activation of T by rhodopsin (R*) with the activation of PDE. These steps include formation of T* (= T α GTP) out of T αβγ GTP released from the activation cycle and the subsequent interaction of T* with PDE. This poses a problem. The duration of an average cycle of activation of T in rods is approximately 5 ms and is determined by the frequency of collisions between R* and T in the photoreceptor membrane. The frequency is roughly proportional to the surface packing density of T in the membrane. As the packing density of PDE is approximately 12 times lower than that of T, it could be expected that the rate of the T*-PDE interaction were an order of magnitude slower than that of R* and T. As modeling shows, this is the case in rods. However, the delay in cones is approximately 3 ms which could be achieved only at a T*-PDE interaction time of less than or equal to 5 ms. This means that either the frequency of the collisions of T* and PDE, or the efficiency of collisions, or both in cones are approximately ten times higher than in rods. This may be a challenge to the present model of the molecular organization of the photoreceptor membrane., Conclusions: The delay of the photoresponse is mainly set by the rate of interaction of T* with PDE. In cones, the delay is shorter than in rods and, moreover, shorter than the duration of the cycle of repetitive activation of T by R*. This poses a problem for the present model of diffusion interaction of phototransduction proteins in the photoreceptor membrane.

The absolute sensitivity of vertebrate retinas is set by a background noise, called dark noise, which originates from several different cell types and is generated by different molecular mechanisms. The major share of dark noise is produced by photoreceptors and consists of two components, discrete and continuous. Discrete noise is generated by spontaneous thermal activations of visual pigment. These events are undistinguishable from real single-photon responses (SPRs) and might be considered an equivalent of the signal. Continuous noise is produced by spontaneous fluctuations of the catalytic activity of the cGMP phosphodiesterase. This masks both SPR and spontaneous SPR-like responses. Circadian rhythms affect photoreceptors, among other systems by periodically increasing intracellular cAMP levels ([cAMP] in ), which increases the size and changes the shape of SPRs. Here, we show that forskolin, a tool that increases [cAMP] in , affects the magnitude and frequency spectrum of the continuous and discrete components of dark noise in photoreceptors. By changing both components of rod signaling, the signal and the noise, cAMP is able to increase the photoreceptor signal-to-noise ratio by twofold. We propose that this results in a substantial improvement of signal detection, without compromising noise rejection, at the rod bipolar cell synapse., (© 2017 Astakhova et al.)

Numerous and profound cyclical changes in retina functioning during a 24-hour daily cycle are largely determined by the influence of two neuromodulators--melatonin and dopamine. Do- pamine and melatonin form a reciprocal pair by mutually inhibiting the synthesis of each other, and they release into extracellular space of the retina approximately in antiphase. Dopamine is cyclically synthesized by special populations of dopaminergic amacrine cells in the retina and its content increases during the day and decreases at night. Like melatonin, dopamine affects all the major cell types of the outer and inner retinal layers. Activation of dopamine D1- and D2-type receptors regulate the activity of protein kinase A and the intracellular concentration of cAMP, and may trigger other regulatory pathways, including activation of phospholipase C. In photorecep- tors, dopamine acting on D2-type receptors reduces cAMP concentration, suppresses melatonin synthesis and regulates the conduction of gap junctions between rods and cones, depending on the phase ofthe light cycle. By decreasing the concentration of cAMP, dopamine could also be a regu- lator of the phototransduction cascade and other cellular functions of the photoreceptor. Here we review some of these possibilities. Key words: dopamine, dopamine receptors, cAMP, retina, photoreceptor, circadian rhythms, protein kinase A.

Until recently, it has generally been believed that cyclic AMP plays an important role in supporting circadian cycles in the vertebrate retina, but does not directly control the photoreceptors' phototransduction cascade. However, the cAMP levels in photoreceptors oscillate during the day/night cycle, and the cAMP turnover in photoreceptors may be light-dependent. Thus it is natural to suggest that the cAMP-dependent protein phosphorylation may be a mechanism of tuning phototransduction to lighting conditions. In the present review, we summarize available information on the structure and operation of the retinal pacemaker, role(s) of cAMP in its functioning, and identified intracellular targets that could be controlled by cAMP. We discuss our recent results that show that cAMP changes do regulate the phototransduction cascade. This regulation may substantially extend the range of photoreceptor's adaptation by increasing its sensitivity at night, and reducing the sensitivity in bright light.

In early studies, both cyclic AMP (cAMP) and cGMP were considered as potential secondary messengers regulating the conductivity of the vertebrate photoreceptor plasma membrane. Later discovery of the cGMP specificity of cyclic nucleotide-gated channels has shifted attention to cGMP as the only secondary messenger in the phototransduction cascade, and cAMP is not considered in modern schemes of phototransduction. Here, we report evidence that cAMP may also be involved in regulation of the phototransduction cascade. Using a suction pipette technique, we recorded light responses of isolated solitary rods from the frog retina in normal solution and in the medium containing 2 µM of adenylate cyclase activator forskolin. Under forskolin action, flash sensitivity rose more than twofold because of a retarded photoresponse turn-off. The same concentration of forskolin lead to a 2.5-fold increase in the rod outer segment cAMP, which is close to earlier reported natural day/night cAMP variations. Detailed analysis of cAMP action on the phototransduction cascade suggests that several targets are affected by cAMP increase: (a) basal dark phosphodiesterase (PDE) activity decreases; (b) at the same intensity of light background, steady background-induced PDE activity increases; (c) at light backgrounds, guanylate cyclase activity at a given fraction of open channels is reduced; and (d) the magnitude of the Ca(2+) exchanger current rises 1.6-fold, which would correspond to a 1.6-fold elevation of [Ca(2+)](in). Analysis by a complete model of rod phototransduction suggests that an increase of [Ca(2+)](in) might also explain effects (b) and (c). The mechanism(s) by which cAMP could regulate [Ca(2+)](in) and PDE basal activity is unclear. We suggest that these regulations may have adaptive significance and improve the performance of the visual system when it switches between day and night light conditions.

Among the GPCR-signaling cascades, phototransduction in vertebrate retinal photoreceptors has been characterized in unprecedented details. It is believed that basic mechanisms of phototransduction and adaptation are reliably and completely established, and phototransduction may serve as a benchmark for understanding other G-protein-coupled systems. In this review, we compare present scheme of phototransduction with other GPCR-cascades in order to reveal their similarities and specific features. We show, based mainly on our physiological and biophysical data, that the existing scheme misses a few important regulations whose molecular basis is unknown. There exists a fast and efficient mechanism that accelerates the turn-off of the activated G-protein (transducin) during light adaptation. A few slowly acting processes result in a long-lasting modification of the cascade's components and regulate the speed of rhodopsin and transducin quenching. Similarly to other GPCR-cascades, one may suggest that there are multiple signalling pathways that start from photoactivated rhodopsin and rely on different secondary messengers (e.g. cAMP vs. cGMP). We also show that rhodopsin in retinal rods may form areas of paracristalline organization, and that the oligomerization might be a mechanism for controlling the amplification of the signalling cascade. The missing mechanisms are by no means minor, and could ensure sensitivity regulation within two orders of magnitude range.

The time course of the light-induced activity of phototrandsuction effector enzyme cGMP-phosphodiesterase (PDE) is shaped by kinetics of rhodopsin and transducin shut-offs. The two processes are among the key factors that set the speed and sensitivity of the photoresponse and whose regulation contributes to light adaptation. The aim of this study was to determine time courses of flash-induced PDE activity in frog rods that were dark adapted or subjected to nonsaturating steady background illumination. PDE activity was computed from the responses recorded from solitary rods with the suction pipette technique in Ca(2+)-clamping solution. A flash applied in the dark-adapted state elicits a wave of PDE activity whose rising and decaying phases have characteristic times near 0.5 and 2 seconds, respectively. Nonsaturating steady background shortens both phases roughly to the same extent. The acceleration may exceed fivefold at the backgrounds that suppress approximately 70% of the dark current. The time constant of the process that controls the recovery from super-saturating flashes (so-called dominant time constant) is adaptation independent and, hence, cannot be attributed to either of the processes that shape the main part of the PDE wave. We hypothesize that the dominant time constant in frog rods characterizes arrestin binding to rhodopsin partially inactivated by phosphorylation. A mathematical model of the cascade that considers two-stage rhodopsin quenching and transducin inactivation can mimic experimental PDE activity quite well. The effect of light adaptation on the PDE kinetics can be reproduced in the model by concomitant acceleration on both rhodopsin phosphorylation and transducin turn-off, but not by accelerated arrestin binding. This suggests that not only rhodopsin but also transducin shut-off is under adaptation control.

The recovery of rod responsiveness after saturating flashes is greatly retarded above a certain critical level of rhodopsin bleaching (approximately 0.1%). A mathematical description of the process of turn-off of the phototransduction cascade allows attributing different phases of the recovery to specific products of rhodopsin photolysis. The fast phase is determined by quenching of metarhodopsin II and activated transducin. The slow phase is controlled by decay of partially inactivated (phosphorylated and arrestin-bound) metarhodopsins, and by regeneration of rhodopsin. The transition between the two regimes of adaptation is rather abrupt, occurring within a few-fold range of stimulus intensity. This marks the border between reversal of light adaptation and dark adaptation, as it is commonly defined.

Thermal activation of the visual pigment constitutes a fundamental constraint on visual sensitivity. Its electrical correlate in the membrane current of dark-adapted rods are randomly occurring discrete "dark events" indistinguishable from responses to single photons. It has been proposed that thermal activation occurs in a small subpopulation of rhodopsin molecules where the Schiff base linking the chromophore to the protein part is unprotonated. On this hypothesis, rates of thermal activation should increase strongly with rising pH. The hypothesis has been tested by measuring the effect of pH changes on the frequency of discrete dark events in red rods of the common toad Bufo bufo. Dark noise was recorded from isolated rods using the suction pipette technique. Changes in cytoplasmic pH upon manipulations of extracellular pH were quantified by measuring, using fast single-cell microspectrophotometry, the pH-dependent metarhodopsin I-metarhodopsin II equilibrium and subsequent metarhodopsin III formation. These measurements show that, in the conditions of the electrophysiological experiments, changing perfusion pH from 6.5 to 9.3 resulted in a cytoplasmic pH shift from 7.6 to 8.5 that was readily sensed by the rhodopsin. This shift, which implies an 8-fold decrease in cytoplasmic [H(+)], did not increase the rate of dark events. The results contradict the hypothesis that thermal pigment activation depends on prior deprotonation of the Schiff base.

Photoreceptors in the isolated turtle retina of two species of turtle, Chelydra serpentina and Pseudemus scripta elegans, were penetrated with double-barrel electrodes. Physiological responses were recorded through one barrel and Neurobiotin tracer was injected from the other. Intracellular injection of Neurobiotin revealed patterns of tracer-coupled photoreceptors. Both the patterns of tracer coupling and the electrophysiology suggest a high degree of specificity of connections. Rods seem to be coupled only to rods and green and red cones seem to be coupled to cones of the same spectral type. Receptive-field profiles, measured with a thin, sharply focused slit of light, often had well-defined peaks and troughs in sensitivity. We have taken advantage of this observation and used the position of a peak in sensitivity to locate the position on the retina of a coupled cell. In one rod, it was possible to correlate physiological and morphological data and to show that the peaks in the physiological receptive field occurred at positions on the retina where there were dye-coupled cells. This provides direct evidence that gap junctions produce the physiological coupling between rods.

Rods in the bull-frog retina contain varying proportions of rhodopsin (lambda max = 502 nm) and porphyropsin (lambda max = 527 nm) in a dorso-ventral gradient from the porphyropsin-rich dorsal rim to the virtually pure rhodopsin fields of the central and ventral retina. We investigated if quantal excitations in the same rod are different depending on whether they are initiated by isomerization of a rhodopsin or a porphyropsin molecule. Current photoresponses were recorded from dark-adapted rods by sucking the outer segment into a recording pipette. The relation between pigment composition and spectral sensitivity was established by comparison with microspectrophotometrically measured absorbance spectra of rods from the same neighbourhood. Rods with suitable porphyropsin: rhodopsin mixtures (ideally between 1:4 and 1:2) were stimulated with flashes of red (608 nm) and blue (465 nm) light, whereby the red light will isomerize porphyropsin much more often than rhodopsin, and the reverse will be true of the blue light. The amplitude and shape of the single-photon response were found to be identical for the "red" and "blue" flash series to within measurement error (ca 10%). This indicates that the quantal responses initiated by the two pigments are identical.

1. The dark current and responses to dim flashes were recorded with the suction pipette technique from single rods in pieces of bull-frog retina taken from either the dorsal porphyropsin or the ventral rhodopsin field. 2. The composition of visual pigment in the rods was determined by microspectrophotometry. Rods from the dorsal pieces contained 70-88% porphyropsin523 mixed with rhodopsin502. The ventral rods contained almost pure rhodopsin, any possible admixture of porphyropsin being below the level of detectability (less than 5%). 3. In most cells, the responses to dim flashes were well fitted by a four-stage linear filter model, with no systematic differences in the response kinetics of porphyropsin and rhodopsin rods. The amplitude of saturated responses varied between 8 and 55 pA and that of responses to single isomerizations between 0.4 and 3.5 pA. 4. In porphyropsin rods, discrete events similar to the response to one photoisomerization were clearly seen in complete darkness. The dark current amplitude histogram was fitted by a convolution of the probability densities for the Gaussian continuous noise component and the averaged dim-flash response waveform. This allows estimation of the frequency and amplitude of discrete events and the standard deviation of the continuous component. The mean frequency of discrete dark events thus obtained from six porphyropsin cells was 0.057 rod-1 s-1 at 18 degrees C. 5. In rhodopsin rods, the dark current amplitude histogram appeared completely symmetrical, indicating that the frequency of discrete events must be lower than 0.005 rod-1 s-1 (except in one rod where it was 0.006 events rod-1 s-1). Per molecule of rhodopsin, the events are then at least 5 times rarer than reported for toad rhodopsin rods at the same temperature. 6. The low rate of isomerization-like 'dark' events in bull-frog rhodopsin rods shows, firstly, that results cannot be generalized across species even for rhodopsins which appear spectrally identical. Secondly, it suggests that these events need not (in an evolutionary sense) constitute an irreducible noise factor which must set the ultimate limit to the sensitivity of dark-adapted vision. 7. The difference between porphyropsin and rhodopsin rods shows that, given (presumably) the same opsin, the pigment utilizing retinal2 and absorbing maximally at longer wavelengths produces more noise. The signal/noise ratio attained in the photoreceptor may be an important factor in the natural selection of visual pigments.