Chirped flicker optoretinography for in vivo characterization of human photoreceptors' frequency response to light.

Flicker electroretinography (ERG) has served as a valuable noninvasive objective tool for investigating retinal physiological function through the measurement of electrical signals originating from retinal neurons in response to temporally modulated light stimulation. Deficits in the response at certain frequencies can be used as effective biomarkers of cone-pathway dysfunction. In this Letter, we present the progress we made on its optical counterpart-photopic flicker optoretinography (f-ORG). Specifically, we focus on the measurement of the response of light-adapted retinal photoreceptors to a flicker stimulus with chirped frequency modulation. In contrast to measurements performed at discrete frequencies, this technique enables a significantly accelerated characterization of photoreceptor outer segment optical path length modulation amplitudes in the nanometer range as a function of stimulus frequency, enabling the acquisition of the characteristic frequency response in less than 2 sec.

Rod and Cone Dark Adaptation in Congenital Aniridia and Its Association With Retinal Structure.

Purpose: To characterize the association between dark-adapted rod and cone sensitivity and retinal structure in PAX6-related aniridia. Methods: Dark-adaptation curves were measured after a 5-minute exposure to bright light with red (625 nm) and green (527 nm) 2° circular light stimuli presented at ≈20° temporal retinal eccentricity in 27 participants with aniridia (nine males; 11-66 years old) and 38 age-matched healthy controls. A two-stage exponential model was fitted to each participant's responses to determine their cone and rod thresholds over time. The thicknesses of macular inner and outer retinal layers were obtained from optical coherence tomography images in 20 patients with aniridia and the 38 healthy controls. Aniridia-associated keratopathy (AAK) grade (0-3) and lens opacities were quantified by clinical examination of the anterior segment. Results: The rod-cone break time was similar between patients with aniridia and healthy controls. Dark-adapted cone and the rod thresholds were higher in aniridia compared with healthy controls. In aniridia, foveal outer retinal layer thickness correlated with both final cone and rod thresholds. A multiple regression model indicated that foveal outer retinal layer thickness and age were the main explanatory variables to predict both final cone and rod thresholds in aniridia when the AAK grade was 2 or less. Conclusions: The results show that both rod- and cone-related functions are affected in PAX6-related aniridia and suggest that retinal anatomical and physiological changes extend beyond the area commonly studied in this condition: the central macula.

Assessment of Scotopic Function in Rod-Cone Inherited Retinal Degeneration With the Scotopic Macular Integrity Assessment.

Purpose: The scotopic macular integrity assessment (S-MAIA) can perform scotopic assessment to detect localized changes to scotopic rod and cone function. This study is an exploratory investigation of the feasibility of using the S-MAIA in a rod-cone dystrophy population to identify the pattern of loss in scotopic photoreceptor function. Methods: Twenty patients diagnosed with a rod-cone dystrophy underwent visual acuity testing, full-field stimulus threshold assessment, and multiple S-MAIA tests after dark adaptation periods of 20 minutes and 45 minutes performed separately. Only right eyes were tested. Three tests were performed following a learning test. A Bland-Altman analysis was used to assess repeatability and agreement between tests after the two time periods. Spatial interpolation maps were created from the group plots to display the pattern of rod and cone loss. Results: Learning effects took place between testing sessions 1 and 2 but not 2 and 3. Limits of agreement were larger in the patient eyes than control eyes, but within previously reported values. Using longer adaptation time of 45 minutes did not offer a significant advantage over 20 minutes. Patterns for the cyan and red sensitivities were different, indicating different patterns of loss for rods and cones. Conclusions: A dark adaptation time of 20 minutes before testing is sufficient for thresholding. The S-MAIA is suitable for use in patients with a logarithm of the minimum angle of resolution vision of at least 0.7 and provides a viable outcome measure for patients with rod-cone dystrophies and preserved central vision. The spatial information about scotopic function from the S-MAIA provides information about disease processes and progression. Translational Relevance: There is a need for scotopic measures for use in clinical trials. Scotopic microperimetry works well in patients with early disease, allowing the extension of recruitment criteria for novel therapies of rod-cone dystrophies.

Glial cells expressing visual cycle genes are vital for photoreceptor survival in the zebrafish pineal gland.

Photoreceptors in the vertebrate eye are dependent on the retinal pigmented epithelium for a variety of functions including retinal re-isomerization and waste disposal. The light-sensitive pineal gland of fish, birds, and amphibians is evolutionarily related to the eye but lacks a pigmented epithelium. Thus, it is unclear how these functions are performed. Here, we ask whether a subpopulation of zebrafish pineal cells, which express glial markers and visual cycle genes, is involved in maintaining photoreceptors. Selective ablation of these cells leads to a loss of pineal photoreceptors. Moreover, these cells internalize exorhodopsin that is secreted by pineal rod-like photoreceptors, and in turn release CD63-positive extracellular vesicles (EVs) that are taken up by pdgfrb-positive phagocytic cells in the forebrain meninges. These results identify a subpopulation of glial cells that is critical for pineal photoreceptor survival and indicate the existence of cells in the forebrain meninges that receive EVs released by these pineal cells and potentially function in waste disposal.

Phototransduction in Anuran Green Rods: Origins of Extra-Sensitivity.

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".

Wearable light spectral sensor optimized for measuring daily α-opic light exposure.

Light has many non-visual effects on human physiology, including alterations in sleep, mood, and alertness. These effects are mainly mediated by photoreceptors containing the photopigment melanopsin, which has a peak sensitivity to short wavelength ('blue') light. Commercially available light sensors are commonly wrist-worn and report photopic illuminance and are calibrated to perceive visual brightness and hence cannot be used to investigate the non-visual impacts of light. In this paper, we report the development of a wearable spectrophotometer designed to be worn as a pendant or affixed to clothing to capture spectral power density data close to eye level in the visible wavelength range 380-780 nm. From this, the relative impact of a given light stimulus can be determined for each photoreceptive input in the human eye by calculating effective illuminances. This device showed high accuracy for all effective illuminances while measuring a range of commonly encountered light sources by calibrating for directional response, dark noise, sensor saturation, non-linearity, stray-light and spectral response. Features of the device include IoT-integration, onboard data storage and processing, Bluetooth Low Energy (BLE) enabled data transfer, and cloud storage in one cohesive unit.

Coming of Age for the Photoreceptor Synapse.

Purpose: To discuss the potential contribution of rod and cone synapses to the loss of visual function in retinal injury and disease. Methods: The published literature and the authors' own work were reviewed. Results: Retinal detachment is used as a case study of rod spherule and cone pedicle plasticity after injury. Both rod and cone photoreceptors terminals are damaged after detachment although the structural changes observed are only partially overlapping. For second-order neurons, only those associated with rod spherules respond consistently to injury by remodeling. Examination of signaling pathways involved in plasticity of conventional synapses and in neural development has been and may continue to be productive in discovering novel therapeutic targets. Rho kinase (ROCK) inhibition is an example of therapy that may reduce synaptic damage by preserving normal synaptic structure of rod and cone cells. Conclusions: We hypothesize that synaptic damage contributes to poor visual restoration after otherwise successful anatomical repair of retinal detachment. A similar situation may exist for patients with degenerative retinal disease. Thus, synaptic structure and function should be routinely studied, as this information may disclose therapeutic strategies to mitigate visual loss.

Glucose-mediated de novo lipogenesis in photoreceptors drives early diabetic retinopathy.

Diabetic retinopathy (DR) is an increasingly frequent cause of blindness across populations; however, the events that initiate pathophysiology of DR remain elusive. Strong preclinical and clinical evidence suggests that abnormalities in retinal lipid metabolism caused by diabetes may account for the origin of this disease. A major arm of lipid metabolism, de novo biosynthesis, is driven by elevation in available glucose, a common thread binding all forms of vision loss in diabetes. Therefore, we hypothesized that aberrant retinal lipid biogenesis is an important promoter of early DR. In murine models, we observed elevations of diabetes-associated retinal de novo lipogenesis ∼70% over control levels. This shift was primarily because of activation of fatty acid synthase (FAS), a rate-limiting enzyme in the biogenic pathway. Activation of FAS was driven by canonical glucose-mediated disinhibition of acetyl-CoA carboxylase, a major upstream regulatory enzyme. Mutant mice expressing gain-of-function FAS demonstrated increased vulnerability to DR, whereas those with FAS deletion in rod photoreceptors maintained preserved visual responses upon induction of diabetes. Excess retinal de novo lipogenesis-either because of diabetes or because of FAS gain of function-was associated with modestly increased levels of palmitate-containing phosphatidylcholine species in synaptic membranes, a finding with as yet uncertain significance. These findings implicate glucose-dependent increases in photoreceptor de novo lipogenesis in the early pathogenesis of DR, although the mechanism of deleterious action of this pathway remains unclear.

Core circadian clock genes Per1 and Per2 regulate the rhythm in photoreceptor outer segment phagocytosis.

Retinal photoreceptors undergo daily renewal of their distal outer segments, a process indispensable for maintaining retinal health. Photoreceptor outer segment (POS) phagocytosis occurs as a daily peak, roughly about 1 hour after light onset. However, the underlying cellular and molecular mechanisms which initiate this process are still unknown. Here we show that, under constant darkness, mice deficient for core circadian clock genes (Per1 and Per2) lack a daily peak in POS phagocytosis. By qPCR analysis, we found that core clock genes were rhythmic over 24 hours in both WT and Per1, Per2 double mutant whole retinas. More precise transcriptomics analysis of laser capture microdissected WT photoreceptors revealed no differentially expressed genes between time points preceding and during the peak of POS phagocytosis. In contrast, we found that microdissected WT retinal pigment epithelium (RPE) had a number of genes that were differentially expressed at the peak phagocytic time point compared to adjacent ones. We also found a number of differentially expressed genes in Per1, Per2 double mutant RPE compared to WT ones at the peak phagocytic time point. Finally, based on STRING analysis, we found a group of interacting genes that potentially drive POS phagocytosis in the RPE. This potential pathway consists of genes such as: Pacsin1, Syp, Camk2b, and Camk2d among others. Our findings indicate that Per1 and Per2 are necessary clock components for driving POS phagocytosis and suggest that this process is transcriptionally driven by the RPE.

Interpretation of anatomic correlates of outer retinal bands in optical coherence tomography.

By providing the sectioning capability to differentiate individual retinal layers, optical coherence tomography (OCT) is revolutionizing eye disease diagnosis and treatment evaluation. A better understanding of the hyper- and hypo-reflective bands in retinal OCT is essential for accurate interpretation of clinical outcomes. In this article, we summarize the interpretations of clinical OCT and adaptive optics (AO) OCT (AO-OCT) of the outer retina in the human eye, and briefly review OCT investigation of the outer retina in animal models. Quantitative analysis of outer retinal OCT bands is compared to established parameters of retinal histology. The literature review and comparative analysis support that both inner/outer segment (IS/OS) junction and IS ellipsoid zone nonexclusively contribute to the second band; and OS, OS tips, and retinal pigment epithelium apical processes contribute to the third band in conventional OCT. In contrast, AO-OCT might predominantly detect the IS/OS junction and OS tip signals at the second and third bands due to its improved sectioning capability and possible AO effect on the sensitivities for recording ballistic and diffusive photons from different regions of the outer retina.

Pigment epithelium-derived factor (PEDF) and derived peptides promote survival and differentiation of photoreceptors and induce neurite-outgrowth in amacrine neurons.

Pigment epithelium-derived factor (PEDF) is a cytoprotective protein for the retina. We hypothesize that this protein acts on neuronal survival and differentiation of photoreceptor cells in culture. The purpose of the present study was to evaluate the neurotrophic effects of PEDF and its fragments in an in vitro model of cultured primary retinal neurons that die spontaneously in the absence of trophic factors. We used Wistar albino rats. Cell death was assayed by immunofluorescence and flow cytometry through TUNEL assay, propidium iodide, mitotracker, and annexin V. Immunofluorescence of cells for visualizing rhodopsin, CRX, and antisyntaxin under confocal microscopy was performed. Neurite outgrowth was also quantified. Results show that PEDF protected photoreceptor precursors from apoptosis, preserved mitochondrial function and promoted polarization of opsin enhancing their developmental process, as well as induced neurite outgrowth in amacrine neurons. These effects were abolished by an inhibitor of the PEDF receptor or receptor-derived peptides that block ligand/receptor interactions. While all the activities were specifically conferred by short peptide fragments (17 amino acid residues) derived from the PEDF neurotrophic domain, no effects were triggered by peptides from the PEDF antiangiogenic region. The observed effects on retinal neurons imply a specific activation of the PEDF receptor by a small neurotrophic region of PEDF. Our findings support the neurotrophic PEDF peptides as neuronal guardians for the retina, highlighting their potential as promoters of retinal differentiation, and inhibitors of retinal cell death and its blinding consequences. Cover Image for this issue: https://doi.org/10.1111/jnc.15089.

Variations in photoreceptor throughput to mouse visual cortex and the unique effects on tuning.

Visual input to primary visual cortex (V1) depends on highly adaptive filtering in the retina. In turn, isolation of V1 computations requires experimental control of retinal adaptation to infer its spatio-temporal-chromatic output. Here, we measure the balance of input to mouse V1, in the anesthetized setup, from the three main photoreceptor opsins-M-opsin, S-opsin, and rhodopsin-as a function of two stimulus dimensions. The first dimension is the level of light adaptation within the mesopic range, which governs the balance of rod and cone inputs to cortex. The second stimulus dimension is retinotopic position, which governs the balance of S- and M-cone opsin input due to the opsin expression gradient in the retina. The fitted model predicts opsin input under arbitrary lighting environments, which provides a much-needed handle on in-vivo studies of the mouse visual system. We use it here to reveal that V1 is rod-mediated in common laboratory settings yet cone-mediated in natural daylight. Next, we compare functional properties of V1 under rod and cone-mediated inputs. The results show that cone-mediated V1 responds to 2.5-fold higher temporal frequencies than rod-mediated V1. Furthermore, cone-mediated V1 has smaller receptive fields, yet similar spatial frequency tuning. V1 responses in rod-deficient (Gnat1-/-) mice confirm that the effects are due to differences in photoreceptor opsin contribution.

Complete congenital stationary night blindness associated with a novel NYX variant (p.Asn216Lys) in middle-aged and older adult patients.

BACKGROUND: Complete congenital stationary night blindness (CSNB) is a retinal disorder thought to be non-progressive. The purpose of this study was to characterize the clinical and genetic findings of middle-aged and older adult patients with X-linked complete CSNB. METHODS: Three male CSNB patients (aged 62, 72, and 51 years) and one unaffected female carrier in a Japanese family were included in this study. Whole-exome sequencing (WES) was performed to determine the disease-causing variants. Co-segregation was confirmed in the family members. We performed a comprehensive ophthalmic examination on each patient. RESULTS: In the 62-year-old patient, a novel hemizygous variant (c.648 C > A; p.Asn216Lys) of the NYX gene was identified by WES analysis. The other two patients carried the variant hemizygously, and the unaffected carrier harbored the variant heterozygously. The clinical and electroretinography (ERG) findings were very similar among all three patients. Fundus images exhibited high myopic chorioretinal atrophy with long axial length. Ultra-wide field fundus autofluorescence images showed no retinal degenerative changes except for changes resulting from high myopia and previous retinal diseases. The ERG findings showed no response in rod ERG, electronegative configuration with preserved a-waves in standard/bright-flash ERG, and preserved responses in cone and 30-Hz flicker ERG, which were compared with age-matched controls with high myopia. CONCLUSIONS: We identified a novel missense NYX variant in a Japanese family with complete CSNB. Our clinical findings indicated that photoreceptor mediated ERG responses are well preserved even in middle-aged and older adult patients.

Photoreceptor Disc Enclosure Is Tightly Controlled by Peripherin-2 Oligomerization.

Mutations in the PRPH2 gene encoding the photoreceptor-specific protein PRPH2 (also known as peripherin-2 or rds) cause a broad range of autosomal dominant retinal diseases. Most of these mutations affect the structure of the light-sensitive photoreceptor outer segment, which is composed of a stack of flattened "disc" membranes surrounded by the plasma membrane. The outer segment is renewed on a daily basis in a process whereby new discs are added at the outer segment base and old discs are shed at the outer segment tip. New discs are formed as serial membrane evaginations, which eventually enclose through a complex process of membrane remodeling (completely in rods and partially in cones). As disc enclosure proceeds, PRPH2 localizes to the rims of enclosed discs where it forms oligomers which fortify the highly curved membrane structure of these rims. In this study, we analyzed the outer segment phenotypes of mice of both sexes bearing a single copy of either the C150S or the Y141C PRPH2 mutation known to prevent or increase the degree of PRPH2 oligomerization, respectively. Strikingly, both mutations increased the number of newly forming, not-yet-enclosed discs, indicating that the precision of disc enclosure is regulated by PRPH2 oligomerization. Without tightly controlled enclosure, discs occasionally over-elongate and form large membranous "whorls" instead of disc stacks. These data show that the defects in outer segment structure arising from abnormal PRPH2 oligomerization are manifested at the stage of disc enclosure.SIGNIFICANCE STATEMENT The light-sensitive photoreceptor outer segment contains a stack of flattened "disc" membranes that are surrounded, or "enclosed," by the outer segment membrane. Disc enclosure is an adaptation increasing photoreceptor light sensitivity by facilitating the diffusion of the second messenger along the outer segment axes. However, the molecular mechanisms by which photoreceptor discs enclose within the outer segment membrane remain poorly understood. We now demonstrate that oligomers of the photoreceptor-specific protein peripherin-2, or PRPH2, play an active role in this process. We further propose that defects in disc enclosure because of abnormal PRPH2 oligomerization result in major structural abnormalities of the outer segment, ultimately leading to loss of visual function and cell degeneration in PRPH2 mutant models and human patients.

Vitamin A1/A2 chromophore exchange: Its role in spectral tuning and visual plasticity.

Vertebrate rod and cone photoreceptors detect light via a specialized organelle called the outer segment. This structure is packed with light-sensitive molecules known as visual pigments that consist of a G-protein-coupled, seven-transmembrane protein known as opsin, and a chromophore prosthetic group, either 11-cis retinal ('A1') or 11-cis 3,4-didehydroretinal ('A2'). The enzyme cyp27c1 converts A1 into A2 in the retinal pigment epithelium. Replacing A1 with A2 in a visual pigment red-shifts its spectral sensitivity and broadens its bandwidth of absorption at the expense of decreased photosensitivity and increased thermal noise. The use of vitamin A2-based visual pigments is strongly associated with the occupation of aquatic habitats in which the ambient light is red-shifted. By modulating the A1/A2 ratio in the retina, an organism can dynamically tune the spectral sensitivity of the visual system to better match the predominant wavelengths of light in its environment. As many as a quarter of all vertebrate species utilize A2, at least during a part of their life cycle or under certain environmental conditions. A2 utilization therefore represents an important and widespread mechanism of sensory plasticity. This review provides an up-to-date account of the A1/A2 chromophore exchange system.

Retinal degeneration-3 protein promotes photoreceptor survival by suppressing activation of guanylyl cyclase rather than accelerating GMP recycling.

Retinal degeneration-3 protein (RD3) deficiency causes photoreceptor dysfunction and rapid degeneration in the rd3 mouse strain and in human Leber's congenital amaurosis, a congenital retinal dystrophy that results in early vision loss. However, the mechanisms responsible for photoreceptor death remain unclear. Here, we tested two hypothesized biochemical events that may underlie photoreceptor death: (i) the failure to prevent aberrant activation of retinal guanylyl cyclase (RetGC) by calcium-sensor proteins (GCAPs) versus (ii) the reduction of GMP phosphorylation rate, preventing its recycling to GDP/GTP. We found that GMP converts to GDP/GTP in the photoreceptor fraction of the retina ∼24-fold faster in WT mice and ∼400-fold faster in rd3 mice than GTP conversion to cGMP by RetGC. Adding purified RD3 to the retinal extracts inhibited RetGC 4-fold but did not affect GMP phosphorylation in wildtype or rd3 retinas. RD3-deficient photoreceptors rapidly degenerated in rd3 mice that were reared in constant darkness to prevent light-activated GTP consumption via RetGC and phosphodiesterase 6. In contrast, rd3 degeneration was alleviated by deletion of GCAPs. After 2.5 months, only ∼40% of photoreceptors remained in rd3/rd3 retinas. Deletion of GCAP1 or GCAP2 alone preserved 68% and 57% of photoreceptors, respectively, whereas deletion of GCAP1 and GCAP2 together preserved 86%. Taken together, our in vitro and in vivo results support the hypothesis that RD3 prevents photoreceptor death primarily by suppressing activation of RetGC by both GCAP1 and GCAP2 but do not support the hypothesis that RD3 plays a significant role in GMP recycling.

Short prolactin isoforms are expressed in photoreceptors of canine retinas undergoing retinal degeneration.

Prolactin (PRL) hormone functions as a pleiotropic cytokine with a protective role in the retina. We recently identified by transcriptome profiling that PRL is one of the most highly upregulated mRNAs in the retinas of mutant rcd1 (PDE6B) and xlpra2 (RPGR) dogs at advanced stages of photoreceptor disease. In the present study, we have identified the expression of a short PRL isoform that lacks exon 1 in canine retinas and analyzed the time-course of expression and localization of this isoform in the retinas of these two models. Using laser capture microdissection to isolate RNA from each of the retinal cellular layers, we found by qPCR that this short PRL isoform is expressed in photoreceptors of degenerating retinas. We confirmed by in situ hybridization that its expression is localized to the outer nuclear layer and begins shortly after the onset of disease at the time of peak photoreceptor cell death in both models. PRL protein was also detected only in mutant dog retinas. Our results call for further investigations into the role of this novel PRL isoform in retinal degeneration.

A cell atlas of the chick retina based on single-cell transcriptomics.

Retinal structure and function have been studied in many vertebrate orders, but molecular characterization has been largely confined to mammals. We used single-cell RNA sequencing (scRNA-seq) to generate a cell atlas of the chick retina. We identified 136 cell types plus 14 positional or developmental intermediates distributed among the six classes conserved across vertebrates - photoreceptor, horizontal, bipolar, amacrine, retinal ganglion, and glial cells. To assess morphology of molecularly defined types, we adapted a method for CRISPR-based integration of reporters into selectively expressed genes. For Müller glia, we found that transcriptionally distinct cells were regionally localized along the anterior-posterior, dorsal-ventral, and central-peripheral retinal axes. We also identified immature photoreceptor, horizontal cell, and oligodendrocyte types that persist into late embryonic stages. Finally, we analyzed relationships among chick, mouse, and primate retinal cell classes and types. Our results provide a foundation for anatomical, physiological, evolutionary, and developmental studies of the avian visual system.

The evolutionary relationships of organisms and of genes have long been studied in various ways, including genome sequencing. More recently, the evolutionary relationships among the different types of cells that perform distinct roles in an organism, have become a subject of inquiry. High throughput single-cell RNA sequencing is a technique that allows scientists to determine what genes are switched on in single cells. This technique makes it possible to catalogue the cell types that make up a tissue and generate an atlas of the tissue based on what genes are switched on in each cell. The atlases can then be compared among species. The retina is a light-sensitive tissue that animals with a backbone, called vertebrates, use to see. The basic plan of the retina is very similar in vertebrates: five classes of neurons ­ the cells that make up the nervous system ­ are arranged into three layers. The chicken is a highly visual animal and it has frequently been used to study the development of the retina, from understanding how unspecialized embryonic cells become neurons to examining how circuits of neurons form. The structure and role of the retina have been studied in many vertebrates, but detailed descriptions of this tissue at the molecular level have been largely limited to mammals. To bridge this gap, Yamagata, Yan and Sanes generated the first cell atlas of the chicken retina. Additionally, they developed a gene editing-based technique based on CRISPR technology called eCHIKIN to label different cell types based on genes each type switched on selectively, providing a means of matching their shape and location to their molecular identity. Using these methods, it was possible to subdivide each of the five classes of neurons in the retina into multiple distinct types for a total of 136. The atlas provided a foundation for evolutionary analysis of how retinas evolve to serve the very different visual needs of different species. The chicken cell types could be compared to types previously identified in similar studies of mouse and primate retinas. Comparing the relationships among retinal cells in chickens, mice and primates revealed strong similarities in the overall cell classes represented. However, the results also showed big differences among species in the specific types within each class, and the genes that were switched on within each cell type. These findings may provide a foundation to study the anatomy, physiology, evolution, and development of the avian visual system. Until now, neural development of the chicken retina was being studied without comprehensive knowledge of its cell types or the developmentally important genes they express. The system developed by Yamagata, Yan and Sanes may be used in the future to learn more about vision and to investigate how neural cell types evolve to match the repertoire of each species to its environment.

Inhibition of PARP activity improves therapeutic effect of ARPE-19 transplantation in RCS rats through decreasing photoreceptor death.

Photoreceptor (PR) dysfunction or death is the key pathological change in retinal degeneration (RD). The death of PRs might be due to a primary change in PRs themselves or secondary to the dysfunction of the retinal pigment epithelium (RPE). Poly(ADP-ribose) polymerase (PARP) was reported to be involved in primary PR death, but whether it plays a role in PR death secondary to RPE dysfunction has not been determined. To clarify this question and develop a new therapeutic approach, we studied the changes in PAR/PARP in the RCS rat, a RD model, and tested the effect of PARP intervention when given alone or in combination with RPE cell transplantation. The results showed that poly(ADP-ribosyl)ation of proteins was increased in PRs undergoing secondary death in RCS rats, and this result was confirmed by the observation of similar changes in sodium iodate (SI)-induced secondary RD in SD rats. The increase in PAR/PARP was highly associated with increased apoptotic PRs and decreased visual function, as represented by lowered b-wave amplitudes on electroretinogram (ERG). Then, as we expected, when the RCS rats were treated with subretinal injection of the PARP inhibitor PJ34, the RD process was delayed. Furthermore, when PJ34 was given simultaneously with subretinal ARPE-19 cell transplantation, the therapeutic effects were significantly improved and lasted longer than those of ARPE-19 or PJ34 treatment alone. These results provide a potential new approach for treating RD.

Photoreceptive Ganglion Cells Drive Circuits for Local Inhibition in the Mouse Retina.

Intrinsically photosensitive retinal ganglion cells (ipRGCs) exhibit melanopsin-dependent light responses that persist in the absence of rod and cone photoreceptor-mediated input. In addition to signaling anterogradely to the brain, ipRGCs signal retrogradely to intraretinal circuitry via gap junction-mediated electrical synapses with amacrine cells (ACs). However, the targets and functions of these intraretinal signals remain largely unknown. Here, in mice of both sexes, we identify circuitry that enables M5 ipRGCs to locally inhibit retinal neurons via electrical synapses with a nonspiking GABAergic AC. During pharmacological blockade of rod- and cone-mediated input, whole-cell recordings of corticotropin-releasing hormone-expressing (CRH+) ACs reveal persistent visual responses that require both melanopsin expression and gap junctions. In the developing retina, ipRGC-mediated input to CRH+ ACs is weak or absent before eye opening, indicating a primary role for this input in the mature retina (i.e., in parallel with rod- and cone-mediated input). Among several ipRGC types, only M5 ipRGCs exhibit consistent anatomical and physiological coupling to CRH+ ACs. Optogenetic stimulation of local CRH+ ACs directly drives IPSCs in M4 and M5, but not M1-M3, ipRGCs. CRH+ ACs also inhibit M2 ipRGC-coupled spiking ACs, demonstrating direct interaction between discrete networks of ipRGC-coupled interneurons. Together, these results demonstrate a functional role for electrical synapses in translating ipRGC activity into feedforward and feedback inhibition of local retinal circuits.SIGNIFICANCE STATEMENT Melanopsin directly generates light responses in intrinsically photosensitive retinal ganglion cells (ipRGCs). Through gap junction-mediated electrical synapses with retinal interneurons, these uniquely photoreceptive RGCs may also influence the activity and output of neuronal circuits within the retina. Here, we identified and studied an electrical synaptic circuit that, in principle, could couple ipRGC activity to the chemical output of an identified retinal interneuron. Specifically, we found that M5 ipRGCs form electrical synapses with corticotropin-releasing hormone-expressing amacrine cells, which locally release GABA to inhibit specific RGC types. Thus, ipRGCs are poised to influence the output of diverse retinal circuits via electrical synapses with interneurons.