Cone Photoreceptors Research Articles

Natural disease history of a canine model of oligogenic RPGRIP1-cone-rod dystrophy establishes variable effects of previously and newly mapped modifier loci.

Canine RPGRIP1-cone-rod dystrophy (CRD), a model for human inherited retinal diseases (IRDs), was originally identified as autosomal recessive early-onset blindness. However, later studies revealed extensive phenotypic variability among RPGRIP1 mutants. This led to the identification of a homozygous MAP9 variant as a modifier associated with early-onset disease. Based on further phenotypic variation affecting cone photoreceptor function, we report mapping of L3 as an additional modifier locus, within a 4.1-Mb locus on canine chromosome 30. We establish the natural disease history of RPGRIP1-CRD based on up to 9-year long-term functional and structural retinal data from 58 dogs including 44 RPGRIP1 mutants grouped according to the modifier status. RPGRIP1 mutants affected by both MAP9 and L3 modifiers exhibited the most severe phenotypes with rapid disease progression. MAP9 alone was found to act as an overall accelerator of rod and cone diseases, while L3 had a cone-specific effect. Ultrastructural analysis of photoreceptors revealed varying degrees of rod and cone damage, while the connecting cilia appeared structurally preserved in all groups. We conclude that RPGRIP1-CRD is an oligogenic disease with at least three loci contributing to the pathogenesis. While the RPGRIP1 variant is required for developing the disease, MAP9 and L3 modifiers exacerbate the phenotype, individually and cumulatively. Oligogenic canine RPGRIP1-CRD illustrates the impact of multiple genetic modifiers on disease phenotype and thus has the potential to reveal new targets for broad-spectrum therapies for oligogenic or polygenic forms of human IRDs.

Preclinical evaluation of ADVM-062, a novel intravitreal gene therapy vector for the treatment of blue cone monochromacy

Blue cone monochromacy (BCM) is a rare X-linked retinal disease characterized by the absence of L- and M-opsin in cone photoreceptors, considered a potential gene therapy candidate. However, most experimental ocular gene therapies utilize subretinal vector injection which would pose a risk to the fragile central retinal structure of BCM patients. Here we describe the use of ADVM-062, a vector optimized for cone-specific expression of human L-opsin and administered using a single intravitreal (IVT) injection. Pharmacological activity of ADVM-062 was established in gerbils, whose cone-rich retina naturally lacks L-opsin. A single IVT administration dose of ADVM-062 effectively transduced gerbil cone photoreceptors and produced a de novo response to long-wavelength stimuli. To identify potential first-in-human doses we evaluated ADVM-062 in non-human primates. Cone-specific expression of ADVM-062 in primates was confirmed using ADVM-062.myc, a vector engineered with the same regulatory elements as ADVM-062. Enumeration of human OPN1LW.myc-positive cones demonstrated that doses ≥3×1010 vg/eye resulted in transduction of 18%-85% of foveal cones. A Good Laboratory Practice (GLP) toxicology study established that IVT administration of ADVM-062 was well tolerated at doses that could potentially achieve clinically meaningful effect, thus supporting the potential of ADVM-062 as a one-time IVT gene therapy forBCM.

A laser-induced mouse model of progressive retinal degeneration with central sparing displays features of parafoveal geographic atrophy

There are no disease-modifying treatments available for geographic atrophy (GA), the advanced form of dry age-related macular degeneration. Current murine models fail to fully recapitulate the features of GA and thus hinder drug discovery. Here we describe a novel mouse model of retinal degeneration with hallmark features of GA. We used an 810 nm laser to create a retinal lesion with central sparing (RLCS), simulating parafoveal atrophy observed in patients with progressive GA. Laser-induced RLCS resulted in progressive GA-like pathology with the development of a confluent atrophic lesion. We demonstrate significant changes to the retinal structure and thickness in the central unaffected retina over a 24-week post-laser period, confirmed by longitudinal optical coherence tomography scans. We further show characteristic features of progressive GA, including a gradual reduction in the thickness of the central, unaffected retina and of total retinal thickness. Histological changes observed in the RLCS correspond to GA pathology, which includes the collapse of the outer nuclear layer, increased numbers of GFAP + , CD11b + and FcγRI + cells, and damage to cone and rod photoreceptors. We demonstrate a laser-induced mouse model of parafoveal GA progression, starting at 2 weeks post-laser and reaching confluence at 24 weeks post-laser. This 24-week time-frame in which GA pathology develops, provides an extended window of opportunity for proof-of-concept evaluation of drugs targeting GA. This time period is an added advantage compared to several existing models of geographic atrophy.

Genetic and Clinical Characterization of Danish Achromatopsia Patients.

Achromatopsia is a rare congenital condition with cone photoreceptor dysfunction causing color blindness, reduced vision, nystagmus and photophobia. New treatments are being developed, but the current evidence is still conflicting regarding possible progression over time, and there is no clear genotype-phenotype correlation. This natural history study aimed to further explore the course of disease and potential clinical differences between various genotypes. The retrospective design allowed for the study of a large cohort with a long follow-up. Patients were identified from the Danish national registries. If not already available, genetic analysis was offered to the patient. Clinical data from 1945-2022 were retrieved from medical records and included best-corrected visual acuity (BCVA), color vision, refractive error, nystagmus, visual fields and fundoscopic findings. We identified variants believed to be disease causing in five of the known achromatopsia genes: CNGA3; CNGB3; GNAT2; PDE6C and PDE6H; and novel variants were identified in CNGB3 and PDE6C. Progressive deterioration of BCVA only attributable to achromatopsia was found in three of 58 patients. Progressive phenotype was seen with variants in CNGB3 and PDE6C. The results indicate that myopia could be more frequently occurring with variants in GNAT2, PDE6C and PDE6H and support the evidence that achromatopsia is a predominantly stationary condition with respect to BCVA. Although a clear genotype-phenotype correlation can still not be concluded, there may be differences in phenotypical characteristics with variants in different genes.

Blimp-1/PRDM1 and Hr3/RORβ specify the blue-sensitive photoreceptor subtype in Drosophila by repressing the hippo pathway.

During terminal differentiation of the mammalian retina, transcription factors control binary cell fate decisions that generate functionally distinct subtypes of photoreceptor neurons. For instance, Otx2 and RORβ activate the expression of the transcriptional repressor Blimp-1/PRDM1 that represses bipolar interneuron fate and promotes rod photoreceptor fate. Moreover, Otx2 and Crx promote expression of the nuclear receptor Nrl that promotes rod photoreceptor fate and represses cone photoreceptor fate. Mutations in these four transcription factors cause severe eye diseases such as retinitis pigmentosa. Here, we show that a post-mitotic binary fate decision in Drosophila color photoreceptor subtype specification requires ecdysone signaling and involves orthologs of these transcription factors: Drosophila Blimp-1/PRDM1 and Hr3/RORβ promote blue-sensitive (Rh5) photoreceptor fate and repress green-sensitive (Rh6) photoreceptor fate through the transcriptional repression of warts/LATS, the nexus of the phylogenetically conserved Hippo tumor suppressor pathway. Moreover, we identify a novel interaction between Blimp-1 and warts, whereby Blimp-1 represses a warts intronic enhancer in blue-sensitive photoreceptors and thereby gives rise to specific expression of warts in green-sensitive photoreceptors. Together, these results reveal that conserved transcriptional regulators play key roles in terminal cell fate decisions in both the Drosophila and the mammalian retina, and the mechanistic insights further deepen our understanding of how Hippo pathway signaling is repurposed to control photoreceptor fates for Drosophila color vision.

Gap junction Delta-2b (gjd2b/Cx35.1) depletion causes hyperopia and visual-motor deficiencies in the zebrafish.

The zebrafish is a powerful model to investigate the developmental roles of electrical synapses because many signaling pathways that regulate the development of the nervous system are highly conserved from fish to humans. Here, we provide evidence linking the mammalian connexin-36 (Cx36) ortholog gjd2b/Cx35.1, a major component of electrical synapses in the zebrafish, with a refractive error in the context of morphological, molecular, and behavioral changes of zebrafish larvae. Two abnormalities were identified. The optical coherence tomography analysis of the adult retina confirmed changes to the refractive properties caused by eye axial length reduction, leading to hyperopic shifts. The gjd2b/Cx35.1 depletion was also correlated with morphological changes to the head and body ratios in larvae. The differential expression of Wnt/ß-catenin signaling genes, connexins, and dopamine receptors suggested a contribution to the observed phenotypic differences. The alteration of visual-motor behavioral responses to abrupt light transitions was aggravated in larvae, providing evidence that cone photoreceptor cell activity was enhanced when gjd2b/Cx35.1 was depleted. The visual disturbances were reversed under low light conditions in gjd2b -/- /Cx35.1-/- larvae. Since qRT-PCR data demonstrated that two rhodopsin genes were downregulated, we speculated that rod photoreceptor cells in gjd2b/Cx35.1-/- larvae were less sensitive to bright light transitions, thus providing additional evidence that a cone-mediated process caused the VMR light-ON hyperactivity after losing Cx35.1 expression. Together, this study provides evidence for the role of gjd2b/Cx35.1 in the development of the visual system and visually guided behaviors.

From mouse to human: Accessing the biochemistry of vision in vivo by two-photon excitation.

The eye is an ideal organ for imaging by a multi-photon excitation approach, because ocular tissues such as the sclera, cornea, lens and neurosensory retina, are highly transparent to infrared (IR) light. The interface between the retina and the retinal pigment epithelium (RPE) is especially informative, because it reflects the health of the visual (retinoid) cycle and its changes in response to external stress, genetic manipulations, and drug treatments. Vitamin A-derived retinoids, like retinyl esters, are natural fluorophores that respond to multi-photon excitation with near IR light, bypassing the filter-like properties of the cornea, lens, and macular pigments. Also, during natural aging some retinoids form bisretinoids, like diretinoid-pyridiniumethanolamine (A2E), that are highly fluorescent. These bisretinoids appear to be elevated concurrently with aging. Vitamin A-derived retinoids and bisretinoidss are detected by two-photon ophthalmoscopy (2PO), using a new class of light sources with adjustable spatial, temporal, and spectral properties. Furthermore, the two-photon (2P) absorption of IR light by the visual pigments in rod and cone photoreceptors can initiate visual transduction by cis-trans isomerization of retinal, enabling parallel functional studies. Recently we overcame concerns about safety, data interpretation and complexity of the 2P-based instrumentation, the major roadblocks toward advancing this modality to the clinic. These imaging and retina-function assessment advancements have enabled us to conduct the first 2P studies with humans.

Self‐powered transparent photodetector for subretinal visual functions of wide‐field‐of‐view and broadband perception

AbstractNatural photoreceptors enable color vision in humans, wherein the eyes detect colors and their corresponding intensities via cone and rod photoreceptors, respectively. Herein, we developed an artificial broadband photoreceptor with light‐color intensity detection similar to that of natural photoreceptors. The developed photoreceptor operates in the self‐powered mode and is capable of broadband perception (365–940 nm). The designed metal‐oxide heterojunction (n‐ZnO/p‐NiO) photoreceptor with a thin tin sulfide layer embedded in between is capable of perceiving various colors. It exhibits good transparency in the visible range and displays excellent integration with flexible substrates, highlighting its potential for use in flexible electronics. The fabricated structure has an exceptional response time (≈1 ms) and a wide‐field‐of‐view (150°) compared to the human eye's sensing range (50–100 ms and 108°). The transparent photoreceptor mimics cones and rods to detect a various wavelength‐dependent signals and explicitly differentiate between the intensities of the detected signals, respectively. This is further illustrated by employing the developed photoreceptor to detect colors in real time by generating unique signals corresponding to each color. The demonstration provides the proof of concept for self‐biased flexible bioelectronics emulating high‐performing visual functions of artificial eyes.image

Reduced inspired oxygen decreases retinal superoxide radicals and promotes cone function and survival in a model of retinitis pigmentosa.

Retinitis pigmentosa (RP) is caused by many different mutations that promote the degeneration of rod photoreceptors and have no direct effect on cones. After the majority of rods have died cone photoreceptors begin to slowly degenerate. Oxidative damage contributes to cone cell death and it has been hypothesized that tissue hyperoxia due to reduced oxygen consumption from the loss of rods is what initiates oxidative stress. Herein, we demonstrate in animal models of RP that reduction of retinal hyperoxia by reducing inspired oxygen to continuous breathing of 11% O2 reduced the generation of superoxide radicals in the retina and preserved cone structure and function. These data indicate that retinal hyperoxia is the initiating event that promotes oxidative damage, loss of cone function, and cone degeneration in the RP retina.

Histological characterization of retinal degeneration in mucopolysaccharidosis type IIIC

Heparan-α-glucosaminide N-acetyltransferase (HGSNAT) participates in lysosomal degradation of heparan sulfate. Mutations in the gene encoding this enzyme cause mucopolysaccharidosis IIIC (MPS IIIC) or Sanfilippo syndrome type C. MPS IIIC patients exhibit progressive neurodegeneration, leading to dementia and death in early adulthood. Currently there is no approved treatment for MPS IIIC. Incidences of non-syndromic retinitis pigmentosa and early signs of night blindness are reported in some MPS IIIC patients, however the majority of ocular phenotypes are not well characterized. The goal of this study was to investigate retinal degeneration phenotype in the Hgsnat knockout mouse model of MPS IIIC and a cadaveric human MPS IIIC eye. Cone and rod photoreceptors in the eyes of homozygous 6-month-old Hgsnat knockout mice and their wild-type counterparts were analyzed using cone arrestin, S-opsin, M-opsin and rhodopsin antibodies. Histological observation was performed on the eye from a 35-year-old MPS IIIC donor. We observed a nearly 50% reduction in the rod photoreceptors density in the Hgsnat knockout mice compared to the littermate wild-type controls. Cone photoreceptor density was unaltered at this age. Severe retinal degeneration was also observed in the MPS IIIC donor eye. To our knowledge, this is the first report characterizing ocular phenotypes arising from deleterious variants in the Hgsnat gene associated with MPS IIIC clinical phenotype. Our findings indicate retinal manifestations may be present even before behavioral manifestations. Thus, we speculate that ophthalmological evaluations could be used as diagnostic indicators of early disease, progression, and end-point evaluation for future MPS IIIC therapies.

Retinal Dysfunction in a Mouse Model of HCN1 Genetic Epilepsy.

Pathogenic variants in HCN1 are associated with a range of epilepsy syndromes including a developmental and epileptic encephalopathy. The recurrent de novo HCN1 pathogenic variant (M305L) results in a cation leak, allowing the flux of excitatory ions at potentials where the wild-type channels are closed. The Hcn1M294L mouse recapitulates patient seizure and behavioral phenotypes. As HCN1 channels are highly expressed in rod and cone photoreceptor inner segments, where they shape the light response, mutated channels are likely to impact visual function. Electroretinogram (ERG) recordings from male and female mice Hcn1M294L mice revealed a significant decrease in the photoreceptor sensitivity to light, as well as attenuated bipolar cell (P2) and retinal ganglion cell responses. Hcn1M294L mice also showed attenuated ERG responses to flickering lights. ERG abnormalities are consistent with the response recorded from a single female human subject. There was no impact of the variant on the structure or expression of the Hcn1 protein in the retina. In silico modeling of photoreceptors revealed that the mutated HCN1 channel dramatically reduced light-induced hyperpolarization, resulting in more Ca2+ flux during the response when compared with the wild-type situation. We propose that the light-induced change in glutamate release from photoreceptors during a stimulus will be diminished, significantly blunting the dynamic range of this response. Our data highlight the importance of HCN1 channels to retinal function and suggest that patients with HCN1 pathogenic variants are likely to have a dramatically reduced sensitivity to light and a limited ability to process temporal information.SIGNIFICANCE STATEMENT Pathogenic variants in HCN1 are emerging as an important cause of catastrophic epilepsy. HCN1 channels are ubiquitously expressed throughout the body, including the retina. Electroretinogram recordings from a mouse model of HCN1 genetic epilepsy showed a marked decrease in the photoreceptor sensitivity to light and a reduced ability to respond to high rates of light flicker. No morphologic deficits were noted. Simulation data suggest that the mutated HCN1 channel blunts light-induced hyperpolarization and consequently limits the dynamic range of this response. Our results provide insights into the role HCN1 channels play in retinal function as well as highlighting the need to consider retinal dysfunction in disease caused by HCN1 variants. The characteristic changes in the electroretinogram open the possibility of using this tool as a biomarker for this HCN1 epilepsy variant and to facilitate development of treatments.

Modeling of dielectric resonator antenna array for retina photoreceptors

The retina encompasses several cone and rod photoreceptors at fovea region i.e., 90 million cells of rod photoreceptors and 4.5million cells of cone photoreceptors. The overall photoreceptors determine the vision of every human. An electromagnetic dielectric resonator antenna has been presented for retina photoreceptors in order to model them at fovea and its peripheral retina with the respected angular spectrum. Three coloring primary system of human eye (R, G, B) can be realized based on the model. Three miscellaneous models i.e., simple, graphene coated, and interdigital models have been presented in this paper. The nonlinear property of interdigital structures is one of the best advantages to use for creating the capacitor. The capacitance property helps improving the upper band of visible spectrum. The absorption of light for graphene as an energy harvesting material and its conversion into electrochemical signals is making it one of the best models. The mentioned three electromagnetic models of human photoreceptors have been expressed as a receiver antenna. The proposed electromagnetic models based on dielectric resonator antenna (DRA) are being analyzed for cones and rods photoreceptors of retina in the human eye by Finite Integral Method (FIM) utilized by CST MWS. The results show that the models are so fine for vision spectrum due to its localized near field enhancement property. The results indicate fine parameters of S11 (return loss below -10 dB) with invaluable resonants in a wide range of frequencies from 405 THz to 790 THz (vision spectrum), appropriate S21 (insertion loss 3-dB bandwidth), very good field distribution of electric and magnetic fields for flowing the power and electrochemical signals. Finally, mfERG clinical and experimental results validate the numeric results by the normalized output to input ratio of these models and it points out that these models can stimulate the electrochemical signals in photoreceptor cells for the best suiting of realizing the new retinal implants.

C-subfamily ATP binding cassette transporters extrude the calcium fluorescent probe fluo-4 from a cone photoreceptor cell line.

Whole transcriptome sequencing has revealed the existence of mRNAs for multiple membrane transporters in photoreceptors. Except for ATP binding cassette (ABC) member A4, involved in the retinoid cycle, an understanding of the function of most transport proteins in photoreceptors is lacking. In this research paper, extrusion of fluo-4, a Ca2+ fluorescent probe, from 661W cells, a cone photoreceptor murine cell line was studied with online fluorometry and immunocytochemistry. Fluo-4 efflux was temperature dependent, required ATP but not extracellular Na+, was not affected by pH in the range 5.4-8.4, and followed saturating kinetics with a Km of nearly 4μM, suggesting it was effected by ABC type transporters. A panel of antagonists showed an inhibitory profile typical of the C subfamily of ABC transporters. Immunofluorescence staining was positive for ABCC3, ABCC4 and ABCC5. These experimental results are compatible with fluo-4 being extruded from 661W cones by one or a combination of C-type ABC transporters. Examination of physicochemical descriptors related to drug membrane permeability and ABC substrate binding region further suggested efflux of fluo-4 by C-type ABC transporters. Possible functions of this transport mechanism in photoreceptors are discussed.

Systematic analysis of CNGA3 splice variants identifies different mechanisms of aberrant splicing

Achromatopsia is an autosomal recessive cone photoreceptor disease that is frequently caused by pathogenic variants in the CNGA3 gene. Here, we present a systematic functional analysis of 20 CNGA3 splice site variants detected in our large cohort of achromatopsia patients and/or listed in common variant databases. All variants were analyzed by functional splice assays based on the pSPL3 exon trapping vector. We demonstrated that ten variants, both at canonical and non-canonical splice sites, induced aberrant splicing, including intronic nucleotide retention, exonic nucleotide deletion and exon skipping, resulting in 21 different aberrant transcripts. Of these, eleven were predicted to introduce a premature termination codon. The pathogenicity of all variants was assessed based on established guidelines for variant classification. Incorporation of the results of our functional analyses enabled re-classification of 75% of variants previously classified as variants of uncertain significance into either likely benign or likely pathogenic. Our study is the first in which a systematic characterization of putative CNGA3 splice variants has been performed. We demonstrated the utility of pSPL3 based minigene assays in the effective assessment of putative splice variants. Our findings improve the diagnosis of achromatopsia patients, who may thus benefit from future gene-based therapeutic strategies.

Novel compound heterozygous variants in the USH2A gene associated with autosomal recessive retinitis pigmentosa without hearing loss.

Background: Retinitis pigmentosa (RP) is a group of progressive inherited retinal dystrophies characterized by the primary degeneration of rod photoreceptors and the subsequent loss of cone photoreceptors because of cell death. It is caused by different mechanisms, including inflammation, apoptosis, necroptosis, pyroptosis, and autophagy. Variants in the usherin gene (USH2A) have been reported in autosomal recessive RP with or without hearing loss. In the present study, we aimed to identify causative variants in a Han-Chinese pedigree with autosomal recessive RP. Methods: A six-member, three-generation Han-Chinese family with autosomal recessive RP was recruited. A full clinical examination, whole exome sequencing, and Sanger sequencing, as well as co-segregation analysis were performed. Results: Three heterozygous variants in the USH2A gene, c.3304C>T (p.Q1102*), c.4745T>C (p.L1582P), and c.14740G>A (p.E4914K), were identified in the proband, which were inherited from parents and transmitted to the daughters. Bioinformatics analysis supported the pathogenicity of the c.3304C>T (p.Q1102*) and c.4745T>C (p.L1582P) variants. Conclusions: Novel compound heterozygous variants in the USH2A gene, c.3304C>T (p.Q1102*) and c.4745T>C (p.L1582P), were identified as the genetic causes of autosomal recessive RP. The findings may enhance the current knowledge of the pathogenesis of USH2A-associated phenotypes, expand the spectrum of the USH2A gene variants, and contribute to improved genetic counseling, prenatal diagnosis, and disease management.

High-Efficiency CRISPR/Cas9-Mediated Correction of a Homozygous Mutation in Achromatopsia-Patient-Derived iPSCs.

Achromatopsia is an autosomal recessive disorder, in which cone photoreceptors undergo progressive degeneration, causing color blindness and poor visual acuity, among other significant eye affectations. It belongs to a group of inherited retinal dystrophies that currently have no treatment. Although functional improvements have been reported in several ongoing gene therapy studies, more efforts and research should be carried out to enhance their clinical application. In recent years, genome editing has arisen as one of the most promising tools for personalized medicine. In this study, we aimed to correct a homozygous PDE6C pathogenic variant in hiPSCs derived from a patient affected by achromatopsia through CRISPR/Cas9 and TALENs technologies. Here, we demonstrate high efficiency in gene editing by CRISPR/Cas9 but not with TALENs approximation. Despite a few of the edited clones displaying heterozygous on-target defects, the proportion of corrected clones with a potentially restored wild-type PDE6C protein was more than half of the total clones analyzed. In addition, none of them presented off-target aberrations. These results significantly contribute to advances in single-nucleotide gene editing and the development of future strategies for the treatment of achromatopsia.

Observations for Sjögren’s Pigment Epithelial Reticular Dystrophy in a 16-Year-Old Boy—An Extremely Rare Retinal Case Report

The purpose of this publication is to present an extremely rare case of Sjögren's pigment epithelial reticular dystrophy. So far, 10 such publications have been found in world literature. A 16-year-old boy was diagnosed due to a slight loss of visual acuity, confirmed in static perimetry/24-2/. Abnormal dense clusters of retinal pigment epithelium (RPE) cells forming a reticular network pattern (resembling a fishing net) with marked knots were detected by fundoscopy in the macular area and the mid-periphery of the retina. No abnormalities were found in the anterior segment, intraocular pressure, kinetic perimetry, Ishihara or Farnsworth D-15 tests or OCT. Fluorescein angiography confirmed blocked fluorescence from the choroidal vessels caused by the pigment in RPE. An autofluorescence test showed hypofluorescent foci corresponding to symmetrical and bilateral retinal hyperpigmentation with an RPE reticular pattern. Multifocal ERG (mfERG) revealed slight cone photoreceptor and bipolar bioelectrical dysfunction. Electrooculography (EOG) showed significant asymmetry (Arden Ratio 1.8), suggesting bioelectrical dysfunction of RPE/photoreceptors. Flash ERG (ERG) revealed only slight increase in implicit time of the a and b waves of the rod and cone responses and exclude cone-rod dystrophies. This article highlights the importance of the results of ophthalmoscopy, fluorescein angiography, autofluorescence, mfERG, fERG, EOG and genetic tests for Sjögren's reticular dystrophy with a pathogenic variant in the region of the C2 gene-c.841_849+19del (dbSNP rs9332736).

Spatial distribution of metabolites in the retina and its relevance to studies of metabolic retinal disorders

IntroductionThe primate retina has evolved regional specialisations for specific visual functions. The macula is specialised towards high acuity vision and is an area that contains an increased density of cone photoreceptors and signal processing neurons. Different regions in the retina display unique susceptibility to pathology, with many retinal diseases primarily affecting the macula.ObjectivesTo better understand the properties of different retinal areas we studied the differential distribution of metabolites across the retina.MethodsWe conducted an untargeted metabolomics analysis on full-thickness punches from three different regions (macula, temporal peri-macula and periphery) of healthy primate retina.ResultsNearly half of all metabolites identified showed differential abundance in at least one comparison between the three regions. Furthermore, mapping metabolomics results from macula-specific eye diseases onto our region-specific metabolite distributions revealed differential abundance defining systemic metabolic dysregulations that were region specific.ConclusionsThe unique metabolic phenotype of different retinal regions is likely due to the differential distribution of different cell types in these regions reflecting the specific metabolic requirements of each cell type. Our results may help to better understand the pathobiology of retinal diseases with region specificity.

Cost-effective sequence analysis of 113 genes in 1,192 probands with retinitis pigmentosa and Leber congenital amaurosis.

Introduction: Retinitis pigmentosa (RP) and Leber congenital amaurosis (LCA) are two groups of inherited retinal diseases (IRDs) where the rod photoreceptors degenerate followed by the cone photoreceptors of the retina. A genetic diagnosis for IRDs is challenging since >280 genes are associated with these conditions. While whole exome sequencing (WES) is commonly used by diagnostic facilities, the costs and required infrastructure prevent its global applicability. Previous studies have shown the cost-effectiveness of sequence analysis using single molecule Molecular Inversion Probes (smMIPs) in a cohort of patients diagnosed with Stargardt disease and other maculopathies. Methods: Here, we introduce a smMIPs panel that targets the exons and splice sites of all currently known genes associated with RP and LCA, the entire RPE65 gene, known causative deep-intronic variants leading to pseudo-exons, and part of the RP17 region associated with autosomal dominant RP, by using a total of 16,812 smMIPs. The RP-LCA smMIPs panel was used to screen 1,192 probands from an international cohort of predominantly RP and LCA cases. Results and discussion: After genetic analysis, a diagnostic yield of 56% was obtained which is on par with results from WES analysis. The effectiveness and the reduced costs compared to WES renders the RP-LCA smMIPs panel a competitive approach to provide IRD patients with a genetic diagnosis, especially in countries with restricted access to genetic testing.

Cone photoreceptors transfer damaged mitochondria to Müller glia.

Mitochondria are vital organelles that require sophisticated homeostatic mechanisms for maintenance. Intercellular transfer of damaged mitochondria is a recently identified strategy broadly used to improve cellular health and viability. Here, we investigate mitochondrial homeostasis in the vertebrate cone photoreceptor, the specialized neuron that initiates our daytime and color vision. We find a generalizable response to mitochondrial stress that leads to loss of cristae, displacement of damaged mitochondria from their normal cellular location, initiation of degradation, and transfer to Müller glia cells, a key non-neuronal support cell in the retina. Our findings show transmitophagy from cones to Müller glia as a response to mitochondrial damage. Intercellular transfer of damaged mitochondria represents an outsourcing mechanism that photoreceptors use to support their specialized function.