In vivo analysis of onset and progression of retinal degeneration in the Nr2e3rd7/rd7 mouse model of enhanced S-cone sensitivity syndrome

The photoreceptor-specific nuclear receptor Nr2e3 is not expressed in Nr2e3rd7/rd7 mice, a mouse model of the recessively inherited retinal degeneration enhanced S-cone sensitivity syndrome (ESCS). We characterized in detail C57BL/6J Nr2e3rd7/rd7 mice in vivo by fundus photography, optical coherence tomography and fluorescein angiography and, post mortem, by histology and immunohistochemistry. White retinal spots and so-called ‘rosettes’ first appear at postnatal day (P) 12 in the dorsal retina and reach maximal expansion at P21. The highest density in ‘rosettes’ is observed within a region located between 100 and 350 µM from the optic nerve head. ‘Rosettes’ disappear between 9 to 12 months. Non-apoptotic cell death markers are detected during the slow photoreceptor degeneration, at a rate of an approximately 3% reduction of outer nuclear layer thickness per month, as observed from 7 to 31 months of age. In vivo analysis of Nr2e3rd7/rd7 Cx3cr1gfp/+ retinas identified microglial cells within ‘rosettes’ from P21 on. Subretinal macrophages were observed in vivo and by confocal microscopy earliest in 12-months-old Nr2e3rd7/rd7 retinas. At P21, S-opsin expression and the number of S-opsin expressing dorsal cones was increased. The dorso-ventral M-cone gradient was present in Nr2e3rd7/rd7 retinas, but M-opsin expression and M-opsin expressing cones were decreased. Retinal vasculature was normal.

EML1 is essential for retinal photoreceptor localization and light detection

Calcium regulates the response sensitivity, kinetics and adaptation in photoreceptors. In striped bass cones, this calcium feedback includes direct modulation of the transduction cyclic nucleotide-gated (CNG) channels by the calcium-binding protein CNG-modulin. However, the possible role of EML1, the mammalian homolog of CNG-modulin, in modulating phototransduction in mammalian photoreceptors has not been examined. Here, we used mice expressing mutant Eml1 to investigate its role in the development and function of mouse photoreceptors using immunostaining, in-vivo and ex-vivo retinal recordings, and single-cell suction recordings. We found that the mutation of Eml1 causes significant changes in the mouse retinal structure characterized by mislocalization of rods and cones in the inner retina. Consistent with the fraction of mislocalized photoreceptors, rod and cone-driven retina responses were reduced in the mutants. However, the Eml1 mutation had no effect on the dark-adapted responses of rods in the outer nuclear layer. Notably, we observed no changes in the cone sensitivity in the Eml1 mutant animals, either in darkness or during light adaptation, ruling out a role for EML1 in modulating cone CNG channels. Together, our results suggest that EML1 plays an important role in retina development but does not modulate phototransduction in mammalian rods and cones.

Interspecies Correlations between Human and Mouse NR2E3-Associated Recessive Disease

NR2E3-associated recessive disease in humans is historically defined by congenital night blinding retinopathy, characterized by an initial increase in short-wavelength (S)-cone sensitivity and progressive loss of rod and cone function. The retinal degeneration 7 (rd7) murine model, harboring a recessive mutation in the mouse ortholog of NR2E3, has been a well-studied disease model and recently evaluated as a therapeutic model for NR2E3-associated retinal degenerations. This study aims to draw parallels between human and mouse NR2E3-related disease through examination of spectral domain optical coherence tomography (SD-OCT) imaging between different stage of human disease and its murine counterpart. We propose that SD-OCT is a useful non-invasive diagnostic tool to compare human clinical dystrophy presentation with that of the rd7 mouse and make inference that may be of therapeutically relevance. Additionally, a longitudinal assessment of rd7 disease progression, utilizing available clinical data from our patients as well as extensive retrospective analysis of visual acuity data from published cases of human NR2E3-related disease, was curated to identify further valuable correlates between human and mouse Nr2e3 disease. Results of this study validate the slow progression of NR2E3-associated disease in humans and the rd7 mice and identify SD-OCT characteristics in patients at or near the vascular arcades that correlate well with the whorls and rosettes that are seen also in the rd7 mouse and point to imaging features that appear to be associated with better preserved S-cone mediated retinal function. The correlation of histological findings between rd7 mice and human imaging provides a solid foundation for diagnostic use of pathophysiological and prognostic information to further define characteristics and a relevant timeline for therapeutic intervention in the field of NR2E3-associated retinopathies.

Quantifying Color Vision Changes Associated with Cataracts Using Cone Contrast Thresholds

PurposeThe cone contrast threshold (CCT) test quantified color vision changes in subjects of all ages and those undergoing cataract surgery.MethodsTwenty-four healthy volunteers from two cohort studies performed CCT using best corrected visual acuity, filters, mydriasis, and pinhole correction. Retrospective cross-sectional study of patients seen in eye clinics evaluated the relationship between age and color vision, and age and lens status in 355 eyes. Lastly, 25 subjects performed CCT before and after cataract surgery.ResultsCCT scores were most reliable in the non-mydriatic condition without pinhole correction. Progressively dense brown filters produced small but significant reductions in S-cone sensitivity. Linear regression analysis of phakic subjects showed a decline for all cone classes with age. Rate of decline was greater for S-cones (slope (95% CI) = −1.09 (−1.23, 0.94)) than M-cones (slope (95% CI) = −0.80 (−0.95, −0.66)) and L-cones (slope (95% CI) = −0.66 (−0.81, - 0.52)). CCT scores increased for S-cones but reduced for L- and M-cones in pseudophakic subjects compared to phakic patients. CCT scores after cataract surgery increased for S-cones, M-cones, and L-cones by 33.0 (p<0.001), 24.9 (p=0.001), and 22.0 (p=0.008).ConclusionsCCT assessment allows for clinically practical quantitation of color and contrast vision improvement after cataract surgery and aging patients who note poor vision despite good visual acuity.Translational RelevanceCCT testing, historically used in research, is now a clinically practical tool to evaluate age and cataract related changes in color and contrast vision and routinely quantify vision beyond black and white visual acuity testing.

Thyroid hormone receptor beta mutations alter photoreceptor development and function in Danio rerio (zebrafish)

We investigate a splice variant of thrb isolated in the retina, trβ2, identifying functional changes in larval and adult mutant zebrafish lacking trβ2. We constructed two CRISPR mutant zebrafish with mutations located in the N-terminus region. The first is a 6BP+1 insertion deletion frameshift resulting in a truncated protein. The second is a 3BP in frame deletion with intact binding domains. ERG recordings showed that the 6BP+1 mutants did not respond to red wavelengths of light while the 3BP mutants did respond. 6BP+1 mutants lacked optomotor and optokinetic responses to red/black and green/black contrasts. Adult 6BP+1 mutants exhibit a loss of red-cone contribution to the ERG, and an increase in green and UV contributions. Anatomical markers show loss of red-cones in the 6BP+1 mutant but increase in blue, green, and UV cone density. Our results confirm trβ2’s role in retinal cone development.Author SummaryThere are four cone photoreceptors responsible for color vision in zebrafish: red, green, blue, and UV. The thyroid hormone receptor trβ2 is localized in the vertebrate retina. We know that it is necessary for the development of long-wavelength-sensitive cones (red), but here we investigate the functional alterations that accompany a loss of trβ2. Our work contributes to the ongoing investigations of retinal development and the involvement of thyroid hormone receptors. Confirming previous morphological findings, we see that the fish become red colorblind when trβ2 is knocked out, but the contributions of the other three cone types shift in response. Our work highlights the plasticity of the retinal circuit as we see changes in opsin peaks and cone sensitivity, increases in contributions of UV cones, and an attempt at a mosaic pattern in the adult retina all in the absence of trβ2 and red cones. We now have an increased understanding of mechanisms underlying retinal development

Mitochondrial absorption of short wavelength light drives primate blue retinal cones into glycolysis which may increase their pace of aging

Photoreceptors have high energy demands and densely packed mitochondria through which light passes before phototransduction. Old world primates including humans have three cone photoreceptor types mediating color vision with short (S blue), medium (M green), and long (L red) wavelength sensitivities. However, S-cones are enigmatic. They comprise <10% of the total cone population, their responses saturate early, and they are susceptible in aging and disease. Here, we show that primate S-cones actually have few mitochondria and are fueled by glycolysis, not by mitochondrial respiration. Glycolysis has a limited ability to sustain activity, potentially explaining early S-cone saturation. Mitochondria act as optical filters showing reduced light transmission at 400–450 nm where S-cones are most sensitive (420 nm). This absorbance is likely to arise in a mitochondrial porphyrin that absorbs strongly in the Soret band. Hence, reducing mitochondria will improve S-cone sensitivity but result in increased glycolysis as an alternative energy source, potentially increasing diabetic vulnerability due to restricted glucose access. Further, glycolysis carries a price resulting in premature functional decline as seen in aged S-cones. Soret band absorption may also impact on mitochondrial rich M and L cones by reducing sensitivity at the lower end of their spectral sensitivity range resulting in increased differentiation from S-cone responses. These data add to the list of unique characteristic of S-cones and may also explain aspects of their vulnerability.