Mutations in cone opsin-encoding genes are associated with colorblindness and retinal disease. In certain forms of blue cone monochromacy (BCM) in humans, a viable cone population that does not express cone opsin is present in the retina. Double cone opsin knockout (DKO) mouse models of BCM have been developed previously using gene trap insertion in one or both opsin genes. Here, we report and characterize a new mouse line that completely lacks both short-wavelength (S) and middle-wavelength (M) sensitive cone opsin genes but maintains a viable cone population to one year old. DKO mice experienced approximately a 30% reduction in cones between 4 and 12 months, similar to the age-related decline in wild-type controls, and did not show signs of retinal degeneration. This mouse will be useful for the development of cone opsin gene therapies directed at treating BCM, in which the expression of L and M opsin is lost, but many cones remain as potential gene therapy targets. It also has implications for understanding the survival of cones without opsin, which differs from the fate of rods without rhodopsin.

Background/Objectives: Ganglion cells are the projection neurons of the retina, and there are multiple types that differ in their morphology, light responses and central projections. Parasol cells are one of the major retinal ganglion cell types in primates. The presynaptic bipolar cells have been well-characterized, but less is known about the amacrine cells that provide the majority of their inputs. The goal of this study was to identify the amacrine cells presynaptic to the OFF subtype of parasol cells. Methods: Central retinal tissue from an adult macaque was processed for serial block-face scanning EM, and a volume of images of the inner retina located 2 mm temporal to the center of the fovea was analyzed. Results: All the OFF parasol cells in the volume were reconstructed. All the synaptic inputs of two OFF parasol cells were analyzed. They received 80% or more of their input from amacrine cells and the remainder from bipolar cells, almost entirely from the Off diffuse type. Many of the presynaptic amacrine cells were reconstructed sufficiently to be classified as wide-field or narrow-field, and the latter type predominated. Five specific types of presynaptic amacrine cells were identified as AII, A4, knotty bistratified type 1, A13 and wiry type 1. Notably, the same types of amacrine cells are also presynaptic to OFF midget ganglion cells, another major type. Conclusions: These findings suggest that differences between the light responses of midget and parasol ganglion cells likely arise from differences in the presynaptic bipolar cell types.

In primates, the OFF midget retinal ganglion cells (OFF mRGCs) have a high spatial density and small dendritic arbors. Their axons provide input to the parvocellular pathway mediating both colour vision and the highest-acuity spatial vision. This study aimed to understand the basis for their light responses by identifying the presynaptic amacrine and bipolar cells. Retinal tissue from an adult macaque was processed for serial block-face scanning electron microscopy, and a volume of images of the inner retina located 2 mm temporal to the centre of the fovea was analysed. Ten OFF mRGCs and many of their presynaptic cells were reconstructed. Both midget and diffuse types of bipolar cells provided excitatory, glutamatergic input. Axons and long dendrites of wide-field amacrine cells made synapses, and we propose that these mediate tonic, GABAergic inhibition. Narrow-field amacrine cells also made synapses onto the OFF mRGCs, and we propose that most of them are glycinergic, inhibitory synapses. One presynaptic narrow-field amacrine cell was the knotty bistratified type 1 (KB1), which contains immunoreactive glycine and vesicular glutamate transporter 3. We propose that they enlarge the receptive field centers of OFF mRGCs via direct, excitatory synapses. The KB1 cell studied most extensively was presynaptic to some of the same types of amacrine cells that made inhibitory synapses onto OFF mRGCs. We propose that the knotty bistratifed type 1cells release glycine at those synapses and disinhibit responses of OFF mRGCs. KEY POINTS: In primates, OFF midget ganglion cells have the highest spatial density of any projection neurons, and they mediate high acuity vision. Ten of these cells and the neurons providing their inputs were reconstructed from a volume of serial ultrathin sections taken 2 mm temporal to the centre of the macaque fovea. They received the majority of their inputs from amacrine cells, local circuit neurons that are typically inhibitory. One of the presynaptic amacrine cells resembled those containing vesicular glutamate transporter 3, and we propose that they provide excitatory input that enlarges the receptive field centers of OFF midget ganglion cells. They also receive excitatory input from both midget and diffuse bipolar cells. The results provide an explanation for some apparent contradictions between anatomical and physiological studies and are potentially important for understanding the etiology of retinal diseases.

Red-green color vision deficiency (CVD) is the most common single-locus genetic disorder in humans, but our ability to accurately diagnose the subtype and severity of red-green CVD in the clinic is limited. The most widely used diagnostic tools are subject to both Type I and Type II errors, while highly accurate tests are expensive, timeconsuming, and not widely available. In this study, we present an iPhone-based color matching test that uses a combination of optics and software to achieve the diagnostic accuracy of an anomaloscope and the convenience of pseudoisochromatic plates. In validation testing, the device correctly diagnosed 100% of participants, as verified by genetic testing. (c) 2024 Optica Publishing Group. All rights, including for text and data mining (TDM), Artificial Intelligence (AI) training, and similar technologies, are reserved.

We are writing to address errors of misrepresentation in the article "ON and OFF receptive field processing in the presence of optical scattering" [Biomed. Opt. Express14, 2618 (2023)10.1364/BOE.489117]. In their investigation of predictions of "contrast theory" to explain the efficacy of diffusion optics technology (DOT), a myopia control lens design [Br. J. Ophthalmol.107, 1709 (2023)10.1136/bjo-2021-321005], Breher et al. incorrectly indicated that our contrast theory proposed that the association between cone opsin gene splicing defects and myopia was due to differential involvement in ON- and OFF-visual pathways. In addition, the Authors write that we have "hypothesized enhanced ON contrast sensitivity in myopes," but we predict the opposite.

Purpose:Complement dysregulation is a key component in the pathogenesis of age-related macular degeneration (AMD) and related diseases such as early-onset macular drusen (EOMD). Although genetic variants of complement factor H (CFH) are associated with AMD risk, the impact of CFH and factor H-like protein 1 (FHL-1) expression on local complement activity in human retinal pigment epithelium (RPE) remains unclear. Methods:We identified a novel CFH variant in a family with EOMD and generated patient induced pluripotent stem cell (iPSC)-derived RPE cells. We assessed CFH and FHL-1 co-factor activity through C3b breakdown assays and measured complement activation by immunostaining for membrane attack complex (MAC) formation. Expression of CFH, FHL-1, local alternative pathway (AP) components, and regulators of complement activation (RCA) in EOMD RPE cells was determined by quantitative PCR, western blot, and immunostaining. Isogenic EOMD (cEOMD) RPE was generated using CRISPR/Cas9 gene editing. Results:The CFH variant (c.351-2A>G) resulted in loss of CFH and FHL-1 expression and significantly reduced CFH and FHL-1 protein expression (∼50%) in EOMD iPSC RPE cells. These cells exhibited increased MAC deposition upon exposure to normal human serum. Under inflammatory or oxidative stress conditions, CFH and FHL-1 expression in EOMD RPE cells paralleled that of controls, whereas RCA expression, including MAC formation inhibitors, was elevated. CRISPR/Cas9 correction restored CFH/FHL-1 expression and mitigated alternative pathway complement activity in cEOMD RPE cells. Conclusions:Identification of a novel CFH variant in patients with EOMD resulting in reduced CFH and FHL-1 and increased local complement activity in EOMD iPSC RPE supports the involvement of CFH haploinsufficiency in EOMD pathogenesis.

Trichromacy is unique to primates among placental mammals, enabled by blue (short/S), green (medium/M), and red (long/L) cones. In humans, great apes, and Old World monkeys, cones make a poorly understood choice between M and L cone subtype fates. To determine mechanisms specifying M and L cones, we developed an approach to visualize expression of the highly similar M- and L-opsin mRNAs. M-opsin was observed before L-opsin expression during early human eye development, suggesting that M cones are generated before L cones. In adult human tissue, the early-developing central retina contained a mix of M and L cones compared to the late-developing peripheral region, which contained a high proportion of L cones. Retinoic acid (RA)-synthesizing enzymes are highly expressed early in retinal development. High RA signaling early was sufficient to promote M cone fate and suppress L cone fate in retinal organoids. Across a human population sample, natural variation in the ratios of M and L cone subtypes was associated with a noncoding polymorphism in the NR2F2 gene, a mediator of RA signaling. Our data suggest that RA promotes M cone fate early in development to generate the pattern of M and L cones across the human retina.

Considerable progress has been made in studying the receptive fields of the most common primate retinal ganglion cell (RGC) types, such as parasol RGCs. Much less is known about the rarer primate RGC types and the circuitry that gives rise to noncanonical receptive field structures. The goal of this study was to analyze synaptic inputs to smooth monostratified RGCs to determine the origins of their complex spatial receptive fields, which contain isolated regions of high sensitivity called "hotspots." Interestingly, smooth monostratified RGCs co-stratify with the well-studied parasol RGCs and are thus constrained to receiving input from bipolar and amacrine cells with processes sharing the same layer, raising the question of how their functional differences originate. Through 3D reconstructions of circuitry and synapses onto ON smooth monostratified and ON parasol RGCs from central macaque retina, we identified four distinct sampling strategies employed by smooth and parasol RGCs to extract diverse response properties from co-stratifying bipolar and amacrine cells. The two RGC types differed in the proportion of amacrine cell input, relative contributions of co-stratifying bipolar cell types, amount of synaptic input per bipolar cell, and spatial distribution of bipolar cell synapses. Our results indicate that the smooth RGC's complex receptive field structure arises through spatial asymmetries in excitatory bipolar cell input which formed several discrete clusters comparable with physiologically measured hotspots. Taken together, our results demonstrate how the striking differences between ON parasol and ON smooth monostratified RGCs arise from distinct strategies for sampling a common set of synaptic inputs.

There is growing interest in developing artificial lighting that stimulates intrinsically photosensitive retinal ganglion cells (ipRGCs) to entrain circadian rhythms to improve mood, sleep, and health. Efforts have focused on stimulating the intrinsic photopigment, melanopsin; however, specialized color vision circuits have been elucidated in the primate retina that transmit blue-yellow cone-opponent signals to ipRGCs. We designed a light that stimulates color-opponent inputs to ipRGCs by temporally alternating short- and long-wavelength components that strongly modulate short-wavelength sensitive (S) cones. Two-hour exposure to this S-cone modulating light produced an average circadian phase advance of 1 h and 20 min in 6 subjects (mean age = 30 years) compared to no phase advance for the subjects after exposure to a 500 lux white light equated for melanopsin effectiveness. These results are promising for developing artificial lighting that is highly effective in controlling circadian rhythms by invisibly modulating cone-opponent circuits.

Aims To evaluate the myopia control efficacy of Diffusion Optics Technology (DOT) spectacle lenses in children over a 4-year treatment period.Methods CYPRESS Part 1 (NCT03623074) was a 3-year multicentre, randomised, controlled, double-masked trial comparing two investigational spectacle lens DOT designs (Test 1, Test 2) and standard single vision Control lenses in 256 North American children aged 6–10 years. Children completing Part 1 (n=200) were invited to enrol in CYPRESS Part 2 (NCT04947735) for an additional 1-year period. In Part 2, Test 1 (n=35) and Control groups (n=42) continued with their original lens assignment and the Test 2 group (n=21) were crossed over to Test 1 (DOT 0.2) lenses. The co-primary endpoints were change from baseline in axial length (AL) and cycloplegic spherical equivalent refraction (cSER).Results Test 1 spectacle lenses demonstrated superiority to the Control in both co-primary endpoints: with a difference between means (Test 1−Control) of −0.13 mm for AL (p=0.018) and 0.33 D for cSER (p=0.008) in Part 1 and −0.05 mm for AL (p=0.038) and 0.13 D for cSER (p=0.043) in Part 2. Comparing treatment effects in Part 1 and 2 suggests that COVID-19 public health restrictions negatively impacted treatment efficacy in study years 2 and 3.Conclusion DOT 0.2 spectacle lenses are safe and effective at reducing myopia progression, with additional benefit evident in year 4 of wear. These results support the hypothesis that a mild reduction in retinal contrast can slow myopia progression in young children. The unprecedented disruption in participant schooling and lifestyle during the COVID-19 pandemic may have depressed treatment efficacy in Part 1.

Diffusion optics Technology (DOT) myopia control spectacle lenses are based on contrast theory. This innovative theory represents a radical departure from the classical concept of visual deprivation myopia. However, traditional theories have evolved, arriving at remarkably similar solutions for myopia control as the DOT lenses. Nonetheless, contrast theory still represents a departure from mainstream theories. Here, in an effort to resolve discrepancies, we review the science behind contrast theory and compare it to more conventional blur and defocus theories. Finally, we consider the implications of the different theories for the rational design of myopia control solutions.

Microglial cells are the primary resident immune cells in the retina. In healthy adults, they are ramified; that is, they have extensive processes that move continually. In adult retinas, microglia maintain the normal structure and function of neurons and other glial cells, but the mechanism underlying this process is not well-understood. In the mouse hippocampus, microglia engulf small pieces of axons and presynaptic terminals via a process called trogocytosis. Here we report that microglia in the adult macaque retina also engulf pieces of neurons and glial cells, but not at sites of synapses. We analyzed microglia in a volume of serial, ultrathin sections of central macaque retina in which many neurons that ramify in the inner plexiform layer (IPL) had been reconstructed previously. We surveyed the IPL and identified the somas of microglia by their small size and scant cytoplasm. We then reconstructed the microglia and studied their interactions with other cells. We found that ramified microglia frequently ingested small pieces of each major type of inner retinal neuron and Müller glial cells via trogocytosis. There were a few instances where the interactions took place near synapses, but the synapses, themselves, were never engulfed. If trogocytosis by retinal microglia plays a role in synaptic remodeling, it was not apparent from the ultrastructure. Instead, we propose that trogocytosis enables these microglia to present antigens derived from normal inner retinal cells and, when activated, they would promote antigen-specific tolerance.