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Though rod and cone photoreceptors use similar phototransduction mechanisms, previous model calculations have indicated that the most important differences in their light responses are likely to be differences in amplification of the G-protein cascade, different decay rates of phosphodiesterase (PDE) and pigment phosphorylation, and different rates of turnover of cGMP in darkness. To test this hypothesis, we constructed TrUx;GapOx rods by crossing mice with decreased transduction gain from decreased transducin expression, with mice displaying an increased rate of PDE decay from increased expression of GTPase-activating proteins (GAPs). These two manipulations brought the sensitivity of TrUx;GapOx rods to within a factor of 2 of WT cone sensitivity, after correcting for outer-segment dimensions. These alterations did not, however, change photoreceptor adaptation: rods continued to show increment saturation though at a higher background intensity. These experiments confirm model calculations that rod responses can mimic some (though not all) of the features of cone responses after only a few changes in the properties of transduction proteins.

PurposeTo investigate if accounting for a cup-to-disc ratio (CDR) genetic risk score (GRS) modified the association between large CDR and cognitive function among women.DesignThis was a retrospective study using data from the Women's Health Initiative.MethodsPatients with glaucoma or ocular hypertension were excluded. Large CDR was defined as ≥ 0.6 in either eye. Cognitive function was measured by the Modified Mini-Mental State Examination (3MSE). We used the combined effects from 13 single nucleotide polymorphisms (SNPs) to formulate the GRS for CDR. We used logistic regression to investigate associations between weighted GRS and large CDR, then a linear regression to assess the association between weighted GRS and 3MSE scores, and between weighted GRS, CDR, and 3MSE scores, adjusted for demographic and clinical characteristics.ResultsFinal analyses included 1,196 White women with mean age of 69.60 ± 3.62 years and 7.27% with large CDR. Mean GRS in women with and without large CDR was 1.51 ± 0.31 vs. 1.41 ± 0.36, respectively (p = 0.004). The odds of large CDR for a one unit increase in GRS was 2.30 (95% CI: (1.22, 4.36), p = 0.011). Adding the CDR GRS in the model with CDR and 3MSE, women with large CDR still had statistically significantly lower 3MSE scores than those without large CDR, yielding a predicted mean difference in 3MSE scores of 0.84 (p = 0.007).ConclusionsIndependent of the CDR GRS, women with large CDR had a lower cognitive function.

PurposeInadequacies in healthcare access and utilization substantially impact outcomes for diabetic patients. The All of Us database offers extensive survey data pertaining to social determinants that is not routinely available in electronic health records. This study assesses whether social determinants were associated with an increased risk of developing proliferative diabetic retinopathy or related complications (e.g. related diagnoses or procedures).MethodsWe identified 729 adult participants in the National Institutes of Health All of Us Research Program data repository with diabetic retinopathy (DR) who answered survey questions pertaining to healthcare access and utilization. Electronic health record data regarding co-morbidities, laboratory values, and procedures were extracted. Multivariable logistic regression with bi-directional stepwise variable selection was performed from a wide range of predictors. Statistical significance was defined as p

BackgroundAcanthamoeba keratitis is challenging to treat and thought to result in poor outcomes, but very few comparative studies exist to assess whether ulcers caused by Acanthamoeba are worse than those caused by bacteria or fungus.MethodsIn a retrospective cohort study, all cases of smear- or culture-proven Acanthamoeba keratitis diagnosed from January 2006 to June 2011 at an eye hospital in South India were identified from the microbiology database. Random samples of the same number of cases of bacterial and fungal keratitis, matched by year, were identified from the same database in order to compare outcomes between the three types of organism. The main outcomes were the time until the following events: re-epithelialization, discontinuation of antimicrobials, perforation/keratoplasty, elevated intraocular pressure, and new cataract.ResultsThe median time until re-epithelialization was 113 days for Acanthamoeba keratitis, 30 days for fungal keratitis, and 25 days for bacterial keratitis, and the median time until discontinuation of antimicrobial therapy was 100 days for Acanthamoeba keratitis, 49 days for fungal keratitis, and 40 days for bacterial keratitis. Compared to the other two organisms, Acanthamoeba ulcers took significantly longer to re-epithelialize (adjusted HR 0.4, 95% CI 0.3 to 0.6 relative to bacterial ulcers and HR 0.3, 95% CI 0.2 to 0.5 relative to fungal ulcers; overall p

PurposeThe phosphodiesterase inhibitor sildenafil is a promising treatment for neurodegenerative disease, but it can cause oxidative stress in photoreceptors ex vivo and degrade visual performance in humans. Here, we test the hypotheses that in wildtype mice sildenafil causes i) wide-spread photoreceptor oxidative stress in vivo that is linked with ii) impaired vision.MethodsIn dark or light-adapted C57BL/6 mice ± sildenafil treatment, the presence of oxidative stress was evaluated in retina laminae in vivo by QUEnch-assiSTed (QUEST) magnetic resonance imaging, in the subretinal space in vivo by QUEST optical coherence tomography, and in freshly excised retina by a dichlorofluorescein assay. Visual performance indices were also evaluated by QUEST optokinetic tracking.ResultsIn light-adapted mice, 1 hr post-sildenafil administration, oxidative stress was most evident in the superior peripheral outer retina on both in vivo and ex vivo examinations; little evidence was noted for central retina oxidative stress in vivo and ex vivo. In dark-adapted mice 1 hr after sildenafil, no evidence for outer retina oxidative stress was found in vivo. Evidence for sildenafil-induced central retina rod cGMP accumulation was suggested as a panretinally thinner, dark-like subretinal space thickness in light-adapted mice at 1 hr but not 5 hr post-sildenafil. Cone-based visual performance was impaired by 5 hr post-sildenafil and not corrected with anti-oxidants; vision was normal at 1 hr and 24 hr post-sildenafil.ConclusionsThe sildenafil-induced spatiotemporal pattern of oxidative stress in photoreceptors dominated by rods was unrelated to impairment of cone-based visual performance in wildtype mice.

PurposeAutorefractors allow non-specialists to quickly assess refractive error, and thus could be a useful component of large-scale vision screening programs. In order to better characterize the role of autorefraction for public health outreach programs in resource-limited settings, the diagnostic accuracy of two autorefractors was assessed relative to subjective refraction in an adult Indian population.MethodsAn optometrist refracted a series of patients aged ≥50 years at an eye clinic in Bangalore, India using the Nidek ARK-900 autorefractor first, followed by the 3nethra Royal autorefractor, and then subjective refraction. The diagnostic accuracy of each autorefractor for myopia, hyperopia, and astigmatism was assessed using subjective refraction as the reference standard, and measures of agreement between refractions were calculated.ResultsA total of 197 eyes in 104 individuals (mean age 63 ± 8 years, 52% female) were evaluated. Both autorefractors produced spherical equivalent estimates that were on average more hyperopic than subjective refraction, with a measurement bias of +0.16 D (95%CI +0.09 to +0.23D) for Nidek and +0.42 D (95%CI +0.28 to +0.54D) for 3nethra. When comparing pairs of measurements from autorefraction and subjective refraction, the limits of agreement were approximately ±1D for the Nidek autorefractor and ±1.75D for the 3Nethra autorefractor. The sensitivity and specificity of detecting ≥1 diopter of myopia were 94.6% (95%CI 86.8-100%) and 92.5% (95%CI 88.9-97.5%) for the Nidek, and 89.2% (95%CI 66.7-97.4) and 77.5% (95%CI 71.2-99.4%) for the 3Nethra. The accuracy of each autorefractor increased at greater levels of refractive error.ConclusionsThe sensitivity and specificity of the Nidek autorefractor for diagnosing refractive error among adults ≥50 years in an urban Indian clinic was sufficient for screening for visually significant refractive errors, although the relatively wide limits of agreement suggest that subjective refinement of the eyeglasses prescription would still be necessary.

PurposeTo investigate any associations between cigarette smoking and retinal microvascular changes in diabetic patients without visible retinopathy.DesignRetrospective, cross-sectional study.Participants1099 eyes from 1099 diabetic patients with no clinical evidence of diabetic retinopathy (DR) were included in this study.MethodsDiabetic patients underwent optical coherence tomography angiography (OCTA) scanning at Zuckerberg San Francisco General Hospital and Trauma Center between April 2018 and September 2019. Patient demographic and clinical information was collected. Standard bivariate statistics and multivariate linear regression were performed.Main outcome measuresOCTA parameters included metrics related to the foveal avascular zone (FAZ; area, perimeter, circularity), perfusion density (PD; full, center, inner), and vessel length density (VLD; full, center, inner).ResultsThe study population included 750 non-smokers and 349 smokers. FAZ perimeter was the only OCTA parameter that was significantly different between the two groups on uncontrolled analysis (P = 0.033). Multivariate regression analyses revealed significant associations between lower VLD full (β = -0.31, P = 0.048), lower VLD inner (β = -0.35, P = 0.046) and a history of smoking. No significant associations between cigarette smoking and either FAZ or PD were detected.ConclusionsOur results suggest that smoking is likely associated with deleterious changes in the retinal microvasculature of patients with a history of diabetes and no visible DR. Based on these findings, diabetic patients with a history of smoking may benefit from higher prioritization in terms of ophthalmic screening.

PurposeTo examine the relationship between pre-corneal and pre-contact lens tear film stability (TFS), and to determine whether pre-corneal TFS is a reliable predictor of subsequent pre-lens TFS after a contact lens is placed on the eye.Methods667 records met inclusion criteria and were extracted from a soft contact lens multi-study database. Multivariable linear mixed effects models were fit to examine the association between pre-corneal and pre-lens TFS, adjusting for potential confounders and accounting for repeated measures. Receiver Operating Characteristic (ROC) analysis was employed to assess the predictive performance of pre-corneal TFS for subsequent pre-lens TFS. TFS was quantified for this analysis as the non-invasive tear breakup time (NITBUT).ResultsPre-corneal NITBUT was significantly related to the pre-lens NITBUT at both 10 min (p

PurposeThe purpose of this study was to examine whether racial and ethnic differences in retinal microvasculature are detectable with quantitative measures derived from optical coherence tomography angiography (OCTA).MethodsOCTA scans and fundus photography were obtained in 447 eyes from 271 patients with and without diabetes between April and October 2018. Fundus photos were graded by the hospital reading center for diabetic retinopathy (DR) severity. Eight OCTA parameters relating to the foveal avascular zone (FAZ), superficial vascular perfusion, and deep vascular perfusion were analyzed for significant differences between race and ethnicity groups, self-reported by patients and organized according to National Center for Health Statistics groupings. Multiple regression was then used to adjust estimates for possible confounding by age, gender, hypertension, and last hemoglobin A1c level.ResultsSignificant differences in FAZ area were found between white and non-white patients. After adjustment, the differences between white and all non-white groups were statistically significant (p

PurposeMitochondrial (mt) DNA damage is associated with age-related macular degeneration (AMD) and other human aging diseases. This study was designed to quantify and characterize mtDNA low-frequency heteroplasmy single nucleotide polymorphisms (SNPs) of three different tissues isolated from AMD subjects using Next Generation Sequencing (NGS) technology.MethodsDNA was extracted from neural retina, [RPE+choroid] and blood from three deceased age-related macular degeneration (AMD) subjects. Entire mitochondrial genomes were analyzed for low-frequency heteroplasmy SNPs using NGS technology that independently sequenced both mtDNA strands. This deep sequencing method (average sequencing depth of 30,000; range 1,000-100,000) can accurately differentiate low-frequency heteroplasmy SNPs from DNA modification artifacts. Twenty-three 'hot-spot' heteroplasmy mtDNA SNPs were analyzed in 222 additional blood samples.ResultsGermline homoplasmy SNPs that defined mtDNA haplogroups were consistent in the three tissues of each subject. Analyses of SNPs with T, m.1284T>C, m.1556C>T, m.7256C>T) were found in additional samples (n = 222). Five heteroplasmy SNPs (m.4104A>G, m.5320C>T, m.5471G>A, m.5474A>G, m.5498A>G) declined with age. Two heteroplasmy SNPs (m.13095T>C, m.13105A>G) increased in AMD compared to Normal samples. In the heteroplasmy SNPs, very few transversion mutations (purine to pyrimidine or vice versa, associated with oxidative damage) were found and the majority were transition changes (purine to purine or pyrimidine to pyrimidine, associated with replication errors).ConclusionWithin an individual, the blood, retina and [RPE+choroid] contained identical homoplasmy SNPs representing inherited germline mtDNA haplogroup. NGS methodology showed significantly more mtDNA heteroplasmy SNPs in blood compared to retina and [RPE+choroid], suggesting the latter tissues have substantial protection. Significantly higher heteroplasmy levels of m.13095T>C and m.13105A>G may represent potential AMD biomarkers. Finally, high levels of transition mutations suggest that accumulation of heteroplasmic SNPs may occur through replication errors rather than oxidative damage.

PurposeFemale mice have been found to be resistant to streptozotocin (STZ)-induced diabetes, and pre-clinical research related to diabetic complications commonly omits females. The purpose of this study was to develop a method to induce diabetes in female mice, and to determine if retinas of diabetic female mice develop molecular changes and histopathological abnormalities comparable to those which develop in male diabetic mice.MethodsTo induce diabetes, animals of both sexes received daily intraperitoneal (i.p.) injection of STZ for 5 consecutive days at 55 mg/kg BW (a dose that is known to induce diabetes in male mice) or for females, 75 mg/kg BW of STZ. Retinal abnormalities that have been implicated in the development of the retinopathy (superoxide generation and expression of inflammatory proteins, iNOS and ICAM-1) were evaluated at 2 months of diabetes, and retinal capillary degeneration was evaluated at 8 months of diabetes.ResultsDaily i.p. injection of STZ for 5 consecutive days at a concentration of 55 mg/kg BW was sufficient to induce diabetes in 100% of male mice, but only 33% of female mice. However, females did become hyperglycemic when the dose of STZ administered was increased to 75 mg/kg BW. The resulting STZ-induced hyperglycemia in female and male mice was sustained for at least 8 months. After induction of the diabetes, both sexes responded similarly with respect to the oxidative stress, expression of iNOS, and degeneration of retinal capillaries, but differed in the limited population evaluated with respect to expression of ICAM-1.ConclusionsThe resistance of female mice to STZ-induced diabetes can be overcome by increasing the dose of STZ used. Female mice can, and should, be included in pre-clinical studies of diabetes and its complications.

We can retain only a portion of the visual information that we encounter within our visual working memory. Which factors influence how much information we can remember? Recent studies have demonstrated that the capacity of visual working memory is influenced by the type of information to be remembered and is greater for real-world objects than for abstract stimuli. One explanation for this effect is that the semantic knowledge associated with real-world objects makes them easier to maintain in working memory. Previous studies have indirectly tested this proposal and led to inconsistent conclusions. Here, we directly tested whether semantic knowledge confers a benefit for visual working memory by using familiar and unfamiliar real-world objects. We found a mnemonic benefit for familiar objects in adults and children between the ages of 4 and 9 years. Control conditions ruled out alternative explanations, namely the possibility that the familiar objects could be more easily labeled or that there were differences in low-level visual features between the two types of objects. Together, these findings demonstrate that semantic knowledge influences visual working memory, which suggests that the capacity of visual working memory is not fixed but instead fluctuates depending on what has to be remembered.

No single neuroimaging technique or sequence is capable of reflecting the functional deficits manifest in MS. Given the interest in imaging biomarkers for short- to medium-term studies, we aimed to assess which imaging metrics might best represent functional impairment for monitoring in clinical trials. Given the complexity of functional impairment in MS, however, it is useful to isolate a particular functionally relevant pathway to understand the relationship between imaging and neurological function. We therefore analyzed existing data, combining multiparametric MRI and OCT to describe MS associated visual impairment. We assessed baseline data from fifty MS patients enrolled in ReBUILD, a prospective trial assessing the effect of a remyelinating drug (clemastine). Subjects underwent 3T MRI imaging, including Neurite Orientation Dispersion and Density Imaging (NODDI), myelin content quantification, and retinal imaging, using OCT. Visual function was assessed, using low-contrast letter acuity. MRI and OCT data were studied to model visual function in MS, using a partial, least-squares, regression analysis. Measures of neurodegeneration along the entire visual pathway, described most of the observed variance in visual disability, measured by low contrast letter acuity. In those patients with an identified history of ON, however, putative myelin measures also showed correlation with visual performance. In the absence of clinically identifiable inflammatory episodes, residual disability correlates with neurodegeneration, whereas after an identifiable exacerbation, putative measures of myelin content are additionally informative.

The healthy cornea is remarkably resistant to infection, quickly clearing deliberately inoculated bacteria such as Pseudomonas aeruginosa and Staphylococcus aureus. Contrasting with the adjacent conjunctiva and other body surfaces, it also lacks a resident viable bacterial microbiome. Corneal resistance to microbes depends on intrinsic defenses involving tear fluid and the corneal epithelium. Dry eye, an ocular surface disease associated with discomfort and inflammation, can alter tear fluid composition and volume, and impact epithelial integrity. We previously showed that experimentally-induced dry eye (EDE) in mice does not increase corneal susceptibility to P. aeruginosa infection. Here, we explored if EDE alters corneal resistance to bacterial colonization. EDE was established in mice using scopolamine injections and dehumidified air-flow, and verified by phenol-red thread testing after 5 and 10 days. As expected, EDE corneas showed increased fluorescein staining versus controls consistent with compromised epithelial barrier function. Confocal imaging using mT/mG knock-in mice with red-fluorescent membranes revealed no other obvious morphological differences between EDE corneas and controls for epithelium, stroma, and endothelium. EDE corneas were imaged ex vivo and compared to controls after alkyne-functionalized D-alanine labeling of metabolically-active colonizing bacteria, or by FISH using a universal 16S rRNA gene probe. Both methods revealed very few viable bacteria on EDE corneas after 5 or 10 days (median of 0, upper quartile of ≤ 1 bacteria per field of view for each group [9-12 eyes per group]) similar to control corneas. Furthermore, there was no obvious difference in abundance of conjunctival bacteria, which included previously reported filamentous forms. Thus, despite reduced tear flow and apparent compromise to corneal barrier function (fluorescein staining), EDE murine corneas continue to resist bacterial colonization and maintain the absence of a resident viable bacterial microbiome.

PurposeTo compare quantitative changes in macular parameters in diabetic patients detected by two optical coherence tomography angiography (OCTA) instruments.Methods80 phakic eyes were classified as no diabetes, diabetes without diabetic retinopathy (DR), mild non-proliferative diabetic retinopathy (NPDR), and severe NPDR or proliferative DR (PDR). OCTA was performed using devices from two manufacturers (Zeiss and Heidelberg). Superficial and deeper vascular skeleton density (SVSD, DVSD), superficial and deeper vessel area density (SVAD, DVAD), choriocapillaris flow voids (CCFV), and choroidal flow voids (CFV) were calculated. Inter-device comparisons were performed using the size comparison index (SCI) and the discrepancy index (DI).ResultsThe two devices were inconsistent in SVSD, DVSD, DVAD, CCFV and CFV parameters (all P < 0.05). In addition, the SCI was positive for DVAD (all P < 0.001) and negative for SVSD, DVSD, CCFV and CFV in all groups (all P 0.05). The mean DI of CCFV was statistically different between the four groups (P < 0.001).ConclusionsThe two instruments were largely inconsistent in the measurement of macular parameters relevant to DR. The choice of imaging device can impact OCTA analytics and should be taken into account when drawing conclusions about DR-related changes.

PURPOSE:To evaluate the surgical technique for subretinal implantation of two sizes of PRIMA photovoltaic wireless microchip in two animal models, and refine these surgical procedures for human trials. METHODS:Cats and Macaca fascicularis primates with healthy retina underwent vitrectomy surgery and were implanted with subretinal wireless photovoltaic microchip at the macula/central retina. The 1.5mm PRIMA chip was initially studied in feline eyes. PRIMA implant (2mm,1.5mm sizes) arrays were studied in primates. Feasibility of subretinal chip implantation was evaluated with a newly-developed surgical technique, with surgical complications and adverse events recorded. RESULTS:The 1.5mm implant was placed in the central retina of 11 feline eyes, with implantation duration 43-106 days. The 1.5mm implant was correctly positioned into central macula of 11 primate eyes, with follow-up periods of minimum 6 weeks (n = 11), 2 years (n = 2), and one eye for 3 years. One primate eye underwent multi-chip 1.5mm implantation using two 1.5mm chips. The 2mm implant was delivered to 4 primate eyes. Optical coherence tomography confirmed correct surgical placement of photovoltaic arrays in the subretinal space in all 26 eyes. Intraoperative complications in primate eyes included retinal tear, macular hole, retinal detachment, and vitreous hemorrhage that resolved spontaneously. Postoperatively, there was no case of significant ocular inflammation in the 1.5mm implant group. CONCLUSIONS:We report subretinal implantation of 1.5mm and 2mm photovoltaic arrays in the central retina of feline and central macula of primate eyes with a low rate of device-related complications. The in vivo PRIMA implantation technique has been developed and refined for use for a 2mm PRIMA implant in ongoing human trials.

Mutations in the gene Centrosomal Protein 290 kDa (CEP290) result in multiple ciliopathies ranging from the neonatal lethal disorder Meckel-Gruber Syndrome to multi-systemic disorders such as Joubert Syndrome and Bardet-Biedl Syndrome to nonsyndromic diseases like Leber Congenital Amaurosis (LCA) and retinitis pigmentosa. Results from model organisms and human genetics studies, have suggest that mutations in genes encoding protein components of the transition zone (TZ) and other cilia-associated proteins can function as genetic modifiers and be a source for CEP290 pleiotropy. We investigated the zebrafish cep290fh297/fh297 mutant, which encodes a nonsense mutation (p.Q1217*). This mutant is viable as adults, exhibits scoliosis, and undergoes a slow, progressive cone degeneration. The cep290fh297/fh297 mutants showed partial mislocalization of the transmembrane protein rhodopsin but not of the prenylated proteins rhodopsin kinase (GRK1) or the rod transducin subunit GNB1. Surprisingly, photoreceptor degeneration did not trigger proliferation of Müller glia, but proliferation of rod progenitors in the outer nuclear layer was significantly increased. To determine if heterozygous mutations in other cilia genes could exacerbate retinal degeneration, we bred cep290fh297/fh297 mutants to arl13b, ahi1, and cc2d2a mutant zebrafish lines. While cep290fh297/fh297 mutants lacking a single allele of these genes did not exhibit accelerated photoreceptor degeneration, loss of one alleles of arl13b or ahi1 reduced visual performance in optokinetic response assays at 5 days post fertilization. Our results indicate that the cep290fh297/fh297 mutant is a useful model to study the role of genetic modifiers on photoreceptor degeneration in zebrafish and to explore how progressive photoreceptor degeneration influences regeneration in adult zebrafish.

The human retina contains three classes of cone photoreceptors each sensitive to different portions of the visual spectrum: long (L), medium (M) and short (S) wavelengths. Color information is computed by downstream neurons that compare relative activity across the three cone types. How cone signals are combined at a cellular scale has been more difficult to resolve. This is especially true near the fovea, where spectrally-opponent neurons in the parvocellular pathway draw excitatory input from a single cone and thus even the smallest stimulus projected through natural optics will engage multiple color-signaling neurons. We used an adaptive optics microstimulator to target individual and pairs of cones with light. Consistent with prior work, we found that color percepts elicited from individual cones were predicted by their spectral sensitivity, although there was considerable variability even between cones within the same spectral class. The appearance of spots targeted at two cones were predicted by an average of their individual activations. However, two cones of the same subclass elicited percepts that were systematically more saturated than predicted by an average. Together, these observations suggest both spectral opponency and prior experience influence the appearance of small spots.

People with schizophrenia (SZ) or bipolar disorder (BD) experience dysfunction in visual processing. Dysfunctional neural tuning, in which neurons and neuronal populations are selectively activated by specific features of visual stimuli, may contribute to these deficits. Few studies have examined this possibility and there are inconsistent findings of tuning deficits in the literature. We utilized an event-related potential (ERP) paradigm to examine neural adaptation for visual objects, a measure of neural tuning whereby neurons respond less strongly to the repeated presentation of the same stimulus. Seventy-seven SZ, 53 BD, and 49 healthy comparison participants (HC) were examined. In three separate conditions, pictures of objects were presented repeatedly: the same object (SS), different objects from the same category (e.g., two different vases; SD), or different objects from different categories (e.g., a barrel and a clock, DD). Mass-univariate cluster-based permutation analyses identified electrodes and time-windows in which there were significant differences between the SS vs. DD and the SD vs. DD conditions. Mean ERP amplitudes were extracted from these clusters and analyzed for group differences. Results revealed a significant condition difference over parieto-occipital electrodes for the SS-DD comparison between 109-164 ms and for the SD-DD comparison between 78-203 ms, with larger amplitudes in the DD compared to either SS or SD condition. However, there were no significant differences in the pattern of results between groups. Thus, while we found neural adaptation effects using this ERP paradigm, we did not find evidence of group differences. Our results suggest that people with SZ or BD may not exhibit deficits in neural tuning for processing of visual objects using this EEG task with rapidly presented stimuli. However, the results are inconsistent with other studies using different methodologies (e.g., fMRI, behavioral tasks) that have found tuning deficits in people with schizophrenia.

Background/aimsTrachoma programs base treatment decisions on the community prevalence of the clinical signs of trachoma, assessed by direct examination of the conjunctiva. Automated assessment could be more standardized and more cost-effective. We tested the hypothesis that an automated algorithm could classify eyelid photographs better than chance.MethodsA total of 1,656 field-collected conjunctival images were obtained from clinical trial participants in Niger and Ethiopia. Images were scored for trachomatous inflammation-follicular (TF) and trachomatous inflammation-intense (TI) according to the simplified World Health Organization grading system by expert raters. We developed an automated procedure for image enhancement followed by application of a convolutional neural net classifier for TF and separately for TI. One hundred images were selected for testing TF and TI, and these images were not used for training.ResultsThe agreement score for TF and TI tasks for the automated algorithm relative to expert graders was κ = 0.44 (95% CI: 0.26 to 0.62, P < 0.001) and κ = 0.69 (95% CI: 0.55 to 0.84, P < 0.001), respectively.DiscussionFor assessing the clinical signs of trachoma, a convolutional neural net performed well above chance when tested against expert consensus. Further improvements in specificity may render this method suitable for field use.

BackgroundNumerous preclinical studies have supported the theory that enhanced activation of mGluR5 signaling, due to the absence or reduction of the FMR1 protein, contributes to cognitive and behavioral deficits in patients with fragile X syndrome (FXS). However multiple phase 2 controlled trials in patients with FXS have failed to demonstrate efficacy of compounds that negatively modulate mGluR5, including two phase 2b randomized controlled trials (RCT) of mavoglurant (AFQ056, Novartis Pharma AG), when the primary measures of interest were behavioral ratings. This has cast some doubt onto the translation of the mGluR5 theory from animal models to humans with the disorder.MethodsWe evaluated social gaze behavior-a key phenotypic feature of the disorder-and sympathetic nervous system influence on pupil size using a previously-validated eye tracking paradigm as a biobehavioral probe, in 57 adolescent or adult patients with FXS at baseline and following three months of blinded treatment with one of three doses of mavoglurant or placebo, within the context of the AFQ056 RCTs.ResultsPatients with FXS treated with mavoglurant demonstrated increased total absolute looking time and number of fixations to the eye region while viewing human faces relative to baseline, and compared to those treated with placebo. In addition, patients had greater pupil reactivity to faces relative to baseline following mavoglurant treatment compared to placebo.DiscussionThe study shows that negative modulation of mGluR5 activity improves eye gaze behavior and alters sympathetically-driven reactivity to faces in patients with FXS, providing preliminary evidence of this drug's impact on behavior in humans with the disorder.

Unihemispheric slow wave sleep (USWS) is a unique form of sleep in which one brain hemisphere maintains low voltage electrical activity indicative of waking while the opposite exhibits slow wave electrical activity indicative of sleep. USWS is present in several marine mammals and in some species of birds. One proposed biological function of USWS is to enable the animal to monitor the environment to detect predators or conspecifics. While asymmetrical eye state was often observed during behavioral sleep in birds and marine mammals, electrophysiological (electroencephalogram, EEG) correlates between the asymmetry of eye state and EEG of two cortical hemispheres have not been reliably established. This study examined the association between eye state and EEG activity during aquatic sleep in two subadult northern fur seals (Callorhinus ursinus), taking advantage of the simultaneous visibility of both eyes when the seals were in the prone position. We found that during USWS the eye contralateral to the sleeping hemisphere was closed on average 99.4±0.1% of the recording time. The eye contralateral to the waking hemisphere opened briefly for on average 1.9±0.1 sec with a rate of 8.2±1.0 per min. This eye was open on average 24.8±2.5% of the USWS time and it was closed no longer than 3 sec, on average 39.4±5.6% of the time. These data indicate that fur seals sleep in seawater by having intermittent visual monitoring. Our findings document the extent of visual monitoring of both eyes during USWS and support the idea that USWS allows intermittent visual vigilance. Thus, USWS serves two functions in the fur seal, facilitating movement and visual vigilance, which may also be the case in cetaceans.

People typically rely heavily on visual information when finding their way to unfamiliar locations. For individuals with reduced vision, there are a variety of navigational tools available to assist with this task if needed. However, for wayfinding in unfamiliar indoor environments the applicability of existing tools is limited. One potential approach to assist with this task is to enhance visual information about the location and content of existing signage in the environment. With this aim, we developed a prototype software application, which runs on a consumer head-mounted augmented reality (AR) device, to assist visually impaired users with sign-reading. The sign-reading assistant identifies real-world text (e.g., signs and room numbers) on command, highlights the text location, converts it to high contrast AR lettering, and optionally reads the content aloud via text-to-speech. We assessed the usability of this application in a behavioral experiment. Participants with simulated visual impairment were asked to locate a particular office within a hallway, either with or without AR assistance (referred to as the AR group and control group, respectively). Subjective assessments indicated that participants in the AR group found the application helpful for this task, and an analysis of walking paths indicated that these participants took more direct routes compared to the control group. However, participants in the AR group also walked more slowly and took more time to complete the task than the control group. The results point to several specific future goals for usability and system performance in AR-based assistive tools.

The retinal rod pathway, featuring dedicated rod bipolar cells (RBCs) and AII amacrine cells, has been intensely studied in placental mammals. Here, we analyzed the rod pathway in a nocturnal marsupial, the South American opossum Monodelphis domestica to elucidate whether marsupials have a similar rod pathway. The retina was dominated by rods with densities of 338,000-413,000/mm². Immunohistochemistry for the RBC-specific marker protein kinase Cα (PKCα) and the AII cell marker calretinin revealed the presence of both cell types with their typical morphology. This is the first demonstration of RBCs in a marsupial and of the integration of RBCs and AII cells in the rod signaling pathway. Electron microscopy showed invaginating synaptic contacts of the PKCα-immunoreactive bipolar cells with rods; light microscopic co-immunolabeling for the synaptic ribbon marker CtBP2 confirmed dominant rod contacts. The RBC axon terminals were mostly located in the innermost stratum S5 of the inner plexiform layer (IPL), but had additional side branches and synaptic varicosities in strata S3 and S4, with S3-S5 belonging to the presumed functional ON sublayer of the IPL, as shown by immunolabeling for the ON bipolar cell marker Gγ13. Triple-immunolabeling for PKCα, calretinin and CtBP2 demonstrated RBC synapses onto AII cells. These features conform to the pattern seen in placental mammals, indicating a basically similar rod pathway in M. domestica. The density range of RBCs was 9,900-16,600/mm2, that of AII cells was 1,500-3,260/mm2. The numerical convergence (density ratio) of 146-156 rods to 4.7-6.0 RBCs to 1 AII cell is within the broad range found among placental mammals. For comparison, we collected data for the Australian nocturnal dunnart Sminthopsis crassicaudata, and found it to be similar to M. domestica, with rod-contacting PKCα-immunoreactive bipolar cells that had axon terminals also stratifying in IPL strata S3-S5.

In retinal raster imaging modalities, fixational eye movements manifest as image warp, where the relative positions of the beam and retina change during the acquisition of single frames. To remove warp artifacts, strip-based registration methods-in which fast-axis strips from target images are registered to a reference frame-have been applied in adaptive optics (AO) scanning light ophthalmoscopy (SLO) and optical coherence tomography (OCT). This approach has enabled object tracking and frame averaging, and methods have been described to automatically select reference frames with minimal motion. However, inconspicuous motion artifacts may persist in reference frames and propagate themselves throughout the processes of registration, tracking, and averaging. Here we test a previously proposed method for removing movement artifacts in reference frames, using biases in stripwise cross-correlation statistics. We applied the method to synthetic retinal images with simulated eye motion artifacts as well as real AO-SLO images of the cone mosaic and volumetric AO-OCT images, both affected by eye motion. In the case of synthetic images, the method was validated by direct comparison with motion-free versions of the images. In the case of real AO images, performance was validated by comparing the correlation of uncorrected images with that of corrected images, by quantifying the effect of motion artifacts on the image power spectra, and by qualitative examination of AO-OCT B-scans and en face projections. In all cases, the proposed method reduced motion artifacts and produced more faithful images of the retina.

PurposeTo evaluate microvasculature dropout in the optic disc (Mvd-D) using optical coherence tomography angiography (OCTA) and investigate factors associated with Mvd-D in primary open-angle glaucoma (POAG) eyes.MethodsOne hundred twenty-three eyes of 123 POAG patients were included from the Diagnostic Innovations in Glaucoma Study. The 3.0×3.0-mm optic nerve head OCTA scans were acquired using a spectral-domain OCT instrument. Images with whole-signal-mode were evaluated. Eyes were classified into 3 categories (Mvd-D, pseudo-Mvd-D, and no Mvd-D). Mvd-D and pseudo-Mvd-D had complete loss of OCTA signals on the temporal side of the optic disc on the en face projection image. They were distinguished base on the visualization of the anterior lamina cribrosa in the horizontal B-scans of that area. No Mvd-D was defined when no complete signal loss of OCTA signals was observed. Covariates including focal lamina cribrosa defects assessed by swept-source OCT and microvasculature dropout in the parapapillary region (Mvd-P) were analyzed.ResultsForty-two, 37, and 44 eyes were identified as having Mvd-D, pseudo-Mvd-D, and no Mvd-D, respectively. The eyes with Mvd-D showed significantly lower intraocular pressure, worse visual field mean deviation, larger cup-to-disc ratio, thinner circumpapillary retinal nerve fiber layer (cpRNFL), and lower circumpapillary vessel density within the RNFL than the eyes with pseudo-Mvd-D or the eyes without Mvd-D. Multivariable logistic regression analysis showed significant associations of Mvd-D with larger cup-to-disc ratio (OR, 1.08; P = 0.001), worse visual field mean deviation (OR, 1.09; P = 0.048), higher prevalence of focal lamina cribrosa defect (OR, 9.05; P = 0.002), and higher prevalence of Mvd-P (OR, 10.33; P

Extracellular vesicles (EVs) released by virtually every cell of all organisms are involved in processes of intercellular communication through the delivery of their functional mRNAs, proteins and bioactive lipids. We previously demonstrated that mouse embryonic stem cell-released EVs (mESEVs) are able to transfer their content to different target retinal cells, inducing morphological and biochemical changes in them. The main objective of this paper is to characterize EVs derived from human embryonic stem cells (hESEVs) and investigate the effects that they have on cultured retinal glial, progenitor Müller cells, which are known to give rise to retinal neurons under specific conditions. This would allow us to establish if hESEVs have a pro-regenerative potential not yet described that could be used in the future for treatment of human retinal degenerative diseases. Initially, we showed that hESEVs are heterogeneous in size, contain mRNAs and proteins involved in the induction and maintenance of stem cell pluripotency and can be internalized by cultured Müller cells. After a single exposure to hESEVs these cells display changes in their gene expression profile, and with multiple exposures they de-differentiate and trans-differentiate into retinal neuronal precursors. hESEVs were then fractionated into microvesicles (MVs) and exosomes (EXOs), which were characterized by size, specific surface proteins and biochemical/molecular components. We demonstrate that despite the similar internalization of non-fractionated hESEVs, MVs and EXOs by Müller progenitor cells, in vitro, only the release of MVs' cargo into the cells' cytoplasm induces specific changes in their levels of pluripotency mRNAs and early retinal proteins. EXOs do not produce any detectable effect. Thus, we conclude that MVs and MVs-containing hESEVs are promising agents that possibly could promote the regeneration of diseased or damaged retinas in vivo through inducing glial Müller cells to become replacement neurons.

PurposeIn this article, we introduce a novel flow chart-based screening tool for the categorization of contact lens-induced dryness (CLIDE) and its impact on daily visual activities: the Berkeley Dry Eye Flow Chart (DEFC).MethodsOne hundred thirty (130) experienced soft contact lens wearers discontinued lens wear for 24 hrs, passed a baseline screening and eye health examination, completed the Ocular Surface Disease Index (OSDI) then were dispensed fresh pairs of their habitual lenses. After 6 hrs of wear, subjects were administered a battery of symptom questionnaires, and underwent non-invasive tear breakup time (NITBUT) measurement, grading of distortion in reflected topographer mires, grading of lens surface wettability, and a fluorescein examination of the ocular surface. Subjects returned after at least 48 hrs and repeated all assessments after 6 hrs of wear of a second fresh pair of habitual lenses.ResultsThe repeatability of the DEFC between visits was within 1%, and Limits of Agreement and Coefficient of Repeatability were comparable to those of the other CLIDE assessments. Higher DEFC score was significantly related to shorter pre-lens NITBUT, higher OSDI score, and higher Visual Analog Scale (VAS) ratings of average and end-of-day severity and frequency of dryness (all p < 0.001). For CLIDE as diagnosed based on DEFC score, the highest sensitivities and specificities were achieved by the OSDI and VAS ratings; pre-lens NITBUT exhibited good sensitivity but poor specificity. The optimum pre-lens NITBUT diagnostic threshold was found to be ≤ 2.0 sec for debilitating CLIDE, and the OSDI threshold was ≥ 11.4.ConclusionsThe DEFC provides a means of quickly categorizing CLIDE patients based on severity and frequency of symptoms, and on the degree to which symptoms impact daily life. The DEFC has several potential advantages as a CLIDE screening and monitoring tool, has good repeatability, and is significantly related to commonly employed clinical assessments for CLIDE.

Mutations in the elongation of very long chain fatty acid 4 (ELOVL4) gene cause Stargardt macular dystrophy 3 (STGD3), a rare, juvenile-onset, autosomal dominant form of macular degeneration. Although several mouse models have already been generated to investigate the link between the three identified disease-causing mutations in the ELOVL4 gene, none of these models recapitulates the early-onset cone photoreceptor cell death observed in the macula of STGD3 patients. To address this specifically, we investigated the effect of mutant ELOVL4 in a mouse model with an all-cone retina. Hence, we bred mice carrying the heterozygously mutated Elovl4 gene on the R91W;Nrl-/- all-cone background and analyzed the retinal lipid composition, morphology, and function over the course of 1 year. We observed a reduction of total phosphatidylcholine-containing very long chain-polyunsaturated fatty acids (PC-VLC-PUFAs) by 39% in the R91W;Nrl-/-;Elovl4 mice already at 6 weeks of age with a pronounced decline of the longest forms of PC-VLC-PUFAs. Total levels of shorter-chain fatty acids (< C26) remained unaffected. However, this reduction in PC-VLC-PUFA content in the all-cone retina had no impact on morphology or function and did not accelerate retinal degeneration in the R91W;Nrl-/-;Elovl4 mice. Taken together, mutations in the ELOVL4 gene lead to cone degeneration in humans, whereas mouse models expressing the mutant Elovl4 show predominant rod degeneration. The lack of a phenotype in the all-cone retina expressing the mutant form of the protein supports the view that aberrant function of ELOVL4 is especially detrimental for rods in mice and suggests a more subtle role of VLC-PUFAs for cone maintenance and survival.

The current research explored toddlers' gaze fixation during a scene showing a person expressing sadness after a ball is stolen from her. The relation between the duration of gaze fixation on different parts of the person's sad face (e.g., eyes, mouth) and theory of mind skills was examined. Eye tracking data indicated that before the actor experienced the negative event, toddlers divided their fixation equally between the actor's happy face and other distracting objects, but looked longer at the face after the ball was stolen and she expressed sadness. The strongest predictor of increased focus on the sad face versus other elements of the scene was toddlers' ability to predict others' emotional reactions when outcomes fulfilled (happiness) or failed to fulfill (sadness) desires, whereas toddlers' visual perspective-taking skills predicted their more specific focusing on the actor's eyes and, for boys only, mouth. Furthermore, gender differences emerged in toddlers' fixation on parts of the scene. Taken together, these findings suggest that top-down processes are involved in the scanning of emotional facial expressions in toddlers.

BackgroundAge-related macular degeneration (AMD) shares many similarities with cardiovascular disease (CVD) pathophysiology. We sought to determine the relationship of AMD to the progression of coronary artery calcium (CAC) using data from the Multi-Ethnic Study of Atherosclerosis (MESA).MethodsOur cohort consisted of 5803 adults aged 45 to 84 years free of known cardiovascular disease (CVD). Retinal photographs were taken during visit 2 (Aug 2002-Jan 2004). CAC was measured with computed tomography at visit 1 (July 2000-Aug 2002) and visit 5 (April 2010-Dec 2011) and changes between visits were determined.ResultsParticipants were categorized as with (n = 244) and without AMD (n = 5559) at visit 2. At visit 5, 92 participants with and 2684 without AMD had CAC scores. Among those with detectable CAC at baseline (>0 at visit 1), CAC progression was greater in persons with compared to those without AMD after multivariable adjustment (530 ± 537 vs. 339 ± 426 Agatston units, P0. The retinal exam might be a useful tool for pre-clinical assessment and prevention of CVD events.

Rbfox1 is a splicing regulator that has been associated with various neurological conditions such as autism spectrum disorder, mental retardation, epilepsy, attention-deficit/hyperactivity disorder and schizophrenia. We show that in adult rodent retinas, Rbfox1 is expressed in all types of retinal ganglion cells (RGCs) and in certain subsets of amacrine cells (ACs), within the inner nuclear (INL) and ganglion cell (GCL) layers. In the INL, all Rbfox1-positive cells were colocalized with GABAergic ACs, however not all GABAergic ACs were immunostained for Rbfox1. In the GCL, a vast majority of GABAergic dACs were Rbfox1-immunopositive. Furthermore, all cholinergic starburst ACs (SACs) in the INL (type a) and in the GCL (type b) were Rbfox1 positive. The expression of Rbfox1 in the retina significantly overlapped with expression of Rbfox2, another member of Rbfox family of proteins. Rbfox2, in addition to RGCs and ACs, was also expressed in horizontal cells. In developing retinas at E12 and E15, Rbfox1 is localized to the cytoplasm of differentiating RGCs and ACs. Between P0 and P5, Rbfox1 subcellular localization switched from cytoplasmic to predominantly nuclear. Downregulation of Rbfox1 in adult Rbfox1loxP/loxP mice had no detectable effect on retinal gross morphology. However, the visual cliff test revealed marked abnormalities of depth perception of these animals. RNA sequencing of retinal transcriptomes of control and Rbfox1 knockout animals identified a number of Rbfox1-regulated genes that are involved in establishing neuronal circuits and synaptic transmission, including Vamp1, Vamp2, Snap25, Trak2, and Slc1A7, suggesting the role of Rbfox1 in facilitating synaptic communications between ACs and RGCs.

ImportanceThe loss of vision following Boston Keratoprosthesis (BKPro) surgery due to glaucoma occurs at a high frequency as diagnosis and management of glaucoma after this procedure pose challenges.ObjectiveTo compare visual outcomes in patients undergoing Boston Keratoprosthesis surgery with and without prior or concurrent glaucoma surgery.Design, setting, and participantsThis is a retrospective, observational cohort study of patients who underwent Boston Type I Keratoprosthesis surgery. 19 eyes of 18 patients who had undergone BKPro and met the inclusion criteria were identified. Twelve eyes received BKPro with prior or concurrent glaucoma surgery (Group 1), and seven eyes were identified undergoing BKPro surgery without prior or concurrent glaucoma surgery (Group 2).Main outcomes and measuresMain outcome included best corrected visual acuity at each follow up.ResultsIn Group 1, mean best corrected visual acuity (BCVA) within a year of BKPro surgery was 20/100 (range 20/40 to Count Fingers (CF); n = 12) and mean BCVA at 1 year from BKPro surgery was 20/115 (range 20/30 to CF; n = 12). 7 out of 12 patients retained or had improved BCVA at 1 year follow up after BKPro implantation, and 5 out of 12 patients had mild BCVA worsening. In Group 2, the mean BCVA within a year of BKPro surgery was 20/140 (ranging from 20/25 to hand motion vision (HM); n = 7) and mean BCVA at 1 year from BKPro surgery was Count Fingers (range 20/60 to Light Perception (LP); n = 6). 4 out of 6 patients lost significant vision at one year after BKPro.Conclusions and relevanceBKPro patients with early glaucoma surgical intervention retained vision significantly better compared to patients with late or no intervention. Our preliminary findings support the recommendation for concurrent or pre-emptive glaucoma surgical intervention in patients undergoing BKPro implantation.

All-trans-retinoic acid (RA) inhibits adipogenesis in established preadipocyte cell lines. Dosing pharmacological amounts of RA reduces weight gain in mice fed a high-fat diet, i.e. counteracts diet-induced obesity (DIO). The aldehyde dehydrogenase Raldh1 (Aldh1a1) functions as one of three enzymes that converts the retinol metabolite retinal into RA, and one of many proteins that contribute to RA homeostasis. Female Raldh1-ablated mice resist DIO. This phenotype contrasts with ablations of other enzymes and binding-proteins that maintain RA homeostasis, which gain adiposity. The phenotype observed prompted the conclusion that loss of Raldh1 causes an increase in adipose tissue retinal, and therefore, retinal functions independently of RA to prevent DIO. A second deduction proposed that low nM concentrations of RA stimulate adipogenesis, in contrast to higher concentrations. Using peer-reviewed LC/MS/MS assays developed and validated for quantifying tissue RA and retinal, we show that endogenous retinal and RA concentrations in adipose tissues from Raldh1-null mice do not correlate with the phenotype. Moreover, male Raldh1-null mice resist weight gain regardless of dietary fat content. Resistance to weight gain occurs during adolescence in both sexes. We show that RA concentrations as low as 1 nM, i.e. in the sub-physiological range, impair adipogenesis of embryonic fibroblasts from wild-type mice. Embryonic fibroblasts from Raldh1-null mice resist differentiating into adipocytes, but retain ability to generate RA. These fibroblasts remain sensitive to an RA receptor pan-agonist, and are not affected by an RA receptor pan-antagonist. Thus, the data do not support the hypothesis that retinal itself represses weight gain and adipogenesis independently of RA. Instead, the data indicate that Raldh1 functions as a retinal and atRA-independent promoter of adiposity during adolescence, and enhances adiposity through pre-adipocyte cell autonomous actions.

ObjectiveTo measure the parapapillary atrophy (PPA) area in en face images obtained with swept-source optical coherence tomography (SS-OCT), and to evaluate its relationship to glaucoma, myopia, and age in non-highly myopic subjects.DesignRetrospective, cross-sectional study.ParticipantsFifty eyes of 30 subjects with open-angle glaucoma (G group) and forty-three eyes of 26 healthy control subjects (C group). Eyes with high myopia (spherical equivalent refractive error ≤ -8 diopters or axial length ≥ 26.5 mm) were excluded.MethodsMean age ± standard deviation was 59.9 ± 12.4 years. The beta zone and the gamma zone PPA areas were measured in en face images reconstructed from three-dimensional SS-OCT images. Relationship between the PPA areas and patient characteristics such as glaucoma, axial length, and age was statistically evaluated using multivariate mixed-effects models.Main outcome measuresAreas of the beta zone and the gamma zone PPA measured on en face OCT images.ResultsAverage ± standard deviation area of the beta and the gamma zone was 0.64 ± 0.79 and 0.16 ± 0.30 mm2, respectively. In multivariate models, the gamma zone significantly correlated with axial length (P = 0.001) but not with glaucoma (P = 0.944). In contrast, the beta zone significantly correlated with age (P = 0.0249) and glaucoma (P = 0.014).ConclusionsEn face images reconstructed from 3D SS-OCT data facilitated measurements of the beta and the gamma PPA zones even in eyes with optic disc distortion. The OCT-defined beta zone is associated with glaucoma and age, whereas the gamma zone correlated with myopia but not with glaucoma. This study confirmed the clinical usefulness of OCT-based classification of the PPA zones in distinguishing glaucomatous damage of the optic nerve from myopic damage in non-highly myopic eyes.

Animal models of ocular hypertension are important tools for glaucoma studies. Both acute transient models and chronic models of ocular hypertension may be useful to investigate specific aspects of neurodegeneration. In this study, we compare the intraocular pressure (IOP) and inner retinal changes induced by 1) laser photocoagulation of both episcleral veins and limbal vessels and 2) circumlimbal suture in CD-1 mice. The suture group is divided into 3 subgroups depending on the level of the immediate IOP spike (acute > 55 mmHg or chronic < 55 mmHg) and time period of monitoring (7 or 28 days). The laser group is followed for 7 days. IOP data show that it peaks at 5 hours and returns to normal level within 7 days in the laser group. In all suture groups, IOP spikes initially and decreases gradually, but it remains significantly elevated at 7 days. In 7 days, the acute suture model generates rapid loss of retinal nerve fiber layer (RNFL) and retinal ganglion cells (RGCs) when compared to the gradual loss by the chronic suture model, possibly due to retinal ischemia and reperfusion within the first few hours after treatment. The laser model falls between the acute suture and chronic suture models resulting in less RNFL and RGC loss than the acute suture model but significantly more loss than the chronic suture model. These results suggest that when using suture models of IOP elevation, it is critical to take the initial IOP spike into consideration and to choose between the acute and chronic models depending on respective research purposes.

Sjögren's syndrome (SS) is a chronic, autoimmune exocrinopathy that leads to severe dryness of the mouth and eyes. Exocrine function is highly regulated by neuronal mechanisms but little is known about the link between chronic inflammation, innervation and altered exocrine function in the diseased eyes and exocrine glands of SS patients. To gain a better understanding of neuronal regulation in the immunopathogenesis of autoimmune exocrinopathy, we profiled a mouse model of spontaneous, autoimmune exocrinopathy that possess key characteristics of peripheral neuropathy experienced by SS patients. Mice deficient in the autoimmune regulator (Aire) gene developed spontaneous, CD4+ T cell-mediated exocrinopathy and aqueous-deficient dry eye that were associated with loss of nerves innervating the cornea and lacrimal gland. Changes in innervation and tear secretion were accompanied by increased proliferation of corneal epithelial basal cells, limbal expansion of KRT19-positive progenitor cells, increased vascularization of the peripheral cornea and reduced nerve function in the lacrimal gland. In addition, we found extensive loss of MIST1+ secretory acinar cells in the Aire -/- lacrimal gland suggesting that acinar cells are a primary target of the disease, Finally, topical application of ophthalmic steroid effectively restored corneal innervation in Aire -/- mice thereby functionally linking nerve loss with local inflammation in the aqueous-deficient dry eye. These data provide important insight regarding the relationship between chronic inflammation and neuropathic changes in autoimmune-mediated dry eye. Peripheral neuropathies characteristic of SS appear to be tightly linked with the underlying immunopathological mechanism and Aire -/- mice provide an excellent tool to explore the interplay between SS-associated immunopathology and peripheral neuropathy.

Task preparation involves multiple component processes, including a general evaluative process that signals the need for adjustments in control, and the engagement of task-specific control settings. Here we examined the dynamics of these different mechanisms in preparing the attentional control system for visual search. We explored preparatory activity using pupil dilation, a well-established measure of task demands and effortful processing. In an initial exploratory experiment, participants were cued at the start of each trial to search for either a salient color singleton target (an easy search task) or a low-salience shape singleton target (a difficult search task). Pupil dilation was measured during the preparation period from cue onset to search display onset. Mean dilation was larger in preparation for the difficult shape target than the easy color target. In two additional experiments, we sought to vary effects of evaluative processing and task-specific preparation separately. Experiment 2 showed that when the color and shape search tasks were matched for difficulty, the shape target no longer evoked larger dilations, and the pattern of results was in fact reversed. In Experiment 3, we manipulated difficulty within a single feature dimension, and found that the difficult search task evoked larger dilations. These results suggest that pupil dilation reflects expectations of difficulty in preparation for a search task, consistent with the activity of an evaluative mechanism. We did not find consistent evidence for relationship between pupil dilation and search performance (accuracy and response timing), suggesting that pupil dilation during search preparation may not be strongly linked to ongoing task-specific preparation.

PurposeCongenital cataracts occur in isolation in about 70% of cases or are associated with other abnormalities such as anterior segment dysgenesis and microphthalmia. We identified a three-generation family in the University of California San Francisco glaucoma clinic comprising three individuals with congenital cataracts and aphakic glaucoma, one of whom also had microphthalmia. The purpose of this study was to identify a possible causative mutation in this family and to investigate its pathogenesis.MethodsWe performed exome sequencing and identified a putative mutation in gap junction protein α8 (GJA8). We used PCR and DNA sequencing of GJA8 in affected and unaffected members of the pedigree to test segregation of the variant with the phenotype. We tested cellular distribution and function of the variant protein by immunofluorescence and intercellular transfer of Neurobiotin in transiently transfected HeLa cells.ResultsExome sequencing revealed a variant in GJA8 (c.658A>G) encoding connexin50 (Cx50) that resulted in a missense change (p.N220D) in transmembrane domain 4. The variant was present in all three affected family members, but was also present in the proband's grandfather who was reported to be unaffected. The mutant protein localized to the plasma membrane and supported intercellular Neurobiotin transfer in HeLa cells.ConclusionsWe identified a variant in transmembrane domain 4 of Cx50 in a family with autosomal dominant congenital cataracts. This variant has been previously identified in other cataract cohorts, but it is also present in unaffected individuals. Our study demonstrates that the mutant protein localized to the plasma membrane and formed functional intercellular channels. These data suggest that GJA8 c.658A>G is most likely a benign rare variant.

BackgroundEach inherited retinal disorder is rare, but together, they affect millions of people worldwide. No treatment is currently available for these blinding diseases, but promising new options-including gene therapy-are emerging. Arguably, the most prevalent retinal dystrophy is Stargardt disease. In each case, the specific combination of ABCA4 variants (> 900 identified to date) and modifying factors is virtually unique. It accounts for the vast phenotypic heterogeneity including variable rates of functional and structural progression, thereby potentially limiting the ability of phase I/II clinical trials to assess efficacy of novel therapies with few patients. To accommodate this problem, we developed and validated a sensitive and reliable composite clinical trial endpoint for disease progression based on structural measurements of retinal degeneration.Methods and findingsWe used longitudinal data from early-onset Stargardt patients from the Netherlands (development cohort, n = 14) and the United Kingdom (external validation cohort, n = 18). The composite endpoint was derived from best-corrected visual acuity, fundus autofluorescence, and spectral-domain optical coherence tomography. Weighting optimization techniques excluded visual acuity from the composite endpoint. After optimization, the endpoint outperformed each univariable outcome, and showed an average progression of 0.41° retinal eccentricity per year (95% confidence interval, 0.30-0.52). Comparing with actual longitudinal values, the model accurately predicted progression (R2, 0.904). These properties were largely preserved in the validation cohort (0.43°/year [0.33-0.53]; prediction: R2, 0.872). We subsequently ran a two-year trial simulation with the composite endpoint, which detected a 25% decrease in disease progression with 80% statistical power using only 14 patients.ConclusionsThese results suggest that a multimodal endpoint, reflecting structural macular changes, provides a sensitive measurement of disease progression in Stargardt disease. It can be very useful in the evaluation of novel therapeutic modalities in rare disorders.

PurposeAssess potential risk factors for severe Meibomian gland atrophy (SMGA) in a young adult population.MethodsCross-sectional study using medical history and ocular surface examination to evaluate relationships with study outcomes: SMGA, tear lipid layer (TLL) thickness, non-invasive (NITBUT) and fluorescein (FTBUT) tear breakup times, and symptoms using the Standard Patient Evaluation of Eye Dryness (SPEED) questionnaire.ResultsOne hundred one participants (101; 202 eyes; Age: mean±SD = 22.3±4.0 years) completed the study. Hormonal birth control (HBC) use was the only significant risk factor for SMGA (p = 0.028). Female HBC users had 4.8 times greater odds of having SMGA compared to female HBC non-users (p = 0.028), but the odds of having SMGA was similar between female HBC non-users and males (p = 0.885). Multivariable analysis suggested that the relationship between SMGA and TLL thickness was dependent on HBC use. Compared to female HBC non-users without SMGA, TLL thickness for HBC users was estimated to be 10 nm thinner if SMGA was absent (p = 0.007) and 21 nm thinner if SMGA was present (p

Retinal OFF bipolar cells show distinct connectivity patterns with photoreceptors in the wild-type mouse retina. Some types are cone-specific while others penetrate further through the outer plexiform layer (OPL) to contact rods in addition to cones. To explore dendritic stratification of OFF bipolar cells in the absence of rods, we made use of the 'cone-full' Nrl-/- mouse retina in which all photoreceptor precursor cells commit to a cone fate including those which would have become rods in wild-type retinas. The dendritic distribution of OFF bipolar cell types was investigated by confocal and electron microscopic imaging of immunolabeled tissue sections. The cells' dendrites formed basal contacts with cone terminals and expressed the corresponding glutamate receptor subunits at those sites, indicating putative synapses. All of the four analyzed cell populations showed distinctive patterns of vertical dendritic invasion through the OPL. This disparate behavior of dendritic extension in an environment containing only cone terminals demonstrates type-dependent specificity for dendritic outgrowth in OFF bipolar cells: rod terminals are not required for inducing dendritic extension into distal areas of the OPL.

PurposeTo compare the diagnostic abilities of vessel density measurements of the optic nerve head (ONH), peripapillary and macular regions on optical coherence tomography (OCT) angiography in eyes with primary open angle glaucoma (POAG) with that of the ONH rim area, peripapillary retinal nerve fiber layer (RNFL) thickness and the macular ganglion cell complex (GCC) thickness measurements.MethodsIn a cross sectional study, 78 eyes of 50 control subjects and 117 eyes of 67 POAG patients underwent vessel density and structural measurements with spectral domain OCT. POAG was diagnosed based on the masked evaluation of optic disc stereo photographs. Area under receiver operating characteristic curves (AUC) and sensitivities at fixed specificities of vessel densities in ONH, peripapillary and macular regions were compared with rim area, RNFL and GCC thickness.ResultsThe AUC (sensitivity at 95% specificity) of average vessel densities within the ONH, peripapillary and macular region were 0.77 (31%), 0.85 (56%) and 0.70 (18%) respectively. The same of ONH rim area, average RNFL and GCC thickness were 0.94 (83%), 0.95 (72%) and 0.93 (62%) respectively. AUCs of vessel densities were significantly lower (p

PurposeTo compare intraoperative angle stability and postoperative outflow of two ab interno trabeculectomy devices that excise the trabecular meshwork with or without active aspiration and irrigation. We hypothesized that anterior segment optical coherence tomography (AS-OCT) allows for a quantitative comparison of intraoperative angle stability in a microincisional glaucoma surgery (MIGS) pig eye training model.MethodsTwelve freshly enucleated porcine eyes were measured with AS-OCT at baseline, at the beginning of the procedure and at its conclusion to determine the anterior chamber depth (ACD) and the nasal angle α in degrees. The right and left eye of pairs were randomly assigned to an active dual blade goniectome (aDBG) and a passive dual blade goniectome (pDBG) group, respectively. The aDBG had irrigation and aspiration ports while the pDBG required surgery under viscoelastic. We performed the procedures using our MIGS training system with a standard, motorized ophthalmic operating microscope. We estimated outflow by obtaining canalograms with fluorescent spheres.ResultsIn aDBG, the nasal angle remained wide open during the procedure at above 90° and did not change towards the end (100±10%, p = 0.9). In contrast, in pDBG, ACD decreased by 51±19% to 21% below baseline (p

Transparency of the human cornea is necessary for vision. Fuchs Endothelial Corneal Dystrophy (FECD) is a bilateral, heritable degeneration of the corneal endothelium, and a leading indication for corneal transplantation in developed countries. While the early onset, and rarer, form of FECD has been linked to COL8A2 mutations, the more common, late onset form of FECD has genetic mutations linked to only a minority of cases. Epigenetic modifications that occur in FECD are unknown. Here, we report on and compare the DNA methylation landscape of normal human corneal endothelial (CE) tissue and CE from FECD patients using the Illumina Infinium HumanMethylation450 (HM450) DNA methylation array. We show that DNA methylation profiles are distinct between control and FECD samples. Differentially methylated probes (10,961) were identified in the FECD samples compared with the control samples, with the majority of probes being hypermethylated in the FECD samples. Genes containing differentially methylated sites were disproportionately annotated to ontological categories involving cytoskeletal organization, ion transport, hematopoetic cell differentiation, and cellular metabolism. Our results suggest that altered DNA methylation patterns may contribute to loss of corneal transparency in FECD through a global accumulation of sporadic DNA methylation changes in genes critical to basic CE biological processes.

γδ T cells can either enhance or inhibit an adaptive immune response, but the mechanisms involved are not fully understood. Given that CD73 is the main enzyme responsible for conversion of AMP into the immunosuppressive molecule adenosine, we investigated its role in the regulatory function of γδ T cells in experimental autoimmune uveitis (EAU). We found that γδ T cells expressed different amounts of CD73 during the different stages of EAU and that low CD73 expression on γδ T cells correlated with enhanced Th17 response-promoting activity. Functional comparison of CD73-deficient and wild-type B6 (CD73+/+) mice showed that failure to express CD73 decreased both the enhancing and suppressive effects of γδ T cells on EAU. We also demonstrated that γδ T cells expressed different amounts of CD73 when activated by different pathways, which enabled them to either enhance or inhibit an adaptive immune response. Our results demonstrate that targeting CD73 expression on γδ T cells may allow us to manipulate their pro- or anti-inflammatory effect on Th17 responses.

PurposeThis study investigated the gene expression of TGFβ isoforms and their receptors in chick retina, retinal pigment epithelium (RPE), and choroid and the effects of short-term imposed optical defocus.MethodsThe expression of TGFβ isoforms (TGF-β1, 2, 3) and TGFβ receptors (TGFBR1, 2, 3) was examined in the retina, RPE, and choroid of young White-Leghorn untreated chicks (19 days-old). The effects on the expression of the same genes of monocular +10 and -10 D defocusing lenses, worn for either 2 or 48 h by age-matched chicks, were also examined by comparing expression in treated and untreated fellow eyes. RNA was purified, characterized and then reverse transcribed to cDNA. Differential gene expression was quantified using real-time PCR.ResultsAll 3 isoforms of TGFβ and all 3 receptor subtypes were found to be expressed in all 3 ocular tissues, with apparent tissue-dependent differences in expression profiles. Data are reported as mean normalized expression relative to GAPDH. Sign-dependent optical defocus effects were also observed. Optical defocus did not affect retinal gene expression but in the RPE, TGF-β2 expression was significantly up-regulated with +10 D lenses, worn for either 2 h (349% increase ± 88%, p < 0.01) or 48 h (752% increase ± 166%, p < 0.001), and in the choroid, the expression of TGF-β3 was up-regulated with -10 D lenses, worn for 48 h (147% increase ± 9%, p < 0.01).ConclusionsThe effects of short term exposure to optical defocus on TGFβ gene expression in the RPE and choroid, which were sign-dependent and isoform specific, provide further supporting evidence for important roles of members of the TGFβ family and these two tissues in local signal cascades regulating ocular growth.

Opsins are photosensitive proteins catalyzing light-dependent processes across the tree of life. For both microbial (type 1) and metazoan (type 2) opsins, photosensing depends upon covalent interaction between a retinal chromophore and a conserved lysine residue. Despite recent discoveries of potential opsin homologs lacking this residue, phylogenetic dispersal and functional significance of these abnormal sequences have not yet been investigated. We report discovery of a large group of putatively non-retinal binding opsins, present in a number of fungal and microbial genomes and comprising nearly 30% of opsins in the Halobacteriacea, a model clade for opsin photobiology. We report phylogenetic analyses, structural modeling, genomic context analysis and biochemistry, to describe the evolutionary relationship of these recently described proteins with other opsins, show that they are expressed and do not bind retinal in a canonical manner. Given these data, we propose a hypothesis that these abnormal opsin homologs may represent a novel family of sensory opsins which may be involved in taxis response to one or more non-light stimuli. If true, this finding would challenge our current understanding of microbial opsins as a light-specific sensory family, and provides a potential analogy with the highly diverse signaling capabilities of the eukaryotic G-protein coupled receptors (GPCRs), of which metazoan type 2 opsins are a light-specific sub-clade.

BackgroundAquaporin 4 (AQP4) is considered a putative autoantigen in patients with Neuromyelitis optica (NMO), an autoinflammatory disorder of the central nervous system (CNS). HLA haplotype analyses of patients with NMO suggest a positive association with HLA-DRB1* 03:01. We previously showed that the human (h) AQP4 peptide 281-300 is the dominant immunogenic determinant of hAQP4 in the context of HLA-DRB1*03:01. This immunogenic peptide stimulates a strong Th1 and Th17 immune response. AQP4281-300-specific encephalitogenic CD4+ T cells should initiate CNS inflammation that results in a clinical phenotype in HLA-DRB1*03:01 transgenic mice.MethodsControlled study with humanized experimental animals. HLA-DRB1*03:01 transgenic mice were immunized with hAQP4281-300, or whole-length hAQP4 protein emulsified in complete Freund's adjuvant. Humoral immune responses to both antigens were assessed longitudinally. In vivo T cell frequencies were assessed by tetramer staining. Mice were followed clinically, and the anterior visual pathway was tested by pupillometry. CNS tissue was examined histologically post-mortem. Flow cytometry was utilized for MHC binding assays and to immunophenotype T cells, and T cell frequencies were determined by ELISpot assay.ResultsImmunization with hAQP4281-300 resulted in an in vivo expansion of antigen-specific CD4+ T cells, and an immunoglobulin isotype switch. HLA-DRB1*03:01 TG mice actively immunized with hAQP4281-300, or with whole-length hAQP4 protein were resistant to developing a neurological disease that resembles NMO. Experimental mice show no histological evidence of CNS inflammation, nor change in pupillary responses. Subsequent analysis reveals that a single amino acid substitution from aspartic acid in hAQP4 to glutamic acid in murine (m)AQP4 at position 290 prevents the recognition of hAQP4281-300 by the murine T cell receptor (TCR).ConclusionInduction of a CNS inflammatory autoimmune disorder by active immunization of HLA-DRB1*03:01 TG mice with human hAQP4281-300 will be complex due to a single amino acid substitution. The pathogenic role of T cells in this disorder remains critical despite these observations.

Various pathological conditions are accompanied by ATP release from the intracellular to the extracellular compartment. Extracellular ATP (eATP) functions as a signaling molecule by activating purinergic P2 purine receptors. The key P2 receptor involved in inflammation was identified as P2X7R. Recent studies have shown that P2X7R signaling is required to trigger the Th1/Th17 immune response, and oxidized ATP (oxATP) effectively blocks P2X7R activation. In this study we investigated the effect of oxATP on mouse experimental autoimmune uveitis (EAU). Our results demonstrated that induced EAU in B6 mice was almost completely abolished by the administration of small doses of oxATP, and the Th17 response, but not the Th1 response, was significantly weakened in the treated mice. Mechanistic studies showed that the therapeutic effects involve the functional change of a number of immune cells, including dendritic cells (DCs), T cells, and regulatory T cells. OxATP not only directly inhibits the T cell response; it also suppresses T cell activation by altering the function of DCs and Foxp3+ T cell. Our results demonstrated that inhibition of P2X7R activation effectively exempts excessive autoimmune inflammation, which may indicate a possible therapeutic use in the treatment of autoimmune diseases.