Retinal Function Research Articles

The effects of time restricted feeding on age-related changes in the mouse retina

Dietary modifications such as caloric restriction (CR) and intermittent fasting (IF) have gained popularity due to their proven health benefits in aged populations. In time restricted feeding (TRF), a form of intermittent fasting, the amount of time for food intake is regulated without restricting the caloric intake. TRF is beneficial for the central nervous system to support brain health in the context of aging. Therefore, we here ask whether TRF also exerts beneficial effects in the aged retina. We compared aged mice (24 months) on a TRF paradigm (access to food for six hours per day) for either 6 or 12 months against young control mice (8 months) and aged control mice on an ad libitum diet. We examined changes in the retina at the functional (electroretinography), structural (histology and fluorescein angiograms) and molecular (gene expression) level. TRF treatment showed amelioration of age-related reductions in both scotopic and photopic b-wave amplitudes suggesting benefits for retinal interneuron signaling. TRF did not affect age-related signs of retinal inflammation or microglial activation at either the molecular or histological level. Our data indicate that TRF helps preserve some aspects of retinal function that are decreased with aging, adding to our understanding of the health benefits that altered feeding patterns may confer.

Thyroid Hormone Signaling in Retinal Development and Function: Implications for Diabetic Retinopathy and Age-Related Macular Degeneration.

Thyroid Hormones (THs) play a central role in the development, cell growth, differentiation, and metabolic homeostasis of neurosensory systems, including the retina. The coordinated activity of various components of TH signaling, such as TH receptors (THRs) and the TH processing enzymes deiodinases 2 and 3 (DIO2, DIO3), is required for proper retinal maturation and function of the adult photoreceptors, Müller glial cells, and pigmented epithelial cells. Alterations of TH homeostasis, as observed both in frank or subclinical thyroid disorders, have been associated with sight-threatening diseases leading to irreversible vision loss i.e., diabetic retinopathy (DR), and age-related macular degeneration (AMD). Although observational studies do not allow causal inference, emerging data from preclinical models suggest a possible correlation between TH signaling imbalance and the development of retina disease. In this review, we analyze the most important features of TH signaling relevant to retinal development and function and its possible implication in DR and AMD etiology. A better understanding of TH pathways in these pathological settings might help identify novel targets and therapeutic strategies for the prevention and management of retinal disease.

Short communication: Maintained visual performance in birds under high altitude hypoxia

Birds are highly dependent on their vision for orientation and navigation. The avian eye differs from the mammalian eye as the retina is avascular, leaving the inner, highly metabolically active layers with a very long diffusion distance to the oxygen supply. During flight at high altitudes, birds face a decrease in environmental oxygen partial pressure, which leads to a decrease in arterial oxygen levels. Since oxygen perfusion to the retina is already limited in birds, we hypothesize that visual function is impaired by low oxygen availability. However, the visual performance of birds exposed to hypoxia has not been evaluated before. Here, we assess the optomotor response (OMR) in zebra finches under simulated high-altitude hypoxia (10%) and show that the OMR is largely maintained under hypoxia with only a modest reduction in OMR, demonstrating that birds can largely maintain visual function at high altitudes. The method of our study does not provide insight into the mechanisms involved, but our findings suggest that birds have evolved physiological mechanisms for retinal function at low tissue oxygen levels.

Small extracellular vesicles derived from microRNA-22-3p-overexpressing mesenchymal stem cells protect retinal ganglion cells by regulating MAPK pathway

Glaucoma is the leading cause of irreversible blindness and is characterized by progressive retinal ganglion cell (RGC) loss and retinal nerve fiber layer thinning. Currently, no existing treatment is effective for the preservation of RGCs. MicroRNA-22-3p (miR22) and small extracellular vesicles derived from mesenchymal stem cells (MSC-sEVs) have neuroprotective effects. In this study, we apply miR22-overexpressing MSC-sEVs in an N-methyl-D-aspartic acid (NMDA)-induced RGC injury model to assess their short-term therapeutic effects and explore the underlying mechanisms. We find that mice in the miR22-sEVs-treated group have thicker retinas, fewer apoptotic cells, more reserved RGCs, better retinal function, and lower expression levels of Bax and caspase-3. MiR22-sEVs treatment promotes viability, inhibits apoptosis and inhibits Bax and caspase-3 expression in RGC-5 cells. MiR22 targets mitogen-activated protein kinase kinase kinase 12 to inhibit apoptosis by regulating the mitogen-activated protein kinase (MAPK) signaling pathway. Collectively, our results suggest that miR22-sEVs ameliorate NMDA-induced RGC injury through the inhibition of MAPK signaling pathway-mediated apoptosis, providing a potential therapy for glaucoma and other diseases that involve RGC damage.

Exploitation of enhanced prime editing for blocking aberrant angiogenesis

IntroductionAberrant angiogenesis plays an important part in the development of a variety of human diseases including proliferative diabetic retinopathy, with which there are still numerous patients remaining a therapeutically challenging condition. Prime editing (PE) is a versatile gene editing approach, which offers a novel opportunity to genetically correct challenging disorders. ObjectivesThe goal of this study was to create a dominant-negative (DN) vascular endothelial growth factor receptor (VEGFR) 2 by editing genomic DNA with an advanced PE system to block aberrant retinal angiogenesis in a mouse model of oxygen-induced retinopathy. MethodsAn advanced PE system (referred to as PE6x) was established within two lentiviral vectors, with one carrying an enhanced PE guide RNA and a canonical Cas9 nickase fused with an optimized reversal transcriptase, and the other conveying a nicking guide RNA and a DN-MLH1 to improve PE efficiency. Dual non-integrating lentiviruses (NILVs) produced with the two lentiviral PE6x vectors were then employed to create a mutation of VEGFR2 T17967A by editing the Mus musculus VEGFR2 locus in vitro and in vivo, leading to generation of a premature stop codon (TAG, K796stop) to produce DN-VEGFR2, to interfere with the wild type VEGFR2 which is essential for angiogenesis. ResultsNILVs targeting VEGFR2 delivered into cultured murine vascular endothelial cells led to 51.06 % VEGFR2 T17967A in the genome analyzed by next generation sequencing and the production of DN-VEGFR2, which was found to hamper VEGF-induced VEGFR2 phosphorylation, as demonstrated by Western blot analysis. Intravitreally injection of the dual NILVs into postnatal day 12 mice in a model of oxygen-induced retinopathy, led to production of retinal DN-VEGFR2 in postnatal day 17 mice which blocked retinal VEGFR2 expression and activation as well as abnormal retinal angiogenesis without interfering with retinal structure and function, as assessed by electroretinography, optical coherence tomography, fundus fluorescein angiography and histology. ConclusionDN-VEGFR2 resulted from editing genomic VEGFR2 using the PE6x system can be harnessed to treat intraocular pathological angiogenesis.

Changes in Retinal Structure and Function in Mice Exposed to Flickering Blue Light: Electroretinographic and Optical Coherence Tomographic Analyses.

The harmful effects of blue light on the retina and health issues attributed to flickering light have been researched extensively. However, reports on the effects of flickering blue light at a frequency in the visible range on the retina are limited. This study aimed to non-invasively investigate the structural and functional changes in mice retinas following exposure to flickering blue light. BALB/c mice were subjected to non-flickering and flickering blue light, and changes in the retinal function and structure were assessed using electroretinography (ERG) and spectral-domain optical coherence tomography (SD-OCT), respectively. Retinal damage progression was monitored on days 3, 7, 14, and 42 following light exposure. Significant reductions in scotopic and photopic ERG responses were observed on day 3 (p<0.05). On day 7, the non-flickering and flickering groups demonstrated different functional changes: the flickering group showed further ERG response reduction, while the non-flickering group showed no reduction or slight improvement that was statistically insignificant (p>0.05). A similar trend lasted by day 14. On day 42, however, the difference between the non-flickering and flickering groups was significant, which was corroborated by the normalized amplitudes at 0, 0.5, and 1 log cd s/m2 (p<0.05). Quantitative and qualitative SD-OCT assays revealed more severe and progressive retinal damage in the flickering group throughout the study. Flickering blue light causes more persistent and severe retinal damage than non-flickering blue light and may be a risk factor for retinal degeneration even at frequencies as low as 20 Hz.

High temporal frequency light response in mouse retina requires FAT3 signaling in bipolar cells.

Vision is initiated by the reception of light by photoreceptors and subsequent processing via downstream retinal neurons. Proper cellular organization depends on the multi-functional tissue polarity protein FAT3, which is required for amacrine cell connectivity and retinal lamination. Here we investigated the retinal function of Fat3 mutant mice and found decreases in physiological and perceptual responses to high frequency flashes. These defects did not correlate with abnormal amacrine cell wiring, pointing instead to a role in bipolar cell subtypes that also express FAT3. The role of FAT3 in the response to high temporal frequency flashes depends upon its ability to transduce an intracellular signal. Mechanistically, FAT3 binds to the synaptic protein PTPσ, intracellularly, and is required to localize GRIK1 to OFF-cone bipolar cell synapses with cone photoreceptors. These findings expand the repertoire of FAT3's functions and reveal its importance in bipolar cells for high frequency light response.

Topical administration of GLP-1 eyedrops improves retinal ganglion cell function by facilitating presynaptic GABA release in early experimental diabetes.

Diabetic retinopathy is a prominent cause of blindness in adults, with early retinal ganglion cell (RGC) loss contributing to visual dysfunction or blindness. In the brain, defects in y-aminobutyric acid (GABA) synaptic transmission are associated with pathophysiological and neurodegenerative disorders, whereas glucagon-like peptide-1 (GLP-1) has demonstrated neuroprotective effects. However, it is not yet clear whether diabetes causes alterations in inhibitory input to RGCs and whether and how GLP-1 protects against neurodegeneration in the diabetic retina through regulating inhibitory synaptic transmission to RGCs. In the present study, we used the patch-clamp technique to record GABA subtype A receptor-mediated miniature inhibitory postsynaptic currents (mIPSCs) in RGCs from streptozotocin-induced diabetes model rats. We found that early diabetes (4 weeks of hyperglycemia) decreased the frequency of GABAergic mIPSCs in RGCs without altering their amplitude, suggesting a reduction in the spontaneous release of GABA to RGCs. Topical administration of GLP-1 eyedrops over a period of 2 weeks effectively countered the hyperglycemia-induced downregulation of GABAergic mIPSC frequency, subsequently enhancing the survival of RGCs. Concurrently, the protective effects of GLP-1 on RGCs in diabetic rats were eliminated by topical administration of exendin-9-39, a specific GLP-1 receptor antagonist, or SR95531, a specific antagonist of the GABA subtype A receptor. Furthermore, extracellular perfusion of GLP-1 was found to elevate the frequencies of GABAergic mIPSCs in both ON- and OFF-type RGCs. This elevation was shown to be mediated by activation of the phosphatidylinositol-phospholipase C/inositol 1,4,5-trisphosphate receptor/Ca2+/protein kinase C signaling pathway downstream of GLP-1 receptor activation. Moreover, multielectrode array recordings revealed that GLP-1 functionally augmented the photoresponses of ON-type RGCs. Optomotor response tests demonstrated that diabetic rats exhibited reductions in visual acuity and contrast sensitivity that were significantly ameliorated by topical administration of GLP-1. These results suggest that GLP-1 facilitates the release of GABA onto RGCs through the activation of GLP-1 receptor, leading to the de-excitation of RGC circuits and the inhibition of excitotoxic processes associated with diabetic retinopathy. Collectively, our findings indicate that the GABA system has potential as a therapeutic target for mitigating early-stage diabetic retinopathy. Furthermore, the topical administration of GLP-1 eyedrops represents a non-invasive and effective treatment approach for managing early-stage diabetic retinopathy.

Relative foveal dark adaptation: a potential method for assessing macular health.

Dark adaptation measures photoreceptor recovery following intense light stimulation. Time to recovery reflects retinal function. We describe a novel method of relative foveal dark adaptation using an iPhone. Data from a small number of healthy subjects were studied to assess reproducibility, effects of age, and consider potential clinical utility. Relative foveal dark adaption was studied in 6 normal subjects across ages from 20 to 81 years and across differing testing conditions. Foveal bleaching is produced by fixating a bright white circle on an iPhone for variable times. After foveal bleaching an annular surround appears to complete a bullseye stimulus with surround initially brighter than centre. As the fovea recovers the centre regains brightness. Relative foveal dark adaptation, the time for the visual anchor to shift from surround to centre, was studied across a range of bleaching times, ages, and testing conditions. Dispersion of dark adaptation times grows with increasing age. Foveal bleaching for 30 s was as effective as longer times. Testing times with a 30 s bleach were less than 1 min. Foveal dark adaptation was reproducible within each subject and was unaffected by ambient room lighting, pupil size, and light attenuation. Repeat, immediately sequential testing was similarly reproducible except after long bleaching. This method of dark adaptation is intuitive, repeatable, and relatively unaffected by testing condition. Testing times are brief, requiring only an iPhone screen positioned at reading distance. Relative foveal dark adaptation may be a useful tool to assess macular health.

Photovoltaic, wireless wide‐field epiretinal prosthesis to treat retinitis pigmentosa

AbstractPurposeTo develop and evaluate a photovoltaic, wireless wide‐field epiretinal prosthesis for the treatment of retinitis pigmentosa.MethodsA mosaic array of thinned silicon‐based photodiodes with integrated thin‐film stimulation electrodes was fabricated with a flexible polyimide substrate film to form a film‐based miniaturized electronic system with wireless optical power and signal transmission and integrated electrostimulation. Manufactured implants were characterized with respect to their optoelectronic performance and biocompatibility following DIN EN ISO 10993.ResultsA 14 mm diameter prosthesis containing 1276 pixels with a maximum sensitivity at a near infrared wavelength of 905 nm and maximized stimulation current density 30–50 μm below the electrodes was developed for direct activation of retinal ganglion cells during epiretinal stimulation. Fabricated prostheses demonstrated mucosal tolerance and the preservation of both metabolic activity, proliferation and membrane integrity of human fibroblasts as well as the retinal functions of bovine retinas. Illumination of the prosthesis, which was placed epiretinally on an isolated perfused bovine retina, with infrared light resulted in electrophysiological recordings reminiscent of an a‐wave (hyperpolarization) and b‐wave (depolarization).ConclusionsA photovoltaic, wireless wide‐field epiretinal prosthesis for the treatment of retinitis pigmentosa using near infrared light for signal transmission was designed, manufactured and its biocompatibility and functionality demonstrated in vitro and ex vivo.

In Search of Mouse Models for Exfoliation Syndrome

Exfoliation syndrome (XFS) is a systemic connective tissue disorder with elusive pathophysiology. We hypothesize that a mouse model with elastic fiber defects caused by lack of lysyl oxidase like 1 (LOXL1 encoded by Loxl1), combined with microfibril deficiency due to Fbn1 mutation (encoding fibrillin-1, Fbn1C1041G/+) will display ocular and systemic phenotypes of XFS. Loxl1-/- was crossed with Fbn1C1041G/+ to create double mutant (dbm) mice. Intraocular pressure (IOP), visual acuity (VA), electroretinogram (ERG) and biometry were characterized in 4 genotypes (wt, Fbn1C1041G/+, Loxl1-/-, dbm) at 16 weeks old. Optic nerve area was measured by ImageJ and axon counting was achieved by AxonJ. Deep whole-body phenotyping was performed in wt and dbm mice. Two-tailed Student's t-test was used for statistical analysis. There was no difference in IOP between the 4 genotypes. VA was significantly reduced only in dbm mice. The majority of biometric parameters showed significant differences in all 3 mutant genotypes compared to wt, and dbm had exacerbated anomalies compared to single mutants. Dbm mice showed reduced retinal function and significantly enlarged ON area when compared with wt. Dbm mice exhibited severe systemic phenotypes related to abnormal elastic fibers, such as pelvic organ prolapse, cardiovascular and pulmonary abnormalities. Ocular and systemic findings in dbm mice support functional overlap between fibrillin-1 and LOXL1, two prominent components of exfoliation material. Although no elevated IOP or reduction of axon numbers was detected in dbm mice at 16-week-old, their reduced retinal function and enlarged ON area indicate early retinal ganglion cell dysfunction. Dbm mice also provide insight on the link between XFS and systemic diseases in humans.

Assessment of Brain-Derived Neurotrophic Factor on Retinal Structure and Visual Function in Rodent Models of Optic Nerve Crush.

The effects of brain-derived neurotrophic factor (BDNF) on retinal ganglion cell (RGC) survival and visual function were assessed in rat and mouse models of optic nerve (ON) crush. ONs were crushed on Day 1, followed by intravitreal injections of a vehicle or BDNF on Days 1 and 8. The spatial frequency threshold was measured using optokinetic tracking on Days 7 and 14. On Day 15, ganglion cell complex (GCC) thickness was quantified using optical coherence tomography. Furthermore, all eyes were enucleated for immunohistochemical analysis of the surviving RGC somas and axons. BDNF significantly reduced the RGC soma in mice and increased GCC thickness in intact eyes, with apparent axonal swelling in both species. It displayed significantly greater RGC soma survival in eyes with ON injury, with moderately thicker axonal bundles in both species and a thicker GCC in rats. Visual function was significantly reduced in all ON-crushed animals, regardless of BDNF treatment. Thus, we obtained a comprehensive analysis of the structural and functional impact of BDNF in intact and ON-crushed eyes in two rodent models. Our results provide a foundation for further BDNF evaluation and the design of preclinical studies on neuroprotectants using BDNF as a reference positive control.

Endothelial dysfunction in retinal vessels of hemodialysis patients compared to healthy controls

Endothelial dysfunction is a key factor promoting atherosclerosis and cardiovascular complications. Hemodialysis patients typically show various cardiovascular complications and impaired retinal venular dilation has been described as a risk factor for mortality. Non-invasive retinal vessel analysis provides insight into the microvasculature and endothelial function. Static retinal vessel analysis determines arteriolar and venular vessel diameters and dynamic retinal vessel analysis measures microvascular function by flicker-light induced stimulation, which results in physiological dilation of retinal vessels. We measured 220 healthy individuals and compared them to our preexisting cohort of hemodialysis patients (275 for static and 214 for dynamic analysis). Regarding static vessel diameters, hemodialysis patients and healthy individuals did not significantly differ between vessel diameters. Dynamic retinal vessel analysis showed attenuated dilation of the arteriole of hemodialysis patients with 1.6% vs 2.3% in healthy individuals (p = 0.009). Case–control matching for age (mean 65.4 years) did not relevantly diminish the difference. Hemodialysis patients also exhibited reduced venular dilation after matching for age (3.2% vs 3.8%, p = 0.019). Hemodialysis patients showed microvascular dysfunction compared to healthy individuals when using dynamic retinal vessel analysis. Further studies should focus on dynamic retinal vessel analysis which can add insights into the microvascular function and risk factors in multimorbid patients.

Effect of Bromfenac on Reducing Neuroinflammation in an Ischemia-Reperfusion Glaucoma Model.

In the context of glaucoma, intraocular pressure (IOP) and age are recognized as the primary factors contributing to its onset and progression. However, significant reductions in IOP fail to completely halt its advancement. An emerging body of literature highlights the role of neuroinflammation in glaucoma. This study aimed to explore Bromfenac's anti-inflammatory properties in mitigating neuroinflammation associated with glaucoma using an ischemia-reperfusion (IR) glaucoma model. Bromfenac's impact on microglia and astrocytes under pressure was assessed via Western blotting and an enzyme-linked immunosorbent assay. Immunohistochemical staining was used to evaluate glial activation and changes in inflammatory marker expression in the IR model. Bromfenac led to the downregulation of inflammatory markers, which were elevated in the conditions of elevated pressure, and necroptosis markers were downregulated in astrocytes. In the IR model, elevated levels of GFAP and Iba-1 indicated glial activation. Following Bromfenac administration, levels of iNOS, COX-2, and PGE2-R were reduced, suggesting a decrease in neuroinflammation. Furthermore, Bromfenac administration in the IR model resulted in the improved survival of retinal ganglion cells (RGCs) and preservation of retinal function, as demonstrated by immunohistochemical staining and electroretinography. In summary, Bromfenac proved effective in diminishing neuroinflammation and resulted in enhanced RGC survival.

Tracing the influence of prenatal risk factors on the offspring retina: Focus on development and putative long-term consequences.

Pregnancy represents a window of vulnerability to fetal development. Disruptions in the prenatal environment during this crucial period can increase the risk of the offspring developing diseases over the course of their lifetime. The central nervous system (CNS) has been shown to be particularly susceptible to changes during crucial developmental windows. To date, research focused on disruptions in the development of the CNS has predominantly centred on the brain, revealing a correlation between exposure to prenatal risk factors and the onset of neuropsychiatric disorders. Nevertheless, some studies indicate that the retina, which is part of the CNS, is also vulnerable to in utero alterations during pregnancy. Such changes may affect neuronal, glial and vascular components of the retina, compromising retinal structure and function and possibly impairing visual function. A search in the PubMed database was performed, and any literature concerning prenatal risk factors (drugs, diabetes, unbalanced diet, infection, glucocorticoids) affecting the offspring retina were included. This review collects evidence on the cellular, structural and functional changes occurring in the retina triggered by maternal risk factors during pregnancy. We highlight the adverse impact on retinal development and its long-lasting effects, providing a critical analysis of the current knowledge while underlining areas for future research. Appropriate recognition of the prenatal risk factors that negatively impact the developing retina may provide critical clues for the design of preventive strategies and for early therapeutic intervention that could change retinal pathology in the progeny.

Formyl Peptide Receptor 1 Inhibits Reparative Angiogenesis and Aggravates Neuroretinal Dysfunction in Ischemic Retinopathy

Purpose Ischemic retinopathy is the major cause of vision-threatening conditions. Inflammation plays an important role in the pathogenesis of ischemic retinopathy. Formyl peptide receptor 1 (FPR1) has been reported to be implicated in the regulation of inflammatory disorders. However, the role of FPR1 in the progression of ischemic retinal injury has not been fully explained. Methods The activation of FPR1 was measured by real-time PCR and western blotting in the retina of OIR. The effect of FPR1 on the expression of inflammatory cytokines and relevant pro-angiogenic factors was assessed between wild-type and FPR1-deficiency OIR mice. The impact of FPR1 on retinal angiogenesis was evaluated through quantifying retinal vaso-obliteration and neovascularization between FPR1+/+ and FPR1–/– OIR mice. At last, the neuronal effect of FPR1 on the ischemic retina was investigated by ERG between wild-type and FPR1-deficient OIR mice. Results The expression of FPR1 significantly increased in the retina of OIR. Furthermore, FPR1 deficiency downregulated pro-inflammatory and pro-angiogenic factors. Ablation of FPR1 suppressed the retinal pathological neovascularization and promoted reparative revascularization, ultimately improving retinal neural function after ischemic injury. Conclusion In ischemic retinopathy, FPR1 aggravates inflammation and inhibits reparative angiogenesis to exacerbate neuronal dysfunction.

Raddeanin A Protects the BRB Through Inhibiting Inflammation and Apoptosis in the Retina of Alzheimer's Disease.

Neuroinflammation and endothelial cell apoptosis are prominent features of blood-brain barrier (BBB) disruption, which have been described in Alzheimer's disease (AD) and can predict cognitive decline. Recent reports revealed vascular β-amyloid (Aβ) deposits, Muller cell degeneration and microglial dysfunction in the retina of AD patients. However, there has been no in-depth research on the roles of inflammation, retinal endothelial cell apoptosis, and blood-retinal barrier (BRB) damage in AD retinopathy. We found that Raddeanin A (RDA) could improve pathological and cognitive deficits in a mouse model of Alzheimer's disease by targeting β-amyloidosis, However, the effects of RDA on AD retinal function require further study. To clarify whether RDA inhibits inflammation and apoptosis and thus improves BRB function in AD-related retinopathy. In vitro we used Aβ-treated HRECs and MIO-M1 cells, and in vivo we used 3×Tg-AD mice to investigate the effect of RDA on BRB in AD-related retinopathy. We found that RDA could improve BRB function in AD-related retinopathy by inhibiting NLRP3-mediated inflammation and suppressing Wnt/β-catenin pathway-mediated apoptosis, which is expected to improve the pathological changes in AD-related retinopathy and the quality of life of AD patients.

The Role of CCL Chemokines in Experimental Staphylococcus aureus Endophthalmitis.

To test the hypothesis that (C-C motif) ligand 2 (CCL2) and CCL3 impact retinal function decline and inflammation during Staphylococcus aureus endophthalmitis. Experimental endophthalmitis was initiated by intravitreal injection of 5000 colony-forming units of S. aureus into the eyes of C57BL/6J, CCL2-/-, or CCL3-/- mice. At 12 and 24hours post-infection, retinal function, bacterial load, and myeloperoxidase levels were quantified. During S. aureus endophthalmitis, we observed a significant improvement in retinal function in CCL2-/- mice relative to C57BL/6J mice at 12hours but not at 24hours. In CCL3-/- mice, retinal function was significantly improved relative to C57BL/6J mice at 12 and 24hours. The absence of CCL2 did not alter intraocular S. aureus intraocular concentrations. However, CCL3-/- mice had significantly lower intraocular S. aureus at 12hours but not at 24hours. No difference in myeloperoxidase levels was observed between C57BL/6J and CCL2-/- mice at 12hours. CCL3-/- mice had almost no myeloperoxidase at 12hours. At 24hours, increased myeloperoxidase was observed in CCL2-/- and CCL3-/- mice relative to C57BL/6J mice. Although the absence of CCL2 resulted in improved retinal function retention at 12hours, CCL3 deficiency resulted in improved retinal function at 12 and 24hours. CCL3 deficiency, but not CCL2 deficiency, resulted in almost no inflammation at 12hours. However, at 24hours, the absence of CCL2 or CCL3 resulted in significantly increased inflammation. These results suggest that, although both CCL2 and CCL3 impact intraocular infection outcomes, CCL3 may have a more significant impact in S. aureus endophthalmitis.

Influencing Factors on Pupillary Light Responses as a Biomarker for Local Retinal Function in a Large Normative Cohort.

Investigating influencing factors on the pupillary light response (PLR) as a biomarker for local retinal function by providing epidemiological data of a large normative collective and to establish a normative database for the evaluation of chromatic pupil campimetry (CPC). Demographic and ophthalmologic characteristics were captured and PLR parameters of 150 healthy participants (94 women) aged 18 to 79 years (median = 46 years) were measured with L-cone- and rod-favoring CPC protocols. Linear-mixed effects models were performed to determine factors influencing the PLR and optical coherence tomography (OCT) data were correlated with the pupillary function volume. Relative maximal constriction amplitude (relMCA) and latency under L-cone- and rod-favoring stimulation were statistically significantly affected by the stimulus eccentricity (P < 0.0001, respectively). Iris color and gender did not affect relMCA or latency significantly; visual hemifield, season, and daytime showed only minor influence under few stimulus conditions. Age had a statistically significant effect on latency under rod-specific stimulation with a latency prolongation ≥60 years. Under photopic and scotopic conditions, baseline pupil diameter declined significantly with increasing age (P < 0.0001, respectively). Pupillary function volume and OCT data were not correlated relevantly. Stimulus eccentricity had the most relevant impact on relMCA and latency of the PLR during L-cone- and rod-favoring stimulation. Latency is prolonged ≥60 years under scotopic conditions. Considering the large study collective, a representative normative database for relMCA and latency as valid readout parameters for L-cone- and rod-favoring stimulation could be established. This further validates the usability of the PLR in CPC as a biomarker for local retinal function.

Inner limiting Membrane Peel Extends In vivo Calcium Imaging of Retinal Ganglion Cell Activity Beyond the Fovea in Non-Human Primate.

High resolution retinal imaging paired with intravitreal injection of a viral vector coding for the calcium indicator GCaMP has enabled visualization of activity dependent calcium changes in retinal ganglion cells (RGCs) at single cell resolution in the living eye. The inner limiting membrane (ILM) is a barrier for viral vectors, restricting transduction to a ring of RGCs serving the fovea in both humans and non-human primates (NHP). We evaluate peeling the ILM prior to intravitreal injection as a strategy to expand calcium imaging beyond the fovea in the NHP eye in vivo. Five Macaca fascicularis eyes (age 3-10y; n=3 individuals; 2M, 1F) underwent vitrectomy and 5 to 6-disc diameter ILM peel centered on the fovea prior to intravitreal delivery of 7m8:SNCG:GCaMP8s. Calcium responses from RGCs were recorded using a fluorescence adaptive optics scanning laser ophthalmoscope. In all eyes GCaMP was expressed throughout the peeled area, representing a mean 8-fold enlargement in area of expression relative to a control eye. Calcium recordings were obtained up to 11 degrees from the foveal center. RGC responses were comparable to the fellow control eye and showed no significant decrease over the 6 months post ILM peel, suggesting that RGC function was not compromised by the surgical procedure. In addition, we demonstrate that activity can be recorded directly from the retinal nerve fiber layer. This approach will be valuable for a range of applications in visual neuroscience including pre-clinical evaluation of retinal function, detecting vision loss, and assessing the impact of therapeutic interventions.