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Purpose: Many older dogs (Canis lupus familiaris) develop cognitive impairment. Dog owners often describe impairments in multiple sensory functions, yet the relationships between sensory and cognitive function in older dogs is not well understood., Methods: We performed assessments of dog vision and hearing, both clinically (n = 91, electroretinography and brainstem auditory evoked potential) and via validated questionnaire (n = 238). We determined prevalence of sole and dual hearing/vision impairments in younger (<8 years) and older (≥8 years) dogs. Impairment cutoffs were determined using data from young dogs. We assessed the relationships between questionnaire-assessed vision and/or hearing impairments and cognitive impairment using logistic regression., Results: Younger and older dog groups had similar distributions of sex and purebred/mixed breed status. Sex had no relationship to prevalence of sensory impairments. Older dogs had higher prevalence of hearing, vision, and dual sensory impairments, assessed both clinically and by questionnaire (P<0.001), and cognitive impairment assessed by questionnaire (P<0.001). Dogs had higher prevalence of reported cognitive impairment when owners reported dual vision and hearing impairments (79-94%, versus 25-27% in dogs with no sensory impairments), which was most consistent in dogs aged ≥8 years. In these older dogs, dual vision/hearing impairments were associated with a significantly increased risk of cognitive impairment (1.8-2.0 odds ratio)., Conclusion: Dogs aged ≥8 years are at higher risk for dual hearing/vision impairments and associated cognitive impairments. The causal relationship between these impairments is not defined, but clinical consideration of these multimorbidity risks should be made in older dogs., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Hopper et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Dog models with spontaneously occurring mutations in retinal dystrophy genes are an invaluable resource for preclinical development of retinal gene therapy. Adeno-associated virus (AAV) vectors have been most successful; to target the outer retina and RPE they are delivered by subretinal injection, causing a temporary retinal detachment with some potential for retinal morbidity. A recent reporter gene study using an AAV2/8 vector in dogs reported transgene expression beyond the boundary of the subretinal bleb. This could be a desirable feature which increases the area of retina treated while minimizing the retinal detachment and any associated morbidity. We performed a detailed study of the lateral spread of transgene expression beyond the subretinal injection site following subretinally delivered AAV vectors in normal dogs. Vectors expressed green fluorescent protein (GFP) using a small chicken beta-actin promoter. AAV2/2 (quadruple tyrosine to phenylalanine (Y-F) capsid mutant), self-complementary (sc) AAV2/8 (single Y-F capsid mutant) and a scAAV2/5 were used. We found that in all eyes GFP expression involved retina beyond the initial post-injection subretinal bleb boundary. In all eyes there was post-injection spread of the retinal detachment within the first 3 days post procedure and prior to retinal reattachment. In 11/16 eyes this accounted for the entire "lateral spread" of GFP expression while in 5/16 eyes a very slight extension of GFP expression beyond the final boundary of the subretinal bleb could be detected. All 3 AAV constructs induced GFP expression in the nerve fiber layer with spread to the optic nerve. Patients treated by subretinal injection should be monitored for possible expansion of the subretinal injection bleb prior to reattachment. Injections in the para-foveal region may expand to lead to a foveal detachment that may be undesirable. Cell-specific promoters may be required to limit spread of expressed transgene to the brain with these AAV serotypes.

Background: Hypoxia plays a key role in ischaemic and neovascular disorders of the retina. Cellular responses to oxygen are mediated by hypoxia-inducible transcription factors (HIFs) that are stabilised in hypoxia and induce the expression of a diverse range of genes. The purpose of this study was to define the cellular specificities of HIF-1alpha and HIF-2alpha in retinal ischaemia, and to determine their correlation with the pattern of retinal hypoxia and the expression profiles of induced molecular mediators., Methodology/principal Findings: We investigated the tissue distribution of retinal hypoxia during oxygen-induced retinopathy (OIR) in mice using the bio-reductive drug pimonidazole. We measured the levels of HIF-1alpha and HIF-2alpha proteins by Western blotting and determined their cellular distribution by immunohistochemistry during the development of OIR. We measured the temporal expression profiles of two downstream mediators, vascular endothelial growth factor (VEGF) and erythropoietin (Epo) by ELISA. Pimonidazole labelling was evident specifically in the inner retina. Labelling peaked at 2 hours after the onset of hypoxia and gradually declined thereafter. Marked binding to Müller glia was evident during the early hypoxic stages of OIR. Both HIF-1alpha and HIF-2alpha protein levels were significantly increased during retinal hypoxia but were evident in distinct cellular distributions; HIF-1alpha stabilisation was evident in neuronal cells throughout the inner retinal layers whereas HIF-2alpha was restricted to Müller glia and astrocytes. Hypoxia and HIF-alpha stabilisation in the retina were closely followed by upregulated expression of the downstream mediators VEGF and EPO., Conclusions/significance: Both HIF-1alpha and HIF-2alpha are activated in close correlation with retinal hypoxia but have contrasting cell specificities, consistent with differential roles in retinal ischaemia. Our findings suggest that HIF-2alpha activation plays a key role in regulating the response of Müller glia to hypoxia.