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J. M. Gross, R. Nuckels, A. Ng; The Vacuolar-ATPase Complex Is Required for the Survival of Retinal and RPE Cells in the Zebrafish Eye. Invest. Ophthalmol. Vis. Sci. 2007;48(13):4488.
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Zebrafish have become increasingly useful as animal models of human visual system disorders owing largely to the ability to perform forward genetic screens and rapidly identify the affected loci. In a retrovirus-based insertional mutagenesis screen we identified six recessive mutants that presented with pigmentation deficits and severe degeneration of the retina and RPE. Each of the mutations disrupted a distinct subunit of the vacuolar-ATPase complex (v-ATPase), or proteins that associate with the v-ATPase. The v-ATPase plays key roles in the acidification of lysosomes and endosomes in numerous biological contexts; however, its role in the eye has not been well studied. Here, we sought to characterize the ocular defects in zebrafish v-ATPase mutants and to determine if they represent useful animal models for studying degenerative conditions of the human eye.
Histological and transmission electron microscopic characterizations of homozygous recessive and heterozygous v-ATPase mutants were performed at several time points. Molecular studies assessed retinal organization, photoreceptor morphogenesis, apoptosis and cell proliferation. Wild-type mRNA distribution was determined by in situ hybridization.
Recessive zebrafish v-ATPase mutants possess numerous apoptotic cells throughout their retinas, with concentrated regions in the outer nuclear layer, ciliary marginal zone and RPE. Photoreceptor outer segment structure is abnormal and large membranous inclusions are present in the RPE suggesting an inability of RPE cells to degrade ingested outer segments. Melanosome formation is also compromised. Preliminary observations suggest that drusen-like deposits may be accumulating in the eyes of aging heterozygous v-ATPase mutants.
The v-ATPase complex is required for cell survival in the zebrafish retina and RPE. Within the RPE, the v-ATPase complex likely functions cell-autonomously to facilitate the pH-dependent degradation of ingested photoreceptor outer segments. Defects in v-ATPase activity results in an accumulation of undigested outer segment material and this might contribute to the RPE and photoreceptor atrophy observed in these mutants.
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