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Aisling Lynch, Lina Sprogyte, Ma'ayan Semo, Amanda-Jayne Francis Carr, Anthony Vugler; RPE65 Fuels the intrinsic pupillary light reflex in mice. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):1710.
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© ARVO (1962-2015); The Authors (2016-present)
The rd12 mouse is a naturally occurring model of Leber’s congenital amarosis type 2 (LCA2) in which a premature stop codon prevents translation of the RPE65 protein. Here we sought to explore residual light responses in these animals using a melanopsin-sensitive assay of behavioural light aversion (BLA), electroretinography (ERG), functional anatomy and in vitro pupillometry.
Adult rd12 and C57 mice of various ages were used in this study. Photoreceptor function was assessed by scotopic and photopic ERG and melanopsin-driven light aversion was measured by BLA for 30 minutes in a light:dark choice box. The intrinsic pupillary light reflex (iPLR) was examined in the enucleated eyes of dark-adapted mice using an established method (Vugler et al., 2014). Whole retinas from other animals were processed by immunohistochemistry for melanopsin, Brn3A, tyrosine hydroxylase (TH) and cfos. For functional anatomy with cfos, animals were either killed in darkness, or following 2h of ambient light.
The scotopic ERG was dramatically reduced in rd12 mice, with a clear but significantly reduced photopic ERG also present. However, this ERG response was insufficient to activate the retinal dopamine system in rd12 mice, as assessed by an absence of light-induced cfos expression in TH cells. Melanopsin expression was also degraded in rd12 retinas, with up to 40% fewer melanopsin cells than C57 mice depending upon age and retinal region. In rd12 mice, there was also a clear reduction in the intensity of melanopsin staining in ipRGC somas and a dramatic decline in ipRGC neurite staining. However, the number of Brn3a positive RGCs appeared stable in the rd12 retina and light-induced cfos was still observed in ipRGCs, with the majority of light-induced activity found in ventral-nasal retina. The rd12 mice showed an absence of BLA and only a tiny, transient iPLR in response to the brightest light. However, this residual iPLR could be fully restored following supplementation with 9-cis-retinal.
We show here that the reduced levels of melanopsin/outer retinal function in rd12 mice are insufficient to drive BLA and that RPE65 is the likely source of retinal for the iPLR. This may involve a previously unrecognised role for RPE65 in the pigmented cells of iris/ciliary body. As residual ERGs in these mice are thought to originate from rods, our data also indicates that light-driven TH cell activation in mice may be largely cone-driven.
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