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A. Wenzel, N. Tanimoto, V. Oberhauser, C. Grimm, M.W. Seeliger, J. von Lintig, C.E. Remé; Role of RPE65 for Cone Function in NRL Deficient Mice . Invest. Ophthalmol. Vis. Sci. 2006;47(13):2041.
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Within the retinal pigment epithelium (RPE), RPE65 catalyzes the conversion of all–trans retinyl esters into 11–cis retinol, a key step of the visual cycle responsible for pigment regeneration in the rod system. Several lines of evidence indicate that the cone system may utilize different routes, possibly involving Mueller glia, to re–isomerize a bleached chromophore. Furthermore, RPE65 may be expressed in cones and therefore could be involved in cone–specific pathways of chromophore regeneration. Using NRL deficient mice, a mouse model with exclusively cone photoreceptors, we analyzed the potential role of RPE65 for cone vision.
Proteins were quantified on Western blots using a Licor Odyssey scanner. Immunohistochemistry was performed on cryostat sections. Morphology was assessed on Epon sections. Retinoids were analyzed in extracts from retina and eyecup by HPLC. ERGs were recorded after stimulation with single flashes or with flicker stimulation.
In isolated retinas of Nrl–/– mice RPE65 immunoreactivity (IR) on Western blots was 5–fold elevated as compared to wild type mice. No difference in the amount of RPE65 was observed when eyecups (containing RPE but no retina) were compared. CRALBP–IR in Mueller cells was elevated 2–fold in the retina and slightly reduced in the eyecup. The Mueller cell markers GFAP and Glutamine Synthase were elevated about 3– and 2–fold, respectively in the retina of NRL deficient mice. Comparing Nrl–/– mice with mice double deficient for Nrl and Rpe65, showed that lack of RPE65 caused: 1) an extreme desensitization of the retina as indicated by a 3–log unit reduction of ERG sensitivity 2) 30–fold elevated amounts of retinyl esters, restricted to the RPE 3) amounts of 11–cis retinal below the limit of detection, and 4) detectable levels of 9–cis retinal.
While elevated levels of RPE65 and Mueller cell markers might indicate that cones in the NRL deficient mouse may in fact utilize pathways for retinoid processing different from rods, the knock out of Rpe65 in Nrl–/– mice affects their cones in a way very similar to that described for rods in Rpe65–/– mice: No detectable 11–cis retinal, increased 9–cis retinal, accumulation of retinyl esters and a vast loss of light sensitivity. Collectively, these data indicate that RPE65 does play similar roles in chromophore regeneration of the rod and the cone system.
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