Interphotoreceptor retinoid-binding protein (IRBP) promotes the release of all-trans retinol from the isolated retina following rhodopsin bleaching illumination

All-trans retinol generated in rod photoreceptors upon the bleaching of rhodopsin is known to move from the rods to the retinal pigment epithelium (RPE), where it is enzymatically converted to 11-cis retinal in the retinoid visual cycle. Interphotoreceptor retinoid-binding protein (IRBP) contained in the extracellular compartment (interphotoreceptor matrix) that separates the retina and RPE has been hypothesized to facilitate this movement of all-trans retinol, but the precise role of IRBP in this process remains unclear. To examine the activity of IRBP in the release of all-trans retinol from the rods, initially dark-adapted isolated retinas obtained from toad (Bufo marinus) eyes were bleached and then incubated in darkness for defined periods (5-180 min) in physiological saline (Ringer solution) supplemented with IRBP (here termed 'IRBP I') at defined concentrations (2-90 μm). Retinoids present in the retina and extracellular medium were then determined by extraction and HPLC analysis. Preparations incubated with ≥10 μm IRBP I showed a pronounced release of all-trans retinol with increasing period of incubation. As determined with 25 μm IRBP I, the increase of all-trans retinol in the extracellular medium was accompanied by a significant decrease in the combined amount of all-trans retinal and all-trans retinol contained in the retina. This effect was not mimicked by unsupplemented Ringer solution or by Ringer solution containing 25 or 90 μm bovine serum albumin. However, incubation with 'IRBP II', a previously described variant of IRBP with altered lectin-binding properties, led to the appearance of substantial all-trans retinol in the extracellular medium. The results suggest that in vivo, IRBP plays a direct role in the release of all-trans retinol from the rods during operation of the visual cycle.