ABSTRACT Vision is one of the fundamental senses that enables the perception of the surrounding environment. In humans, this process is initiated in the retina, which is lined with millions of photoreceptors expressing opsins bound to the visual chromophore, 11-cis-retinal (11-cis-RAL). Absorption of a photon by the opsin-bound visual chromophore causes its isomerization to an all-trans configuration. This initial photochemical reaction triggers the activation of a signal transduction cascade that eventually leads to the transmission of visual information to the brain, while the activated opsin is rendered insensitive to further light stimulation. Sustained vision thus requires continuous renewal of the visual chromophore following light exposure, a process thought to be accomplished through the canonical retinoid (visual) cycle. Here, we propose to explore a non-canonical complementary pathway for the photic generation of 11-cis-RAL. Our long-term goal is to obtain a better understanding of the retinoid cycle and other pathways capable of regenerating the visual chromophore and to develop interventions that stop the progression of human retinal diseases related to these metabolic events. Accordingly, we propose four thematically and experimentally linked Specific Aims to: 1) improve our mechanistic insights regarding a photochemical reaction catalyzed by retinal pigment epithelium (RPE)-retinal G protein-coupled receptor (RGR) that can generate 11-cis-RAL. We will determine the quantum yield (Φ) of recombinant RGR and compare its rate of photoisomerization with the activity of RPE65, the enzymatic isomerase of the RPE visual cycle. We also will test the influence of retinol dehydrogenase (RDH) and cellular- retinaldehyde-binding protein (CRALBP encoded by Rlbp1 gene) as modulators of the RGR-based photoisomerase activity; 2) determine the role of RGR in RPE and Müller cells (MC). To accomplish this objective, we will investigate conditional Rgr-/- mice that permit the temporal regulation of the expression of RGR; 3) establish the contribution of RPE and MC separately to both the visual cycle as well as the photochemical production of 11-cis-RAL using the traceable artificial chromophore 9-cis-RAL; and 4) establish the role of CRALBP in RPE and MC during the regeneration of rod and cone visual pigments, by employing genetic and pharmacological approaches. We will evaluate the effects of RPE/MC-specific Rlbp1 deletion on the regeneration of rod and cone visual pigments, along with photoreceptor function in the presence and absence of potent inhibitors of RPE65. In summary, recent advances pertaining to the photic generation of 11-cis-RAL in the RPE have added a new and novel physiological element to our understanding of the cellular pathways capable of regenerating the visual chromophore. However, these advances have yet to be fully integrated with what we know about the canonical visual cycle. Here, we will provide a more unified perspective o...