PURPOSE. The purpose of this study was to detect and establish the cellular localizations of nicotinic acetylcholine receptor (nAChR) subunits in Rhesus monkey retina. METHODS. Retinas were dissected from the eyes of monkeys killed after unrelated experiments. RNA was extracted and analyzed by RT-PCR, using primers designed against human sequences of α3-α7, α9, and β2-β4 nAChR subunits. The RTPCR products were separated by gel electrophoresis and sequenced. Frozen sections of postmortem fixed monkey eyes were immunolabeled with well-characterized and specific monoclonal antibodies against the α3, α4, α6, α7, β2, or β4 nAChR subunits and visualized with fluorescence labeling. RESULTS. Products of the predicted size for the α3-α7, α9, and β2-β4 nAChR subunits were detected by RT-PCR in Rhesus monkey retina. Homology between transcripts from monkey retina and human nucleotide sequences ranged from 93 to 99%. Immunohistochemical studies demonstrated that neurons in various cell layers of monkey retina expressed α3, α4, α7, or β2 nAChR subunits and cells with the morphology of microglia were immunoreactive for the α6 or α4 nAChR subunits. CONCLUSIONS. nAChR subunits are expressed in the monkey retina and localize to diverse retinal neurons as well as putative microglia. Besides mediating visual processing, retinal nAChRs may influence refractive development and ocular pathologies such as neovascularization. Copyright © Association for Research in Vision and Ophthalmology.