Recently, a number of laboratories have been interested in developing cell lines of ocular tissues to understand the pathogenesis of ocular diseases. Toward this end, we report here the generation of cell lines of human retina by transfection of simian virus SV40 T antigen gene. Established retinal cells grow as a monolayer and exhibit limited serum dependence. Phase-contrast and electron microscopic studies revealed distinct morphological cell types. Immunofluorescence studies showed that the established retinal cells were positive for neuron-specific enolase, neurofilament protein, glycine receptor, synaptophysin, and secretogranin. Cells were negative for glial fibrillary acidic protein, glutamine synthetase, galactocerebroside, and carbonic anhydrase II. In addition to neuronal features, a small percentage of flat cells were, however, positive for cellular retinaldehyde binding protein, and cells with the phenotype of rod and cone photoreceptor coexpressed opsin and interphotoreceptor retinoid-binding protein. An important feature of this cell line is that addition of phorbol ester and cAMP induced dramatic changes, with 100% of the cells extending long, thin neuritic processes. Thus, the established retinal cells would be useful for studies dealing with differentiation and plasticity of the cells of the nervous system.