Prostaglandins are locally produced mediators of physiologic function which act through specific receptors to influence cellular response pathways. We have conducted a series of experiments designed to obtain a preliminary pharmacologic characterization of the prostaglandin E2 receptor subtype mediating increased synthesis of adenosine 3',5'-cyclic monophosphate (cyclic AMP) in an SV40 transformed line of human non-pigmented ciliary epithelium (ODMC2). Human non-pigmented epithelial (NPE) cells were grown to confluency, preincubated with indomethacin (to prevent endogenous PGE2 synthesis) and with isobutyl-methyl-xanthine (to prevent breakdown of cAMP), challenged with PGE2 or receptor-selective prostanoids and extracted. Prostanoid-stimulated cAMP synthesis was determined using a protein binding assay and normalized to cellular protein. Prostanoid agonists known to be relatively selective for the EP2 receptor subtype stimulated cAMP synthesis by human NPE cells. EP1 and EP3, IP, FP and DP selective prostanoids had negligible effects on stimulation of cAMP synthesis. The results of these studies lead us to conclude that human NPE cells express receptors of the EP2 subtype which mediate PGE2 stimulated synthesis of cyclic AMP.