Adenylate cyclase activity in rabbit retinal homogenates can be stimulated directly by forskolin or through a receptor-mediated mechanism by vasoactive intestinal peptide (VIP). In contrast the alpha 2-adrenoceptor agonists clonidine and UK-14,304 reduce the basal cAMP level slightly. This was more evident following application of forskolin and VIP where the decrease of cAMP caused by clonidine and UK-14,304 is dose-dependent. The alpha 2-adrenoceptor agonist response is blocked by pertussis toxin and is insensitive to the phosphodiesterase inhibitor, isobutylmethylxanthine, suggesting the involvement of a Gi-protein. Clonidine and UK-14,304 attenuation of elevated cAMP levels can be inhibited by the alpha 2-receptor antagonist yohimbine and phentolamine but not by the specific alpha 1-receptor antagonist, prazosin. Serotonergic, cholinergic and beta-adrenergic receptor antagonists were without effect. The results demonstrate that alpha 2-adrenergic receptors in the retina exert inhibitory effects on adenylate cyclase activity mediated by an inhibitory guanine nucleotide regulating protein.