The effects of cholera toxin on rat parotid gland function were determined in order to further characterize the relationship between cyclic AMP and exocytosis in this tissue. Cholera toxin induced the release of alpha-amylase from rat parotid minces in vitro. This release was accompanied by an activation of adenylate cyclase, elevated cyclic AMP levels, an elevated protein kinase activity ratio, and changes in the degree of phosphorylation of three endogenous phosphoproteins. Two of the phosphoproteins became more phosphorylated upon cholera toxin stimulation while the phosphorylation of the other decreased. The effects of cholera toxin on endogenous phosphoprotein labelling appeared to mimic those of the beta-adrenergic agonist isoproterenol but were of a smaller magnitude. These results are consistent with cyclic AMP functioning as a major mediator of exocytosis in this gland exerting its effects, at least in part, via activation of cyclic AMP dependent protein kinase. The mechanism by which an increased cyclic AMP level results in the decreased phosphorylation of an endogenous phosphoprotein is not known.