In rat pinealocytes, protein kinase C (PKC) is involved in the α1-adrenergic-mediated potentiation of β-adrenergic-stimulated cyclic nucleotide responses; however, the specific PKC isozyme(s) involved in the potentiation mechanism remain unknown. In the present study, we compared the effects of two PKC inhibitors, calphostin C, a specific inhibitor of PKC, and Gö6976, a selective inhibitor of PKCα and PKCβ1, on the adrenergic-stimulated cyclic nucleotide accumulation in rat pinealocytes. Surprisingly, Gö6976 was found to have an enhancing effect on basal cyclic GMP and isoproterenol-stimulated cyclic AMP and cyclic GMP accumulation, an effect not shared by calphostin C. Gö6976 also increased the norepinephrine- and ionomycin-induced potentiation of isoproterenol-stimulated cyclic AMP and cyclic GMP accumulation, whereas the effect of calphostin C was inhibitory. The enhancing effect of Gö6976 was abolished in the presence of isobutylmethylxanthine or zaprinast, but not rolipram, suggesting that this effect of Gö6976 may be mediated through type V or the retinal type of phosphodiesterase. Based on these observations, we propose that some of the PKC isozyme(s) inhibited by calphostin C are involved in the potentiation of β-adrenergic-stimulated cyclic nucleotide responses and that they act by enhancing synthesis. However, PKC isozymes inhibited by Gö6976 appear to be basally active and tonically inhibit cyclic nucleotide accumulation through their stimulatory action on phosphodiesterase.