Effects of protein kinase inhibitor (1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H7) on protein kinase C activity and adrenergic stimulation of cAMP and cGMP in rat pinealocytes

Protein kinase C is thought to be involved in the adrenergic regulation of pineal function. In this tissue, norepinephrine increases cAMP and cGMP accumulation through a synergistic dual receptor mechanism involving alpha 1- and beta-adrenergic receptors; the available evidence indicates that the alpha 1-adrenergic stimulation activates protein kinase C, and that this potentiates beta-adrenergic stimulation of pineal cAMP. The role of protein kinase C in the regulation of cGMP is unclear. In the present report, we determined whether an inhibitor of protein kinase C, 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H7), inhibits pineal protein kinase C and the adrenergic stimulation of pineal cAMP and cGMP. H7 (10(-4) M) reduced soluble protein kinase C activity by 40%. Treatment of intact pinealocytes with H7 for 0-240 min reduced the effects of subsequent norepinephrine (NE) stimulation of cAMP and cGMP accumulation by at least 25%. H7 also inhibited 25-30% the maximum stimulation of both cAMP and cGMP produced by concurrent treatment with isoproterenol and two agents which elevate intracellular Ca2+, ouabain and A23187. However, H7 did not reduce the effects of selective beta-adrenergic activation, indicating that H7 was probably inhibiting the effects of NE by blocking alpha 1-adrenergic potentiation of beta-adrenergic stimulation, not beta-adrenergically activated mechanisms. H7 also reduced the stimulation of cAMP accumulation produced by the combined treatment of isoproterenol and an activator of protein kinase C, 4-beta-phorbol 12-myristate, 13-acetate, which is consistent with the view that H7 is acting by inhibiting protein kinase C activity. These observations are in agreement with the conclusion that potentiation of beta-adrenergic stimulation of cAMP by alpha 1-adrenergic agonists, protein kinase C activators, or [Ca2+]i elevating agents involves protein kinase C. In addition, these results are of special interest because they point to the possibility that protein kinase C is involved in the regulation of cGMP accumulation.