SYNERGISTIC ANTIPROLIFERATIVE ACTIONS OF CYCLIC ADENOSINE-3',5'-MONOPHOSPHATE, INTERLEUKIN-1-BETA, AND ACTIVATORS OF CA2+ CALMODULIN-DEPENDENT PROTEIN-KINASE IN PRIMARY HEPATOCYTES/

cAMP and Ca2+ acted together with the acute phase cytokine interleukin -1 beta (IL-1 beta) to inhibit hepatocyte DNA replication. At sub-basa l activity of cAMP-dependent protein kinase (PKA), neither IL-1 beta n or the Ca2+-elevating hormone vasopressin affected hepatocyte prolifer ation. Basal level of PKA activity permitted IL-1 beta action. Increas ed PKA activity also permitted vasopressin action and sensitized furth er towards IL-1 beta, which acted at 10-50 pM concentrations. Vasopres sin acted via Ca2+/calmodulin-dependent protein kinase II (CaMKII), an d its action was mimicked by the serine/threonine phosphatase inhibito r microcystin, which activates CaMKII. Inhibitors (KN93 and KT5926) of CaMKII selectively counteracted the effects of vasopressin and microc ystin on hepatocyte proliferation at concentrations similar to those r equired to inhibit CaMKII in vitro. Two-dimensional gel electrophoresi s of P-32-prelabeled hepatocytes revealed a common set of proteins pho sphorylated in response to vasopressin and microcystin. Their phosphor ylation was counteracted by CaMKII inhibitor (KT5926). Phosphorylation of the CaMKII substrate phenylalanine hydroxylase (PAH; EC 1.14.16.1) was used as an endogenous marker of CaMKII activation. It was found t hat treatment of the cells with vasopressin or microcystin increased t he phosphorylation of PAH, and that the vasopressin-induced PAH phosph orylation was inhibited by KT5926. In conclusion, the Ca2+-elevating h ormone vasopressin potentiated the antiproliferative effects of cAMP a nd IL-1 beta through CaMKII activation.