Involvement of protein kinase C in the regulation of Na+/Ca2+ exchanger in bovine adrenal chromaffin cells.

1. The Na(+)/Ca(2+) exchanger (NCX) exchanges Na+ and Ca(2+) bidirectionally through the forward mode (Ca(2+) extrusion) or the reverse mode (Ca(2+) influx). The present study was undertaken to clarify the role of protein kinase C (PKC) in the regulation of NCX in bovine adrenal chromaffin cells. The Na(+)-loaded cells were prepared by treatment with 100 micromol/L ouabain and 50 micromol/L veratridine. Incubation of Na(+)-loaded cells with Na(+)-free solution in the presence of the Ca(2+) channel blockers nicardipine (3 micromol/L) and omega-conotoxin MVIIC (0.3 micromol/L) caused Ca(2+) uptake and catecholamine release. 2. The Na(+)-dependent Ca(2+) uptake and catecholamine release were inhibited by 2-[4-[(2,5-difluorophenyl)methoxy]phenoxy]-5-ethoxyaniline (SEA0400; 1 micromol/L) and 2-[2-[4-(4-nitrobenzyloxy)phenyl]isothiourea (KB-R7943; 10 micromol/L), both NCX inhibitors. These results indicate that the Na(+)-dependent responses are mostly due to activation of the NCX working in the reverse mode. 3. In addition, we examined the effects of PKC inhibitors and an activator on the NCX-mediated Ca(2+) uptake and catecholamine release. Bisindolylmaleimide I (0.3-10 micromol/L) and chelerythrine (3-100 micromol/L), both PKC inhibitors, inhibited NCX-mediated responses. In contrast, phorbol 12,13-dibutyrate (0.1-10 micromol/L), a PKC activator, enhanced the responses. Bisindolylmaleimide I and chelerythrine, at effective concentrations for inhibition of Na(+)-dependent catecholamine release, had a little or no effect on high K(+)-induced catecholamine release in intact cells or on Ca(2+)-induced catecholamine release in beta-escin-permeabilized cells. 4. These results suggest that PKC is involved in the activation of NCX in bovine adrenal chromaffin cells.