Effects of phorbol esters (stimulators of protein kinase C, PK-C) on the L-type Ca2+ current (ICa) and the intracellular Ca2+ level ([Ca]i) were investigated using single rabbit sino-atrial (SA) node cells. 2. In whole-cell patch-clamp experiments, both phorbol esters (1 microM). 4-beta-phorbol-12,13-dibutyrate (PDB) and 12-O-tetradecanoyl-phorbol-13-acetate (TPA), inhibited the ICa. Phorbol esters did not affect the voltages of half-maximum activation and inactivation for ICa. 3. The time-course of inactivation phase for ICa was two exponentials. The fast component was decreased, whereas the slow component was increased by phorbol esters. 4. Addition of H-7 (10 microM), an inhibitor of PK-C, did not antagonize, but rather increased both components. In contrast, 4-alpha-phorbol-12,13-didecanoate (PDD), an inactive analog, did not affect the ICa amplitude and the inactivation process. 5. The phorbol esters elicited a transient inward current by repetitive clamp pulses. 6. In SA node cells loaded with fura-2 (a Ca(2+)-sensitive fluorescent dye), both TPA and PDB at 0.3 microM in the presence of isoproterenol (ISP, 0.2 microM) elevated the [Ca]i. PDD did not affect [Ca]i. 7. These results indicate that PK-C stimulation by phorbol esters elevates [Ca]i (which is potentiated by ISP), possibly resulting in cellular calcium overload.
