Elevation of intracellular Ca2+ concentration by protein kinase C stimulation in isolated single rabbit sino-atrial node cells.
1. 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.
Department of Pharmacology, Nara Medical University, Japan.
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Satoh
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism; D004594:Electrophysiology; D016257:Fura-2 / Q000494:pharmacology; D066298:In Vitro Techniques; D007545:Isoproterenol / Q000494:pharmacology; D007546:Isoquinolines / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010879:Piperazines / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011817:Rabbits; D012849:Sinoatrial Node / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D013268:Stimulation, Chemical; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D007546:Isoquinolines; D010879:Piperazines; D015240:Phorbol 12,13-Dibutyrate; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D011493:Protein Kinase C; D007545:Isoproterenol; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D016257:Fura-2