Protein kinase C mediates insulin-inhibited Ca2+ transport and contraction of vascular smooth muscle.
Insulin acutely inhibits contraction of primary cultured vascular smooth muscle (VSM) cells from canine femoral artery by inhibiting contractile agonist-induced Ca2+ influx. Insulin also inhibits contraction at step(s) distal to intracellular Ca2+ concentration (Ca2+i) by stimulating cyclic guanosine monophosphate (GMP) production. We wished to see whether these effects of insulin are mediated by protein kinase C (PKC). Ca2+ influx was assessed by measuring the rate of fluorescence quenching of intracellular fura 2 by extracellular Mn2+. We found that 10 micromol/L serotonin (5-HT) stimulated Mn2+ influx 3-fold, and 1 nmol/L insulin inhibited the 5-HT-stimulated component of Mn2+ influx by 63% (P < .05), but insulin had no effect in the presence of 1 micromol/L staurosporine, an inhibitor of PKC. In the absence of insulin, preincubating cells with 0.1 micromol/L phorbol 12-myristate 13-acetate (PMA) for 5 min inhibited the 5-HT-stimulated component of Mn2+ influx by 69% (P < .05). Insulin inhibited cell contraction induced by raising Ca2+i to supraphysiologic levels with ionomycin by 75% (P < .05). We also noted that 10(-6) mol/L calphostin C, another PKC inhibitor, or 16-h preincubation with PMA completely blocked this effect of insulin. Finally, 10-min exposure to insulin or PMA increased cyclic GMP production in ionomycin-treated cells by 50% and 64%, respectively (both P < .05). We conclude that insulin inhibits VSM cell contraction by inhibiting 5-HT-stimulated Ca2+ influx and also at step(s) distal to Ca2+i by a PKC-dependent mechanism.
Kahn
A M
AM
Department of Medicine, The University of Texas Health Science Center, Houston 77225, USA.
Allen
J C
JC
Seidel
C L
CL
Song
T
T
D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D002273:Carcinogens / Q000494:pharmacology; D006152:Cyclic GMP / Q000378:metabolism; D004285:Dogs; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D005263:Femoral Artery / Q000201:enzymology; D005456:Fluorescent Dyes; D016166:Free Radical Scavengers / Q000494:pharmacology; D016257:Fura-2; D007004:Hypoglycemic Agents / Q000494:pharmacology*; D007328:Insulin / Q000494:pharmacology*; D008297:Male; D008345:Manganese / Q000494:pharmacology; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000201:enzymology*; D009281:Naphthalenes / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012701:Serotonin / Q000494:pharmacology; D019311:Staurosporine / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014661:Vasoconstriction / Q000187:drug effects / Q000502:physiology
D002273:Carcinogens; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D016166:Free Radical Scavengers; D007004:Hypoglycemic Agents; D007328:Insulin; D009281:Naphthalenes; D012701:Serotonin; D008345:Manganese; D011493:Protein Kinase C; D006152:Cyclic GMP; D019311:Staurosporine; C058819:calphostin C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D016257:Fura-2