INSULIN STIMULATION OF INTRACELLULAR FREE CA2+ RECOVERY AND CA2+-ATPASE GENE-EXPRESSION IN CULTURED VASCULAR SMOOTH-MUSCLE CELLS - ROLE OF GLUCOSE-6-PHOSPHATE

We have previously reported that insulin accelerates recovery of intracellular Ca2+ concentrations ([Ca2+](i)) from pressor agonist-induced Ca2+ loads and stimulates both plasmalemmal and sarcoplasmic-reticulum Ca2+-ATPase gene expression in cultured and freshly isolated vascular smooth-muscle cells (VSMCs), suggesting that insulin attenuation of vascular tone may result from modulation of [Ca2+](i). Accordingly, we have now evaluated the linkage between this insulin-regulation of VSMC [Ca2+](i) and classical actions of insulin (i.e. glucose transport and metabolism). Cultured VSMCs were incubated in the presence or absence of insulin in a medium containing either pyruvate, glucose, 3-O-methylglucose or 2-deoxyglycose. Insulin caused an 87% increase in [Ca2+](i) recovery rate after stimulation with arginine-vasopressin (P < 0.01) and caused a marked increase in Ca2+-ATPase mRNA and protein levels in the presence of glucose. Comparable increases in both [Ca2+](i) recovery and Ca2+-ATPase expression were found when glucose was replaced by 2-deoxyglucose. In contrast, no stimulation was found in either the glucose-free or 3-O-methylglucose-containing medium. As both glucose analogues are transported, but only a-deoxyglucose is phosphorylated, this indicates that glucose transport and metabolism to glucose 6-phosphate is essential for insulin regulation of VSMC [Ca2+](i), possibly via a glucose-6-phosphate-dependent carbohydrate-response element in the Ca2+-ATPase gene.