HIGH GLUCOSE ATTENUATES PEPTIDE AGONIST-EVOKED INCREASES IN CYTOSOLIC-FREE [CA2-MUSCLE CELLS(] IN RAT AORTIC SMOOTH)

Incubation of cultured rat aortic smooth muscle cells (ASMCs) in a med ium containing high glucose concentrations (25 mM) did not affect the basal cytosolic free calcium ([Ca2+](i)) but led to significant reduct ions in peak [Ca2+](i) response evoked by arginine vasopressin, angiot ensin II, and endothelin-1 (ET-1). This was observed in both the prese nce and absence of extracellular Ca2+ Maintenance of rat ASMCs in a me dium containing mannose (an osmotic control for high glucose) did not affect either the basal or peptide agonist-evoked increase in [Ca2+](i ). However, pretreatment with either the nonselective protein kinase C (PKC) inhibitor staurosporine or the selective PKC inhibitor 2,6-diam ino-N-([1-(1-oxotridecyl)-2 piperidinyl] methyl) hexanamide reversed t he attenuating effect of high glucose on peak [Ca2+](i) response evoke d by ET-1. Also, short-term incubation of ASMCs with the active phorbo l ester, phorbol 12-myristate 13-acetate, led to a reduction in peak [ Ca2+](i) response to all three agonists, whereas the inactive phorbol ester, 4 alpha-phorbol 12,l3-didecanoate, which does not activate PKC, had no such effect. Although high-glucose treatment of rat ASMCs led to significant reductions in the maximal number of binding sites to th e extent of 39% of [I-125]ET-1 specific binding, no significant differ ences in the affinity (K-d similar to 110 pM) characteristics were evi dent between control and high-glucose treatment groups. It is proposed that incubation of rat ASMCs with high glucose enhances the de novo s ynthesis of diacylglycerol and activates membrane-bound PKC and that t his, in turn, impairs agonist-mediated intracellular Ca2+ mobilization .