Methods and Results-We examined the effect of insulin on AT(1) receptor gene expression in cultured vascular smooth muscle cells (VSMCs). A 24-hour incubation with insulin (100 nmol/L) produced a 2-fold increase in AT(1) receptor density on VSMCs, as assessed by radioligand binding assays. This enhanced AT(1) receptor expression was caused by a time- and concentration-dependent upregulation of the AT(1) receptor mRNA levels, as assessed by Northern analysis. The maximal effect was detected after a 24-hour incubation of cells with 100 nmol/L insulin (270+/-20%). AT(1) receptor upregulation was caused by a stabilization of the AT(1) receptor mRNA, because the AT(1) receptor mRNA half-life was prolonged from 5 hours under basal conditions to 10 hours after insulin stimulation. In contrast, insulin had no influence on AT(1) receptor gene transcription, as assessed by nuclear run-on assays. The insulin-induced AT(1) receptor upregulation was followed by an increased functional response, because angiotensin II evoked a significantly elevated intracellular release of calcium in cells that were preincubated with 100 nmol/L insulin for 24 hours. The insulin-induced AT(1) receptor upregulation was dependent on tyrosine kinases, as assessed by experiments with the tyrosine kinase inhibitor genistein. Furthermore, experiments using the intracellular calcium chelator bis(2-amino-5-methylphenoxy)ethane-N,N,N',N'-tetraacetic acid tetraacetoxymethyl ester suggest that intracellular calcium release may be involved in AT(1) receptor regulation.

Conclusions-Insulin-induced upregulation of the AT(1) receptor by posttranscriptional mechanisms may explain the association of hyperinsulinemia with hypertension and arteriosclerosis, because activation of the AT(1) receptor plays a key role in the regulation of blood pressure and fluid homeostasis.