Angiotensin-(1-7) downregulates the angiotensin II type 1 receptor in vascular smooth muscle cells

Angiotensin (Ang)-(1-7) is a biologically active peptide of the renin-angio tensin system that has both vasodilatory and antiproliferative activities t hat are opposite the constrictive and proliferative effects of angiotensin II (Ang II). We studied the actions of Ang-(1-7) on the Ang II type 1 (AT(1 )) receptor in cultured rat aortic vascular smooth muscle cells to determin e whether the effects of Ang-(1-7) are due to its regulation of the AT(1) r eceptor. Ang -(1-7) competed poorly for [I-125]Ang II binding to the AT(1) receptor on vascular smooth muscle cells, with an IC50 of 2.0 mu mol/L comp ared with 1.9 nmol/L for Ang II. The pretreatment of vascular smooth muscle cells with Ang-(1-7) followed by treatment with acidic glycine to remove s urface-bound peptide resulted in a significant decrease in [I-125]Ang II bi nding; however, reduced Ang II binding was observed only at micromolar conc entrations of Ang-(1-7). Scatchard analysis of vascular smooth muscle cells pretreated with 1 mu mol/L Ang-(1-7) showed that the reduction in Ang II b inding resulted from a loss of the total number of binding sites [B-max 437 .7 +/- 261.5 fmol/mg protein in Ang-(1-7)pretreated cells compared with 607 .5 +/- 301.2 fmol/mg protein in untreated cells, n=5, P <0.05] with no sign ificant effect on the affinity of Ang II for the AT(1) receptor. Pretreatme nt with the AT(1) receptor antagonist L-158,809 blocked the reduction in [I -125]Ang II binding by Ang-(1-7) or Ang II. Pretreatment of vascular smooth muscle cells with increasing concentrations of Ang-(1-7) reduced Ang II-st imulated phospholipase C activity; however, the decrease was significant (8 1.2 +/-6.4%, P <0.01, n=5) only at 1 mu mol/L Ang-(1-7). These results demo nstrate that pharmacological concentrations of Ang-(1-7) in the micromolar range cause a modest downregulation of the AT(1) receptor on vascular cells and a reduction in Ang II-stimulated phospholipase C activity. Because the antiproliferative and vasodilatory effects of Ang-(1-7) are observed at na nomolar concentrations of the heptapeptide, these responses to Ang-(1-7) ca nnot be explained by competition of Ang-(1-7) at the AT(1) receptor or Ang- (1-7)-mediated downregulation of the vascular AT(1) receptor.