Downregulation of the AT(1A) receptor by pharmacologic concentrations of angiotensin-(1-7)

Angiotensin (Ang)-(1-7), the amino terminal heptapeptide fragment of Ang II , is an endogenous Ang peptide with vasodilatory and antiproliferative acti ons. Because Ang II causes vasoconstriction and promotes growth through act ivation of Ang type 1 (AT(1)) receptors, we investigated whether the action s of Ang-(1-7) are due to its regulation of these receptors. Studies were p erformed in CHO cells stably transfected with the AT(1A) receptor. Ang-(1-7 ) competed poorly with [I-125]-Ang II for the AT(1A) binding site and was i neffective at shifting the ICS, for Ang II competition with [I-125]-Ang II for binding to the AT(1A) receptor. However, if CHO-AT(1A) cells were pretr eated with Ang-(1-7) and then treated with acidic glycine to remove surface -bound ligand, the heptapeptide caused a concentration-dependent reduction in Ang II binding, with a maximal inhibition to 67.8 +/- 4.6% of total (p < 0.05) at 1 <mu>M Ang-(1-7) compared with a reduction to 24% of total by 10 nM Ang II. Ang-(1-7) pretreatment caused a small but significant decrease in the affinity of [I-125]-Ang II for the AT(1A) receptor and a significant reduction in the total number of binding sites. The Ang-(1-7)-induced redu ction in binding was rapid (occurring as early as 5 min after exposure to t he peptide), was maintained for 30 min during continued exposure of the cel ls to Ang-(1-7), and rapidly recovered after removal of the heptapeptide. T he AT(1) receptor antagonist L-158,809 reduced the Ang-(1-7)-induced downre gulation of the AT(1A) receptor, suggesting that interactions with AT(1A) r eceptors mediate the regulatory events. Pretreatment with 1 muM or 10 muM A ng-(1-7) significantly reduced inositol phosphate production in response to 10 nM Ang II. The decrease in binding and responsiveness of the AT(1A) rec eptor after exposure to micromolar concentrations of Ang-(1-7) suggests tha t the heptapeptide downregulates the AT(1A) receptor to reduce responses to Ang II. Because downregulation of the receptor only occurred at micromolar concentrations of the heptapeptide, our findings suggest that Ang-(1-7) is not a potent antagonist at the AT(1A) receptor. However, when the balance between Ang II and Ang-(1-7) is shifted in favor of Ang-(1-7), such as duri ng inhibition of Ang-converting enzyme, some contribution of this mechanism may come into play.