Angiotensin-(1-7) is a novel peptide of the renin-angiotensin system that counteracts the presser and proliferative responses to angiotensin II. We now report that cultured bovine aortic endothelial cells contain a saturable, high-affinity [I-125]angiotensin-(1-7) binding site with an affinity of 19.3 +/- 10.7 nmol/L and a density of 1351 +/- 710 fmol/mg protein. Angiotensin-(1-7) competed at a second lower-affinity site, with an IC50 of 2.9 mu mol/L. The high-affinity angiotensin II receptor antagonist sarcosine(1)-isoleucine(8)-angiotensin II blocked [I-125]angiotensin-(1-7) binding to bovine aortic endothelial cells at both a high- (IC50 = 1.3 nmol/L) and a low-affinity (IC50 = 6.2 mu mol/L) binding site. In contrast, D-alanine(7)-angiotensin-(1-7) completely blocked [I-125]angiotensin-(1-7) binding, with an IC50 of 19.8 nmol/L, suggesting that D-alanine(7)-angiotensin-(1-7) may selectively block responses to angiotensin-(1-7) in endothelial cells.