CHARACTERIZATION OF ENDOTHELIN RECEPTOR SUBTYPES IN ISOLATED RAT RENAL PREGLOMERULAR MICROVESSELS

Endothelin (ET) increases renal vascular resistance by constriction of post- and preglomerular vessels in the rat. However, ET receptor subt ypes in renal microvessels have not been clearly defined. Radioligand binding experiments were performed in isolated arcuate and interlobula r arteries as well as branching afferent arterioles to characterize I- 125-ET-1 binding sites. Competitive inhibition assays were performed w ith ET-1, ET-2, ET-3, sarafotoxin 6b (S6b), BQ-123 (a preferential ET( A) receptor antagonist) and 4-Ala-ET-1 (a preferential ET(B) receptor agonist). Saturation data revealed a single class of high affinity bin ding sites with a k(d) of 0.31 +/- 0.03 nM and a B-max of 1336 +/- 181 fmol/mg protein. Competitive inhibition of I-125-ET-1 binding showed that all assayed compounds displaced I-125-ET-1 in a dose-related mann er. ET-1 displaced 100% of I-125-ET-1 binding, displaying monophasic c urves with B-max and k(d) values of 1369 +/- 170 fmol/mg protein and 0 .35 +/- 0.04 nM, respectively. ET-2's displacement curves were similar to those of ET-1. ET-3 and S6b inhibited 100% of I-125-ET-1 binding i n a biphasic manner, suggesting these peptides bind both high and low affinity sites. BQ-123 displaced about 50% of I-ET-1 in a monophasic m anner, indicating a single high affinity binding site. 4-Ala-ET-1 disp laced I-125-ET-1 in a clearly biphasic manner with an almost equal pro portion of high and low affinity binding sites. Our results suggest th at both ET(A) and ET(B) receptors are expressed in rat renal preglomer ular vessels in almost equal proportions. However, the characteristics of competitive inhibition of I-125-ET-1 binding by several agents can not be fully explained by a two-receptor model.