Multiple subtypes of endothelin receptors in porcine tissues: characterization by ligand binding, affinity labeling and regional distribution

To clarify the existence and the distribution of endothelin (ET) receptor subtypes, we have examined the pharmacological properties and the molecular weight ( M r ) of 125 I-ET-1 and 125 I-ET-3 binding sites in various tissues of pigs. ET-1 and ET-2 showed almost identical potencies in displacing the bound 125 I-ET-1 in all the tissues examined. ET-3, sarafotoxin S6b (SRT-b) and sarafotoxin S6c (SRT-c) displaced the 125 I-ET-1 with the same sensitivity as ET-1 (IC 50 = 0.1–1.4 nM) in brain, kidney, liver and adrenal, whereas the three peptides showed very weak competition (IC 50 = 40–500 nM) against 125 I-ET-1 binding in cardiac atria, aorta, lung, stomach and uterus. The computer analyses of the binding data suggested the presence of high ( K d 1 = 0.04–0.29 nM) and low ( K d 2 = 60–190 nM) affinity binding sites for ET-3 and SRT-b in lung and stomach. 125 I-ET-3 bound to the high affinity sites in lung and stomach was displaced by ET/SRT isopeptides almost equipotently. Two proteins with M r of 47,000 and 35,000 were affinity-labeled with 125 I-ET-1 in cerebellum, while a protein with M r of 123,000, in addition to the two proteins, was predominantly labeled in lung. The above findings indicated that two distinct subclasses of ET receptors, namely, ET-1-specific and ET/SRT family-common receptors were distributed in various proportions in mammalian tissues, and suggested that their molecular forms are also different.