S-0139 (27-O-3-[2-(3-carboxy-acryloylamino)-5-hydroxyphenyl]-acryloyloxy myricerone, sodium salt) is a highly specific nonpeptide endothelin ET A receptor antagonist. The binding of [ 3 H]S-0139 was compared to that of [ 1 2 5 I]endothelin-1 to characterize the binding of the antagonist in porcine aortic smooth muscle membranes. Scatchard analysis revealed a single class of [ 3 H]S-0139 binding sites with a K d value of 0.61+/-0.10 nM and a B m a x of 0.72+/-0.16 pmol/mg protein. These sites were saturable and reversible. [ 1 2 5 I]Endothelin-1 also showed binding with high affinity (K d =0.12+/-0.02 nM) to a homogenous population of binding sites, whose B m a x (0.71+/-0.20 pmol/mg protein) was almost the same as that for [ 3 H]S-0139. In both cases, the binding could be displaced by known endothelin receptor ligands and their IC 5 0 values in each case showed a very close correlation (r=0.986). The potency of seven endothelin receptor antagonists to displace [ 3 H]S-0139 binding also correlated highly to the potency for inhibiting the endothelin-1-induced increase in cytosolic Ca 2 + concentration (r=0.949). Myriceric acid A showed a more potent functional activity than expected from its binding affinity, but this seemed to result from the different assay conditions, such as incubation time. Together, the results suggest that S-0139 labels only endothelin ET A receptor binding sites in porcine aortic smooth muscle.