BINDING CHARACTERIZATION OF [H-3]S-0139, AN ANTAGONIST OF THE ENDOTHELIN ETA-RECEPTOR SUBTYPE

S-0139 arboxy-acryloylamino)-5-hydroxyphenyl]-acryloyloxy myricerone, sodium salt) is a highly specific nonpeptide endothelin ETA receptor a ntagonist. The binding of [H-3]S-0139 was compared to that of [I-125]e ndothelin-1 to characterize the binding of the antagonist in porcine a ortic smooth muscle membranes. Scatchard analysis revealed a single cl ass of [H-3]S-0139 binding sites with a K-d value of 0.61 +/- 0.10 nM and a B-max of 0.72 +/- 0.16 pmol/mg protein. These sites were saturab le and reversible. [I-125]Endothelin-1 also showed binding with high a ffinity (K-d = 0.12 +/- 0.02 nM) to a homogenous population of binding sites, whose B-max (0.71 +/- 0.20 pmol/mg protein) was almost the sam e as that for [H-3]S-0139. In both cases, the binding could be displac ed by known endothelin receptor ligands and their IC50 values in each case showed a very close correlation (r = 0.986). The potency of seven endothelin receptor antagonists to displace [H-3]S-0139 binding also correlated highly to the potency for inhibiting the endothelin-1-induc ed increase in cytosolic Ca2+ concentration (r = 0.949). Myriceric aci d A showed a more potent functional activity than expected from its bi nding affinity, but this seemed to result from the different assay con ditions, such as incubation time. Together, the results suggest that S -0139 labels only endothelin ETA receptor binding sites in porcine aor tic smooth muscle. (C) 1998 Elsevier Science B.V.