1. In homogenates of human saphenous vein, [125I]-ET-1 and [125I]-S6b each labelled a single population of high affinity binding sites with K(D) values of 0.64 +/- 0.11 nM and 0.55 +/- 0.08 nM respectively. Hill slopes were close to one. However, the density of receptors labelled by [125I]-ET-1 was significantly greater than that by [125I]-S6b (187.6 +/- 23.0 compared to 91.7 +/- 23.6 fmol mg-1 protein, P < 0.02). 2. BQ123, an ET(A-)selective antagonist, inhibited specific [125I]-ET-1 and [125I]-S6b binding with equal affinity. BQ123 competed in a biphasic manner for both [125I]-ET-1 (0.1 nM) and [125I]-S6b (0.1 nM) with ET(A) K(D) values of 0.55 +/- 0.17 nM and 0.52 +/- 0.02 nM and ET(B) K(D) values of 14.4 +/- 2.60 microM and 11.2 +/- 0.31 microM respectively. S6b monophasically inhibited 0.1 nM [125I]-ET-1 (K(D) 1.16 +/- 0.9 nM) but competed for 0.25 nM [125I]-ET-1 in a biphasic manner (K(D) high affinity site 1.99 +/- 0.84 nM, K(D) low affinity site 0.68 +/- 0.63 microM, ratio 67% : 33%). 3. BQ123 antagonized the vasoconstrictor responses of ET-1 with a pK(B) value of 6.47 whereas BQ123 exhibited 50 fold higher affinity against S6b-mediated vasoconstriction with a pK(B) value of 8.18. Regression slopes were 0.80 +/- 0.13 and 1.08 +/- 0.11 respectively. 4. In desensitization experiments, S6b (300 nM) did not contract preparations which were no longer responsive to ET-1 whereas a small contraction to ET-1 (300 nM) was obtained in preparations rendered unresponsive to S6b. 5. Medial sections of non-diseased human aorta, which express only ET(A) receptors, were used to compare dissociation rates of the two agonists. The time course for the dissociation of [125I]-ET-1 and [125I]-S6b was similar with 20-30% of each ligand dissociating at 4 h. 6. These data suggest that whilst BQ123, in common with other endothelin antagonists, is a much more potent blocker of S6b contractile responses than of ET-1 contractile responses, this is not reflected by the equal affinity of BQ123 determined in competition binding experiments against both [125I]-ET-1 and [125I]-S6b. This discrepancy in antagonist potency is probably not due to a marked difference in the rate of dissociation of [125I]-ET-1 and [125I]-S6b from endothelin receptors. One possible explanation is that ET-1 is activating an additional population of receptors which may have lower affinity for BQ123. This is suggested by the discrepancy in receptor density identified by [125I]-ET-1 and [125I]-S6b.