The contractile responses to endothelin-1 (ET-1), endothelin-3 (ET-3), and sarafotoxin S6c (STXc) were best-fitted by a two-site model in the rabbit isolated saphenous vein. Agonists were equipotent at the high-affinity site (pD2s 12.0 +/- 0.2, 12.2 +/- 0.2, and 12.3 +/- 0.3, respectively), characteristic of an ETB receptor, whereas ET-3 and STXc were significantly more potent than ET-1 as vasoconstrictors at the low-affinity site (pD2s 10.2 +/- 0.3, 10.6 +/- 0.3, and 9.1 +/- 0.1, respectively), characteristic of an ETC-like receptor. The high affinity, ETB-mediated response was inhibited by SB 209670, Ro 47-0203, BQ-788, and Ro 46-2005 (Kbs of 0.15, 14, 110, and 280 nM against STXc, respectively) but was insensitive to PD 142893, RES-701 and BQ-123 (Kbs > or = 10 microM), consistent with this response being mediated by the ETB2-like receptor subtype. The low-affinity ETC-like-mediated component was inhibited by SB 209670, Ro 47-0203, BQ-788, Ro 46-2005, PD 142893, and RES-701 (Kbs 38 nM, 62 nM, 670 nM, 580 nM, 1.7 microM, and 2.0 microM, respectively), but was insensitive to BQ-123 (Kb > or = 10 microM). ET-3 produced endothelium-dependent vasorelaxation in the presence of active tone. Antagonist IC50s were approximated as being: SB 209670 3 nM; BQ-788 and RES 701,300 nM; Ro 46-2005 and PD 142893 3 microM; BQ-123 > or = 10 microM. These values were consistent with vasorelaxation being mediated by an ETB1-like receptor. Therefore, three pharmacologically distinct ET receptor subtypes have been identified in rabbit saphenous vein. Two contractile receptors, ETB2-like and ETC-like, are present on the vascular smooth-muscle receptor, whereas an ETB1-like receptor, which mediates the vasodilator actions of this peptide family, is present on the endothelium.