PLATELET AND VASCULAR THROMBOXANE-A2 PROSTAGLANDIN-H2 RECEPTORS - EVIDENCE FOR DIFFERENT SUBCLASSES IN THE RAT

Thromboxane A2 (TXA2) and its precursor prostaglandin H-2 (PGH2) induce platelet aggregation and vascular contraction through shared cell surface receptors commonly referred to as TXA2 or TXA2/PGH2 receptors. Whether different subclasses of TXA2/PGH2 receptors exist in platelets and vascular smooth muscle cells is controversial. In this study, TXA2 receptors on washed rat and human platelets and cultured rat aortic smooth muscle cells (RASMC) were characterized using radioligand competition binding assays with the I-125-labeled TXA2/PGH2 receptor agonist [1S-(1-alpha,2-beta-(5Z),3-alpha-(1E,3R*),4-alpha)]-7-[3-(3-hydroxy-4-(4'-iodophenoxy)-1-butenyl)-7-oxabicyclo-[2.2.1]-heptan-2-yl-5-heptenoic acid (I-BOP) and various agonists and antagonists. Scatchard analyses of equilibrium binding data revealed K(d) values of 205 +/- 68 pM (N = 6), 2.2 +/- 0.3 nM (N = 9) and 310 +/- 60 pM (N = 7) and B(max) values of 1.3 +/- 0.45 fmol/10(6) platelets, 2.8 +/- 0.2 fmol/10(6) platelets and 20.9 +/- 2.2 fmol/10(6) cells for rat and human platelets and RASMC, respectively. Concentration-dependent increases in intracellular free Ca2+ concentrations induced by I-BOP were observed in RASMC loaded with the calcium sensitive dye fura-2. The IC50 values for various TXA2/PGH2 analogues in competition binding assays with I-125-BOP were determined. Based on their IC50 values, the rank orders were I-BOP < L657925 < ONO11113 less-than-or-equal-to SQ29548 < PTA-TPO < PTA-NO less-than-or-equal-to L657926 less-than-or-equal-to I-PTA-OH < PTA-OH[2] = meta-I-PTA-PO less-than-or-equal-to ONO11120[2] = ONO11120[1] < PTA-OH[1] in rat platelets. I-BOP < SQ29548 < PTA-TPO = L657925 less-than-or-equal-to ONO11113 < I-PTA-OH < PTA-NO less-than-or-equal-to meta-I-PTA-PO less-than-or-equal-to PTA-OH[2] < ONO11120[2] less-than-or-equal-to ONO11120[1] < L657926 less-than-or-equal-to PTA-OH[1] in human platelets, and I-BOP < L657925 < ONO11113 less-than-or-equal-to SQ29548 < ONO11120[2] less-than-or-equal-to L657926 less-than-or-equal-to PTA-OH[2] < PTA-TPO < ONO11120[1] < I-PTA-OH < meta-I-PTA-PO < PTA-NO < PTA-OH[1] in RASMC. For the structurally dissimilar compounds I-BOP, SQ29548, ONO-11113, L657925 and L657926, there were no significant differences in the rank order potencies between the three tissues. In contrast, the rank order potencies for the 13-azapinane TXA2 analogs were significantly different in rat platelets compared to RASMC, and human platelets compared to RASMC, but not significantly different between rat and human platelets. The ratios of IC50 values for the two sets of epimeric pairs of 13-azapinane TXA2 analogs (ONO11120[1] and [2] and PTA-OH[1] and [2] were significantly different (P < 0.05) for rat and human platelets compared to RASMC but not different between rat and human platelets. The results are consistent with the notion that TXA2/PGH2 receptors on platelets are different from those on RASMC, and 13-azapinane TXA2 analogs may be useful compounds for discriminating between the two classes of TXA2/PGH2 receptors.