CHARACTERIZATION OF RAT GLOMERULAR THROMBOXANE-A2 RECEPTORS - COMPARISON TO RAT PLATELETS

This study was designed to characterize rat glomerular thromboxane A2 (TxA2) receptors and compare them to rat platelet TxA2 receptors. The radioligand binding characteristics of the receptors were characterized using [I-125][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-2yl]-5-heptenoic acid ([I-125]BOP), a TxA2 agonist. Equilibrium binding with [I-215]BOP, as well as competitive binding assays between [I-125]BOP and 13-azapinane TxA2 receptors antagonists, were performed in rat glomerular membranes (RGM) and washed rat platelets (WRP). [I-125]BOP identified a single class of TxA2 receptor sites in glomerular membranes with a K(d) of 318 +/- 55 pM and a B(max) of 260 +/- 62 fmol/mg protein (n = 14). [I-125]BOP was displaced by the TxA2 agonist 15S-hydroxy-11-alpha,9-alpha(epoxymethano)-prosta-5Z,13E-dienoic acid (U-46,619) (IC50 = 22 +/- 6 nM, n = 3), the antagonist SQ-29,548 (IC50 = 41 +/- 7 nM, n = 4), and stereoselectively by the antagonists (-)-9-chlorobenzyl-6-fluoro-1,2,3,4-tetrahydrocarbazol-1-yl acetic acid (L-657,925) (IC50 = 0.27 +/- 0.04 nM, n = 3) and (+)-9-chlorobenzyl-6-fluoro-1,2,3,4-tetrahydrocarbazol-1-yl acetic acid (L-657,926) (IC50 = 124 +/- 0 nM, n = 2). The ability of six 13-azapinane TxA2 antagonists to compete with [I-125]BOP was evaluated. The rank orders for the 13-azapinanes showed no significant correlation between RGM and WRP. Phosphoinositide hydrolysis in whole glomeruli was stimulated with I-BOP (EC50 = 1.99 +/- 0.43 nM, n = 4) and stereoselectively blocked by L-657,925 (lC50 = 27.4 +/- 5.5 nM, n = 4) and L-657,926 (IC50 = 811 +/- 116 nM, n = 3). These results indicate that functional TxA2 receptors are present in glomeruli and that different subtypes of TxA2 receptors exist in rat glomeruli and platelets.