CHARACTERIZATION OF THE VASCULAR THROMBOXANE-A2 PROSTAGLANDIN ENDOPEROXIDE RECEPTOR IN RABBIT AORTA - REGULATION BY DEXAMETHASONE

Recently, we have shown that dexamethasone treatment of rabbits specifically reduces vascular smooth muscle responsiveness to agonists that interact with the vascular thromboxane A2/prostaglandin H2 (TXA2/PGH2) receptor. One potential site at which dexamethasone can influence prostanoid-mediated vasoconstriction may be at the level of the vascular TXA2/PGH2 receptor. Therefore, we characterized the vascular TXA2/PGH2 receptor in rabbit aortic membranes and examined the influence of dexamethasone treatment on vascular TXA2/PGH2 receptor affinity and number. The binding of [125I][1S-(1α,2β(5Z),3α(1E,3R)4α)]-7-[3-(3-hydroxy-4- (p-iodophenoxy)-1-butenyl)-7-oxabicyclo[2.2.1]heptan-2-yl]-5- heptanoic acid ([125I] BOP), a potent TXA2/PGH2 receptor agonist, to rabbit aortic membranes was saturable displaceable, and dependent on protein concentration. Scatchard analysis of equilibrium binding data disclosed one class of high affinity binding sites with a K(d) of 0.44 ± 0.13 nM and a B(max) of 114.4 ± 5.2 fmol/mg protein (n = 7). Removal of the endothelium before membrane preparation did not significantly alter the affinity or number of binding sites for [125I]BOP. Kinetic analysis of the rates of [125I]BOP association/dissociation yielded a K(d) of 0.62 nM. The ability of various agonists at the TXA2/PGH2 receptor to displace [125I]BOP from vascular membranes correlated well with their contractile potencies in rabbit aortic rings. Moreover, stereospecific displacement of [125I]BOP binding in aortic membranes and inhibition of U46619-mediated aortic contractions were obtained with the stereoiomers L657925(-) and L657926(+). Collectively, these data suggest that this binding site represents the functionally relevant vascular TXA2/PGH2 receptor. In functional experiments, [127I]BOP induced concentration-dependent contractions of the rabbit aorta, which were reduced by 52% in vessels from dexamethasone-treated rabbits. Binding experiments performed in aortic membranes from dexamethasone-treated rabbits revealed a 25% reduction in vascular TXA2/PGH2 receptor number with no change in affinity. Thus, the dexamethasone-induced decrease in TXA2/PGH2 receptor number in aortic membranes from dexamethasone-treated rabbits may contribute to the accompanying decrease in vascular responsiveness to TXA2/PGH2 receptor agonists.