Vol. 286, Issue 2, 1094-1102, August 1998

Pharmacology of [³H]Prostaglandin E₁/[³H]Prostaglandin E₂ and [³H]Prostaglandin F₂ Binding to EP₃ and FP Prostaglandin Receptor Binding Sites in Bovine Corpus Luteum: Characterization and Correlation with Functional Data

N. A. Sharif, S. X. Xu, G. W. Williams, J. Y. Crider, B. W. Griffin and T. L. Davis

Molecular Pharmacology Unit, Alcon Laboratories Inc., Fort Worth, Texas

Specific binding of [³H]prostaglandin (PG) E₁, [³H]PGE₂ and [³H]PGF₂ to washed total particulate homogenates of bovine corpus luteum comprised 60 to 82% of total binding. Scatchard analysis of competition data revealed the presence of an apparent single population of binding sites for [³H]PGE₁ and [³H]PGE₂ with dissociation constants (K_ds) of 2.76 to 3.39 nM and apparent receptor density (B_max) of 1.5 to 1.56 pmol/g wet weight (n = 3-4). However, [³H]PGF₂ appeared to interact with two classes/states of binding sites (K_d1 = 6.51 ± 0.65 nM, B_max1 = 2.33 ± 0.26 pmol/g wet weight; K_d2 = 986 ± 269 nM; B_max2 = 44.8 ± 11.3 pmol/g wet weight, n = 11). Specific [³H]PGE₁ and [³H]PGE₂ binding was most potently (nanomolar affinity) inhibited by PGs with high selectivity for the EP₃ receptor subtype (e.g., GR63799, sulprostone, enprostil) but was weakly (K_is > 1 µM) influenced by EP₁-selective (SC-19220), FP-selective (fluprostenol, PHXA85), DP-selective (BWA868C; ZK118182), IP-selective (iloprost) and TP-selective (U46619) PGs. Specific [³H]PGF₂ binding was potently displaced by FP-selective agents such as fluprostenol, PHXA85 and cloprostenol with nanomolar affinities (n = 3-25), but weakly (K_is > 1 µM) by other PGs showing high selectivity for other PG receptor subtypes mentioned above. The relative specificities and potencies of EP₃- and FP-selective PGs tested in the binding assays were confirmed using various functional assays. These studies have provided strong pharmacological evidence for the similarity of [³H]PGE₁ and [³H]PGE₂ binding to EP₃ receptors and for [³H]PGF₂ binding to FP receptors in washed bovine corpus luteum homogenates.