Abstract
Specific binding of [3H]prostaglandin (PG) E1, [3H]PGE2 and [3H]PGF2α 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 (Kds) of 2.76 to 3.39 nM and apparent receptor density (Bmax) of 1.5 to 1.56 pmol/g wet weight (n = 3–4). However, [3H]PGF2α appeared to interact with two classes/states of binding sites (Kd1 = 6.51 ± 0.65 nM, Bmax1 = 2.33 ± 0.26 pmol/g wet weight;Kd2 = 986 ± 269 nM; Bmax2 = 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 (Kis > 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]PGF2α binding was potently displaced by FP-selective agents such as fluprostenol, PHXA85 and cloprostenol with nanomolar affinities (n = 3–25), but weakly (Kis > 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]PGF2α binding to FP receptors in washed bovine corpus luteum homogenates.
Footnotes
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Send reprint requests to: Dr. N. A. Sharif, Head, Molecular Pharmacology Unit, Alcon Laboratories, Inc. (R2–19), 6201 South Freeway, Fort Worth, TX 76134-2099.
- Abbreviations:
- BCLM
- bovine corpus luteum
- PG
- prostaglandin
- PGI2
- prostacyclin
- TXA2
- thromboxane A2
- PI
- phosphoinositide
- Received July 25, 1997.
- Accepted April 17, 1998.
- The American Society for Pharmacology and Experimental Therapeutics
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