Specific binding of gonadotrophin-releasing hormone and an agonist to human corpus luteum homogenates: characterization, properties, and luteal phase levels.
The binding of radiolabeled GnRH and GnRH agonist (GnRHA) was studied in homogenates of human luteal tissue using a polyethylene glycol precipitation technique. GnRHA binding to human luteal homogenates was dependent on pH, was inhibited by a number of divalent metal ions, and was of moderately high affinity (Ka = 3 X 10(7) M-1). Binding of both radiolabeled GnRH and GnRHA increased linearly with increasing homogenate concentration, and binding of either tracer was parallel with different corpus luteum (CL) homogenates. Moreover, similar concentrations of unlabeled GnRH or GnRHA blocked the binding of either tracer, indicating that both hormones bound to a common site. The rate of dissociation of bound ligand was very low at 0 C, but was extremely rapid at physiological temperatures (t1/2 = 2.5-3.5 min). Human luteal GnRHA-binding sites were partially solubilized by treatment with Triton X-100 and were sensitive to trypsin and pronase, but not hyaluronidase. GnRHA binding to homogenates of human CL obtained at each stage of the luteal phase varied markedly, with levels ranging from less than 20 to greater than 1600 pg/mg DNA for CL from the (midluteal) stage of the menstrual cycle. This variability could not be accounted for by assay irreproducibility, loss of binding activity during storage, or occupancy of binding sites by endogenous ligand. These observations may help to explain negative reports from other groups, which demonstrated little or no specific binding of GnRH to human luteal tissue and no effect of GnRH on steroidogenesis using dispersed human luteal cells in vitro. We conclude that human luteal homogenates possess specific, moderately high affinity binding sites for GnRH and its agonist analogs.
D000328:Adult; D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D002064:Buserelin / Q000378:metabolism*; D002614:Chelating Agents / Q000494:pharmacology; D006063:Chorionic Gonadotropin / Q000378:metabolism; D003338:Corpus Luteum / Q000378:metabolism*; D005260:Female; D007987:Gonadotropin-Releasing Hormone / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007985:Leydig Cells / Q000378:metabolism; D008183:Luteal Phase; D008297:Male; D008670:Metals / Q000494:pharmacology; D010902:Pituitary Gland / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D011919:Rats, Inbred Strains; D013048:Specimen Handling; D013696:Temperature
D002614:Chelating Agents; D006063:Chorionic Gonadotropin; D008670:Metals; D007987:Gonadotropin-Releasing Hormone; D002064:Buserelin