RADIOLIGAND BINDING ASSAY OF PROGESTERONE RECEPTORS IN THE PRIMATE CORPUS-LUTEUM AFTER IN-VIVO TREATMENT WITH THE 3-BETA-HYDROXYSTEROID DEHYDROGENASE INHIBITOR, TRILOSTANE

We and others have detected progesterone receptors (PR) in the primate (macaque and human) corpus luteum by immunocytochemistry. However, we have been unable to measure PR in the corpus luteum by conventional l igand binding assay, presumably because high endogenous concentrations of progesterone (P) in luteal tissue prevented specific binding of ra diolabeled hormone to PR during the assays. To test this hypothesis, w e treated monkeys with the 3 beta-hydroxysteroid dehydrogenase inhibit or, Trilostane, to reduce levels of endogenous P before conducting bin ding assays for PR in luteal tissue. To obtain adequate tissue for sat uration analysis, rhesus monkeys (n = 6) were superovulated by treatin g them with hFSH beginning at menses (day 1) for 6 days, then with hFS H and hLH (days 6-9), followed by hCG (day 10). Trilostane (600 mg) wa s given 5 days after hCG treatment (day 15), and binding assays were c onducted 18 h later. Trilostane significantly reduced mean (+/-SE) ser um levels of P from 97.8 +/- 16 to 2.7 +/- 1.3 nmol/L within 18 h (P < 0.001). Strong nuclear staining of PR was detected by immunocytochemi stry in both Trilostane-treated and control (not Trilostane-treated) t issue, but ligand binding was measurable only in Trilostane-treated mo nkeys. Scatchard transformations of saturation curves revealed high af finity binding of [H-3] R5020 in luteal cytosol and nuclear extracts w ith an approximate dissociation constant (K-d) of 4.8 and 1.37 nmol/L, respectively. Also, the PR-specific monoclonal antibody, JZB-39, shif ted a [H-3] R5020-bound luteal macromolecule on sucrose gradients. Mea n cytosolic and nuclear binding of [H-3]R5020 were 0.31 +/- 0.09 and 0 .06 +/- 0.02 fmol/mu g DNA, respectively. Similar binding of [H-3]R502 0 was demonstrated in corpora lutes obtained during spontaneous menstr ual cycles after Trilostane treatment (n = 3). These results show uneq uivocally that the PR in macaque luteal tissue can bind P with high af finity and suggest a receptor-mediated action of P in the primate corp us luteum.