PITUITARY ADENYLATE-CYCLASE ACTIVATING POLYPEPTIDE STIMULATES RAT LEYDIG-CELL STEROIDOGENESIS THROUGH A NOVEL TRANSDUCTION PATHWAY

The aim of the present study was to evaluate the effects of pituitary adenylate cyclase activating polypeptide (PACAP) on testosterone produ ction in isolated adult rat Leydig cells and its possible mechanisms o f action. PACAP-38 stimulated testosterone secretion in a dose-depende nt manner with a minimal and a maximal efficacious dose of 1.0 nM and 100 nM, respectively. PACAP-27 was without effect on testosterone secr etion at any dose tested. Similarly. vasoactive intestinal peptide did not stimulate steroidogenesis nor interfere with PACAP-38 activity, a s well as preincubation of Leydig cells with the vasoactive intestinal peptide-antagonist [Lys(1), Pro(2,5) Arg(3,4), Tyr(6)]- vasoactive in testinal peptide. Removal of extracellular Ca2+ did not inhibit the st imulatory effects of PACAP-38 on Leydig cell testosterone production. Neither PACAP-38 nor PACAP-27 modified intracellular free Ca2+ and cAM P levels at any dose tested thus excluding a role for Ca2+ and cAMP in the stimulatory effects of PACAP. PACAP-38 was able to induce a plasm a membrane depolarization that was dependent on an influx of Na+ from the extracellular medium as confirmed by the monitoring of intracellul ar Na+ with the Na+-sensitive fluorescent dye sodium benzofuran isopht alate. When Na+ was removed from the extracellular medium, PACAP-38 di d not stimulate testosterone production, demonstrating that Na+ influx through the plasma membrane is strictly related to the stimulatory ef fects of this peptide. In addition, preincubation of Leydig cells in t he presence of pertussis-toxin (500 ng/ml for 5 h) significantly reduc ed PACAP-38-stimulated effects both on plasma membrane depolarization and testosterone secretion. These results demonstrate that PACAP-38 st imulates testosterone secretion in isolated adult rat Leydig cells thr ough the interaction with a novel PACAP receptor subtype coupled to a pertussis toxin sensitive G protein whose activation induces a Na+-dep endent depolarization of the plasma membrane and testosterone producti on.