Identification of functional binding sites for progesterone in rat Leydig cell plasma membrane

Steroid hormones influence cell functions by binding to intracellular recep tors and then acting within the nucleus. There is now evidence that steroid s affect cell functions also via interaction with plasma membrane receptors in a number of different cell types. In this: regard, progesterone appears to be one of the most active steroids. In this paper, we evaluate the effe cts of progesterone on rat Leydig cell functions, determining variations of ion homeostasis and testosterone production. This steroid was able to effe ct a depolarization of the plasma membrane that was due to an influx of sod ium (Na+) from the external medium since it was absent when extracellular N a+ was iso-osmotically substituted with choline chloride or sucrose. The de termination of intracellular sodium concentration ([Na+](i)) with the Na+-s ensitive fluorescent dye sodium-benzofuran-isophtalate (SBFI) confirmed the se observations. Progesterone did not modify Leydig cell intracellular calc ium concentration ([Ca2+](i)) at any dose tested. Furthermore, using a cell impermeant progesterone conjugate, we demonstrated that progesterone was a ble to stimulate Leydig cell steroidogenesis in a dose-dependent manner. Th e exclusion of calcium (Ca2+) from the extracellular medium did not modi fy the depolarizing action of progesterone and its steroidogenetic effect whi le in Na+-free medium (sucrose supplemented) progesterone-stimulated effect s were completely blunted. Finally, using fluorescence microscopy with a fl uorescein isothiocyanate-coupled cell impermeant progesterone conjugate, we identified plasma membrane binding sites for progesterone in rat Leydig ce lls. These results suggest that rat Leydig cells possess progesterone recep tors located on the plasma membrane, which when occupied achieves a plasma membrane depolarization, dependent on an influx of Na+ from the external me dium, and the subsequent activation of steroidogenesis. (C) 1999 Elsevier S cience Inc. All rights reserved.