Isolation method of Leydig cells from mature male Djungarian hamsters (Phodopus sungorus) and their steroidogenic activity in vitro

For studies addressing the functions of Leydig cells, isolated cells are of ten better suited than intact animals. Here, the isolation procedure of Ley dig cells from adult male Djungarian hamsters (Phodopus sungorus) is descri bed. Cells were isolated using a procedure involving enzymatic dissociation and Percoll-gradient centrifugation. For each experiment, approximate to 4 .4 x 10(6) Leydig cells from six animals were obtained. The cells showed hi gh steroidogenic responsiveness to physiological (ovine luteinizing hormone (oLH) and human chorionic gonadotropin (hCG)) and nonphysiological (forsko lin) stimuli in vitro. Approximately 98% of cells were viable as assessed b y trypan blue exclusion, and the purity varied from 80 to 95% as tested by 3 beta-hydroxysteroid dehydrogenase activity. Leydig cells were also identi fied by a bright yellow halo under phase-contrast microscopy. They containe d numerous lipid droplets and showed round nuclei and prominent nucleoli. T he cells responded to oLH, hCG and forskolin with an increased testosterone production in a dose-dependent manner. Dose-response curves in these studi es suggest that Leydig cells of Djungarian hamsters undergo desensitization , probably due to down regulation of their LH/CG receptors.