Steroidogenesis in interstitial cells and microsomal fraction of immature pig testes

Testicular interstitial cells (greater than 90% viable) obtained from 6-day-old and 3-6-week-old piglets were capable of producing dehydroepiandrosterone (DHEA, 5-10 ng/500,000 cells) and responded to hCG (60 mi.u./ml), dibutyryl-cAMP (1 mmol/l) and cholera toxin (5 ng/ml) with a 2-3-fold increase in DHEA. Aminoglutethimide (100 mumol/l) abolished the response. Testosterone was produced in comparatively minor quantities (less than 0.3 ng/500,000 cells) and was unaffected by stimulation or inhibition. When cells from both age groups were incubated with [14C]- or [3H]-pregnenolone (360 and 3.0 nmol/l), 17-hydroxypregnenolone (15%) and DHEA (5-10%) were the major metabolites on the androgen pathway and 5,16-androstadien-3 beta-ol (andien-beta, 5-10%) and 4,16-androstadien-3-one (dienone, 5-10%) on the 16-androstene pathway. Stimulation and inhibition of endogenous steroidogenesis did not alter the metabolism of exogenous pregnenolone, the same metabolites being found in the same proportions at similar times. Microsomal enzyme activities accurately reflected the metabolic profile of pregnenolone metabolism seen in intact cells, with low activities for 17 beta-HSD, 3 beta-HSD-isomerase, and 16-ene-5 alpha-reductase being observed. Since steroidogenic capacity, enzyme complement and pregnenolone metabolism were the same in testes from both age groups, the differences in Leydig cell activity observed in vivo would not appear to be consequences of changes in steroidogenic enzymes or responsiveness to gonadotrophin stimulation. The lack of effect of stimulation and inhibition of steroidogenesis on the cellular metabolism of exogenous pregnenolone suggests that the endogenous and exogenous supplies of pregnenolone are metabolized by different populations of enzymes. The relative magnitudes of these populations indicate that most of the steroidogenic enzymes in the interstitial cells are not involved in the normal response to trophic stimulation.