Developmental changes in glucocorticoid receptor and 11β- hydroxysteroid dehydrogenase oxidative and reductive activities in rat Leydig cells

Abstract

Glucocorticoids directly regulate testosterone production in Leydig cells through a glucocorticoid receptor (GR)-mediated repression of the genes that encode testosterone biosynthetic enzymes. The extent of this action is determined by the numbers of GR within the Leydig cell, the intracellular concentration of glucocorticoid, and 11β-hydroxysteroid dehydrogenase (11βHSD) activities that interconvert corticosterone (in the rat) and its biologically inert derivative. 11-dehydrocorticosterone. As glucocorticoid levels remain stable during pubertal development, GR numbers and 11βHSD activities are the primary determinants of glucocorticoid action. Therefore, in the present study, levels of GR and 11βHSD messenger RNA (mRNA) and protein were measured in rat Leydig cells at three stages of pubertal differentiation: mesenchymal-like progenitors (PLC) on day 21 immature Leydig cells (ILC) that secrete 5α-reduced androgens on day 35, and adult Leydig cells (ALC) that are fully capable of testosterone biosynthesis on day 90. Numbers of GR measured by [3H]dexamethasone binding, in purified cells were 6.34 ± 0.27 (x103 sites/cell: mean P SE) for PLC, 30.45 ± 0.74 for ILC, and 32.54 ± 0.84 for ALC. Although GR binding was lower in PLC, steady state levels for GR mRNA were equivalent at all three stages (P > 0.05). Oxidative and reductive activities of 11βHSD were measured by assaying the conversion of radiolabeled substrates in incubations of intact Leydig cells. Both oxidative and reductive activities were barely detectable in PLC, intermediate in ILC, and highest in ALC. The ratio of the two activities favored reduction in PLC and ILC and oxidation in ALC (oxidation/reduction, 0.33 ± 0.33 for PLC, 0.43 ± 0.05 for ILC, and 2.12 ± 0.9 for ALC, with a ratio of i indicating equivalent rates for both activities). The mRNA and protein levels of type I 11βHSD in Leydig cells changed in parallel with 11βHSD reductive activity, which increased gradually during the transition from PLC to ALC, compared with the sharp rise that was seen in oxidative activity. We conclude that Leydig cells at all developmental stages have GR and that their ability to respond to glucocorticoid diminishes as net 11βHSD activity switches from reduction to oxidation. This provides a mechanism for the Leydig cell to regulate its intracellular concentration of corticosterone, thereby varying its response to this steroid during pubertal development.