Glucocorticoid enhancement of glucocorticoid production by cultured ovine adrenocortical cells

The present study examines the effect of chronic treatment with glucocorticoids on the steroidogenic activity of ovine adrenocortical cels in vitro. Cells cultured in the presence of 10-9 to 10-5 M dexamethasone produced more glucocorticosteroids in response to ACTH(1-24), forskolin or 8 BrcAMP than did control cells. Such an enhancing effect required more than 5 h of treatment and was maximal at 30 h; it was both concentration-dependent and steroid-specific. The maximal secretion of corticosteroids was observed when cells were exposed to 10-7 M dexamethasone; with higher concentrations the response to ACTH(1-24) decreased steadily; the ED(50) was 2.8+/-0.8 nM. Cortisol and corticosterone enhanced ACTH(1-24)-induced steroidogenesis to the same extent as dexamethasone, but at concentrations roughly 100-fold higher than for dexamethasone. Testosterone and 17 beta-oestradiol had no enhancing effect. Dexamethasone not only enhanced the maximal steroidogenic response to ACTH(1-24) but also decreased its ED(50) 3-fold. Treatment of cultures with the antiglucocorticoid RU 38486 resulted in a dose-dependent, time-dependent, decrease in ACTH(1-24)-induced corticosteroid output. Moreover, RU 38486 antagonized the enhancing effect of dexamethasone. The production of corticosteroids by dexamethasone-treated cells incubated in the presence of 22(R)-hydroxycholesterol or of exogenous pregnenolone was similar to that of control cells. The enhancing effect of dexamethasone was also observed when cultures were performed in the absence of insulin and/or in serum-free media. These data suggest that chronic exposure to glucocorticoids is necessary for the full steroidogenic activity of ovine adrenocortical cells. Moreover, they indicate that glucocorticoids exert their effect at least at two different levels in the cell: (i) on the adenylate cyclase system and (ii) at step(s) beyond cAMP but before pregnenolone formation.