Glucocorticoids are used clinically to accelerate pulmonary surfactant production, though this treatment has consistently been found to be only≈50% effective in preventing respiratory distress syndrome. Since previous studies (Torday, Endocrinology 126:3240, 1990) had suggested that fetal adrenal androgens (FAA) block steroid-stimulated fetal lung maturation, we have hypothesized that optimal steroid stimulation requires concomitant steroid suppression of the fetal pituitary adrenal axis, eliminating FAA from the fetal circulation. To test this hypothesis we treated pregnant rats with dihydrotestosterone (DHT, 1 mg/kg/day) from day 15-20; fetal rat lung fibroblasts (FRLF) were harvested on day 21 and cultured for 24 hours in serum-free medium containing either Dexamethasone [DEX, 1×10-8M], DHT (1×10-7M), DEX + DHT, DHT + Flutamide (Flut, 1×10-6M, anti-androgen), or DEX + DHT + Flut. FRLF were subsequently assayed for their ability to take up 3H-trioleate, a measure of their capacity to provide substrate for surfactant phospholipid synthesis. In vivo DHT treatment resulted in a 10-fold decrease in TG uptake compared to day 21 control FRLF; incubation of DHT-pretreated FRLF with DEX stimulated the TG uptake rate 4-fold resulting in a TG uptake rate comparable to that of day 21 FRLFs. However, in the presence of DHT, DEX only stimulated TG uptake by ≈90%, which is comparable to activity of FRLF from previable rat fetuses (d18-19). Flut, which was added in an attempt to“neutralize” the androgen effect, did not significantly increase TG uptake. These data suggest that in order to effectively stimulate fetal lung maturation glucocorticoids must inhibit FAA, consistent with companion clinical studies. Supported by the Wyeth Pediatrics Neonatology Research Grants Program.