To determine whether the initiation of fetal lung surfactant phospholipid production and the activation of the gene for the 35-kD surfactant-associated protein are dependent on circulating corticosteroids, we cultured dexamethasone-responsive explants of 15- to 17-d fetal rat lung in medium with 1% FCS (controls), charcoal-stripped 1% FCS, or a variety of glucocorticoid antagonists. The steroid antagonist RU 486 almost completely abolished specific cytoplasmic and nuclear dexamethasone binding in the explants but had no glucocorticoid-agonist activity. There was a significant increase in disaturated phosphatidylcholine synthesis during 7 d in culture in control explants (78%) and in those cultured with Charcoal-stripped serum (83%), RU 486 (82%), or the other glucocorticoid antagonists--clotrimazole, cortexelone, and 11-ketoprogesterone. Specific mRNA for surfactant-associated protein A was not detectable in preculture 17-d lung tissue, but accumulated to the same extent in cultures with or without RU 486 in the medium. These findings support the view that expression of the genes responsible for the synthesis of the various components of surfactant is not induced by glucocorticoids, but by signals contained within the lung tissue itself. The role of circulating hormones is later acceleration and modulation of surfactant production.