In full-term newborn rats, propylthiouracil (PTU) treatment has been previously shown to decrease susceptibility to 02-induced lung damage and improve survival during hyperoxic exposure. However, no differences were found in lung antioxidant enzyme (AOE) activity responses to hyperoxia compared with 02-exposed untreated (control) term rats. To further explore possible pulmonary protective effects of PTU treatment in prematurely delivered animals, we administered PTU (0.015%) in drinking water to timed-pregnant rats for the final 10 d of gestation prior to delivery 1 d before term, and during lactation; control pregnant/nursing rats received untreated water. Both groups of 21-d premature rat pups were randomized to either >95% 02or room air exposure after birth for up to 14 d. The left lungs of 7-d exposure pups were used to quantitate the concentrations of AOE mRNA by solution hybridization; the right lungs of the same pups were assayed for AOE activities. PTU treatment resulted in survival rates of 02-exposed preterm rat pups that were consistently higher at all time periods in hyperoxia including 7 d [PTU, 67 of 82 (82%)versuscontrol pups, 58 of 113 (51%);p< 0.001] and 14 d [PTU, 31 of 39 (79%)versuscontrol, 15 of 66 (23%);p< 0.001]. Further evidence of increased tolerance to >95% 02in PTU pups included a significant decrease in the incidence of microscopic intraalveolar edema and a significant increase in lung tissue surfactant-related phospholipids compared with 02-exposed control pups. At 7 d in high 02, the PTU-treated pups showed greater increases in the lung AOE mRNA levels and AOE activities of catalase and glutathione peroxidase in response to hyperoxia compared with the untreated control 02group. Thus, we conclude that PTU treatment protects premature rats against 02-induced lung injury and lethality during prolonged hyperoxic challenge. The protective action of PTU may be related, at least in part, to the enhanced pulmonary AOE gene expression in the treated rat pups with resultant increases in protective AOE activity levels in response to neonatal lung oxidant challenge.