We asked whether lung injury and surfactant metabolism differed in preterm lambs after a 1-h period of hyperventilation to P(CO2) values of 25-30 mm Hg. The lambs then were surfactant treated and conventionally ventilated (CV) or high-frequency oscillatory ventilated (HFOV) for an additional 1 or 8 h. The results were compared with lambs that were not hyperventilated or surfactant treated but were ventilated with CV or HFOV. The 1-h hyperventilation resulted in increased alveolar protein, increased recovery of intravascular [131I]albumin in the lungs, and an increase in tumor necrosis factor-alpha mRNA. There were no differences between CV or HFOV in alveolar or total lung recoveries of saturated phosphatidylcholine (Sat PC), tracer doses of [14C]Sat PC and [125I]surfactant protein-B, or in percent Sat PC in large aggregate surfactant in surfactant-treated lambs. The lambs not hyperventilated or treated with surfactant had lower large aggregate pools and lower recoveries of [14C]Sat PC and [125I]surfactant protein-B in total lungs than for the surfactant-treated lungs, but there were no differences between the CV and HFOV groups. Hyperventilation followed by surfactant treatment resulted in a mild injury, but the subsequent use of CV or HFOV did not result in differences in surfactant metabolism.