PURPOSE: The authors investigated the effect of gestational age on lung development and maturation after in utero tracheal occlusion (TO) in a rabbit model of congenital diaphragmatic hernia (CDH). METHODS: In 46 fetal rabbits, CDH was created at 23 days' gestational age (GA; term, 31 days), corresponding to the pseudoglandular phase of lung development. A second intervention was performed at either 26, 27, or 28 days on 6 fetuses in each GA group. At that time, either TO (CDH + TO), or a sham operation (CDH + sham) was performed. Nonoperated littermates served as internal normal controls (CTR). All fetuses were delivered by cesarean section at 30 days GA to assess lung response by lung-to-body-weight ratio, pulmonary morphometry, and the density of type II pneumocytes. RESULTS: After TO, the lungs were significantly larger than in CDH animals; their weight was proportional to the duration of TO. Pulmonary morphometry in TO fetuses was comparable with that of controls. The density of type II cells was inversely related to the gestational age at which TO was performed, with normal values with TO at GA at 28 days. CONCLUSION: Timing of TO is critical to subsequent pulmonary development: early in gestation TO leads to pulmonary overgrowth and type II pneumocyte depletion, whereas normal values are obtained when TO is delayed till 28 of 32 days.