To investigate the role of nitric oxide (NO) in hyperoxic lung injury, the 3 day old preterm rats were randomly assigned to four groups: group I (hyperoxia group), group Ⅱ (hyperoxia+N w nitro L arginine methyl ester (L NAME) group), group Ⅲ (air group), and group Ⅳ (air+L NAME) group. Group Ⅰ and Ⅱ were exposed to ≥90 % O 2 for 3 or 7 days. Group Ⅱ and Ⅳ received subcutaneous L NAMEy on daily basis (20 mg/kg). After 3 day or 7 day exposure, the lung wet weight/dry weight ratio (W/D), total protein and malondialdehyde (MDA) in bronchoalveolar lavage fluid (BALF) and lung pathology were examined in all groups. NO content, expression of endothelial NOS (eNOS) and inducible NOS (iNOS) in lungs were measured in group Ⅰ and Ⅲ. Our results showed that after 3 day exposure, group Ⅰ appeared acute lung injury characterized by the increase of MDA content ( P <0.01) and the presence of hyperaemia, red cell extravasation and inflammatory infiltration; after 7 day exposure, except MDA, total protein and W/D were also increased in comparison with group Ⅲ ( P <0.01, 0.05), pathological changes were more severe than those after 3 day exposure. After 3 and 7 day exposure, total protein in group Ⅱ was significantly increased as compared with group Ⅰ ( P <0.01 for both). The pulmonary acute inflammatory changes were more obvious in group Ⅱ than in group Ⅰ. Occasionally, mild hemorrhage was detected in the lungs of group Ⅳ. BALF protein content in group IV was higher than that in group Ⅲ after 7 day exposure ( P <0.01). After 3 and 7 day exposure, NO content in BALF were all significantly elevated in group Ⅰ as compared with group Ⅲ ( P <0.01 for all). In the lungs of group Ⅰ, strong immunostaining for iNOS was observed in airway and alveolar epithelia, inflammatory cells, which were stronger than those in group Ⅲ. Expression of iNOS in rats after 7 day hyperoxic exposure was stronger than that after 3 day exposure. Shortly after 7 day exposure, stronger immunostaining for eNOS in airway epithelia in group Ⅰ than that in group Ⅲ was seen. Our study suggested that treatment with L NAME worsened acute hyperoxic lung injury in preterm rats and also had a deleterious effect on the rats exposed to air, indicating that endogenous nitric oxide may play a protective role in rats under both physiological and hyperoxic status. Hyperoxia can significantly upregulate the expression of iNOS and eNOS in inflammatory cells, epithelia in the lungs of preterm rats, promote NO generation, which suggests that endogenous NO may mediate the hyperoxic pulmonary damage. Over stimulation of iNOS may contribute to the pathogenesis of hyperoxic lung injury. NO may have dual roles in pulmonary oxygen toxicity. To investigate the role of nitric oxide (NO) in hyperoxic lung injury, the 3 day old preterm rats were randomly assigned to four groups: group I (hyperoxia group), group II (hyperoxia + N w nitro L arginine methyl ester (L NAME Group II and IV were exposed to ≧ 90% O 2 for 3 or 7 days. Group Ⅱ and Ⅳ received subcutaneous L NAMEy on daily (group), group Ⅲ (air group), and group Ⅳ (air + L NAME) group. after 3 days or 7 day exposure, the lung wet weight / dry weight ratio (W / D), total protein and malondialdehyde (MDA) in bronchoalveolar lavage fluid (BALF) and lung pathology were examined in all groups. NO content, expression of endothelial NOS (eNOS) and inducible NOS (iNOS) in lungs were measured in group I and III. Our results showed that after 3 day exposure, group Ⅰ Acute lung injury characterized by the increase of MDA content (P <0.01) and the presence of hyperaemia, red cell extravasation and inflammatory infiltration; afte r 7 day exposure, except MDA, total protein and W / D were also increased in comparison with group III (P <0.01, 0.05), pathological changes were more severe than those after 3 day exposure. After 3 and 7 day exposure, total Protein in group Ⅱ was significantly increased as compared with group Ⅰ (P <0.01 for both). The pulmonary acute inflammatory changes were more obvious in group Ⅱ than in group Ⅰ. Occasionally, mild hemorrhage was detected in the lungs of group Ⅳ. BALF After 3 and 7 day exposure, NO content in BALF were all significantly elevated in group I as compared with group III (P <0.01 for all). In the lungs of group I, strong immunostaining for iNOS was observed in airway and alveolar epithelia, inflammatory cells, which were stronger than those in group III. Expression of iNOS in rats after 7 days hyperoxic exposure was stronger than that after after 3 day exp osure. Shortly after 7 day exposure, stronger immunostaining for eNOS in airway epithelia in group I than that in group III was seen. Our study suggested that that treatment with L NAME worsened acute hyperoxic lung injury in preterm rats and also had a deleterious effect on the rats exposed to air, indicating that endogenous nitric oxide may play a protective role in rats under both physiological and hyperoxic status. Hyperoxia can significantly upregulate the expression of iNOS and eNOS in inflammatory cells, epithelia in the lungs of preterm rats, promote NO generation, which suggests that endogenous NO may mediate the hyperoxic pulmonary damage. Over stimulation of iNOS may contribute to the pathogenesis of hyperoxic lung injury. NO may have dual roles in pulmonary oxygen toxicity.