Endoglin is a TGF- superfamily receptor critical for endothelial cell function. Mutations in this gene are associated with hereditary hemorrhagic telangiectasia type I (HHT1), and clinical signs of disease are generally more evident later in life. We previously showed that systemic vessels of adult Eng heterozygous (Eng^+/-) mice exhibit increased vasorelaxation due to uncoupling of endothelial nitric oxide synthase (eNOS). We postulated that these changes may develop with age and evaluated pulmonary arteries from newborn and adult Eng^+/- mice for eNOS-dependent, acetylcholine (ACh-induced) vasorelaxation, compared to that of age-matched littermate controls. While ACh-induced vasorelaxation was similar in all newborn mice, it was significantly increased in the adult Eng^+/- versus control vessels. The vasodilatory responses were inhibited by L-NAME suggesting eNOS-dependence. eNOS uncoupling was observed in lung tissues of adult, but not newborn heterozygous mice and was associated with increased production of reactive O₂ species (ROS) in adult Eng^+/- versus control lungs. Interestingly ROS generation was higher in adult than newborn mice and so were the levels of NADPH oxidase 4 and SOD 1,2,3 isoforms. However, enzyme protein levels and NADPH activity were normal in adult Eng^+/- lungs indicating that the developmental maturation of ROS generation and scavenging cannot account for the increased vasodilatation observed in adult Eng^+/- mice. Our data suggest that eNOS-dependent H₂O₂ generation in Eng^+/- lungs accounts for the heightened pulmonary vasorelaxation. To the extent that these mice mimic human HHT1, age-associated pulmonary vascular eNOS uncoupling may explain the late childhood and adult onset of clinical lung manifestations.