Inhaled nitric oxide (iNO), a selective pulmonary vasodilator, is increasingly being used for the treatment of persistent pulmonary hypertension of the newborn (PPHN). iNO has also been proposed as an adjunctive therapy for respiratory distress syndrome (RDS) in premature infants with surfactant deficiency (J Pediatr 126:450-453, 1995). Studies performed in animals and human adults demonstrate that iNO increases bleeding time (BT) and inhibits platelet aggregation. The use of iNO may therefore increase the risk of bleeding in the neonatal population. We tested the hypothesis that iNO increases bleeding tendencies by studying the effects of iNO on bleeding time and platelet aggregation in 9 infants (gestational age > 29 weeks, postnatal age 14 ± 3 days) with PPHN caused by RDS, meconium aspiration syndrome or pneumonia. Bleeding times (using a modified template technique) were obtained at three time points: following exposure to iNO at 40 ppm for 30 minutes (T1), and 60 minutes (T2) and 24 hours (T3) after discontinuation of iNO. Platelet counts and blood for platelet aggregation studies (performed by stimulation with epinephrine, ADP, collagen and arachidonic acid) were obtained at times T1 and T3. The BT at T3, 24 hours off iNO, was 86 ± 19 seconds (mean ± SEM). When compared to BT value at T3, BT at T1 was significantly increased (159 ± 38 seconds, p<0.05), while that at T2 was increased to 114 ± 18 seconds (p=0.08). No differences were seen among the platelet counts or platelet aggregation studies at any time point. Each of the infants had similar degrees of illness at the three different time points studied, as reflected by their degree of ventilatory support required. These studies demonstrate that iNO at 40 ppm increases the bleeding time in neonates, and that this effect persists 60 minutes after the discontinuation of iNO. These findings suggest that the use of iNO in neonates may place them at risk for hemorrhagic complications.