Nitric oxide (NO) or endothelium-derived relaxing factor may play an important role in modulating pulmonary vascular resistance (PVR), although previous studies have produced conflicting results. Endogenous NO inhibition causes an increase in PVR in intact animals but not in saline-perfused isolated lungs. We hypothesized that blood is essential for NO to serve as a modulator of PVR. Therefore, the effects of endogenous NO inhibition (N omega-nitro-L-arginine methyl ester [L-NAME]) were determined in isolated rat lungs as related to the presence of different blood components under normoxic conditions and after 1 wk of hypoxia (fraction of inspired oxygen [FIO 2]= 10%). Exogenously administered inhaled NO was evaluated in isolated lungs from normoxic and hypoxic rats. In normoxic rats, L-NAME (10-100 micro M) caused a dose-dependent increase in PVR in whole (hematocrit [Hct] 40%) and diluted (Hct 12%) blood-perfused lungs. L-NAME (10-800 micro M) had no effect in isolated lungs perfused with a modified salt solution of equal viscosity to blood either alone, or containing plasma (50%) or free oxyhemoglobin (10 micro M). In whole blood perfused lungs, L-NAME (100 micro M) increased PVR more in hypoxic versus normoxic isolated lungs (141% vs 100%). Inhaled NO decreased PVR in isolated lungs from hypoxic rats and partially reversed the effects of L-NAME, but had no effect in normoxic lungs. In conclusion, endogenous and inhaled NO modulate PVR in isolated rat lungs and this role is increased by prolonged hypoxia. The response to inhibition of endogenous NO is dependent on the presence of red blood cells and is independent of the changes in viscosity or the presence of oxyhemoglobin or plasma.