The effect of nitric oxide (NO) exposure and sulfhydryl-reactive chemicals on l-arginine transport in pulmonary artery endothelial cells was evaluated. Exposure of pulmonary artery endothelial cells to 7.5 ppm (0.4 μM) NO for 4 h resulted in a significant (p < 0.05) reduction of Na + -dependent but not Na + -independentl -arginine transport. More prolonged exposure for 12-24 h reduced both Na + -dependent and Na + -independent transport of l-arginine with maximal loss of transport after 18 h of exposure (p < 0.02 for both). Similarly, incubation of cells in the presence of 50-200 μM S-nitroso-acetyl-penicillamine (SNAP) (but not 500 μM each of nitrate or nitrite) for 2 h also reduced both the Na + -dependent and Na + -independent transport of l-arginine (p < 0.05 for all concentrations). The SNAP-induced reduction of l-arginine transport was blocked by the NO scavenger oxyhemoglobin. When cell monolayers were exposed to varying concentrations of the sulfhydryl reactive chemicals N-ethylmaleimide (NEM) and acrolein, a dose-dependent reduction of l-arginine transport by both Na + -dependent and Na + -independent processes was observed. Na + -dependent l-arginine transport was more susceptible to inhibition by exposure to NO and to sulfhydryl reactive chemicals. Incubation of cells with 0.5 mM of the thiol-containing agent N-acetyl-l-cysteine prior to and during NEM or acrolein exposure blocked NEM and acrolein-induced reduction of l-arginine transport by both Na + -dependent and Na + -independent processes. Similarly, NO-induced reductions of Na + -dependent and Na + -independent l-arginine transport were reversed to control levels 24 h after termination of NO exposure. Treatment with the disulfide reducing agent dithiothreitol after exposure to NO resulted in partial reversal of the decreases in l-arginine transport. These results demonstrate that exposure to exogenous NO is responsible for reversible reductions of plasma membrane-dependent l-arginine transport mediated by both the Na + -dependent (system B o , + ) and the Na + -independent (system y + ) transport processes. Modulation of the sulfhydryl status of plasma membrane proteins involved in l-arginine transport, such as l-arginine transporters and/or Na + + -ATPase, may be responsible, at least in part, for reductions in overall l-arginine transport in pulmonary artery endothelial cells.