Nitric oxide synthase (NOS) inhibition has been shown in humans to attenuate exercise-induced increases in muscle glucose uptake. We examined the effect of infusing the NO precursor L-arginine (L-Arg) on glucose kinetics during exercise in humans. Nine endurance-trained males cycled for 120 min at 72±1% VO₂ peak followed immediately by a 15 min "all-out" cycling performance bout. A [6,6-²H]glucose tracer was infused throughout exercise and either saline alone (Control, CON) or saline containing L-Arg HCl (L-Arg, 30 g at 0.5g/min) was co-infused in a double blind, randomized order during the last 60 min of exercise. L-Arg augmented the increases in glucose Ra, glucose Rd and glucose clearance rate (L-Arg: 16.1±1.8 ml.min^-1.kg^-1, CON: 11.9±0.7 ml.min^-1.kg^-1 at 120 min. P<0.05) during exercise with a net effect of reducing plasma glucose concentration during exercise. L-Arg infusion had no significant effect on plasma insulin concentration but attenuated the increase in non-esterified fatty acid and glycerol concentrations during exercise. L-Arg infusion had no effect on cycling exercise performance. In conclusion, L-Arg infusion during exercise significantly increases skeletal muscle glucose clearance in humans. Since plasma insulin concentration was unaffected by L-Arg infusion, greater NO production may have been responsible for this effect.