The present study investigated the effect of one bout of moderate-intensity exercise on the adrenergic receptor-dependent and -independent vasoconstrictor response in rat aortas, and the role of nitric oxide (NO) bioavailability on these vasomotor responses. One group of rats was submitted to a 60 min of exercise at approximately 60% of maximal exercise capacity on a treadmill (exercise group) and the other one was placed in the treadmill without running (control group). Immediately after this period, both groups were euthanized and the thoracic aorta was removed to evaluate the vasoconstrictor response to norepinephrine and potassium chloride, and to evaluate the vascular nitrite and nitrate concentration. One bout of exercise attenuated the maximal contractile response to both norepinephrine and potassium chloride compared to control group. These differences on vascular reactivity were not observed in endothelium-denuded aortic rings and aortic rings pre-incubated with a nitric oxide synthesis inhibitor. Additionally, exercise group increased NO bioavailability (nitrite and nitrate concentration) as compared to control group. These results demonstrate that one bout of moderate-intensity exercise is able to attenuate adrenergic receptor-dependent and -independent vasoconstrictor response in rat aorta, mainly by increasing vascular NO bioavailability.