Background-From animal studies it emerged that nitric oxide is important for the modulation of CO 2 -mediated cerebral blood flow (CBF chemoregulation) but not for the pressor-dependent mechanism (mechanoregulation). This hypothesis was tested in 18 healthy subjects. Methods and Results-Peak velocity (PV), diastolic velocity (DV), and mean velocity (MV) were measured by transcranial Doppler of the middle cerebral artery. Chemoregulation was assessed during normocapnia, hypocapnia, and after inhaled mixture of 95% O 2 ϩ5% CO 2 . Mechanoregulation was evaluated by incremental doses of phenylephrine. Measurements were repeated during infusion of sodium nitroprusside (SNP). Regional cerebrovascular resistance (CVR) was calculated as mean blood pressure (BP)/MV. SNP infusion decreased mean BP by 7 mm Hg and CVR decreased from 1.38Ϯ0.08 to 1.29Ϯ0.09 mm Hg/cm ⅐ s Ϫ1 ; Pϭ0.01, resulting in unaffected CBF. Phenylephrine (25 to 250 g) caused a similar increase in BP in a dose-response fashion before and during SNP infusion. Despite the increments in BP and CVR, CBF remained unaffected. During hyperventilation (end-tidal CO 2 Ϸ24 mm Hg), CVR increased by 75Ϯ3% and PV and DV decreased by 27Ϯ2% and 43Ϯ2%, respectively (PϽ0.001 for all). SNP infusion blunted the vasoconstrictive effect of hypocapnia; CVR increased only by 57Ϯ5%, and PV and DV decreased by 23Ϯ2% and 35Ϯ3%, respectively, (PϽ0.05 for all). Similarly, SNP augmented the vasodilatory effect of hypercapnia. Conclusions-Exogenous nitric oxide donor affects the basal cerebral vascular tone without affecting the CBF mechanoregulation. However, it selectively affects only the chemoregulatory mechanism (CO 2 -dependent). Thus, the CO 2 -NO axis is a cardinal pathway for CBF regulation in humans.