Background: The potential biological significance of hydrogen sulfide (H₂S) has attracted growing interests in recent years, but its role in the myogenic response of rat cerebral arterioles has not been explored. Methods and Results: Rats were injected with NaHS (an H₂S donor, 2-200μmol·kg^-1·day^-1, i.p.) or saline for 3 weeks. MBP was measured with a tail-cuff method. Cerebral arterioles were isolated and cannulated in an organ bath system, and vessel diameters were measured with an image-shearing device. Changes in diameter in response to stepwise increases in intravascular pressure (20-120mmHg) were investigated under no-flow conditions. After the treatments, plasma H₂S increased and MBP decreased significantly. NaHS reduced the myogenic response in a dose-dependent manner. This effect was markedly attenuated by glibenclamide, a K_ATP channel blocker. Blockade of nitric oxide (NO) production with NG-nitro-L-arginine methyl ester (L-NAME, a NO synthase inhibitor) enhanced, whereas removal of the endothelium abolished the inhibitory role of NaHS on the myogenic response. Conclusions: For the first time it has been demonstrated that H₂S decreases the myogenic response of cerebral arterioles in vivo, and this effect is endothelium-dependent and partially mediated by K_ATP channels. (Circ J 2012; 76: 1012-1019)