Carbon monoxide (CO) is generated endogenously by the enzyme heme oxygenase. Although CO is a known vasodilator, cellular signaling mechanisms are poorly understood and are a source of controversy. The goal of the present study was to investigate mechanisms of CO dilation in porcine cerebral arterioles. Data indicate that exogenous or endogenously produced CO is a potent activator of large-conductance Ca²⁺-activated K⁺ (K_Ca) channels and Ca²⁺ spark–induced transient K_Ca currents in arteriole smooth muscle cells. In contrast, CO is a relatively poor activator of Ca²⁺ sparks. To understand the apparent discrepancy between potent effects on transient K_Ca currents and weak effects on Ca²⁺ sparks, regulation of the coupling relationship between these events by CO was investigated. CO increased the percentage of Ca²⁺ sparks that activated a transient K_Ca current (ie, the coupling ratio) from ≈62% in the control condition to 100% and elevated the slope of the amplitude correlation between these events ≈2.6-fold, indicating that Ca²⁺ sparks induced larger amplitude transient K_Ca currents in the presence of CO. This signaling pathway for CO is physiologically relevant because ryanodine, a ryanodine-sensitive Ca²⁺ release channel blocker that inhibits Ca²⁺ sparks, abolished CO dilation of pial arterioles in vivo. Thus, CO dilates cerebral arterioles by priming K_Ca channels for activation by Ca²⁺ sparks. This study presents a novel dilatory signaling pathway for CO in the cerebral circulation and appears to be the first presents of a vasodilator that acts by increasing the effective coupling of Ca²⁺ sparks to K_Ca channels.