We examined the effect of intraluminal flow of physiological saline on the membrane potential of vascular smooth muscle cells in isolated rabbit cerebral arteries. Intraluminal flow (20 μL/min) caused a depolarization of 4.8±0.7 mV in muscle cells with a resting membrane potential of −62.5±1.2 mV (n=19). However, when cells were depolarized to −48.7±1.8 mV using histamine and serotonin, the response to intraluminal flow was the opposite, a hyperpolarization of 5.6±1.0 mV (n=9). These opposing effects of flow on membrane potential appear to balance at −57.8±1.1 mV (n=31). Our results suggest that intraluminal flow may affect the level of basal tone present in arteries in vivo through modulating the membrane potential of vascular smooth muscle cells by concurrently activated depolarization and hyperpolarization.