In vascular injury, the mechanisms responsible for contractile smooth muscle phenotype are altered, leading to vascular smooth muscle proliferation and vascular diseases such as atherosclerosis and neointimal hyperplasia. We previously reported a change in ion handling when the cells switch phenotype (Cheong et al, 2005; Kumar et al, 2006). This study presents data suggesting a novel role for the voltage-gated potassium channel K_V1.3 in mediating vascular proliferation. Vascular smooth muscle cells were obtained from aorta dissected post mortem from male eight-week old mice (Fountain et al, 2004) or from discarded human saphenous veins obtained at coronary artery bypass graft surgery with ethical consent. Quantitative real-time RT-PCR analysis of mouse vascular smooth muscle cell mRNA revealed upregulation of KCNA3 gene (encoding K_V1.3) in proliferating cells and loss of other K_V1 mRNA species. As a model of vascular injury, a linear wound was made in the culture, removing cells from a predefined region. Cell response to the injury was inhibited by the K_V1.3 blockers margatoxin and correolide compound C in mouse and human vascular smooth muscle cells. Ionic currents measured by whole-cell patch-clamp in human smooth muscle cells were consistent with the presence of active K_V1.3 channels. Margatoxin and correolide compound C, as well as Psora-4 (a third KV1.3 blocker) inhibited a component of the outward current elicited by a 500ms depolarising step to +40mV from a holding potential of -60mV. The threshold of activation of the current was near -40mV. To assess more directly the potential relevance to smooth muscle adaptation in situ, we grew neointimal formations within segments of human saphenous vein as previously described (Cheong et al, 2005). The new cells in the neointimal formations are almost exclusively alpha-actin positive smooth muscle cells and expressed K_V1.3 protein at higher levels than cells in pre-existing vein. Furthermore, using paired vein segments from the same patient, we reveal that treatment with margatoxin or correolide compound C significantly reduced the neointimal growth compared to vehicle control. Together, these data indicate that K_V1.3 is upregulated in proliferating smooth muscle cells and that blockers of K_V1.3 may have therapeutic potential as agents acting against diseases caused or exacerbated by smooth muscle cell proliferation.