Vascular smooth muscle cell (VSMC) migration plays a key role in tissue repair after arterial wall injury. VSMC migration requires integration of chemical and mechanical signaling mechanisms. Recently, we showed that epithelial Na(+) channel (ENaC) proteins are expressed in VSMCs and that ENaC inhibition abolishes pressure-induced constriction in isolated artery segments. However, whether ENaC proteins play a role in VSMC migration is unknown. The goal of this study was to determine whether ENaC molecules are required for VSMC migration. Using RT-PCR, immunoblotting, and immunolabeling, we detected expression of alpha-, beta-, and gammaENaC transcripts and proteins in cultured VSMCs (SV40-LT and A10 cells). Of the three proteins, betaENaC was the most readily detected in both cell lines by immunolocalization and Western blotting. Inhibition of ENaC activity with 1 microM benzamil blunted VSMC migration associated with wound healing (40.3% at 8 h and 26.2% at 24 h) and in response to the chemotactic stimulant platelet-derived growth factor-BB (38.1%). Furthermore, silencing ENaC gene expression with small interfering RNA blunted VSMC migration. These data indicate that expression of ENaC proteins is required for normal VSMC migration and suggest a potential new role for ENaC proteins in vascular tissue repair.