We have previously shown that intercellular communication in the vascular wall is regulated through an interaction between the Na,K-pump and the Na/Ca-exchanger. The interaction controls [Ca]_i in discrete sites near the sarcolemma, which modulates gap junction conductance. We hypothesize here that this mechanism modulates synchronized smooth muscle cell (SMC) oscillatory activity in blood vessels, i.e. vasomotion. We transfected SMCs in culture and rat mesenteric small arteries (MSA) in vivo with siRNA against the a2 isoform of the Na,K-pump and with eGFP-siRNA. Measurement of membrane capacitance (Cm) in pairs of SMCs were used for assessment of electrical coupling. Isometric force development of transfected arteries were obtained in vitro 3 days after transfection. Non-transfected and eGFP-siRNA transfected SMCs were well coupled electrically: SMCs pair Cm (90.4±9.4pF, n=7) was twice that of single cell Cm. Pairs of SMCs were uncoupled in the presence of 10 mM ouabain (46.1±7.2pF). The gap junction uncoupler b-glycyrrhetinic acid (bGA) had no additive effect (41.9±4.9pF). Downregulation of the a2 Na,K-pump significantly reduced coupling (33.4±4.1pF) and in these cells ouabain had no effect (34.1±4.2pF). Transfection of MSA in vivo downregulated the a2 Na,K-pump mRNA expression to 12±3% (n=8) compared to non-transfected controls. a2 Na,K-pump mRNA in eGFP-siRNA transfected arteries did not differ from non-transfected controls (81±21%, n=8). Arteries with reduced a2 Na,K-pump had significantly reduced vasomotion amplitude (26.9±25.5%, n=7) relative to non-transfected controls. Vasomotion amplitude in eGFP-siRNA transfected arteries was not changed (91.9±13.9%, n=8) compared to non-transfected controls. The data confirms the importance of the a2 isofom of the Na,K-pump for intercellular coupling in the vascular wall and demonstrates its involvement in the regulation of vasomotion.