Human airway epithelial cells express β-adrenergic receptors (β-AR) that regulate mucociliary clearance by stimulating transepithelial anion transport and ciliary beat frequency. Previous studies using airway epithelial cells showed that stimulation with isoproterenol increased cell migration and wound repair by a cAMP-dependent mechanism. In the present study, impedance sensing arrays were used to measure cell migration and epithelial restitution following wounding of confluent normal human bronchial epithelial (NHBE) cells and Calu-3 cells by electroporation. Stimulation with epinephrine or the β₂-selective agonist salbutamol significantly delayed wound closure and reduced the mean surface area of lamellipodia protruding into the wound. Treatment with the β-AR bias agonist carvedilol or isoetharine also produced a delay in epithelial restitution similar in magnitude to epinephrine and salbutamol. Measurements of extracellular-signal-regulated kinase (ERK) phosphorylation following salbutamol or carvedilol stimulation showed no significant change in the level of phosphorylation compared to untreated control cells. However, inhibition of PP2A phosphatase activity completely blocked the delay in wound closure produced by β-agonists. In CFTR silenced Calu-3 cells, salbutamol did not inhibit wound repair, suggesting that β-agonist stimulation and loss of CFTR expression inhibit cell migration through a common pathway. Confocal images of the basal membrane of Calu-3 cells labeled with anti-β₁ integrin antibody showed that treatment with epinephrine or carvedilol reduced the level of activated integrin in the membrane. These findings suggest that β-AR agonists delay airway epithelial repair by a G-protein and cAMP-independent mechanism involving the PP2A phosphatase and a reduction in β₁ integrin activation in the basal membrane.