Modafinil inhibits KCₐ3.1 currents and muscle contraction via a cAMP-dependent mechanism

Modafinil has been used as a psychostimulant for the treatment of narcolepsy. However, its primary mechanism of action remains elusive. Therefore, we examined the effects of modafinil on KCₐ3.1 channels and vascular smooth muscle contraction. KCₐ3.1 currents and channel activity were measured using a voltage-clamp technique and inside-out patches in mouse embryonic fibroblast cell line, NIH-3T3 fibroblasts. Intracellular adenosine 3′,5′-cyclic monophosphate (cAMP) concentration was measured, and the phosphorylation of KCₐ3.1 channel protein was examined using western blotting in NIH-3T3 fibroblasts and/or primary cultured mouse aortic smooth muscle cells (SMCs). Muscle contractions were recorded from mouse aorta and rat pulmonary artery by using a myograph developed in-house. Modafinil was found to inhibit KCₐ3.1 currents in a concentration-dependent manner, and the half-maximal inhibition (IC₅₀) of modafinil for the current inhibition was 6.8±0.7nM. The protein kinase A (PKA) activator forskolin also inhibited KCₐ3.1 currents. The inhibitory effects of modafinil and forskolin on KCₐ3.1 currents were blocked by the PKA inhibitors PKI₁₄–₂₂ or H-89. In addition, modafinil relaxed blood vessels (mouse aorta and rat pulmonary artery) in a concentration-dependent manner. Modafinil increased cAMP concentrations in NIH-3T3 fibroblasts or primary cultured mouse aortic SMCs and phosphorylated KCₐ3.1 channel protein in NIH-3T3 fibroblasts. However, open probability and single-channel current amplitudes of KCₐ3.1 channels were not changed by modafinil. From these results, we conclude that modafinil inhibits KCₐ3.1 channels and vascular smooth muscle contraction by cAMP-dependent phosphorylation, suggesting that modafinil can be used as a cAMP-dependent KCₐ3.1 channel blocker and vasodilator.