Abstract

The activation of potassium channels and the ensuing hyperpolarization in skeletal myoblasts are essential for myogenic differentiation. However, the effects of K⁺ channel opening in myoblasts on skeletal muscle mass are unclear. Our previous study revealed that pharmacological activation of intermediate conductance Ca²⁺-activated K⁺ channels (IK_Ca channels) increases myotube formation. In this study, we investigated the effects of 5,6-dichloro-1-ethyl-1,3-dihydro-2H-benzimidazol-2-one (DCEBIO), a Ca²⁺-activated K⁺ channel opener, on the mass of skeletal muscle. Application of DCEBIO to C2C12 cells during myogenesis increased the diameter of C2C12 myotubes in a concentration-dependent manner. This DCEBIO-induced hypertrophy was abolished by gene silencing of IK_Ca channels. However, it was resistant to 1 µM but sensitive to 10 µM TRAM-34, a specific IK_Ca channel blocker. Furthermore, DCEBIO reduced the mitochondrial membrane potential by opening IK_Ca channels. Therefore, DCEBIO should increase myotube mass by opening of IK_Ca channels distributed in mitochondria. Pharmacological studies revealed that mitochondrial reactive oxygen species (mitoROS), Akt, and mammalian target of rapamycin (mTOR) are involved in DCEBIO-induced myotube hypertrophy. An additional study demonstrated that DCEBIO-induced muscle hypertrophic effects are only observed when applied in the early stage of myogenic differentiation. In an in vitro myotube inflammatory atrophy experiment, DCEBIO attenuated the reduction of myotube diameter induced by endotoxin. Thus, we concluded that DCEBIO increases muscle mass by activating the IK_Ca channel/mitoROS/Akt/mTOR pathway. Our study suggests the potential of DCEBIO in the treatment of muscle wasting diseases.