The Molecular Mechanism Of The Regulation Of AKT By Electrical Pulse In Mouse Skeletal Muscle Cells

Objective:Skeletal muscle is the biggest target tissue of insulin,which plays an important role in glucose uptake and insulin sensitivity of the whole body.Glucose transporter 4(GLUT4)is a transmembrane protein,mediated glucose uptake in skeletal muscle cells.Increase of GLUT4 translocation stimulated by exercise is good for skeletal muscle to uptake and utilize glucose and has an important significance in the treatment of insulin resistance and type 2 diabetes.However,the underlying molecular mechanism is still unclear.We found electrical pulse stimulation(EPS)-induced skeletal muscle cells contraction increased Akt phosphorylation.So in this study,we explore the molecular mechanism of EPS-stimulated Akt phosphorylation.Content: The study includes three parts: The first part:EPS induced C2C12-GLUT4 HA myotubes contraction.To detect whether EPS could induce GLUT4 translocation and increase Akt phosphorylation.Akt inhibitors we used to test the effects of EPS-stimualted GLUT4 translocation and the phosphorylation of Akt and TBC1D4.The second part : To study the mechanism of EPS-increased Akt phosphorylation.The third part:C57BL/6 mice were treated with different stimulus to study Rac1 activity and Akt phosphorylation in vivo.Methods: Cultured C2C12-GLUT4 HA skeletal muscle cells were differentiated into mature contractile myotubes.C2C12-GLUT4 HA cells were treated with EPS for 1 h(20 V,1 Hz,24 ms).Enzyme linked immunosorbent assay(ELISA)was used to detect the GLUT4 HA levels on plasma membrane.Chemical inhibitors,activators and si RNA of AMPK or Rac1 were used to study the mechanism of EPS-induced Akt phosphorylation.C57BL/6 mice were treated with different stimulus(insulin,AICAR,Ad-AMPK-CA,exercise)to detect Rac1 activity and Akt phosphorylation in vivo.Results: 1.EPS increased Akt phosphorylation and induced GLUT4 HA translocation in an Akt-dependent way in C2C12-GLUT4 HA skeletal muscle cells.2.EPS and AICAR increased Rac1 activity and Akt phosphorylation;PAK and Akt phosphorylation were inhibited by AMPK inhibitor Compound C.Both si Rac1 and Rac1 inhibitor Ⅱ inhibited EPS-and AICAR-induced Akt phosphorylation.3.Rac1 was activated by AICAR,Ad-AMPK-CA and exercise in vivo.Conclusion: EPS promots GLUT4 translocation by stimulating Akt phosphorylation via AMPK-Rac1 signaling pathways in C2C12-GLUT4 HA skeletal muscle cells.