NAD(P)H oxidase‐dpendent H2O2 production and RyR‐IP3R activation are required for insulin induced GLUT4 translocation and glucose uptake in skeletal muscle cells

Abstract

Insulin‐regulated GLUT4 traffic has been widely explored, but the role of calcium and ROS signaling in this process is poorly understood.We studied insulin‐induced GLUT4 traffic in GLUT4myc skeletal muscle cells.Insulin‐induced exofacial exposure of myc epitope, glucose uptake and Akt activation were independent of extracellular calcium. The intracellular calcium chelator BAPTA‐AM inhibited Akt, p70S6K and ERK1 activation but not p‐ERK2. p‐ERK1/2 was inhibited by 2‐APB, an IP3R inhibitor. Cytosol directed parvalbumin reduced insulin‐dependent exposure of myc. Moreover, ryanodine and 2‐ APB partly inhibited myc exposure with an additive effect. The calcium ionophore ionomycin induced an increase in extracellular exposure of myc and was additive to insulin. In myotubes maintained in calcium‐free media, the effect of insulin was totally inhibited by ionomycin. Insulin increased H2O2 production measured with the cytosolic molecular sensor HyPer which was inhibited by NAC. p47 subunit of NADP(H)ox labeled differentiated myotubes. Both antioxidant agents, NAC and trolox partly reduced the externalization of myc as did the NADP(H)ox inhibitor apocynin and overexpression of catalase.These data suggest that insulin induces an increase in NADP(H)ox‐dependent H2O2 production and RyR/IP3R calcium channels activation to foster GLUT4 translocation.FONDAP 15010006, FONDECYT 3110170