Inhibition of GLUT4 translocation to the sarcolemma by knockdown of Akt2.

(a) Expression vectors for GLUT4myc7-GFP, Myr-p110α, Rac1(G12V), together with either one of two siRNA duplexes against mouse Akt2 (#1 and #2) and a mixture of NC siRNA duplexes, were introduced into gastrocnemius muscle of wild-type mice. Insulin (175.5 μg/kg body weight) was administered intravenously. Endogenous Akt2 was visualized by immunofluorescent staining with an anti-Akt2 antibody. Myr-p110α and Rac1(G12V) were visualized by immunofluorescent staining with an anti-HA antibody. GLUT4myc7-GFP translocated to the sarcolemma was visualized by immunofluorescent staining with an anti-Myc antibody before cell permeabilization. Scale bar, 20 μm. (b) Translocation of GLUT4myc7-GFP to the sarcolemma shown in (a) was quantified. Gray, orange, and blue bars represent the treatment with NC, #1, and #2 siRNA duplexes, respectively. Data are shown as means ± S.E. (n = 6). *P < 0.001.