Insulin-induced cortical actin remodeling promotes GLUT4 insertion at muscle cell membrane ruffles

Insulin stimulates glucose uptake by recruiting glucose transporter 4 (GLUT 4) from an intracellular compartment to the cell surface; this phenomenon i s defective in type 2 diabetes. Here we examine the involvement of actin fi laments in GLUT4 translocation and their possible defects in insulin resist ance, using L6 myotubes expressing myc-tagged GLUT4. Insulin caused membran e ruffling, a dynamic distortion of the myotube dorsal surface. Fluorescenc e microscopy and immunogold staining of surface GLUT4myc coupled to backsca tter electron microscopy revealed a high density of this protein in membran e ruffles. The t-SNARES syntaxin4 and SNAP-23 were also abundant in these r egions. Below the membrane, GLUT4 and the vesicular protein VAMP2, but not VAMP3, colocalized with the actin structures supporting the membrane ruffle s. GLUT4myc externalization and membrane ruffles were reduced by jasplakino lide and by swinholide-A, drugs that affect actin filament stability and pr event actin branching, respectively. Insulin resistance generated by prolon ged (24 hours) exposure of myotubes to high glucose and insulin diminished the acute insulin-dependent remodeling of cortical actin and GLUT4myc trans location, reminiscent of the effect of swinholide-A. We propose that GLUT4 vesicle incorporation into the plasma membrane involves insulin-dependent c ortical actin remodeling and that defective actin remodeling contributes to insulin resistance.