DISRUPTION OF GLUT1 GLUCOSE CARRIER TRAFFICKING IN L6E9 AND SOL8 MYOBLASTS BY THE PHOSPHATIDYLINOSITOL 3-KINASE INHIBITOR WORTMANNIN

In this study we have used wortmannin, a highly specific inhibitor of phosphatidylinositol (PI) 3-kinase, to assess the role of this enzyme on GLUT1 glucose carrier distribution and glucose transport activity i n myoblasts from two skeletal-muscle cell lines, L6E9 and Sol8. As det ected in L6E9 cells, myoblasts exhibited basal and insulin-stimulated PI 3-kinase activities. Incubation of intact myoblasts with wortmannin resulted in a marked inhibition of both basal and insulin-stimulated PI 3-kinase activities. L6E9 and Sol8 myoblasts showed basal and insul in-stimulated glucose transport activities, both of them inhibited by wortmannin in a dose-dependent manner (IC50 approximate to 10-20 nM). Concomitantly, immunofluorescence analysis revealed that 1 h treatment with wortmannin led to a dramatic intracellular accumulation of GLUT1 carriers (the main glucose transporter expressed in L6E9 and Sol8 myo blasts) in both cell systems. The effect of wortmannin on GLUT1 cellul ar redistribution was independent of the presence of insulin. The cell ular distribution of two structural plasma-membrane components such as beta(1)-integrin or the alpha(1) subunit of the Na+-K+-ATPase were un affected by wortmannin in both the absence and the presence of insulin . As a whole, our results indicate that PI 3-kinase is necessary to ba sal and insulin-stimulated glucose transport in L6E9 and Sol8 myoblast s. Moreover, immunofluorescence assays suggest that in both cellular m odels there is a constitutive GLUT 1 trafficking pathway (independent of insulin) that involves PI 3-kinase and which, when blocked, locks G LUT1 in a perinuclear compartment.