Elmendorf, Jeffrey S.Habegger, Kirk M.Roach, Peter J.Brozinick, Joseph T.Sturek, Michael S.Considine, Robert V.2010-01-132010-01-132009-10https://hdl.handle.net/1805/2032http://dx.doi.org/10.7912/C2/1850Indiana University-Purdue University Indianapolis (IUPUI)Study has shown that plasma membrane (PM) cholesterol and cortical filamentous actin (F-actin) influence skeletal muscle glucose transport. Of fundamental and clinical interest is whether diabetogenic insults promote membrane/cytoskeletal dysfunction amendable for therapy. As exposure to excess fatty acid (FA)s induce glucose intolerance by mechanisms imperfectly understood, we tested if PM cholesterol/F-actin changes could contribute to FA-induced glucose transporter GLUT4 dysregulation in skeletal muscle. High-fat fed, insulin-resistant animals displayed elevated levels of skeletal muscle PM cholesterol and a loss in cortical F-actin, compared to normal-chow fed animals. Consistent with a PM cholesterol component of glucose intolerance, human skeletal muscle biopsies revealed an inverse correlation between PM cholesterol and whole-body glucose disposal. Mechanistically, exposure of L6 myotubes to the saturated FA palmitate induced an increase in PM cholesterol that destabilized actin filaments and decreased insulin-stimulated PM GLUT4 and glucose transport, which could be reversed with cholesterol lowering. Next, study tested if the lipid-lowering action of the antidiabetic AMP-activated protein kinase (AMPK) had a beneficial influence on PM cholesterol balance. Consistent with AMPK inhibition of 3-hydroxy-3-methylglutaryl CoA reductase, a rate-limiting enzyme of cholesterol synthesis, we found that AMPK activation promoted a significant reduction in PM cholesterol and amplified basal and insulin-stimulated GLUT4 translocation. A similar loss of PM cholesterol induced by β-cyclodextrin caused an analogous enhancement of GLUT4 regulation. Interestingly, PM cholesterol replenishment abrogated the AMPK effect on insulin, but not basal, regulation of GLUT4 translocation. Conversely, AMPK knockdown prevented the enhancement of both basal and insulin-stimulated GLUT4 translocation. As a whole these studies show PM cholesterol accrual and cortical F-actin loss uniformly in skeletal muscle from glucose-intolerant mice, swine, and humans. In vivo and in vitro dissection demonstrated this membrane/cytoskeletal derangement induces insulin resistance and is promoted by excess FAs. Parallel studies unveiled that the action of AMPK entailed lowering PM cholesterol that enhanced the regulation of GLUT4/glucose transport by insulin. In conclusion, these data are consistent with PM cholesterol regulation being an unappreciated aspect of AMPK signaling that benefits insulin-stimulated GLUT4 translocation during states of nutrient excess promoting PM cholesterol accrual.en-USGlucose IntoleranceGlucose TransportSkeletal MuscleCholesterolMembraneGlucoseMusculoskeletal systemThesis