Regulation of glucose transporter isoform 4 (Glut4) expression and function in primary and cultured skeletal muscle

Skeletal muscle is the largest tissue in the human body and the predominant site for insulin stimulated glucose clearance. This process requires the tissue specific expression and movement of glucose transporter isoform 4 (Glut4) from an intracellular vesicular pool to the cell surface. The perturbation of this process at various levels disrupts cellular glucose utilization and causes pathophysiological defects such as type 2 diabetes mellitus. Thus, understanding factors that regulate Glut4 expression and the efficiency of Glut4 translocation in muscle is important. Denervation of rat hindlimb skeletal muscle is a model for insulin resistance, one that is similar to what is seen in humans as a result of spinal cord injury. Glut4 expression in denervated rat muscle is decreased by 80%. Using microarray mRNA profiling, the orphan nuclear receptor, Nur77, was identified as a possible regulator of Glut4 and other genes linked to glucose utilization. This role of Nur77 was confirmed by in vivo knock-down by electroporating Nur77-directed shRNA into skeletal muscle. This resulted in the down regulation of Glut4 and genes involved in glycolysis. Conversely, the ectopic expression of Nur77 in denervated rat muscle "rescued" the expression of Glut4 and the same set of glycolytic genes. These results demonstrate that Nur77 is a mediator of neuromuscular signaling in the control of metabolic genes required for muscle glucose utilization. Following its tissue expression in muscle, Glut4 function requires that it undergoes translocation to the cell surface, a process acutely regulated by exercise (contraction) as well as by insulin. For Glut4 translocation, an in vitro model to study muscle metabolism was generated by ectopic expression of human Glut4 in a skeletal muscle cell line, which allows broader and more facile experimental manipulations than muscle in situ. The signaling pathway for exercise in these cells was mimicked by AMP-dependent kinase (AMPK) activation and studied in comparison and in combination with the insulin signaling pathway. A synergistic effect on Glut4 translocation by activation of AMPK plus insulin stimulation was observed in 3 independent experimental approaches, and was attributed to the extended activation of the insulin signaling pathway by AMPK.