Imaging GLUT4 translocation in human skeletal muscle fibres — ASN Events
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  3. Imaging GLUT4 translocation in human skeletal muscle fibres

Imaging GLUT4 translocation in human skeletal muscle fibres (#234)

Christopher S Shaw 1 , Helen Bradley 2 , Oliver J Wilson 2 , Anton JM Wagenmakers 2
  1. Institute of Sport, Exercise and Active Living, Victoria University, Melbourne, Australia
  2. School of Sport and Exercise Sciences, University of Birmingham, Birmingham, UK

Defects in insulin-mediated GLUT4 translocation and skeletal muscle glucose uptake precede the development of type 2 diabetes 1. Insulin and contraction-mediated GLUT4 translocation involves activation of specific signalling events and docking, fusion and dispersal of GLUT4-containing vesicles with the sarcolemmal and T-tubule cell surface membranes 2,3. However, these processes remain poorly understood in human skeletal muscle. This study aimed to investigate the translocation of GLUT4 and the localisation of the membrane fusion protein SNAP23 in human skeletal muscle in response to muscle contraction and elevations in plasma insulin concentrations following glucose feeding.

Muscle biopsies were taken from ten insulin sensitive males (age 21±1 years, BMI 23.1±1.0 kg/m2) before and immediately after 30 minutes of exercise performed at 65% VO2max and before and ninety minutes following consumption of a 75 g glucose bolus. GLUT4 and SNAP23 localisation was investigated using immunofluorescence in combination with dystrophin as a marker of the plasma membrane. Images were viewed using confocal immunofluorescence microscopy and Pearson’s correlation coefficient was used to quantify colocalisation.

The specificity of GLUT4 and SNAP23 antibodies was confirmed through a combination of GFP transfection, western blotting and peptide competition experiments. Basal GLUT4 staining presented distinct spots which were particularly abundant in perinuclear and (sub)sarcolemmal regions. Following exercise, colocalisation of GLUT4 with the plasma membrane marker dystrophin increased (P<0.01), which is indicative of GLUT4 translocation. GLUT4 and dystrophin colocalisation ninety minutes after glucose ingestion was unchanged (P>0.05). SNAP23 staining was associated with plasma membrane regions and was not changed by either stimulus (P>0.05).

GLUT4 translocation to the plasma membrane in response to muscle contraction was confirmed using immunofluorescence techniques in human skeletal muscle, which occurred without a change in SNAP23 localisation. GLUT4 translocation in response to physiological increases in plasma insulin concentrations requires further investigation.

  1. Petersen KF, Dufour S, Befroy D, Garcia R, Shulman GI (2004) Impaired mitochondrial activity in the insulin-resistant offspring of patients with type 2 diabetes. N Engl J Med 350:664-671.
  2. Lauritzen HPMM, Galbo H, Toyoda T, Goodyear LJ (2010) Kinetics of contraction-induced GLUT4 translocation in skeletal muscle fibers from living mice. Diabetes 59:2134-2144.
  3. Lauritzen HPMM, Ploug T, Prats C, Tavaré JM, Galbo H (2006) Imaging of insulin signalling in skeletal muscle of living mice shows major role of T-tubules. Diabetes 55:1300-1306.