Exofacial Epitope–Specific Antibodies Detect GLUT4 Translocation in Adult Human, Rat, and Mouse Skeletal Muscle

by sugaradmin



Skeletal muscle glucose transporter 4 (GLUT4) translocation to the plasma membrane determines glucose uptake in response to insulin and exercise and is disrupted in insulin resistance, making its experimental measurement critical. Confocal light microscopy is widely used for this purpose because of its ability to provide quantitative, high-resolution spatial information from small tissue amounts. However, conventional immunofluorescence colocalization microscopy lacks sensitivity and specificity in the detection of GLUT4 translocation. We validated the use of exofacial epitope–specific GLUT4 antibodies to quantify sarcolemmal GLUT4 translocation in fixed, nonpermeabilized adult human and rodent muscle fibers. Across human, mouse, and rat muscles, these antibodies sensitively detected stimulus-induced GLUT4 translocation, and labeling was abolished in muscle-specific GLUT4-knockout muscle, confirming specificity. Importantly, this study includes the first unambiguous visualization of endogenous GLUT4 translocation in intact human skeletal muscle fibers after insulin stimulation and exercise. In TBC1D4-knockout rats, insulin-stimulated GLUT4 translocation was absent despite wild-type–level GLUT4 expression, confirming an essential role for TBC1D4 in this process. Thus, exofacial GLUT4 antibodies provide a straightforward, sensitive, and specific approach to quantify endogenous GLUT4 translocation in fixed adult skeletal muscle.

Article Highlights
  • Reliable quantification of glucose transporter 4 (GLUT4) translocation in intact skeletal muscle is essential for understanding insulin and exercise responses but remains technically challenging.
  • We aimed to test whether exofacial GLUT4 antibodies can specifically detect sarcolemmal GLUT4 translocation in fixed, nonpermeabilized muscle fibers from humans and rodents.
  • GLUT4 translocation in response to insulin, AMPK activation, and exercise was detectable in human and rodent muscles. Insulin-stimulated translocation correlated with 2-deoxyglucose uptake and was abolished in TBC1D4-knockout muscle.
  • Exofacial GLUT4 antibodies enable straightforward, specific quantification of endogenous GLUT4 translocation in rodent and human muscles in healthy and insulin-resistant states.





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