Electroporated GLUT4‐7myc‐GFP detects in vivo glucose transporter 4 translocation in skeletal muscle without discernible changes in GFP‐patterns

New Findings

What is the central question of this study?

Resolving the mechanism(s) leading to glucose transporter 4 (GLUT4) translocation to the muscle surface membrane has great therapeutic potential. However, the measurement of GLUT4 translocation is technically challenging. Here, we asked whether electroporation of GLUT4‐7myc‐GFP into skeletal muscle could be used as a tool to study GLUT4 translocation in vivo.

What is the main finding and its importance?

By acutely inducing GLUT4‐7myc‐GFP expression in skeletal muscle, we verified that in vivo exercise and AICAR stimulation increased the GLUT4 presence in the sarcolemma measured as myc‐signal. Importantly, the increased myc‐signal in the sarcolemma was not accompanied by major visual changes in the distribution of the GFP signal.

Abstract

Insulin and exercise lead to translocation of the glucose transporter 4 (GLUT4) to the surface membrane of skeletal muscle fibres. This process is pivotal for facilitating glucose uptake into skeletal muscle. To study this, a robust assay to directly measure the translocation of GLUT4 in adult skeletal muscle is needed. Here, we aimed to validate a simple GLUT4 translocation assay using a genetically encoded biosensor in mouse skeletal muscle. We transfected GLUT4‐7myc‐GFP into mouse muscle to study live GLUT4 movement and to evaluate GLUT4 insertion in the muscle surface membrane following in vivo running exercise and pharmacological activation of AMP activated protein kinase (AMPK). Transfection led to expression of GLUT4‐7myc‐GFP that were dynamic in live flexor digitorum brevis fibres and which, upon insulin stimulation, exposed the myc‐epitope extracellularly. Running exercise, as well as AMPK‐activation by 5‐Aminoimidazole‐4‐carboxamide ribonucleotide, induced ∼125% and ∼100% increase in extracellularly exposure of GLUT4 in the surface membrane of tibialis anterior muscle. Interestingly, the clear increase in surface‐exposed GLUT4 content by insulin, exercise or AMPK activation was not accompanied by any discernible reorganization of the GLUT4‐GFP signal. In conclusion, we provide a detailed description of an easy to use translocation assay to study GLUT4 accumulation at the surface membrane by exercise and exercise‐mimicking stimuli. Notably, our analyses revealed that increased GLUT4 surface membrane accumulation was not accompanied by a discernible change in the GLUT4 localization pattern.

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