Abstract

Objective. Insulin stimulates muscle glucose transport by translocation of GLUT4 to sarcolemma and t-tubules. Despite muscle glucose uptake plays a major role in insulin resistance and type II diabetes, the temporal and spatial changes in insulin signalling and GLUT4 translocation during these conditions are not well described.

Research Design and Methods. We used time-lapse confocal imaging of GFP-ARNO (evaluation of PI3-K activation) and GLUT4-GFP transfected quadriceps muscle in living, anaesthetized mice, either muscle denervated or high-fat fed. T-tubules were visualized with dye Sulforhodamine B. In incubated muscle glucose transport was measured by 2-deoxy-D-[³H]-glucose uptake, and functional detubulation was carried out by osmotic shock. Muscle fibers were immunostained for insulin receptors.

Results. Denervation and high-fat diet reduced insulin-mediated glucose transport. In denervated muscle insulin-stimulated PIP3 production was abolished in t-tubules, while PIP3 production at sarcolemma was increased 2.6-fold. Correspondingly, GLUT4-GFP translocation to t-tubules was abolished, while translocation to sarcolemma was increased 2.3-fold. In high-fat fed mice a ∼65% reduction in both insulin-induced t-tubular PIP3 production and GLUT4-GFP translocation was seen. Sarcolemma was less affected with reductions of ∼40 % in PIP3 production and ∼15 % in GLUT4-GFP translocation. Access to t-tubules was not compromised and insulin receptor distribution in sarcolemma and t-tubules was unaffected by denervation or high-fat feeding. Detubulation of normal muscle reduced basal and abolished insulin-induced glucose transport.

Conclusions. Our findings demonstrate, for the first time, that impaired insulin signalling and GLUT4 translocation is compartmentalized in muscle and primarily localized to t-tubules and not sarcolemma during insulin resistance.