Immunoelectron microscopic evidence that GLUT4 translocation explains the stimulation of glucose transport in isolated rat white adipose cells

We used an improved cryosectioning technique in combination with quantitati ve immunoelectron microscopy to study GLUT4 compartments in isolated rat wh ite adipose cells, We provide clear evidence that in unstimulated cells mos t of the GLUT4 localizes intracellularly to tubulovesicular structures clus tered near small stacks of Golgi and endosomes, or scattered throughout the cytoplasm, This localization is entirely consistent with that originally d escribed in brown adipose tissue, strongly suggesting that the GLUT4 compar tments in white and brown adipose cells are morphologically similar Further more, insulin induces parallel increases (with similar magnitudes) in gluco se transport activity, approximately 16-fold, and cell-surface GLUT4, appro ximately 12-fold, Concomitantly, insulin decreases GLUT4 equally from all i ntracellular locations, in agreement with the concept that the entire cellu lar GLUT4 pool contributes to insulin-stimulated exocytosis, In the insulin -stimulated state, GLUT4 molecules are not randomly distributed on the plas ma membrane, but neither are they enriched in caveolae, Importantly, the to tal number of GLUT4 C-terminal epitopes detected by the immuno-gold method is not significantly different between basal and insulin-stimulated cells, thus arguing directly against a reported insulin-induced unmasking effect. These results provide strong morphological evidence (1) that GLUT4 compartm ents are similar in all insulin-sensitive cells and (2) for the concept tha t GLUT4 translocation almost fully accounts for the increase in glucose tra nsport in response to insulin.