IDENTIFICATION AND CHARACTERIZATION OF AN EXERCISE-SENSITIVE POOL OF GLUCOSE TRANSPORTERS IN SKELETAL-MUSCLE

Augmentation of glucose transport into skeletal muscle by GLUT4 transl ocation to the plasma and T-tubule membranes can be mediated independe ntly by insulin and by contraction/exercise. Available data suggest th at separable pools of intracellular GLUT4 respond to these two stimuli . To identify and characterize these pools, we fractionated skeletal m uscle membranes in a discontinuous sucrose density gradient, Fractions of 32 and 36% sucrose exhibited the highest enrichment of GLUT4 and w ere independently responsive to insulin and exercise, respectively. Th e combination of the two stimuli depleted both GLUT4 fractions simulta neously. Both vesicle populations contained the gp160 aminopeptidase, whose expression had previously been shown to be specific to muscle an d fat and restricted to GLUT4 vesicles in the latter tissue. In muscle , gp160 translocates exactly as does GLUT4 in response to insulin and exercise. The contraction- and insulin sensitive GLUT4 pools also cont ained secretory component-associated membrane protein/glucose transpor ter vesicle triplet but not GLUT1 and caveolin, Immunoadsorption of th e two pools followed by silver staining did not reveal any obvious dif ference in their major protein components. On the other hand, sediment ational analysis in sucrose velocity gradients revealed that the insul in sensitive GLUT4 vesicles had a larger sedimentation coefficient tha n the exercise-sensitive vesicles, Thus, the separation of the two int racellular GLUT4 pools should be useful in dissecting what are likely to be different signal transduction pathways that mediate their transl ocation to the cell surface.