REGULATION OF CELL-SURFACE GLUT4 IN SKELETAL-MUSCLE OF TRANSGENIC MICE

Marked overexpression of the glucose transporter GLUT4 in skeletal mus cle membrane fractions of GLUT4 transgenic (TG) mice is accompanied by disproportionately small increases in basal and insulin-stimulated gl ucose transport activity. Thus we have assessed cell surface GLUT4 by photolabelling with the membrane-impermeant reagent -trifluoroethyl)be nzoyl]-1,3-bis(D-mannos-4-yloxy) (ATB-BMPA) and measured the correspon ding glucose transport activity using 2-deoxyglucose in isolated exten sor digitorum longus (EDL) muscles from non-transgenic (NTG) and GLUT4 TG mice in the absence and presence of 13.3 nM (2000 mu units/ml) ins ulin, without or with hypoxia as a model of muscle contraction. TG mic e displayed elevated rates of glucose trans port activity under basal and insulin-stimulated conditions, and in the presence of insulin plus hypoxia, compared with NTG mice. Photoaffinity labelling of cell surf ace GLUT4 indicated corresponding elevations in plasma membrane GLUT4 in the basal and insulin-stimulated states, and with insulin plus hypo xia, but no difference in cell surface GLUT4 during hypoxia stimulatio n. Subcellular fractionation of hindlimb muscles confirmed the previou sly observed 3-fold overexpression of GLUT4 in the TG compared with th e NTG mice. These results suggest that: (1) alterations in glucose tra nsport activity which occur with GLUT4 overexpression in EDL muscles a re directly related to cell surface GLUT4 content, regardless of the l evels observed in the corresponding subcellular membrane fractions, (2 ) while overexpression of GLUT4 influences both basal and insulin-stim ulated glucose transport activity, the response to hypoxia/contraction -stimulated glucose transport is unchanged, and (3) subcellular fracti onation provides little insight into the subcellular trafficking of GL UT4, and whatever relationship is demonstrated in EDL muscles from NTG mice is disrupted on GLUT4 overexpression.