In vitro analysis of the glucose-transport system in GLUT4-null skeletal muscle.
We have characterized the glucose-transport system in soleus muscle from female GLUT4-null mice to determine whether GLUT1, 3 or 5 account for insulin-stimulated glucose-transport activity. Insulin increased 2-deoxyglucose uptake 2.8- and 2.1-fold in soleus muscle from wild-type and GLUT4-null mice, respectively. Cytochalasin B, an inhibitor of GLUT1- and GLUT4-mediated glucose transport, inhibited insulin-stimulated 2-deoxyglucose uptake by >95% in wild-type and GLUT4-null soleus muscle. Addition of 35 mM fructose to the incubation media was without effect on insulin-stimulated 3-O-methylglucose transport activity in soleus muscle from either genotype, whereas 35 mM glucose inhibited insulin-stimulated (20 nM) 3-O-methylglucose transport by 65% in wild-type and 99% in GLUT4-null mice. We utilized the 2-N-4-1-(1-azi-2,2,2-triflu oroethyl)benzoyl-1, 3-bis(D-mannose-4-yloxy)-2-propylamine (ATB-BMPA) exofacial photolabel to determine if increased cell-surface GLUT1 or GLUT4 content accounted for insulin-stimulated glucose transport in GLUT4-null muscle. In wild-type soleus muscle, cell-surface GLUT4 content was increased by 2.8-fold under insulin-stimulated conditions and this increase corresponded to the increase in 2-deoxyglucose uptake. No detectable cell-surface GLUT4 was observed in soleus muscle from female GLUT4-null mice under either basal or insulin-stimulated conditions. Basal cell-surface GLUT1 content was similar between wild-type and GLUT4-null mice, with no further increase noted in either genotype with insulin exposure. Neither GLUT3 nor GLUT5 appeared to account for insulin-stimulated glucose-transport activity in wild-type or GLUT4-null muscle. In conclusion, insulin-stimulated glucose-transport activity in female GLUT4-null soleus muscle is mediated by a facilitative transport process that is glucose- and cytochalasin B-inhibitable, but which is not labelled strongly by ATB-BMPA.
Ryder
J W
JW
Department of Clinical Physiology, Karolinska Hospital, S-171 76, Stockholm, Sweden.
Kawano
Y
Y
Chibalin
A V
AV
Rincón
J
J
Tsao
T S
TS
Stenbit
A E
AE
Combatsiaris
T
T
Yang
J
J
Holman
G D
GD
Charron
M J
MJ
Zierath
J R
JR
D019325:3-O-Methylglucose / Q000378:metabolism; D000345:Affinity Labels; D000818:Animals; D001386:Azides; D001693:Biological Transport, Active / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism; D003571:Cytochalasin B / Q000494:pharmacology; D003847:Deoxyglucose / Q000378:metabolism; D004187:Disaccharides; D005260:Female; D015870:Gene Expression; D005947:Glucose / Q000378:metabolism*; D051272:Glucose Transporter Type 1; D051274:Glucose Transporter Type 3; D051275:Glucose Transporter Type 4; D051296:Glucose Transporter Type 5; D006027:Glycosides; D066298:In Vitro Techniques; D007328:Insulin / Q000494:pharmacology; D007700:Kinetics; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D018345:Mice, Knockout; D009004:Monosaccharide Transport Proteins / Q000235:genetics* / Q000378:metabolism*; D009124:Muscle Proteins; D018482:Muscle, Skeletal / Q000187:drug effects / Q000378:metabolism; D009419:Nerve Tissue Proteins; D011437:Propylamines
D000345:Affinity Labels; D001386:Azides; D004187:Disaccharides; D051272:Glucose Transporter Type 1; D051274:Glucose Transporter Type 3; D051275:Glucose Transporter Type 4; D051296:Glucose Transporter Type 5; D006027:Glycosides; D007328:Insulin; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; D009419:Nerve Tissue Proteins; D011437:Propylamines; C495678:Slc2a1 protein, mouse; C495689:Slc2a3 protein, mouse; C495693:Slc2a4 protein, mouse; C065412:2-N-(4-(1-azitrifluoroethyl)benzoyl)-1,3-bis-(mannos-4-yloxy)-2-propylamine; D019325:3-O-Methylglucose; D003571:Cytochalasin B; D003847:Deoxyglucose; D005947:Glucose