In vitro analysis of the glucose-transport system in GLUT4-null skeletal muscle

Jeffrey W. Ryder, Yuichi Kawano, Alexander V. Chibalin, Jorge Rincón, Tsu Shuen Tsao, Antine E. Stenbit, Terry Combatsiaris, Jing Yang, Geoffrey D. Holman, Maureen J. Charron, Juleen R. Zierath

Research output: Contribution to journalArticle

40 Scopus citations

Abstract

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-trifluoroethyl)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.

Original languageEnglish (US)
Pages (from-to)321-328
Number of pages8
JournalBiochemical Journal
Volume342
Issue number2
DOIs
StatePublished - Sep 1 1999

Keywords

  • GLUT1
  • GLUT3
  • GLUT4 knock-out
  • GLUT5
  • Insulin

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Ryder, J. W., Kawano, Y., Chibalin, A. V., Rincón, J., Tsao, T. S., Stenbit, A. E., Combatsiaris, T., Yang, J., Holman, G. D., Charron, M. J., & Zierath, J. R. (1999). In vitro analysis of the glucose-transport system in GLUT4-null skeletal muscle. Biochemical Journal, 342(2), 321-328. https://doi.org/10.1042/0264-6021:3420321