Mechanism of insulin action on glucose transport in the isolated rat adipose cell. Enhancement of the number of functional transport systems
Wardzala, L.J.; Cushman, S.W.; Salans, L.B.
Journal of Biological Chemistry 253(22): 8002-8005
ISSN/ISBN: 0021-9258 PMID: 711732 Accession: 005867849
[3H]Cytochalasin B binding and its competitive inhibition by D-glucose were used to quantitate the number of glucose transport systems in plasma membranes prepared from isolated rat epididymal adipose cells. Curve-fitting analysis of equilibrium cytochalasin B binding in the absence and presence of 500 mM D-glucose permitted identification of 3 classes of binding sites, only one of which is inhibited by D-glucose. Cytochalasin E reduced cytochalasin B binding to those sites which were not sensitive to D-glucose inhibition, thereby permitting a more precise determination of the characteristics of the D-glucose-sensitive site. This site comprised less than 25% of the total number of sites and bound cytochalasin B with a Kd of approximately 120 nM. Since the inhibition constant of cytochalasin B for D-glucose uptake by adipose cell plasma membranes was similar in value, this site was tentatively identified as the glucose transport system. Plasma membranes prepared from isolated adipose cells which have not been preincubated with insulin bound approximately 4 pmol of cytochalasin B/mg of membrane protein to the D-glucose-inhibitable binding site. If 7.0 nM (1000 microunits/ml) insulin was present during preincubation, cytochalasin B binding to the plasma membranes was increased 4-fold without alterations either in this site's dissociation constant or in the characteristics of the other 2 cytochalasin B binding sites. Insulin apparently stimulated glucose transport in isolated rat adipose cells by activating identical, but basally inactive, glucose transport systems.