Abstract
Differentiated rat L6 skeletal muscle cell cultures maintained in glucose-deficient medium containing 25 mM xylose displayed a rapid, reversible, time- and concentration-dependent 3-5-fold increase in glucose transport activity. Glucose deprivation in the continuous presence of insulin (24 h) resulted in an overall 9-10-fold stimulation of glucose transport activity. In contrast, acute (30 min) and chronic (24 h) insulin treatment of L6 cells maintained in high glucose (25 mM)-containing medium resulted in a 1.5- and 4-fold induction of glucose transport activity, respectively. Acute glucose deprivation and/or insulin treatment had no significant effect on the total amount of glucose transporter protein, whereas the long-term insulin- and glucose-dependent regulation of glucose transport activity directly correlated with an increase in the cellular expression of the glucose transporter protein. In situ hybridization of the L6 cells demonstrated a 3-, 4-, and 6-fold increase in glucose transporter mRNA induced by glucose deprivation, insulin, and glucose deprivation plus insulin treatments, respectively. Similarly, Northern blot analysis of total RNA isolated from glucose-deprived, insulin, and glucose-deprived plus insulin-treated cells resulted in a 4-, 3-, and 9-fold induction of glucose transporter mRNA, respectively. The continuous presence of insulin in the medium, either in the presence or absence of glucose, resulted in a transient alteration of the glucose transporter mRNA. The relative amount of the glucose transporter mRNA was maximally increased at 6-12 h which subsequently returned to the basal steady-state level within 48 h. These data demonstrate a role for insulin and glucose in the overall regulation of glucose transporter gene expression which may account for the alteration of glucose transporter activity of muscle tissue observed in pathophysiological states such as type II diabetes mellitus.
Highlights
We have previously observed that glucose starvation of primary rat brain glial cultures results in a time- ancdoncentration-dependent increase in glucose transport activity, protein, andmRNA [41].In order to fully examine this regulatory property of the glucose transporter in an insulin-responsive cell type, the differentiated rat L6 skeletal muscle cells were maintained for h invarious amounts of glucoseplus xylose such that thetotal initialmedium monosaccharide concentration was mM
We determined the effect of D-glucose deprivation in conjunction with insulin treatment on glucose transport activity
Incubation of the L6 cells with 25 mM glucoseplus 100 nM insulin for 24 h resulted in a 3-fold increase in 2-DG (Fig. 2 A ) and an approximate 2-fold increase in 3-OMG (Fig. 223) transport activities compared to control cells maintained in the absence of insulin
Summary
We have previously observed that glucose starvation of primary rat brain glial cultures results in a time- ancdoncentration-dependent increase in glucose transport activity, protein, andmRNA [41].In order to fully examine this regulatory property of the glucose transporter in an insulin-responsive cell type, the differentiated rat L6 skeletal muscle cells were maintained for h invarious amounts of glucoseplus xylose such that thetotal initialmedium monosaccharide concentration was mM. Half-maximal stimulation was found to occur a t 3.5 mM glucose whereas maximal stimulation required the complete absence of exogenously added glucose This starvation-induced increase in glucose transport activity was found to be readily reversible (Fig. 1B).Readdition of mM glucose to the glucosedeprived L6 cells resulted inthereturnto basal glucose transport activity within 2-4 h. Incubation of L6 cells in the absence of glucose but in the continuous presence of insulin for 24 h resulted in a 13-fold increase in 2-DG (Fig. 2 A ) anda 14-fold increase in 3-OMG (Fig. 2B)transport activities. These data demonstrate that under these experimental conditions 2-DG transport is an accurate measure of glucose transport activity and thagt lucose deprivation in the continuous presence of insulin markedly enhances glucose transport activity. A secondphase of insulin-stimulated glucosetransport activity occurred at 4 h which gradually increased over the 24-h i40 B, I
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