The effect of magnesium deficiency on glucose disposal, glucose-stimulated insulin secretion and insulin action on skeletal muscle was investigated in rats which were fed a low magnesium-containing diet for 4 days. Control rats were fed a standard diet. Compared to the control rats, the rats fed with low magnesium diet presented: 1) lower serum magnesium levels (0.45 +/- 0.02 vs 0.78 +/- 0.01 mmol/l, p < 0.001), 2) higher basal serum glucose (6.8 +/- 0.02 vs 5.5 +/- 0.2 mmol/l, p < 0.05) and similar basal serum insulin, 3) 40% reduction (p < 0.001) in the glucose disappearance rate after its i.v. administration, and 4) 45% reduction (p < 0.05) in the glucose-stimulated insulin secretion. The insulin action upon the glucose uptake by skeletal muscle was determined by means of hindquarter perfusions. Compared with control rats, magnesium-deficient rats presented: 1) normal basal glucose uptake, 2) lower stimulatory effect on the glucose uptake by insulin at the concentrations of 5 x 10(-10) mol/l (3.0 +/- 0.9 vs 5.4 +/- 0.6, p < 0.05) and 5 x 10(-9) mol/l (6.3 +/- 0.5 vs 8.0 +/- 0.5, p < 0.05), 3) normal glucose uptake at a maximal insulin concentration of 1 x 10(-7) mol/l, and 4) 50% reduction in the insulin sensitivity (ED50: 1.3 +/- 0.3 vs 0.55 +/- 0.1 mol/l, p < 0.05). In partially purified insulin receptors prepared from gastrocnemius muscle, 125I-insulin binding was similar in both groups of rats. However, the autophosphorylation of the beta-subunit of the insulin receptor was significantly reduced by 50% in magnesium-deficient rats and the tyrosine kinase activity of insulin receptors toward the exogenous substrate Poly Glu4; Tyr 1 was also reduced (p < 0.05) by hypomagnesaemia. The abundance of the insulin-sensitive glucose transporter protein (muscle/fat GLUT4), measured by Western blot analysis using polyclonal antisera, was similar in muscles of control and hypomagnesaemic rats. These findings indicate that hypomagnesaemia has a deleterious effect on glucose metabolism due to an impairment of both insulin secretion and action. The insulin resistance observed in skeletal muscle of magnesium-deficient rats may be attributed, at least in part, to a defective tyrosine kinase activity of insulin receptors.

SummaryThe effect of magnesium deficiency on glucose disposal, glucose-stimulated insulin secretion and insulin action on skeletal muscle was investigated in rats which were fed a low magnesium-containing diet for 4 days. Control rats were fed a standard diet. Compared to the control rats, the rats fed with low magnesium diet presented: 1) lower serum magnesium levels (0.45 + 0.02 vs 0.78 + 0.01 mmol/1, p < 0.001), 2) higher basal serum glucose (6.8 +__ 0.2 vs 5.5 + 0.2 mmol/1, p < 0.05) and similar basal serum insulin, 3) 40 % reduction (p < 0.001) in the glucose disappearance rate after its i.v. administration, and 4) 45 % reduction (p < 0.05) in the glucose-stimulated insulin secretion. The insulin action upon the glucose uptake by skeletal muscle was determined by means of hindquarter perfusions. Compared with control rats, magnesium-deficient rats presented: 1) normal basal glucose uptake, 2) lower stimulatory effect on the glucose uptake by insulin at the concentrations of 5 x 10 -l~ mol/1 (3.0 + 0.9 vs 5.4 + 0.6, p < 0.05) and 5 x 10 -9 tool/1 (6.3 + 0.5 vs 8.0 + 0.5, p < 0.05), 3) normal glucose uptake at a maximal insulin concentration of i x 10 .7 mol/1, and 4) 50 % reduction in the insulin sensitivity (ED50:1.3 +_ 0.3 vs 0.55 + 0.1 mol/1, p < 0.05). In partially purified insulin receptors prepared from gastrocnemius muscle, 125I-insulin binding was similar in both groups of rats. However, the autophosphorylation of the fi-subunit of the insulin receptor was significantly reduced by 50 % in magnesium-deficient rats and the tyrosine kinase activity of insulin receptors toward the exogenous substrate Poly Glu4: Tyr 1 was also reduced (p < 0.05) by hypomagnesaemia. The abundance of the insulin-sensitive glucose transporter protein (muscle/fat GLUT4), measured by Western blot analysis using polyclonal antisera, was similar in muscles of control and hypomagnesaemic rats. These findings indicate that hypomagnesaemia has a deleterious effect on glucose metabolism due to an impairment of both insulin secretion and action. The insulin resistance observed in skeletal muscle of magnesium-deficient rats may be attributed, at least in part, to a defective tyrosine kinase activity of insulin receptors.

In 3T3-L1 adipocytes we have examined the effect of tri-iodothyronine (T 3 ) on glucose transport, total protein content and subcellular distribution of GLUT1 and GLUT4 glucose transporters. Cells incubated in T 3 -depleted serum were used as controls. Cells treated with T 3 (50 nM) for three days had a 3·6-fold increase in glucose uptake (P<0·05), and also presented a higher insulin sensitivity, without changes in insulin binding. The two glucose carriers, GLUT1 and GLUT4, increased by 87% (P<0·05) and 90% (P<0·05), respectively, in cells treated with T 3 . Under non-insulin-stimulated conditions, plasma membrane fractions obtained from cells exposed to T 3 were enriched with both GLUT1 (3·29 0·69 vs 1·20 0·29 arbitrary units (A.U.)/5 µg protein, P<0·05) and GLUT4 (3·50 1·16 vs 0·82 0·28 A.U./5 µg protein, P<0·03). The incubation of cells with insulin produced the translocation of both glucose transporters to plasma membranes, and again cells treated with T 3 presented a higher amount of GLUT1 and GLUT4 in the plasma membrane fractions (P<0·05 and P<0·03 respectively). These data indicate that T 3 has a direct stimulatory effect on glucose transport in 3T3-L1 adipocytes due to an increase in GLUT1 and GLUT4, and by favouring their partitioning to plasma membranes. The effect of T 3 on glucose uptake induced by insulin can also be explained by the high expression of both glucose transporters.

Sulfonylureas are drugs widely used in the treatment of patients with type 2 diabetes mellitus. In addition to their pancreatic effect of stimulating insulin secretion, many studies suggest that sulfonylureas also have extrapancreatic actions. We have previously reported that gliclazide, a second-generation sulfonylurea, stimulates the glucose uptake by rat hindquarter skeletal muscle directly and immediately by promoting the translocation of glucose transporter 4 to the plasma membrane. The aim of our study was to approach the gliclazide intracellular signaling pathway. For this purpose, we incubated clamped and isolated soleus muscle from rat with gliclazide. The following results were obtained: 1) gliclazide stimulates insulin receptor substrate (IRS)-1-phosphatidylinositol 3 (PI3)-kinase-associated activity, and this activity is necessary for gliclazide-stimulated glucose transport; 2) gliclazide treatment produces a gradual translocation of the diacylglycerol (DAG)-dependent isoforms protein kinase C (PKC) alpha, theta, and epsilon from cytosolic to membrane fraction that is dependent on PI3-kinase and phospholipase C (PLC)-gamma activation; and 3) PKC and PLC-gamma activation is necessary for gliclazide-stimulated glucose transport. We propose a hypothetical signaling pathway by which gliclazide could stimulate IRS-1 that would allow its association with PI3-kinase, promoting its activation. PI3-kinase products could induce PLC-gamma activation, whose hydrolytic activity could activate the DAG-dependent isoforms PKC alpha, theta, and epsilon.

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