Muscle Glycogen Synthase Activity Research Articles

Acquired defects of glycogen synthase activity in cultured human skeletal muscle cells: influence of high glucose and insulin levels.

To determine whether defects of muscle glycogen synthase (GS) activity can be acquired by exposure to elevated glucose or insulin levels, human skeletal muscle cells obtained by needle biopsy from normal control subjects were grown in culture for 4-6 weeks followed by 4 days of fusion and differentiation in media containing either normal (5.5 mmol/l glucose and 22 pmol/l insulin) or increased concentrations of glucose (20 mmol/l), insulin (30 micromol/l), or both. After fusion in normal media, acute stimulation by 33 nmol/l insulin for 1 h increased GS fractional velocity (FV) approximately twofold (from 9.01 +/- 1.26 to 16.31 +/- 2.40, P < 0.05). Increasing the media glucose concentration alone to 20 mmol/l during fusion had no effect on basal FV but caused a marginal impairment of the insulin-stimulated GS response (from 8.51 +/- 1.33 to 12.99 +/- 1.90, P = 0.08). Increasing the media insulin concentration to 30 micromol/l during fusion at 5.5 mmol/l glucose also did not alter basal GS FV (10.61 +/- 1.69%) but completely abolished the normal insulin-stimulated increase in GS activity (to 11.63 +/- 1.55%, NS). The combination of high insulin (30 micromol/l) and high glucose (20 mmol/l) during fusion had no greater effect on the FV of either basal (11.66 +/- 2.16%, NS) or insulin-stimulated (9.20 +/- 1.80%, NS) GS activity than high insulin alone. Fusion in hyperinsulinemic media altered the kinetic parameters of GS with a near doubling of the basal Km0.1 and Vmax0.1 for uridinediphospho-glucose. Hyperinsulinemia also totally prevented the normal insulin-stimulated threefold increase in the Vmax0.1 and the 65% decrease in the A0.5 for glucose-6-phosphate. GS mRNA and protein expression, determined by RNase protection assay and immunoblotting, respectively, were unaffected by changes in media conditions. We conclude that exposure of human skeletal muscle cells primarily to high insulin induces severe insulin resistance through multiple acquired posttranslational defects, which affect both the kinetic characteristics and absolute activity of the GS enzyme.

Inhibition of muscle glycogen synthase activity and non-oxidative glucose disposal during hypoglycaemia in normal man.

The purpose of the present study was to evaluate the role of muscle glycogen synthase activity in the reduction of glucose uptake during hypoglycaemia. Six healthy young men were examined twice; during 120 min of hyperinsulinaemic (1.5 mU.kg-1. min-1) euglycaemia followed by: 1)240 min of graded hypoglycaemia (plasma glucose nadir 2.8 mmol/l) or 2) 240 min of euglycaemia. At 350-360 min a muscle biopsy was taken and indirect calorimetry was performed at 210-240 and 330-350 min. Hypoglycaemia was associated with markedly increased levels of adrenaline, growth hormone and glucagon and also with less hyperinsulinaemia. During hypoglycaemia the fractional velocity for glycogen synthase was markedly reduced; from 29.8 +/- 2.3 to 6.4 +/- 0.9%, p < 0.05. Total glucose disposal was decreased during hypoglycaemia (5.58 +/- 0.55 vs 11.01 +/- 0.75 mg.kg-1. min-1 (euglycaemia); p < 0.05); this was primarily due to a reduction of non-oxidative glucose disposal (2.43 +/- 0.41 vs 7.15 +/- 0.7 mg.kg-1 .min-1 (euglycaemia); p < 0.05), whereas oxidative glucose disposal was only suppressed to a minor degree. In conclusion hypoglycaemia virtually abolishes the effect of insulin on muscle glycogen synthase activity. This is in keeping with the finding of a marked reduction of non-oxidative glucose metabolism.

Polymorphism of the glycogen synthase gene in hypertensive and normotensive subjects.

Hypertension and non-insulin-dependent diabetes mellitus (NIDDM) are characterized by a strong genetic component and impaired ability to store glucose as glycogen in skeletal muscle. Impaired insulin activation and altered genetic control of muscle glycogen synthase, the rate-limiting enzyme for glucose storage in skeletal muscle, could provide an explanation for this insulin resistance. We examined whether there is an association between the glycogen synthase gene (Xba I polymorphism) and hypertension in 304 nondiabetic subjects. We examined glucose tolerance with an oral glucose tolerance test and glucose storage in skeletal muscle with the euglycemic insulin clamp technique in combination with indirect calorimetry. The Xba I A2 allele of the glycogen synthase gene was enriched in subjects with hypertension and a family history of NIDDM (48%) compared with normotensive subjects without a family history of NIDDM (6%, P < .0001). The presence of the A2 versus the A1 allele was associated with decreased rates of insulin-stimulated glucose storage in hypertensive subjects (11.2 +/- 2.3 versus 16.9 +/- 2.6 mumol/kg lean body mass per minute, P = .029) but not in normotensive subjects (28.0 +/- 4.6 versus 29.6 +/- 3.7 mumol/kg lean body mass per minute). In conclusion, Xba I polymorphism of the glycogen synthase gene identifies a subgroup of hypertensive subjects with a family history of NIDDM. The data suggest that a locus in the glycogen synthase gene region on chromosome 19 may serve as a "thrifty gene," increasing susceptibility for insulin resistance when exposed to other environmental or genetic factors.

Long-term effects of fluoxetine on glycemic control in obese patients with non-insulin-dependent diabetes mellitus or glucose intolerance: Influence on muscle glycogen synthase and insulin receptor kinase activity

Fluoxetine (F) is a specific serotonin-reuptake inhibitor that has been shown to promote weight loss and improve glycemic control in obese diabetic patients. To study its long-term metabolic effect, 40 obese patients with non-insulin-dependent diabetes mellitus (NIDDM) or impaired glucose tolerance (IGT) were included in a 12-month, randomized, placebo-controlled study. Patients were assigned to receive either 60 mg F or placebo (P) daily in conjunction with a 5.0-MJ/d diet (> 50% carbohydrate). Both groups showed a significant weight loss, with a nadir after 6 months without group differences (mean ± SD: F, 10.1 ± 10.0 kg; P, 9.4 ± 11.5 kg). Fifteen patients from the F group and 14 from the P group completed the 12-month study without weight loss differences. Glycemic regulation improved along with the weight loss, but with a larger decline in plasma C-peptide and fasting glucose levels in the F group ( P < .05). Total skeletal muscle glycogen synthase (GS) activity increased by 31% in the F group ( P < .01) and by 17% in the P group (nonsignificant) after 6 months of treatment, but was still less than the activity in normal-weight controls (aged 28.0 ± 6.3 years; body mass index, 23.5 ± 2.2). After adjustment for fasting glucose, insulin, weight loss, and diabetic state, a positive effect of F remained on the total GS activity, which accounted for 27% of the variation ( P < .05). The waist to hip ratio was reduced in P subjects as compared with F subjects ( P < .05). Fat-free mass (FFM) tended to be more reduced in the F group as compared with P subjects (4.9 v 1.9 kg), although the difference did not reach statistical significance. In conclusion, F seems to improve insulin sensitivity beyond the effect mediated through weight loss by a possible effect on GS activity in skeletal muscle tissue.

Unchanged gene expression of glycogen synthase in muscle from patients with NIDDM following sulphonylurea-induced improvement of glycaemic control

We have previously shown that the mRNA expression of muscle glycogen synthase is decreased in non-insulin-dependent diabetic (NIDDM) patients; the objective of the present protocol was to examine whether the gene expression of muscle glycogen synthase in NIDDM is affected by chronic sulphonylurea treatment. Ten obese patients with NIDDM were studied before and after 8 weeks of treatment with a weight-maintaining diet in combination with the sulphonylurea gliclazide. Gliclazide treatment was associated with significant reductions in HbA1C (p=0.001) and fasting plasma glucose (p=0.005) as well as enhanced beta-cell responses to an oral glucose load. During euglycaemic, hyperinsulinaemic clamp (2 mU x kg-1 x min-1) in combination with indirect calorimetry, a 35% (p=0.005) increase in whole-body insulin-stimulated glucose disposal rate, predominantly due to an increased non-oxidative glucose metabolism (p=0.02) was demonstrated in teh gliclazide-treated patients when compared to pre-treatment values. In biopsies obtained from vastus lateralis muscle during insulin infusion, the half-maximal activation of glycogen synthase was achieved at a significantly lower concentration of the allosteric activator glucose 6-phosphate (p=0.01). However, despite significant increases in both insulin-stimulated non-oxidative glucose metabolism and muscle glycogen synthase activation in gliclazide-treated patients no changes were found in levels of glycogen synthase mRNA or immunoreactive protein in muscle. In conclusion, improved blood glucose control in gliclazide-treated obese NIDDM patients has no impact on the gene expression of muscle glycogen synthase.

Regulation of hexokinase II and glycogen synthase mRNA, protein, and activity in human muscle.

Insulin regulates the activity of key enzymes of glucose metabolism in skeletal muscle by altering transcription or translation or by producing activity-altering modifications of preexisting enzyme molecules. Because of the small size of percutaneous muscle biopsies, these phenomena have been difficult to study in humans. This study was performed to determine how physiological hyperinsulinemia regulates the activities of hexokinase (HK), glycogen synthase (GS), and GLUT-4 in human skeletal muscle in vivo. We determined mRNA abundance, protein content, and activities for these proteins in muscle biopsies before and after a hyperinsulinemic clamp in normal subjects. HK I, HK II, GS, and GLUT-4 were expressed in muscle. HK II accounted for 80% of total HK activity and was increased by insulin from a basal value of 2.11 +/- 0.26 to 3.35 +/- 0.47 pmol.min-1.mg protein-1 (P < 0.05); HK I activity was unaffected. Insulin increased GS activity from 3.85 +/- 0.82 to 6.06 +/- 0.49 nmol.min-1.mg-1 (P < 0.01). HK II mRNA was increased 3.3 +/- 1.3-fold (P < 0.05) by insulin infusion. HK I, GS, and GLUT-4 mRNA and protein were unaffected. Because insulin infusion increased HK II but not GS mRNA, we conclude that HK II and GS may be regulated by insulin by different mechanisms in human skeletal muscle.

Determinants of insulin-stimulated skeletal muscle glycogen metabolism in man.

To examine factors which influence skeletal muscle glycogen synthesis in man, we related insulin sensitivity measured by euglycaemic insulin clamp in 43 healthy males to muscle glycogen synthase (GS) activity, GS protein content (Western blot), glycogen concentrations and fibre composition. Insulin increased muscle glycogen content (P < 0.05) and the change in glycogen content correlated with the GS protein content (r = 0.90, P = 0.01). GS protein concentration correlated inversely with age (r = -0.69, P = 0.04). Non-oxidative glucose disposal was inversely related to per cent type 2B fibres (r = -0.52, P < 0.05). The influence of age on these relationships was separately studied in young (n = 12, age = 26 +/- 2 years) and elderly (n = 15, age = 56 +/- 2 years) males. Insulin increased GS activity significantly only in young subjects (from 17.8 +/- 3.0 to 25.3 +/- 3.2 nmol mg protein-1 min-1; P = 0.015). GS activity and non-oxidative glucose disposal correlated in young (r = 0.69, P = 0.01) but not in the elderly (r = 0.064, P = 0.82) males, and this relationship was not influenced by the degree of obesity. In conclusion, muscle fibre type and GS activity are both determinants of muscle glycogen metabolism in healthy, normoglycaemic males. The close relationship between non-oxidative glucose metabolism and GS activity in young males is altered in ageing.

Unchanged gene expression of glycogen synthase in muscle from patients with NIDDM following sulphonylurea-induced improvement of glycaemic control

We have previously shown that the mRNA expression of muscle glycogen synthase is decreased in non-insulin-dependent diabetic (NIDDM) patients; the objective of the present protocol was to examine whether the gene expression of muscle glycogen synthase in NIDDM is affected by chronic sulphonylurea treatment. Ten obese patients with NIDDM were studied before and after 8 weeks of treatment with a weight-maintaining diet in combination with the sulphonylurea gliclazide. Gliclazide treatment was associated with significant reductions in HbA1c (p=0.01) and fasting plasma glucose (p=0.005) as well as enhanced beta-cell responses to an oral glucose load. During euglycaemic, hyperinsulinaemic clamp (2 mU · kg−1 · min−1) in combination with indirect calorimetry, a 35% (p=0.005) increase in whole-body insulin-stimulated glucose disposal rate, predominantly due to an increased non-oxidative glucose metabolism (p=0.02) was demonstrated in the gliclazide-treated patients when compared to pre-treatment values. In biopsies obtained from vastus lateralis muscle during insulin infusion, the half-maximal activation of glycogen synthase was achieved at a significantly lower concentration of the allosteric activator glucose 6-phosphate (p=0.01). However, despite significant increases in both insulin-stimulated non-oxidative glucose metabolism and muscle glycogen synthase activation in gliclazide-treated patients no changes were found in levels of glycogen synthase mRNA or immunoreactive protein in muscle. In conclusion, improved blood glucose control in gliclazide-treated obese NIDDM patients has no impact on the gene expression of muscle glycogen synthase.

Isolation and Characterization of the Human Muscle Glycogen Synthase Gene

Impaired glycogen synthase (GS) activity in skeletal muscle has been considered to be an inherited trait in patients with non-insulin-dependent diabetes mellitus (NIDDM). We therefore isolated the human muscle GS gene from genomic libraries and determined the genomic structure. The entire coding region, the 5'-flanking region, and the exon-intron boundaries were sequenced. The gene consists of 16 exons spanning approximately 27 kb of DNA and exists as a single copy in the human genome. The negatively charged parts with all known phosphorylation sites were coded by the first and the last exon. A single transcription initiation site was located 167 nucleotides upstream of the initiation codon. All of the exons and the putative promoter region were analyzed by single-strand conformation polymorphism in 30 insulin-resistant Finnish NIDDM patients, and three polymorphic sites were found. A missense mutation Gly464/Ser in exon 11 was found in 2 of 228 NIDDM patients screened but in 0 of 154 control subjects. These two patients were characterized further by severe insulin resistance and premature arteriosclerosis. The characterization of the genomic structure of the human muscle GS gene will facilitate studies of its role in the development of insulin resistance and NIDDM.

Heterogeneous responses to an intravenous glucose load

The aim of this study was to determine how the insulin sensitive enzymes pyruvate dehydrogenase (PDH) complex and glycogen synthase (GS) of different tissues respond to an endogenous pulse of insulin elicited by an intravenous infusion of glucose. An infusion of glucose (0.5 g/kg) into conscious, unrestrained animals via an indwelling cannula rapidly elevated plasma insulin concentration (to approx. 600 μU/ml after 10 min). The animals were sacrificed at selected time points after the commencement of infusion. Samples of heart, red quadriceps muscle, white adipose tissue (WAT) and brown adipose tissue (BAT) were excised and assayed for PDH complex and GS activities. The glucose infusion elicited a rapid (<5–10 min) increase in both PDH complex and GS activities in heart, BAT and WAT. The maximum rise in the activity of PDH and GS above basal were (respectively) 2- and 8-fold for heart, 5.5- and 5-fold for BAT, and 3.5- and 4-fold for WAT. The return of PDH complex activity to basal values was also very rapid (occurring over the next 20 min). The glucose infusion also stimulated GS activity in red quadriceps muscle but was, however without effect on PDH complex activity in this tissue. We conclude that although insulin stimulates PDH and GS with the same time course and magnitude in many insulin sensitive tissues, the time course and magnitude of insulin stimulation of these enzymes can vary between tissues. These results may mean that the stimulation of PDH complex and GS by insulin occurs via different receptor-effector pathways.

Influence of Exercise on Insulin Sensitivity

An important determinant of insulin sensitivity in humans, the insulin-induced glucose uptake in skeletal muscle (directed into glycogen synthesis), has been found to be markedly enhanced in endurance-trained individuals compared with sedentary controls. This enhanced insulin action is accompanied by a decreased insulin response during an oral glucose load. Available evidence indicates that the enhanced insulin action is not a genuine training effect, but merely a lasting effect of the previous exercise session(s). In the young, insulin sensitivity increases with a single exercise session, but the increase may appear only after several training sessions in middle-aged or older individuals. The effect disappears after 3-6 days. The exercise-induced reduction in muscle glycogen is an important factor underlying the increased insulin sensitivity in the period after exercise. There are reports that endurance and strength training enhances whole-body insulin sensitivity to a similar extent, and available evidence suggests that, provided total energy expenditure is held constant, low-intensity and high-intensity exercise are equally effective in promoting insulin sensitivity. Differences in sex steroids may explain the observation of a markedly higher insulin sensitivity in women than in men. Adaptations, which might underlie the increased insulin sensitivity in trained individuals, include increases in levels of the glucose transporter protein GLUT-4 and in muscle glycogen synthase activity, a decrease in the serum triglyceride concentration and, possibly, an increase in the muscle capillary network.

Insulin-resistant glucose metabolism in patients with microvascular angina—syndrome X

Studies in patients with microvascular angina (MA) or the cardiologic syndrome X have shown a hyperinsulinemic response to an oral glucose challenge, suggesting insulin resistance and a role for increased serum insulin in coronary microvascular dysfunction. The aim of the present study was to examine whether patients with MA are insulin-resistant. Nine patients with MA and seven control subjects were studied. All were sedentary and glucose-tolerant. Coronary arteriography was normal in all participants, and exercise-induced coronary ischemia was demonstrated in all MA patients. A euglycemic, hyperinsulinemic clamp was performed in combination with indirect calorimetry. Biopsy of vastus lateralis muscle was taken in the basal state and after 4 hours of euglycemia and hyperinsulinemia (2 mU · kg −1 · min −1). The fasting level of “true” serum insulin was significantly higher (43 ± 6 v 22 ± 3 pmol/L, P < .02) and the rate of insulin-stimulated glucose disposal to peripheral tissues was lower in patients with MA (13.4 ± 1.0 v 18.2 ± 1.4 mg · kg fat-free mass [FFM] −1 · min −1, P < .02) due to a decrease in nonoxidative glucose metabolism (8.4 ± 0.9 v 12.5 ± 1.3 mg · kg FFM −1 · min −1, P < .02). No difference was found in glucose or lipid oxidation rates between the two groups. In patients with MA, as well as in the pooled group of all participants, a positive correlation was demonstrated between the insulin-stimulated increment in fractional activity of skeletal muscle glycogen synthase (GS) and the increment in nonoxidative glucose metabolism after in vivo insulin exposure (MA: r = .73, P = .03; all participants: r = .73, P = .005), indicating a reduced in vivo activation of GS in MA patients. In conclusion, (1) MA is part of the insulin resistance syndrome, and (2) the insulin resistance is predominantly localized to the glycogen synthesis pathway in skeletal muscle tissue.

Stimulation of muscle glucose disposal by insulin in humans is a function of the preexisting plasma insulin level.

To examine whether tissue sensitivity to insulin is dependent upon the prevailing plasma insulin concentration, the ability of acute hyperinsulinemia to stimulate glucose disposal was investigated in six normal subjects before and after prolonged reduction of the plasma insulin concentration. Glucose turnover ([6,6-2H2]glucose), whole body glucose oxidation and nonoxidative glucose disposal (indirect calorimetry), and glycogen synthase activity in muscle were determined in the postabsorptive and in the insulin-stimulated states (euglycemic hyperinsulinemic clamp: 3 mU.kg-1.min-1) before and after a 4-day subcutaneous infusion of the somatostatin analogue octreotide (200 micrograms/24 h). Constant octreotide infusion 1) decreased postabsorptive and meal-stimulated plasma insulin levels by approximately 30-40% but did not significantly alter overall glucose tolerance, free fatty acid, growth hormone, and glucagon levels and 2) was associated with significant increases in insulin-mediated whole body glucose disposal (pre-drug: 10.29 +/- 0.49 vs. postdrug: 11.42 +/- 0.72 mg.kg-1.min-1, P < 0.04), nonoxidative glucose disposal (6.82 +/- 0.57 vs. 7.68 +/- 0.62, P < 0.03), and fractional glycogen synthase activity (0.14 +/- 0.03 vs. 0.20 +/- 0.04 mU/mg protein, P < 0.02). In contrast, infusion of saline instead of octreotide for 4 days to control subjects did not alter any of the metabolic parameters. We conclude that lowering the plasma insulin concentration over a prolonged period of time increases insulin sensitivity. The effects of insulin to stimulate whole body glucose utilization, nonoxidative glucose disposal, and glycogen synthase activity in muscle are therefore functions of the preexisting plasma insulin concentration.

Effect of glucose phosphorylation on the activation by insulin of skeletal muscle glycogen synthase

The effect of insulin injection on skeletal muscle glycogen synthase activation was studied in anesthetized, normal, fed rats. Insulin stimulated the conversion of glycogen synthase to the active I form, increased the concentration of glucose 6-phosphate, and activated glycogen synthase phosphatase. A close correlation between glucose 6-phosphate concentrations, per cent glycogen synthase in the active I form, and phosphatase activity was found. When boiled extracts of muscle from control and insulin-injected animals were added to glycogen pellets containing phosphatase 1 G, the difference in phosphatase activity between muscle extracts from insulin-injected and control rats was restored, indicating that the phosphatase was activated by heat-stable factors. Deproteinized muscle extracts from control and insulin-injected rats, at concentrations equivalent to those present in muscle, were tested for the activation of glycogen synthase by purified protein phosphatases 1 and 2A. The activation with the insulin extracts was four-fold larger than with the control extracts. When the extracts from insulin-injected rats were treated with glucose 6-phosphatase, the difference in activation with the control rat extracts was canceled. It would appear that, as in other insulin sensitive tissues, in skeletal muscle the increase in glucose 6-phosphate subsequent to the activation of glucose transport by insulin contributes to the activation of glycogen synthase.

Counterregulation of Hypoglycemia: Skeletal Muscle Glycogen Metabolism During Three Hours of Physiological Hyperinsulinemia in Humans

We examined the role of skeletal muscle in counterregulation of hypoglycemia (3.4 +/- 0.1 mmol/l) in 12 nondiabetic individuals (age 26 +/- 1 years, body mass index 24.2 +/- 0.7 kg/m2) during physiological hyperinsulinemia (280 +/- 25 pmol/l) compared with euglycemia (4.8 +/- 0.1 mmol/l). During hypoglycemia, hepatic glucose output (3-[3H]-glucose) was greater (7.72 +/- 2.72 mumol.kg-1.min-1, P < 0.01), glucose uptake was approximately 49% lower (21.20 +/- 3.55 mumol.kg-1.min-1, P < 0.005), and glucose clearance was reduced (P < 0.002) compared with euglycemia. Rates of flux of plasma-derived glucosyl units through glycolysis were similar in the two experiments, while glycogen synthetic rates were significantly reduced during hypoglycemia (P < 0.01) and accounted entirely for the reduction in glucose disposal. The insulin-induced activation of skeletal muscle glycogen synthase (reflected by Km decline by approximately 50% from 0.408 +/- 0.056 mmol/l and fractional velocity increase by approximately twofold from 21.8 +/- 2.7%) was completely abolished in hypoglycemia. In concert, glycogen phosphorylase activity increased during hypoglycemia by approximately 40% (P = 0.0001). Hypoglycemia resulted in seven- to eightfold increments in plasma epinephrine (P < 0.0001) and growth hormone (P < 0.001) and 40-60% increments in plasma glucagon (P < 0.005) and cortisol (P < 0.05). We conclude that, in this model of mild hypoglycemia of moderate duration, the majority of the glucose made available during the counterregulatory process (approximately 60-70%) is due to the limitation of glucose disposal, mostly via decreased glycogen synthetic activity in skeletal muscle.

Role of prostaglandins in mediating alterations in glucose metabolism during acute endotoxemia in the rat.

We determined the role of prostanoids in mediating alterations in glucose metabolism during lipopolysaccharide (LPS)-induced (1 mg/kg; LD10) acute endotoxemia in chronically catheterized awake rats. Basal glucose turnover (Rt, infusion of [5-3H]glucose), in vivo insulin action on overall glucose utilization under normoglycemic conditions (euglycemic clamp), whole body glycolysis, and muscle glycogen synthesis were determined in four groups of rats. These groups received 1) LPS (LPS rats, n = 6); 2) saline (control rats, n = 6); 3) indomethacin and LPS (INDO and LPS rats, n = 6); or 4) saline and indomethacin (INDO control rats, n = 6). In the fasted rats, LPS induced hyperthermia, hypotension, and hyperglycemia. These changes were associated with glycogen depletion in both skeletal muscle and liver and with increased Rt. During hyperinsulinemia, whole body glucose disposal was decreased by 37% due to decreased muscle glycogen synthesis and glycogen synthase activity whereas the rate of whole body glycolysis was normal. INDO abolished hyperthermia, hypotension, and hyperglycemia but did not improve whole body insulin sensitivity, muscle glycogen synthesis, or glycogen synthase activity. These data indicate that prostanoids mediate hypotension, transient fasting hyperglycemia, and fever during LPS-induced acute endotoxemia. They do not, however, explain insulin resistance under these conditions.

Multiple defects of both hepatic and peripheral intracellular glucose processing contribute to the hyperglycaemia of NIDDM

Non-insulin-dependent diabetic (NIDDM) patients were studied during a modified euglycaemic state when fasting hyperglycaemia was normalized by a prior (−210 to −150 min) — and later withdrawn (−150–0 min) — intravenous insulin infusion. Glucose metabolism was assessed in NIDDM patients (n=10) and matched control subjects (n=10) using tritiated glucose turnover rates, indirect calorimetry and skeletal muscle glycogen synthase activity determinations. Total and non-oxidative exogenous glycolytic flux rates were measured using appearance rates of tritiated water. A+180 min euglycaemic hyperinsulinaemic (40 mU·m−2·min−1) clamp was performed to determine the insulin responsiveness of the various metabolic pathways. Plasma glucose concentration increased spontaneously during baseline measurements in the NIDDM patients (−120 to 0 min: 4.8±0.3 to 7.0±0.3 mmol/l; p<0.01), and was primarily due to an elevated rate of hepatic glucose production (3.16±0.13 vs 2.51±0.16 mg·kg FFM−1·min−1; p<0.01). In the NIDDM subjects baseline glucose oxidation was decreased (0.92±0.17 vs 1.33±0.14 mg·kg FFM−1·min−1; p<0.01) in the presence of a normal rate of total exogenous glycolytic flux and skeletal muscle glycogen synthase activity. The simultaneous finding of an increased lipid oxidation rate (1.95±0.13 vs 1.61±0.07 mg·kg FFM−1·min−1; p=0.05) and increased plasma lactate concentrations (0.86±0.05 vs 0.66±0.03 mmol/l; p=0.01) are consistent with a role for both the glucose-fatty acid cycle and the Cori cycle in the maintenance and development of fasting hyperglycaemia in NIDDM during decompensation. Insulin resistance was demonstrated during the hyperinsulinaemic clamp in the NIDDM patients with a decrease in the major peripheral pathways of intracellular glucose metabolism (oxidation, storage and muscle glycogen synthase activity), but not in the pathway of non-oxidative glycolytic flux which was not completely suppressed during insulin infusion in the NIDDM patients (0.55±0.15 mg·kg FFM−1·min−1; p<0.05 vs 0; control subjects: 0.17±0.29; NS vs 0). Thus, these data also indicate that the defect(s) of peripheral (skeletal muscle) glucose processing in NIDDM goes beyond the site of glucose transport across the cell membrane.

Role of basal insulin in maintenance of intracellular glucose metabolic pathways in non—insulin-dependent diabetes mellitus

Impairments of both basal and insulin-stimulated oxidative (G ox) and nonoxidative (N ox) glucose metabolism are documented to exist in non—insulin-dependent diabetes mellitus (NIDDM). Although these defects have been well characterized during insulin stimulation, little is known about the effects of basal insulin or its deficiency on intracellular glucose metabolism in NIDDM. To determine the physiological significance of basal insulin in the maintenance of glucose metabolism in NIDDM, we studied nine subjects with NIDDM in the basal and insulin-deficient state produced by 3 hours of somatostatin (SRIF) infusion (0.08 pmol/kg/min). Glucose turnover rates were quantified by [3- 3H]glucose turnover, and substrate oxidation was assessed by a combination of indirect calorimetry and urinary nitrogen measurements. Skeletal muscle glycogen synthase (GS) and pyruvate dehydrogenase (PDH) activities were also measured in the basal state and during SRIF infusion. Basal glucose levels were maintained during SRIF infusion by exogenous glucose infusion (12.5 ± 0.9 mmol/L in the basal state v 12.8 ± 0.8 during SRIF infusion, P = NS). During the last hour of SRIF infusion, plasma C-peptide levels declined by 88% from 0.73 ± 0.11 to 0.09 ± 0.02 nmol/L ( P < .001), and serum insulin concentrations were undetectable (<14 pmol/L). During insulinopenic conditions, rates of glucose uptake (GU) were decreased by 12% from basal level of 2.26 ± 0.13 to 1.99 ± 0.12 mg/kg/min ( P < .05), and were entirely accounted for by reduced rates of G ox (1.01 ± 0.10 to 0.65 ± 0.14 mg/kg/min, P < .01). Corresponding measurements of active PDH (PDHa) activity in skeletal muscle were similarly decreased from 0.50 ± 0.14 to 0.31 ± 0.09 nmol/min/mg protein ( P < .05) during insulinopenia, whereas rates of fat oxidation (F ox) were increased from 0.87 ± 0.05 to 1.05 ± 0.07 mg/kg/min ( P < .01). The reduction in energy expenditure (EE) from lower G ox was completely offset by increased energy from enhanced F ox and resulted in unchanged total EE (1.42 ± 0.10 v 1.41 ± 0.11 kcal/min, P = NS). Although skeletal muscle GS activity was significantly decreased (2.6% ± 0.7% to 1.8% ± 0.8%, P < .05), calculated rates of N ox remained unchanged (1.24 ± 0.17 to 1.34 ± 0.21 mg/kg/min, P = NS) and were significantly correlated with non—insulin-mediated GU (NIMGU) during insulinopenia ( r = .76, P < .05). In conclusion, approximately 35% of basal G ox in NIDDM was dependent on basal insulin action. Basal N ox was independent of basal insulin and was determined by the rate of NIMGU. Basal insulin regulates intracellular glucose oxidation by multiple mechanisms that include effects on glucose uptake, free fatty acid (FFA) substrate availability, and PDH enzyme activity.

Impaired skeletal muscle glycogen synthase activation by insulin in the Goto-Kakizaki (G/K) rat

Impaired skeletal muscle glycogen synthase activation by insulin in the Goto-Kakizaki (G/K) rat

Mechanisms of fatty acid-induced inhibition of glucose uptake.

Increased plasma FFA reduce insulin-stimulated glucose uptake. The mechanisms responsible for this inhibition, however, remain uncertain. It was the aim of this study to determine whether the FFA effect was dose dependent and to investigate its mechanism. We have examined in healthy volunteers (13 male/1 female) the effects of three steady state plasma FFA levels (approximately 50, approximately 550, approximately 750 microM) on rates of glucose uptake, glycolysis (both with 3-3H-glucose), glycogen synthesis (determined with two independent methods), carbohydrate (CHO) oxidation (by indirect calorimetry), hepatic glucose output, and nonoxidative glycolysis (glycolysis minus CHO oxidation) during euglycemic-hyperinsulinemic clamping. Increasing FFA concentration (from approximately 50 to approximately 750 microM) decreased glucose uptake in a dose-dependent fashion (from approximately 9 to approximately 4 mg/kg per min). The decrease was caused mainly (approximately 2/3) by a reduction in glycogen synthesis and to a lesser extent (approximately 1/3) by a reduction in CHO oxidation. We have identified two independent defects in glycogen synthesis. The first consisted of an impairment of muscle glycogen synthase activity. It required high FFA concentration (approximately 750 microM), was associated with an increase in glucose-6-phosphate, and developed after 4-6 h of fat infusion. The second defect, which preceded the glycogen synthase defect, was seen at medium (approximately 550 microM) FFA concentration, was associated with a decrease in muscle glucose-6-phosphate concentration, and was probably due to a reduction in glucose transport/phosphorylation. In addition, FFA and/or glycerol increased insulin-suppressed hepatic glucose output by approximately 50%. We concluded that fatty acids caused a dose-dependent inhibition of insulin-stimulated glucose uptake (by decreasing glycogen synthesis and CHO oxidation) and that FFA and/or glycerol increased insulin-suppressed hepatic glucose output and thus caused insulin resistance at the peripheral and the hepatic level.