Glycogen Synthesis In Skeletal Muscle Research Articles

Mechanisms of insulin resistance in non-oxidative glucose metabolism: the role of glycogen synthase.

Insulin-mediated non-oxidative glucose metabolism is more or less identical to glycogen synthesis in skeletal muscle and that is why this pathway is specifically discussed in this paper. All three major steps in non-oxidative glucose processing--glucose transport, phosphorylation and glycogen synthesis--are found to be reduced in response to insulin in insulin-resistant type 2 diabetic subjects compared with controls. The insulin-signalling cascade from the insulin receptor to PI-3-K was also found to be abnormal, resulting in a severely reduced phosphorylation degree of the IRS-1 (IRS-2?)-PI-3-K complex, which can explain both reduced glucose transport and glycogen synthesis. The most pronounced finding in our studies is reduced glycogen synthase activation by insulin which is found in prediabetic subjects with normal glucose tolerance as well as in type 2 diabetics, but more severely. This defect was not reversible after treatment (normalization of blood glucose) and is therefore a candidate for the primary defect which is likely to be of genetic origin, but also could be caused by genetic imprinting, intrauterine malnutrition and social inheritance (obesity). Most of the abnormalities in non-oxidative glucose metabolism may be of secondary origin due to hyperglycemia itself or obesity. Both events may stimulate production of glucosamine, malonyl CoA and intramuscular triglyceride accumulation. These metabolites can theoretically induce most of the defects in glucose processing and furthermore impair insulin signalling. Whether the primary defect in activation of glycogen synthase is due to an abnormality in the enzyme complex itself or in the insulin signalling cascade still has to be investigated.

Glycogen turnover in skeletal muscle is stimulated along with glucose uptake in vivo during contraction

We have shown previously that glycogen synthesis in the heart can be stimulated in vivo by epinephrine. Our aim in this study was to determine whether glycogen synthesis in skeletal muscle can be similarly affected during increased energy expenditure. Left sciatic nerves of anesthetized fasted rats were electrically stimulated to allow left hindlimb muscles to contract for 5, 10, and 20 min. Glycogen contents in the contracting muscles at the end of electrical stimulation were found to be ~40% less than resting muscles in the right hindlimbs in all three groups of rats. Accompanying the enhanced glycogenolysis was increased incorporation of the intravenously infused [3- 3H]-glucose into glycogen. The rate of tritium incorporation into glycogen in the contracting muscle was found to be 34-fold greater than resting muscles. Glucose utilization was determined by the phosphorylation of the intravenously injected [ 14C]-2-deoxyglucose in the skeletal muscle. The rate of accumulation of [ 14C]-2-deoxyglucose-6-phosphate in the contracting muscles was found to be 28-fold greater than resting muscles. Glycogen synthesis and glucose uptake indexes, calculated by dividing the radioactivity in [ 3H]-glycogen and [ 14C]-2-DGP by the mean specific activity of their respective precursors in the plasma, were not found to be significantly different in the contracting muscle. In conclusion, our data indicate that: (i) glycogenesis and glycogenolysis can be stimulated concurrently in the skeletal muscle; and (ii) glucose utilization in the skeletal muscle during contraction may be mediated through glycogen turnover.

Islet amyloid polypeptide decreases the effects of insulin-like growth factor-I on glucose transport and glycogen synthesis in skeletal muscle

Previous studies have shown that islet amyloid polypeptide (IAPP) is co-secreted with insulin from the β-cell. IAPP reduces insulin-stimulated rates of glycogen synthesis in skeletal muscle but the mechanisms are unclear. Insulin-like growth factor I (IGF-I) is an important regulator of glucose metabolism in skeletal muscle and acts through its own receptor, which has many structural and functional similarities with the insulin receptor. Despite this, the effects of IGF-I on glucose utilization are not identical to those of insulin. The aim of the study was to determine the effects of IAPP on IGF-I-stimulated rates of glucose transport and metabolism (measured by 3- O-methyl[ 3H]glucose and [U- 14C]glucose, respectively) in rat soleus muscle, and compare them with those stimulated by insulin. IAPP (10 nM) decreased the sensitivity of 3- O-methylglucose transport, the flux of glucose to hexosemonophosphate and the sensitivity of glycogen synthesis to IGF-I. In contrast, IAPP had no effect on IGF-I-stimulated rates of lactate formation (i.e. glycolysis). IAPP decreased the sensitivity of 3- O-methylglucose transport and glycogen synthesis to insulin. It is concluded that IAPP blunts the stimulation of glucose uptake and deposition by IGF-I or insulin in skeletal muscle. These observations expand those made initially for IAPP and insulin and suggest that IAPP affects IGF-I- or insulin-stimulated glucose metabolism in muscle by a mechanism which is common for both hormones. These experiments may serve as a framework for future studies in order to clarify the mechanisms by which IAPP affects glucose metabolism in skeletal muscle.

Effects of FFA on insulin-stimulated glucose fluxes and muscle glycogen synthase activity in rats.

To examine effects of free fatty acids (FFA) on insulin-stimulated glucose fluxes, euglycemic hyperinsulinemic (86 pmol . kg-1 . min-1) clamps were performed for 5 h in conscious rats with (n = 8) or without (n = 8) lipid-heparin infusion. Glucose infusion rate required to maintain euglycemia was not different between the two groups during the first 2 h of clamps but became significantly lower with lipid-heparin infusion in the 3rd h and thereafter. To investigate changes in intracellular glucose metabolism during lipid-heparin infusion, additional clamps (n = 8 each) were performed for 1, 2, 3, or 5 h with an infusion of [3-3H]glucose. Insulin-stimulated whole body glucose utilization (Rd), glycolysis, and glycogen synthesis were estimated on the basis of tracer concentrations in plasma during the final 40 min of each clamp. Similar to changes in glucose infusion rate, Rd was not different between the two groups in the 1st and 2nd h but was significantly lower with lipid-heparin infusion in the 3rd h and thereafter. Whole body glycolysis was significantly lower with lipid-heparin infusion in all time periods, i.e., 1st, 2nd, 3rd, and 5th h of clamps. In contrast, whole body glycogen synthesis was higher with lipid-heparin infusion in the 1st and 2nd h but lower in the 5th h. Similarly, accumulation of [3H]glycogen radioactivity in muscle glycogen was significantly higher with lipid-heparin during the 1st and 2nd h but lower during the 3rd and 5th h. Glucose 6-phosphate (G-6-P) concentrations in gastrocnemius muscles were significantly higher with lipid-heparin infusion throughout the clamps. Muscle glycogen synthase (GS) activity was not altered with lipid-heparin infusion at 1, 2, and 3 h but was significantly lower at 5 h. Thus increased availability of FFA significantly reduced whole body glycolysis, but compensatory increase in skeletal muscle glycogen synthesis in association with accumulation of G-6-P masked this effect, and Rd was not affected in the early phase (within 2 h) of lipid-heparin infusion. Rd was reduced in the later phase (>2 h) of lipid-heparin infusion, when glycogen synthesis was reduced in association with reduced skeletal muscle GS activity.

Neonatal de-afferentation of capsaicin-sensitive sensory nerves increases in vivo insulin sensitivity in conscious adult rats.

Sensory neuropeptides, released from the peripheral nervous system, might modulate glucose homeostasis by antagonizing insulin action. The effects of de-afferentation of functional small diameter unmyelinated C-fibres (sensory nerves) on in vivo insulin-mediated intracellular glucose metabolism were investigated by using euglycaemic insulin (6 and 18 mU/kg x min) clamps with [3-(3)H]-glucose infusion in 24 adult rats, treated neonatally with either capsaicin (CAP) (50 mg/kg) or vehicle (CON). Following the clamp, skeletal muscle groups, liver and adipose tissue were freeze-clamped. At plasma insulin levels of approximately 90 mU/l, CAP-rats showed a 21% increase in whole body glucose uptake compared with CON (24.4 +/- 1.6 vs 20.1 +/- 0.8 mg/kg min, p < 0.02), which was paralleled by a 20% increase in whole body glycolysis (12.6 +/- 0.8 vs 10.5 +/- 0.5 mg/ kg.min p < 0.05) (concentration of 3H2O in plasma). Whole body skeletal muscle glycogenesis was increased by 80% in CAP-rats (5.7 +/- 0.7 vs 3.1 +/- 0.7 mg/kg x min, p < 0.05) with increased muscle glycogen synthase activity. Whole body (muscle, liver and adipose tissue combined) de novo lipogenesis also was increased in CAP-rats compared with CON (0.69 +/- 0.10 vs 0.44 +/- 0.06 mg/kg x min, p < 0.05) (incorporation of [3-(3)H]-glucose counts into glycogen or fat). Hepatic glucose production was lower in CAP-rats compared with CON (0.6 +/- 0.6 vs 2.1 +/- 0.7 mg/kg x min, p < 0.05). Plasma glucagon, corticosterone, epinephrine and norepinephrine levels were reduced in CAP-rats: 43 +/- 2 compared with 70 +/- 6 pg/ml, 855 +/- 55 compared with 1131 +/- 138 nmol/l, 513 +/- 136 compared with 1048 +/- 164 pmol/l and 928 +/- 142 compared with 1472 +/- 331 pmol/l, respectively, p < 0.05. At plasma insulin levels of approximately 400 mU/l, CAP-rats showed no differences in peripheral and hepatic insulin action compared with CON. We conclude that the removal of endogenous sensory neuropeptides, by de-afferentation of capsaicin-sensitive sensory nerves, increases in vivo insulin sensitivity, but not responsiveness: 1) primarily through an increased sensitivity of skeletal muscle glycogen synthesis to insulin; 2) through a reduction in the levels of counter-regulatory hormones, thereby creating a milieu which favours overall in vivo insulin sensitivity with respect to glucose uptake, glucose production, glycolysis, glycogenesis and lipogenesis.

Pivotal role of leptin in insulin effects.

The OB protein, also known as leptin, is secreted by adipose tissue, circulates in the blood, probably bound to a family of binding proteins, and acts on central neural networks regulating ingestive behavior and energy balance. The two forms of leptin receptors (long and short forms) have been identified in various peripheral tissues, a fact that makes them possible target sites for a direct action of leptin. It has been shown that the OB protein interferes with insulin secretion from pancreatic islets, reduces insulin-stimulated glucose transport in adipocytes, and increases glucose transport, glycogen synthesis and fatty acid oxidation in skeletal muscle. Under normoglycemic and normoinsulinemic conditions, leptin seems to shift the flux of metabolites from adipose tissue to skeletal muscle. This may function as a peripheral mechanism that helps control body weight and prevents obesity. Data that substantiate this hypothesis are presented in this review.

Defect of an Early Event of Glucose Metabolism in Skeletal Muscle of the Male Otsuka Long-Evans Tokushima Fatty (OLETF) Rat, a Non-Insulin- Dependent Diabetes Mellitus (NIDDM) Model

Otsuka Long-Evans Tokushima Fatty (OLETF) rats showed that the distribution of plasma membrane content of insulin-regulated glucose transporter in skeletal muscle was reminiscent of that in human non-insulin-dependent diabetes mellitus (NIDDM). To obtain more information on the cellular mechanisms of muscle insulin resistance, hexokinase activities were measured in the skeletal muscle of OLETF rats. The results showed that the activity of the type II enzyme in the diabetic rats was significantly decreased (P<0.05) compared with Long-Evans Tokushima Otsuka (LETO) control rats. No significant differences in the activity of the type I hexokinase were observed between these rats. Western blot analysis showed that the protein content of the type II in OLETF rats was also significantly lower than that in LETO rats (P<0.05). After insulin stimulation, the intramuscular content of glucose 6-phosphate, which regulates glycogen synthesis in skeletal muscle, was significantly decreased in OLETF rats (P<0.01). However, glycogen synthase activityin vitroand intramuscular lactate concentration in these rats did not show significant differences. These results suggest that the G6P content of the diabetic rats is decreased as a result of an impaired early event of glucose metabolism, indicating that the molecular defects of skeletal muscle in OLETF rats are similar to those in NIDDM patients.

Time course of insulin action on tissue-specific intracellular glucose metabolism in normal rats.

We investigated the time course of insulin action in conscious rats exposed to constant physiological hyperinsulinemia (approximately 100 mU/l) while maintaining euglycemia (approximately 100 mg/dl) for 0, 0.5, 2, 4, 8, or 12 h. [3-3H]glucose was infused to quantitate whole body glucose disposal (rate of disappearance, Rd), glycolysis (generation of 3H2O in plasma), hepatic glucoses production (HGP), and skeletal muscle and liver glycogen synthesis ([3-3H]glucose incorporation into glycogen and time-dependent change in tissue glycogen concentration). The basal Rd, which equals HGP, was 6.0 +/- 0.3 mg.kg-1.min-1. With increased duration of hyperinsulinemia from 0 to 0.5 to 2 to 4 h, Rd increased from 6.0 +/- 0.3 to 21.0 +/- 1.1 to 24.1 +/- 1.5 to 26.6 +/- 0.6 mg.kg-1.min-1 (P < 0.05 for 2 and 4 h vs. 0.5 h). During the first 2 h the increase in Rd was explained by parallel increases in glycolysis and glycogen synthesis. From 2 to 4 h the further increase in Rd was entirely due to an increase in glycolysis without change in glycogen synthesis. From 4 to 8 to 12 h of hyperinsulinemia, Rd decreased by 19% from 26.6 +/- 0.6 to 24.1 +/- 1.1 to 21.6 +/- 1.8 mg.kg-1.min-1 (P < 0.05 for 8 h vs. 4 h and 12 h vs. 8 h). The progressive decline in Rd, in the face of constant hyperinsulinemia, occurred despite a slight increase (8-14%) in glycolysis and was completely explained by a marked decrease (64%) in muscle glycogen synthesis. In contrast, liver glycogen synthesis increased fourfold, indicating an independent regulation of muscle and liver glycogen synthesis by long-term hyperinsulinemia. In the liver, during the entire 12-h period of insulin stimulation, the contribution of the direct (from glucose) and the indirect (from C-3 fragments) pathways to net glycogen formation remained constant at 77 +/- 5 and 23 +/- 5%, respectively. HGP remained suppressed throughout the 12-h period of hyperinsulinemia.

Involvement of integrins and the cytoskeleton in modulation of skeletal muscle glycogen synthesis by changes in cell volume

Muscle glycogen synthesis is modulated by physiologically relevant changes in cell volume. We have investigated the possible involvement of integrin-extracellular matrix interactions in this process using primary cultures of rat skeletal muscle subject to hypo- or hyper-osmotic exposure with integrin binding peptide GRGDTP to disrupt integrin actions and the inactive analogue GRGESP as control. Osmotically induced increases (77%) and decreases (34%) in glycogen synthesis ( d-[ 14C]-glucose incorporation into glycogen) were prevented by GRGDTP (but not GRGESP) without affecting glucose transport. Cytoskeletal disruption with cytochalasin D or colchicine had similar effects to GRGDTP. Osmotically induced modulation of muscle glycogen synthesis involves integrin-extracellular matrix interactions and cytoskeletal elements, possibly as components of a cell-volume `sensing' mechanism.

Effects of Increased Glucokinase Gene Copy Number on Glucose Homeostasis and Hepatic Glucose Metabolism

The relationship between glucokinase (GK) gene copy number and glucose homeostasis was studied in transgenic mice with additional copies of the entire GK gene locus (Niswender, K. D., Postic, C., Jetton, T. L., Bennett, B. D., Piston, D. W., Efrat, S., and Magnuson, M. A. (1997) J. Biol. Chem. 272, 22564-22569). The plasma glucose concentration was reduced by 25 +/- 3% and 37 +/- 4% in mice with one or two extra copies of the gene locus, respectively. The basis for the hypoglycemic phenotype was determined using metabolic tracer techniques in chronically cannulated, conscious mice with one extra GK gene copy. Under basal conditions (6-h fasted) transgenic mice had a lower blood glucose concentration (-12 +/- 1%) and a slightly higher glucose turnover rate (+8 +/- 3%), resulting in a significantly higher glucose clearance rate (+21 +/- 2%). Plasma insulin levels were not different, suggesting that increased glucose clearance was due to augmented hepatic, not islet, GK gene expression. Under hyperglycemic clamp conditions the transgenic mice had glucose turnover and clearance rates similar to the controls, but showed a lower plasma insulin response (-48 +/- 5%). Net hepatic glycogen synthesis was markedly elevated (+360%), whereas skeletal muscle glycogen synthesis was decreased (-40%). These results indicate that increased GK gene dosage leads to increased hepatic glucose metabolism and, consequently, a lower plasma glucose concentration. Increased insulin secretion was not observed, even though the transgene is expressed in islets, because hypoglycemia causes a down-regulation in islet GK content (Niswender, K. D., Postic, C., Jetton, T. L., Bennett, B. D., Piston, D. W., Efrat, S., and Magnuson, M. A. (1997) J. Biol. Chem. 272, in press).

Skeletal muscle triglyceride levels are inversely related to insulin action.

In animal studies, increased amounts of triglyceride associated with skeletal muscle (mTG) correlate with reduced skeletal muscle and whole body insulin action. The aim of this study was to test this relationship in humans. Subjects were 38 nondiabetic male Pima Indians (mean age 28 +/- 1 years). Insulin sensitivity at physiological (M) and supraphysiological (MZ) insulin levels was assessed by the euglycemic clamp. Lipid and carbohydrate oxidation were determined by indirect calorimetry before and during insulin administration. mTG was determined in vastus lateralis muscles obtained by percutaneous biopsy. Percentage of body fat (mean 29 +/- 1%, range 14-44%) was measured by underwater weighing. In simple regressions, negative relationships were found between mTG (mean 5.4 +/- 0.3 micromol/g, range 1.3-1.9 micromol/g) and log10M (r = -0.53, P < or = 0.001), MZ (r = -0.44, P = 0.006), and nonoxidative glucose disposal (r = -0.48 and -0.47 at physiological and supraphysiological insulin levels, respectively, both P = 0.005) but not glucose or lipid oxidation. mTG was not related to any measure of adiposity. In multiple regressions, measures of insulin resistance (log10M, MZ, log10[fasting insulin]) were significantly related to mTG independent of all measures of obesity (percentage of body fat, BMI, waist-to-thigh ratio). In turn, all measures of obesity were related to the insulin resistance measures independent of mTG. The obesity measures and mTG accounted for similar proportions of the variance in insulin resistance in these relationships. The results suggest that in this human population, as in animal models, skeletal muscle insulin sensitivity is strongly influenced by local supplies of triglycerides, as well as by remote depots and circulating lipids. The mechanism(s) underlying the relationship between mTG and insulin action on skeletal muscle glycogen synthesis may be central to an understanding of insulin resistance.

Modulation of glycogen synthesis in rat skeletal muscle by changes in cell volume.

1. The hypothesis that cellular hydration state modulates muscle glycogen synthesis was tested by measuring the incorporation of [14C]glucose into glycogen (glycogen synthesis) in primary rat myotubes after experimentally induced volume changes. 2. Glycogen synthesis in myotubes increased (by 75%, P < 0.01) after swelling induced by 60 min exposure to hyposmotic media (170 mosmol kg-1) relative to isosmotic control (300 mosmol kg-1) values, it decreased (by 31%, P < 0.05) after shrinkage induced by 60 min exposure to hyperosmotic (430 mosmol kg-1) media. Myotube 2-deoxy-D-glucose (0.05 mM) uptake was unaffected by changes in external osmolality. 3. Wortmannin (100 nM; 60 min), a phosphatidylinositol 3-kinase inhibitor, decreased basal glycogen synthesis by 28% whereas rapamycin (100 nM; 60 min), which blocks the activation of p70 S6 kinase, had no effect. Both wortmannin (100 nM; 60 min) and rapamycin (100 nM; 60 min) blocked the changes in glycogen synthesis resulting from hypo- and hyperosmotic exposure. 4. Myotube glycogen synthesis is modulated by volume changes independently of changes in glucose uptake. The phenomenon may be physiologically important in promoting glycogen storage during circumstances of myofibrillar swelling, e.g. after feeding or exercise.

Diverse effects of Glut 4 ablation on glucose uptake and glycogen synthesis in red and white skeletal muscle.

The ability of muscles from Glut 4-null mice to take up and metabolize glucose has been studied in the isolated white EDL and red soleus muscles. In EDL muscles from male or female Glut 4-null mice, basal deoxyglucose uptake was lower than in control muscles and was not stimulated by insulin. In parallel, glycogen synthesis and content were decreased. Soleus muscles from male Glut 4-null mice took up twice more deoxyglucose in the absence of insulin than control muscles, but did not respond to insulin. In females, soleus deoxyglucose uptake measured in the absence of hormone was similar in Glut 4-null mice and in control mice. This uptake was stimulated twofold in Glut 4-null mice and threefold in control mice. Basal glycogen synthesis was increased by 4- and 2.2-fold in male and female null mice, respectively, compared to controls, and insulin had no or small (20% stimulation over basal) effect. These results indicate that while EDL muscles behaved as expected, soleus muscles were able to take up a large amount of glucose in the absence (males) or the presence of insulin (females). Whether this is due to a change in Glut 1 intrinsic activity or targeting and/or to the appearance of another glucose transporter remains to be determined.

Plasma Free Fatty Acids Decrease Insulin-Stimulated Skeletal Muscle Glucose Uptake by Suppressing Glycolysis in Conscious Rats

The effects of elevated plasma free fatty acid (FFA) levels on insulin -stimulated whole-body and skeletal muscle glucose transport, glucose uptake, glycolysis, and glycogen synthesis were studied in conscious rats during hyperinsulinemic-euglycemic clamps with (n = 26) or without (n = 23) Intralipid and heparin infusion. Whole-body and skeletal muscle glucose uptake, glycolysis, and glycogen synthesis were estimated using D-[3-3H]glucose and 2-[14C]deoxyglucose (study 1), and glucose transport activity was assessed by analyzing plasma kinetics of L-[14C]glucose and 3-O-[3H]-methylglucose (study 2). Plasma FFA levels decreased during the clamps without intralipid but increased above basal during the clamps with Intralipid infusion (P < 0.01 for both). Elevated plasma FFA levels decreased insulin-stimulated whole-body glucose uptake by approximately 15% and approximately 20% during physiological and maximal insulin clamps, respectively (P < 0.01). Similarly, insulin-stimulated glucose uptake was also decreased in individual skeletal muscles with Intralipid infusion (P < 0.05). The most profound effect of elevated plasma FFA levels was a 30-50% suppression of insulin-stimulated glycolysis in whole body and individual skeletal muscles in both clamps. In contrast, physiological insulin-stimulated glycogen synthesis was increased with elevated plasma FFA levels in whole body and individual skeletal muscles (P < 0.05). Glucose-6-phosphate (G-6-P) levels were increased in soleus and extensor digitorum longus (EDL) muscles with Intralipid infusion in both clamps (P < 0.05). Intralipid infusion did not alter the time profiles of plasma L-glucose and 3-O-methylglucose after an intravenous injection during maximal insulin clamps, and compartmental analysis indicated no significant effect of elevated FFA levels on glucose transport activity in insulin-sensitive tissues (P > 0.05). Thus, elevated plasma FFA decreased insulin-stimulated glucose uptake in skeletal muscle by suppressing glycolysis and increasing G-6-P levels. These findings suggest that the classic glucose-fatty acid cycle was the predominant mechanism underlying the inhibitory effect of FFA on skeletal muscle glucose uptake.

The effect of (steroid) immunosuppression on skeletal muscle glycogen metabolism in patients after kidney transplantation.

To examine the mechanisms by which immunosuppression by steroids impairs glycogen synthesis in human skeletal muscle, we measured glycogen synthase protein content and activity in muscle samples from 14 patients receiving corticosteroid therapy after kidney transplantation and in 20 healthy control subjects. A percutaneous muscle sample was taken before and at the end of a euglycemic hyperinsulinemic insulin clamp. Insulin-stimulated glucose disposal was reduced by 33% in kidney transplant patients compared with healthy controls (33.8 +/- 4.2 vs. 50.5 +/- 2.7 mumol (kg LBM)-1 min-1; P<0.01), primarily due to a decrease in nonoxidative glucose metabolism (14.2 +/- 3.3 vs. 32.3 +/- 2.7 mumol (kg LBM)-1 min-1; P<0.001). Glycogen synthase activity measured at both 0.1 mmol/L (17.6 +/- 2.6 vs. 24.0 +/- 2.2 nmol min-1 mg protein-1; P<0.05), and at 10 mmol/L glucose 6-phosphate (24.1 +/- 3.5 vs. 33.7 +- 2.4 nmol min-1 mg protein-1; P<0.05) and glycogen synthase protein concentrations (8.8 +/- 1.8 vs. 18.9 +/- 1.9 relative units per ng DNA; P<0.01) were lower in kidney transplant patients compared with controls. Glycogen synthase protein correlated with nonoxidative glucose metabolism (r=0.42; P=0.04). Alpha-actinin (used as a control of general protein degradation) was lower in kidney transplant patients compared with controls (4.4 +/- 0.8 vs. 9.6 +/- 1.1 cpm/ng DNA; P<0.01). In conclusion, corticosteroids cause insulin resistance, which correlates with impaired activation of glycogen synthase and decreased enzyme protein content. The decrease in glycogen synthase protein may reflect increased degradation rather than a defect in translation.

Analysis of candidate genes for insulin resistance in essential hypertension.

To clarify the genetic basis of insulin resistance in hypertension, case-control association studies were performed to examine candidate genes for insulin resistance in hypertension. Since the main site of insulin resistance in hypertension is glycogen synthesis in skeletal muscle, genes that encode molecules involved in this pathway, i.e. insulin receptor (INSR), insulin-responsive glucose transporter (GLUT4) and glycogen synthase (GSY), were studied. In addition, since recent studies suggest the contribution of beta3 adrenergic receptor to the insulin resistance syndrome, the gene encoding beta3 adrenergic receptor (ADRB3) was also studied. Frequency of homozygotes for common C allele of a microsatellite polymorphism in the INSR gene was higher in the hyperinsulinemia group, but not in the normoinsulinemia group of hypertensive patients than in normotensive control subjects. Insulin sensitivity, however, was not significantly different between hypertensive patients with C/C genotype and those without this genotype. No significant differences were observed in the distribution of alleles or genotypes of the GLUT4, GSY and ADRB3 genes between hyperinsulinemia and normoinsulinemia groups of hypertensive patients or between these groups and the control group. These data suggest that the INSR polymorphism is associated with hyperinsulinemia, but not with insulin resistance, in hypertension.

Alcohol and glucose metabolism in skeletal muscles in the rat.

Ethanol is known to cause an acute and profound insulin resistance in man and the rat primarily via effects on glucose utilization. This paper examines the nature of these inhibitory effects on whole-body glucose utilization using the euglycaemic hyperinsulinaemic clamp in the conscious unrestrained rat. We confirm that ethanol infusion causes an acute insulin resistance, the rate of glucose infusion required to maintain euglycaemia (GIR) being decreased markedly by ethanol. To ensure that the GIR is a measure of whole-body glucose disposal, glucose turnover and hepatic glycogen levels were measured. These studies showed that ethanol totally suppressed hepatic glucose production. The reduction in GIR is associated with marked decreases in glucose uptake and glycogen synthesis in most skeletal muscles. In oxidative but not in non-oxidative muscles, the activation of glycogen synthase in response to insulin was decreased by ethanol, suggesting that a defect in glycogen synthase activation may be responsible for the decrease in glycogen synthesis. The basis of the inhibitory effects of ethanol on insulin-stimulated glucose metabolism in muscle is unknown, but may involve membrane-associated impairments in insulin signalling and/or the glucose transport system.

Acute Effects of Insulin-like Growth Factor I on Interorgan Glucose and Lactate Flux in Protein-Catabolic Dogs

Insulin-like growth factor-I (IGF-I) is a potent protein-anabolic hormone with a glucose-lowering effect and is therefore a possible agent for treating catabolic patients. In this study we investigated the effect of recombinant human (rh) IGF-I on the interorgan flux of glucose under hypo- and normoglycemic conditions in catabolic, anaesthetized, and catheterized dogs. We administered a primed (40 μg/kg) continuous (1.5 μg · kg-1 · min-1) infusion of rhIGF-I (Kabi Biopharma, Stockholm, Sweden) for 180 min together with either a saline (0.9% NaCl) or an amino acid solution (2.2 mg AA · kg-1 · min-1 solution of Vamin, Kabi Nutrition, Stockholm, Sweden). RhIGF-I administration lowered plasma glucose levels for approximately 50% of the baseline (P < 0.001) and stimulated glucose uptake from skeletal muscle about twofold (P < 0.01), but did not modify glucose balances across the gut and liver. The same effects were found when infusing rhIGF-I together with AA. A co-infusion of rhIGF-I and glucose to maintain normoglycemic conditions stimulated glucose uptake from skeletal muscle by about fivefold (P < 0.001) and glucose uptake across the gut by about 50%, but reduced the hepatic glucose liberation (-65%; P < 0.01). The rhIGF-I infusion did not alter arterial lactate levels, but stimulated lactate release from skeletal muscle (P < 0.05) and lactate uptake across the liver (P < 0.05). We conclude that rhIGF-I reduces plasma glucose levels mainly by stimulating glucose uptake across skeletal muscle. Because muscular glucose uptake was higher than lactate release, rhIGF-I possibly stimulated glycogen synthesis or ATP formation in skeletal muscle and therefore may have positive effects on the peripheral energy deficit in catabolic state.

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.

Cloning of a human insulin-stimulated protein kinase (ISPK-1) gene and analysis of coding regions and mRNA levels of the ISPK-1 and the protein phosphatase-1 genes in muscle from NIDDM patients.

Complementary DNA encoding three catalytic subunits of protein phosphatase 1 (PP1 alpha, PP1 beta, and PP1 gamma) and the insulin-stimulated protein kinase 1 (ISPK-1) was analyzed for variations in the coding regions related to insulin-resistant glycogen synthesis in skeletal muscle of 30 patients with non-insulin-dependent diabetes mellitus (NIDDM). The human ISPK-1 cDNA was cloned from T-cell leukemia and placental cDNA libraries and mapped to the short arm of the human X chromosome. Single-strand conformation polymorphism (SSCP) analysis identified a total of six variations in the coding regions of the PP1 genes: two in PP1 alpha at codons 90 and 255; one in PP1 beta at codon 67; and three in PP1 gamma at codons 11,269, and 273, respectively. All were, however, silent single nucleotide substitutions. SSCP analysis of the ISPK-1 gene identified one silent polymorphism at codon 266 and one amino acid variant at codon 38 (Ile-->Ser). This variant was primarily found in one male NIDDM patient. This subject, however, did not exhibit an impairment of muscle insulin-stimulated glycogen synthase activation. No significant differences were found in mRNA levels in muscle of the four genes between 15 NIDDM patients and 14 healthy subjects. Our findings suggest that 1) genetic abnormalities in the coding regions of PP1 alpha, PP1 beta, PP1 gamma, and ISPK-1 are unlikely to be frequently occurring causes of the reduced insulin-stimulated activation of the glycogen synthesis in muscle from the analyzed group of NIDDM patients; 2) the mRNA levels of PP1 alpha, PP1 beta, PP1 gamma, and ISPK-1 are normal in muscle from the NIDDM patients; and 3) putative inherited defects in insulin-stimulated activation of muscle glycogen synthesis in patients with insulin-resistant NIDDM may be located further upstream of ISPK-1 in the insulin action cascade.