Acute Hypoglycemia in Healthy Humans Impairs Insulin-Stimulated Glucose Uptake and Glycogen Synthase in Skeletal Muscle: A Randomized Clinical Study

Insulin promotes dephosphorylation and activation of glycogen synthase (GS) by inactivating glycogen synthase kinase (GSK) 3 through phosphorylation. Insulin also promotes glucose uptake and glucose 6-phosphate (G-6- P) production, which allosterically activates GS. The relative importance of these two regulatory mechanisms in the activation of GS in vivo is unknown. The aim of this study was to investigate if dephosphorylation of GS mediated via GSK3 is required for normal glycogen synthesis in skeletal muscle with insulin. We employed GSK3 knockin mice in which wild-type GSK3α and -β genes are replaced with mutant forms (GSK3α/βS21A/S21A/S9A/S9A), which are nonresponsive to insulin. Although insulin failed to promote dephosphorylation and activation of GS in GSK3α/βS21A/S21A/S9A/S9Amice, glycogen content in different muscles from these mice was similar compared with wild-type mice. Basal and epinephrine-stimulated activity of muscle glycogen phosphorylase was comparable between wild-type and GSK3 knockin mice. Incubation of isolated soleus muscle in Krebs buffer containing 5.5 mM glucose in the presence or absence of insulin revealed that the levels of G-6- P, the rate of [14C]glucose incorporation into glycogen, and an increase in total glycogen content were similar between wild-type and GSK3 knockin mice. Injection of glucose containing 2-deoxy-[3H]glucose and [14C]glucose also resulted in similar rates of muscle glucose uptake and glycogen synthesis in vivo between wild-type and GSK3 knockin mice. These results suggest that insulin-mediated inhibition of GSK3 is not a rate-limiting step in muscle glycogen synthesis in mice. This suggests that allosteric regulation of GS by G-6- P may play a key role in insulin-stimulated muscle glycogen synthesis in vivo.

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Regulation of hexokinase II activity and expression in human muscle by moderate exercise

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1998.274.2.e304 ◽

1998 ◽

Vol 274 (2) ◽

pp. E304-E308 ◽

Cited By ~ 23

Author(s):

Janice A. Koval ◽

Ralph A. DeFronzo ◽

Robert M. O’Doherty ◽

Richard Printz ◽

Hossein Ardehali ◽

...

Keyword(s):

Skeletal Muscle ◽

Glucose Uptake ◽

Glycogen Synthase ◽

Particulate Fraction ◽

Moderate Intensity ◽

Vastus Lateralis Muscle ◽

28S Rrna ◽

Moderate Exercise ◽

Hexokinase Ii ◽

Single Bout

A single bout of exercise increases the rate of insulin-stimulated glucose uptake and metabolism in skeletal muscle. Exercise also increases insulin-stimulated glucose 6-phosphate in skeletal muscle, suggesting that exercise increases hexokinase activity. Within 3 h, exercise increases hexokinase II (HK II) mRNA and activity in skeletal muscle from rats. It is not known, however, if a single bout of moderate-intensity exercise increases HK II expression in humans. The present study was undertaken to answer this question. Six subjects had percutaneous biopsies of the vastus lateralis muscle before and 3 h after a single 3-h session of moderate-intensity aerobic (60% of maximal oxygen consumption) exercise. Glycogen synthase, HK I, and HK II activities as well as HK I and HK II mRNA content were determined from the muscle biopsy specimens. The fractional velocity of glycogen synthase was increased by 446 ± 84% after exercise ( P < 0.005). Hexokinase II activity in the soluble fraction of the homogenates increased from 1.2 ± 0.4 to 4.5 ± 1.6 pmol ⋅ min−1 ⋅ μg−1( P < 0.05) but was unchanged in the particulate fraction (4.3 ± 1.3 vs. 5.3 ± 1.5). HK I activity in neither the soluble nor particulate fraction changed after exercise. Relative to a 28S rRNA control signal, HK II mRNA increased from 0.091 ± 0.02 to 0.195 ± 0.037 ( P < 0.05), whereas HK I mRNA was unchanged (0.414 ± 0.061 vs. 0.498 ± 0.134, P < 0.20). The increase in HK II activity after moderate exercise in healthy subjects could be one factor responsible for the enhanced rate of insulin-stimulated glucose uptake seen after exercise.

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Muscle-specific overexpression of wild type and R225Q mutant AMP-activated protein kinase γ3-subunit differentially regulates glycogen accumulation

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00073.2006 ◽

2006 ◽

Vol 291 (3) ◽

pp. E557-E565 ◽

Cited By ~ 27

Author(s):

Haiyan Yu ◽

Michael F. Hirshman ◽

Nobuharu Fujii ◽

Jason M. Pomerleau ◽

Lauren E. Peter ◽

...

Keyword(s):

Skeletal Muscle ◽

Protein Kinase ◽

Glucose Uptake ◽

Glycogen Content ◽

Specific Activity ◽

Glycogen Synthase ◽

Underlying Mechanism ◽

Wild Type ◽

Glycogen Accumulation ◽

Amp Activated Protein Kinase

AMP-activated protein kinase (AMPK) is a heterotrimeric complex that works as an energy sensor to integrate nutritional and hormonal signals. The naturally occurring R225Q mutation in the γ3-subunit in pigs is associated with abnormally high glycogen content in skeletal muscle. Becauses skeletal muscle accounts for most of the body's glucose uptake, and γ3 is specifically expressed in skeletal muscle, it is important to understand the underlying mechanism of this mutation in regulating glucose and glycogen metabolism. Using skeletal muscle-specific transgenic mice overexpressing wild type γ3 (WTγ3) and R225Q mutant γ3 (MUTγ3), we show that both WTγ3 and MUTγ3 mice have 1.5- to 2-fold increases in muscle glycogen content. In WTγ3 mice, increased glycogen content was associated with elevated total glycogen synthase activity and reduced glycogen phosphorylase activity, whereas alterations in activities of these enzymes could not explain elevated glycogen in MUTγ3 mice. Basal, 5-aminoimidazole- AICAR- and phenformin-stimulated AMPKα2 isoform-specific activities were decreased only in MUTγ3 mice. Basal rates of 2-DG glucose uptake were decreased in both WTγ3 and MUTγ3 mice. However, AICAR- and phenformin-stimulated 2-DG glucose uptake were blunted only in MUTγ3 mice. In conclusion, expression of either wild type or mutant γ3-subunit of AMPK results in increased glycogen concentrations in muscle, but the mechanisms underlying this alteration appear to be different. Furthermore, mutation of the γ3-subunit is associated with decreases in AMPKα2 isoform-specific activity and impairment in AICAR- and phenformin-stimulated skeletal muscle glucose uptake.

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Glucose infusion causes insulin resistance in skeletal muscle of rats without changes in Akt and AS160 phosphorylation

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00133.2007 ◽

2007 ◽

Vol 293 (5) ◽

pp. E1358-E1364 ◽

Cited By ~ 34

Author(s):

Andrew J. Hoy ◽

Clinton R. Bruce ◽

Anna Cederberg ◽

Nigel Turner ◽

David E. James ◽

...

Keyword(s):

Insulin Resistance ◽

Skeletal Muscle ◽

Glucose Uptake ◽

Insulin Signaling ◽

Glycogen Synthase ◽

Glycogen Synthesis ◽

Glucose Infusion ◽

Metabolic Effects ◽

Synthesis Rate ◽

Muscle Sample

Hyperglycemia is a defining feature of Type 1 and 2 diabetes. Hyperglycemia also causes insulin resistance, and our group (Kraegen EW, Saha AK, Preston E, Wilks D, Hoy AJ, Cooney GJ, Ruderman NB. Am J Physiol Endocrinol Metab Endocrinol Metab 290: E471–E479, 2006) has recently demonstrated that hyperglycemia generated by glucose infusion results in insulin resistance after 5 h but not after 3 h. The aim of this study was to investigate possible mechanism(s) by which glucose infusion causes insulin resistance in skeletal muscle and in particular to examine whether this was associated with changes in insulin signaling. Hyperglycemia (∼10 mM) was produced in cannulated male Wistar rats for up to 5 h. The glucose infusion rate required to maintain this hyperglycemia progressively lessened over 5 h (by 25%, P < 0.0001 at 5 h) without any alteration in plasma insulin levels consistent with the development of insulin resistance. Muscle glucose uptake in vivo (44%; P < 0.05) and glycogen synthesis rate (52%; P < 0.001) were reduced after 5 h compared with after 3 h of infusion. Despite these changes, there was no decrease in the phosphorylation state of multiple insulin signaling intermediates [insulin receptor, Akt, AS160 (Akt substrate of 160 kDa), glycogen synthase kinase-3β] over the same time course. In isolated soleus strips taken from control or 1- or 5-h glucose-infused animals, insulin-stimulated 2-deoxyglucose transport was similar, but glycogen synthesis was significantly reduced in the 5-h muscle sample (68% vs. 1-h sample; P < 0.001). These results suggest that the reduced muscle glucose uptake in rats after 5 h of acute hyperglycemia is due more to the metabolic effects of excess glycogen storage than to a defect in insulin signaling or glucose transport.

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Skeletal muscle glucose uptake, glycogen synthase activity and GLUT 4 content during hypoglycaemia in type 1 diabetic subjects

CrossRef Listing of Deleted DOIs ◽

10.1080/003655101316911413 ◽

2001 ◽

Vol 61 (5) ◽

pp. 371-381 ◽

Cited By ~ 2

Author(s):

L. Ørskov ◽

O. Schmitz ◽

J. F. Bak ◽

S. Lund ◽

A. Kaal ◽

...

Keyword(s):

Skeletal Muscle ◽

Glucose Uptake ◽

Glycogen Synthase ◽

Glut 4 ◽

Skeletal Muscle Glucose Uptake ◽

Glycogen Synthase Activity

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Skeletal muscle glucose metabolism in obesity-prone and obesity-resistant rats

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1993.264.6.r1224 ◽

1993 ◽

Vol 264 (6) ◽

pp. R1224-R1228 ◽

Cited By ~ 2

Author(s):

M. J. Pagliassotti ◽

K. A. Shahrokhi ◽

J. O. Hill

Keyword(s):

Skeletal Muscle ◽

Body Weight ◽

Weight Gain ◽

Glucose Metabolism ◽

Glucose Uptake ◽

Glycogen Synthase ◽

Body Weight Gain ◽

Glycogen Synthesis ◽

Insulin Resistant ◽

Wide Range

Ad libitum access to a high-fat (HF) diet produces a wide range of weight gain in rats. Rats most susceptible to weight gain on such a diet (obesity prone; OP) are more insulin resistant after 4-5 wk of diet exposure than are those most resistant (obesity resistant; OR) to weight gain. To investigate whether skeletal muscle glucose metabolism contributes to insulin resistance in this model, insulin-stimulated glucose metabolism was assessed in the perfused hindquarter of rats exposed to either a low-fat (LF, n = 6) or HF diet for 5 wk. Delineation of OP (n = 6) and OR (n = 6) rats was based on body weight gain. OP rats gained 60% more body weight while eating only 10% more energy than OR rats. Single-pass perfusions were carried out for 2 h in the presence of glucose, insulin, and [U-14C]glucose. Insulin-stimulated glucose uptake (mumol.100 g-1.min-1) was 14.2 +/- 0.9 in LF, 11.1 +/- 0.8 in OR, and 6.2 +/- 0.6 in OP. Glucose oxidation (mumol.100 g-1.min-1) was 1.7 +/- 0.3 and 1.2 +/- 0.3 in LF and OR, respectively, but was 0.2 +/- 0.1 in OP. Net glycogen synthesis was significantly reduced in OP compared with OR and LF despite similar glycogen synthase I activity. Muscle triglyceride concentration was not significantly different in OR and OP rats. These results demonstrate significant defects in skeletal muscle glucose uptake and disposal in rats most susceptible to HF diet-induced obesity. Clearly, the heterogeneous response to a HF diet involves not only body weight gain but also skeletal muscle fuel metabolism.

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Quantification, Variability, and Reproducibility of Basal Skeletal Muscle Glucose Uptake in Healthy Humans Using 18F-FDG PET/CT

Journal of Nuclear Medicine ◽

10.2967/jnumed.115.159715 ◽

2015 ◽

Vol 56 (10) ◽

pp. 1520-1526 ◽

Cited By ~ 6

Author(s):

O. Gheysens ◽

A. Postnov ◽

C. M. Deroose ◽

C. Vandermeulen ◽

J. de Hoon ◽

...

Keyword(s):

Skeletal Muscle ◽

Glucose Uptake ◽

Fdg Pet ◽

Healthy Humans ◽

Pet Ct ◽

Skeletal Muscle Glucose Uptake ◽

18F Fdg ◽

Fdg Pet Ct

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Cellular depletion of atypical PKCλ is associated with enhanced insulin sensitivity and glucose uptake in L6 rat skeletal muscle cells

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00171.2010 ◽

2010 ◽

Vol 299 (3) ◽

pp. E402-E412 ◽

Cited By ~ 19

Author(s):

Clare Stretton ◽

Ashleigh Evans ◽

Harinder S. Hundal

Keyword(s):

Skeletal Muscle ◽

Protein Kinase ◽

Insulin Receptor ◽

Glucose Uptake ◽

Glucose Transport ◽

Insulin Signaling ◽

Glycogen Synthase ◽

Muscle Cells ◽

Skeletal Muscle Cells ◽

Dominant Negative Mutant

Atypical protein kinase C (aPKC) isoforms (λ and ζ) have been implicated in the control of insulin-stimulated glucose uptake in adipose and skeletal muscle, but their precise role in this process remains unclear, especially in light of accumulating evidence showing that, in response to numerous stimuli, including insulin and lipids such as ceramide, activation of aPKCs acts to negatively regulate key insulin-signaling molecules, such as insulin receptor substrate-1 (IRS-1) and protein kinase B (PKB)/cAMP-dependent PKC (Akt). In this study, we have depleted PKCλ in L6 skeletal muscle cells using RNA interference and assessed the effect this has upon insulin action. Muscle cells did not express detectable amounts of PKCζ. Depletion of PKCλ (>95%) had no significant effect on the expression of proteins participating in insulin signaling [i.e., insulin receptor, IRS-1, phosphatidylinositol 3-kinase (PI 3-kinase), PKB, or phosphate and tensin homolog deleted on chromosome 10] or those involved in glucose transport [Akt substrate of 160 kDa, glucose transporter (GLUT)1, or GLUT4]. However, PKCλ-depleted muscle cells exhibited greater activation of PKB/Akt and phosphorylation of its downstream target glycogen synthase kinase 3, in the basal state and displayed greater responsiveness to submaximal doses of insulin with respect to p85-PI 3-kinase/IRS-1 association and PKB activation. The increase in basal and insulin-induced signaling resulted in an associated enhancement of basal and insulin-stimulated glucose transport, both of which were inhibited by the PI 3-kinase inhibitor wortmannin. Additionally, like RNAi-mediated depletion of PKCλ, overexpression of a dominant-negative mutant of PKCζ induced a similar insulin-sensitizing effect on PKB activation. Our findings indicate that aPKCs are likely to play an important role in restraining proximal insulin signaling events but appear dispensable with respect to insulin-stimulated glucose uptake in cultured L6 muscle cells.

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Functional Studies of Akt Isoform Specificity in Skeletal Muscle in Vivo; Maintained Insulin Sensitivity Despite Reduced Insulin Receptor Substrate-1 Expression

Molecular Endocrinology ◽

10.1210/me.2006-0154 ◽

2007 ◽

Vol 21 (1) ◽

pp. 215-228 ◽

Cited By ~ 70

Author(s):

Mark E. Cleasby ◽

Tracie A. Reinten ◽

Gregory J. Cooney ◽

David E. James ◽

Edward W. Kraegen

Keyword(s):

Insulin Resistance ◽

Skeletal Muscle ◽

Glucose Metabolism ◽

Insulin Receptor ◽

Glucose Uptake ◽

Glycogen Synthase ◽

Insulin Receptor Substrate ◽

Insulin Receptor Substrate 1 ◽

Short Hairpin

Abstract The phosphoinositide 3-kinase/Akt pathway is thought to be essential for normal insulin action and glucose metabolism in skeletal muscle and has been shown to be dysregulated in insulin resistance. However, the specific roles of and signaling pathways triggered by Akt isoforms have not been fully assessed in muscle in vivo. We overexpressed constitutively active (ca-) Akt-1 or Akt-2 constructs in muscle using in vivo electrotransfer and, after 1 wk, assessed the roles of each isoform on glucose metabolism and fiber growth. We achieved greater than 2.5-fold increases in total Ser473 phosphorylation in muscles expressing ca-Akt-1 and ca-Akt-2, respectively. Both isoforms caused hypertrophy of muscle fibers, consistent with increases in p70S6kinase phosphorylation, and a 60% increase in glycogen accumulation, although only Akt-1 increased glycogen synthase kinase-3β phosphorylation. Akt-2, but not Akt-1, increased basal glucose uptake (by 33%, P = 0.004) and incorporation into glycogen and lipids, suggesting a specific effect on glucose transport. Consistent with this, short hairpin RNA-mediated silencing of Akt-2 caused reductions in glycogen storage and glucose uptake. Consistent with Akt-mediated insulin receptor substrate 1 (IRS-1) degradation, we observed approximately 30% reductions in IRS-1 protein in muscle overexpressing ca-Akt-1 or ca-Akt-2. Despite this, we observed no decrease in insulin-stimulated glucose uptake. Furthermore, a 68% reduction in IRS-1 levels induced using short hairpin RNAs targeting IRS-1 also did not affect glucose disposal after a glucose load. These data indicate distinct roles for Akt-1 and Akt-2 in muscle glucose metabolism and that moderate reductions in IRS-1 expression do not result in the development of insulin resistance in skeletal muscle in vivo.

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Invited Review: Intracellular signaling in contracting skeletal muscle

Journal of Applied Physiology ◽

10.1152/japplphysiol.00167.2002 ◽

2002 ◽

Vol 93 (1) ◽

pp. 369-383 ◽

Cited By ~ 161

Author(s):

Kei Sakamoto ◽

Laurie J. Goodyear

Keyword(s):

Skeletal Muscle ◽

Protein Kinase ◽

Physical Exercise ◽

Glucose Uptake ◽

Intracellular Signaling ◽

Glycogen Synthase ◽

Contractile Activity ◽

Glycogen Synthesis ◽

Mitogen Activated Protein Kinase ◽

Molecular Signaling

Physical exercise is a significant stimulus for the regulation of multiple metabolic and transcriptional processes in skeletal muscle. For example, exercise increases skeletal muscle glucose uptake, and, after exercise, there are increases in the rates of both glucose uptake and glycogen synthesis. A single bout of exercise can also induce transient changes in skeletal muscle gene transcription and can alter rates of protein metabolism, both of which may be mechanisms for chronic adaptations to repeated bouts of exercise. A central issue in exercise biology is to elucidate the underlying molecular signaling mechanisms that regulate these important metabolic and transcriptional events in skeletal muscle. In this review, we summarize research from the past several years that has demonstrated that physical exercise can regulate multiple intracellular signaling cascades in skeletal muscle. It is now well established that physical exercise or muscle contractile activity can activate three of the mitogen-activated protein kinase signaling pathways, including the extracellular signal-regulated kinase 1 and 2, the c-Jun NH2-terminal kinase, and the p38. Exercise can also robustly increase activity of the AMP-activated protein kinase, as well as several additional molecules, including glycogen synthase kinase 3, Akt, and the p70 S6 kinase. A fundamental goal of signaling research is to determine the biological consequences of exercise-induced signaling through these molecules, and this review also provides an update of progress in this area.

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