Local hindlimb antioxidant infusion does not affect muscle glucose uptake during in situ contractions in rat

2010 ◽  
Vol 108 (5) ◽  
pp. 1275-1283 ◽  
Author(s):  
T. L. Merry ◽  
R. M. Dywer ◽  
E. A. Bradley ◽  
S. Rattigan ◽  
G. K. McConell
Keyword(s):  
Skeletal Muscle ◽  
Glucose Uptake ◽  
Ex Vivo ◽  
Moderate Intensity ◽  
Ampk Signaling ◽  
Epigastric Artery ◽  
Hindlimb Muscle ◽  
Skeletal Muscle Glucose Uptake ◽  
Amp Activated Protein Kinase

There is evidence that reactive oxygen species (ROS) contribute to the regulation of skeletal muscle glucose uptake during highly fatiguing ex vivo contraction conditions via AMP-activated protein kinase (AMPK). In this study we investigated the role of ROS in the regulation of glucose uptake and AMPK signaling during low-moderate intensity in situ hindlimb muscle contractions in rats, which is a more physiological protocol and preparation. Male hooded Wistar rats were anesthetized, and then N-acetylcysteine (NAC) was infused into the epigastric artery (125 mg·kg−1·h−1) of one hindlimb (contracted leg) for 15 min before this leg was electrically stimulated (0.1-ms impulse at 2 Hz and 35 V) to contract at a low-moderate intensity for 15 min. The contralateral leg did not receive stimulation or local NAC infusion (rest leg). NAC infusion increased ( P < 0.05) plasma cysteine and cystine (by ∼360- and 1.4-fold, respectively) and muscle cysteine (by 1.5-fold, P = 0.001). Although contraction did not significantly alter muscle tyrosine nitration, reduced (GSH) or oxidized glutathione (GSSG) content, S-glutathionylation of protein bands at ∼250 and 150 kDa was increased ( P < 0.05) ∼1.7-fold by contraction, and this increase was prevented by NAC. Contraction increased ( P < 0.05) skeletal muscle glucose uptake 20-fold, AMPK phosphorylation 6-fold, ACCβ phosphorylation 10-fold, and p38 MAPK phosphorylation 60-fold, and the muscle fatigued by ∼30% during contraction and NAC infusion had no significant effect on any of these responses. This was despite NAC preventing increases in S-glutathionylation with contraction. In conclusion, unlike during highly fatiguing ex vivo contractions, local NAC infusion during in situ low-moderate intensity hindlimb contractions in rats, a more physiological preparation, does not attenuate increases in skeletal muscle glucose uptake or AMPK signaling.

Glucose uptake during contraction in isolated skeletal muscles from neuronal nitric oxide synthase μ knockout mice

2015 ◽  
Vol 118 (9) ◽  
pp. 1113-1121 ◽  
Author(s):  
Yet Hoi Hong ◽  
Tony Frugier ◽  
Xinmei Zhang ◽  
Robyn M. Murphy ◽  
Gordon S. Lynch ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Skeletal Muscle ◽  
Nitric Oxide Synthase ◽  
Glucose Uptake ◽  
Ex Vivo ◽  
Organ Bath ◽  
Skeletal Muscle Glucose Uptake ◽  
Nos Inhibitor ◽  
Amp Activated Protein Kinase ◽  
Oxide Synthase

Inhibition of nitric oxide synthase (NOS) significantly attenuates the increase in skeletal muscle glucose uptake during contraction/exercise, and a greater attenuation is observed in individuals with Type 2 diabetes compared with healthy individuals. Therefore, NO appears to play an important role in mediating muscle glucose uptake during contraction. In this study, we investigated the involvement of neuronal NOSμ (nNOSμ), the main NOS isoform activated during contraction, on skeletal muscle glucose uptake during ex vivo contraction. Extensor digitorum longus muscles were isolated from nNOSμ−/−and nNOSμ+/+mice. Muscles were contracted ex vivo in a temperature-controlled (30°C) organ bath with or without the presence of the NOS inhibitor NG-monomethyl-l-arginine (L-NMMA) and the NOS substrate L-arginine. Glucose uptake was determined by radioactive tracers. Skeletal muscle glucose uptake increased approximately fourfold during contraction in muscles from both nNOSμ−/−and nNOSμ+/+mice. L-NMMA significantly attenuated the increase in muscle glucose uptake during contraction in both genotypes. This attenuation was reversed by L-arginine, suggesting that L-NMMA attenuated the increase in muscle glucose uptake during contraction by inhibiting NOS and not via a nonspecific effect of the inhibitor. Low levels of NOS activity (∼4%) were detected in muscles from nNOSμ−/−mice, and there was no evidence of compensation from other NOS isoform or AMP-activated protein kinase which is also involved in mediating muscle glucose uptake during contraction. These results indicate that NO regulates skeletal muscle glucose uptake during ex vivo contraction independently of nNOSμ.

No effect of NOS inhibition on skeletal muscle glucose uptake during in situ hindlimb contraction in healthy and diabetic Sprague-Dawley rats

2015 ◽  
Vol 308 (10) ◽  
pp. R862-R871 ◽  
Author(s):  
Yet Hoi Hong ◽  
Andrew C. Betik ◽  
Dino Premilovac ◽  
Renee M. Dwyer ◽  
Michelle A. Keske ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Blood Flow ◽  
Glucose Uptake ◽  
Femoral Artery ◽  
Systemic Blood Pressure ◽  
Sprague Dawley ◽  
Capillary Recruitment ◽  
Sprague Dawley Rats ◽  
Skeletal Muscle Glucose Uptake

Nitric oxide (NO) has been shown to be involved in skeletal muscle glucose uptake during contraction/exercise, especially in individuals with Type 2 diabetes (T2D). To examine the potential mechanisms, we examined the effect of local NO synthase (NOS) inhibition on muscle glucose uptake and muscle capillary blood flow during contraction in healthy and T2D rats. T2D was induced in Sprague-Dawley rats using a combined high-fat diet (23% fat wt/wt for 4 wk) and low-dose streptozotocin injections (35 mg/kg). Anesthetized animals had one hindlimb stimulated to contract in situ for 30 min (2 Hz, 0.1 ms, 35 V) with the contralateral hindlimb rested. After 10 min, the NOS inhibitor, NG-nitro-l-arginine methyl ester (l-NAME; 5 μM) or saline was continuously infused into the femoral artery of the contracting hindlimb until the end of contraction. Surprisingly, there was no increase in skeletal muscle NOS activity during contraction in either group. Local NOS inhibition had no effect on systemic blood pressure or muscle contraction force, but it did cause a significant attenuation of the increase in femoral artery blood flow in control and T2D rats. However, NOS inhibition did not attenuate the increase in muscle capillary recruitment during contraction in these rats. Muscle glucose uptake during contraction was significantly higher in T2D rats compared with controls but, unlike our previous findings in hooded Wistar rats, NOS inhibition had no effect on glucose uptake during contraction. In conclusion, NOS inhibition did not affect muscle glucose uptake during contraction in control or T2D Sprague-Dawley rats, and this may have been because there was no increase in NOS activity during contraction.

Skeletal Muscle Glucose Uptake During Exercise: How is it Regulated?

Physiology ◽  
2005 ◽  
Vol 20 (4) ◽  
pp. 260-270 ◽  
Author(s):  
Adam J. Rose ◽  
Erik A. Richter
Keyword(s):  
Skeletal Muscle ◽  
Protein Kinase ◽  
Glucose Uptake ◽  
Intracellular Signaling ◽  
Surface Membrane ◽  
Capillary Perfusion ◽  
Subsequent Increase ◽  
Dependent Protein ◽  
Skeletal Muscle Glucose Uptake ◽  
Amp Activated Protein Kinase

The increase in skeletal muscle glucose uptake during exercise results from a coordinated increase in rates of glucose delivery (higher capillary perfusion), surface membrane glucose transport, and intracellular substrate flux through glycolysis. The mechanism behind the movement of GLUT4 to surface membranes and the subsequent increase in transport by muscle contractions is largely unresolved, but it is likely to occur through intracellular signaling involving Ca2+-calmodulin-dependent protein kinase, 5′-AMP-activated protein kinase, and possibly protein kinase C.

Downstream mechanisms of nitric oxide-mediated skeletal muscle glucose uptake during contraction

2010 ◽  
Vol 299 (6) ◽  
pp. R1656-R1665 ◽  
Author(s):  
Troy L. Merry ◽  
Gordon S. Lynch ◽  
Glenn K. McConell
Keyword(s):  
Nitric Oxide ◽  
Skeletal Muscle ◽  
Glucose Uptake ◽  
P38 Mapk ◽  
Ex Vivo ◽  
Protein S ◽  
Dependent Protein ◽  
No Signaling ◽  
Skeletal Muscle Glucose Uptake ◽  
Nos Inhibitor

There is evidence that nitric oxide (NO) is required for the normal increases in skeletal muscle glucose uptake during contraction, but the mechanisms involved have not been elucidated. We examined whether NO regulates glucose uptake during skeletal muscle contractions via cGMP-dependent or cGMP-independent pathways. Isolated extensor digitorum longus (EDL) muscles from mice were stimulated to contract ex vivo, and potential NO signaling pathways were blocked by the addition of inhibitors to the incubation medium. Contraction increased ( P < 0.05) NO synthase (NOS) activity (∼40%) and dichlorofluorescein (DCF) fluorescence (a marker of oxidant levels; ∼95%), which was prevented with a NOS inhibitor NG-monomethyl-l-arginine (l-NMMA), and antioxidants [nonspecific antioxidant, N-acetylcysteine (NAC); thiol-reducing agent, DTT], respectively. l-NMMA and NAC both attenuated glucose uptake during contraction by ∼50% ( P < 0.05), and their effects were not additive. Neither the guanylate cyclase inhibitor 1 H-[1,2,4]oxadiazolo-[4,3- a]quinoxalin-1-one, which prevents the formation of cGMP, the cGMP-dependent protein (PKG) inhibitor Rp-8-bromo-β-phenyl-1,N2-ethenoguanosine 3′,5′-cyclic monophosphorothioate sodium salt nor white light, which breaks S-nitrosylated bonds, affects glucose uptake during contraction; however, DTT attenuated ( P < 0.05) contraction-stimulated glucose uptake (by 70%). NOS inhibition and antioxidant treatment reduced contraction-stimulated increases in protein S-glutathionylation and tyrosine nitration ( P < 0.05), without affecting AMPK or p38 MAPK phosphorylation. In conclusion, we provide evidence to suggest that NOS-derived oxidants regulate skeletal muscle glucose uptake during ex vivo contractions via a cGMP/PKG-, AMPK-, and p38 MAPK-independent pathway. In addition, it appears that NO and ROS may regulate skeletal muscle glucose uptake during contraction through a similar pathway.

Knockout of the predominant conventional PKC isoform, PKCα, in mouse skeletal muscle does not affect contraction-stimulated glucose uptake

2009 ◽  
Vol 297 (2) ◽  
pp. E340-E348 ◽  
Author(s):  
Thomas E. Jensen ◽  
Stine J. Maarbjerg ◽  
Adam J. Rose ◽  
Michael Leitges ◽  
Erik A. Richter
Keyword(s):  
Skeletal Muscle ◽  
Protein Kinase ◽  
Glucose Uptake ◽  
Ex Vivo ◽  
Elongation Factor ◽  
Wild Type ◽  
Calphostin C ◽  
Eukaryotic Elongation Factor ◽  
In The Wild ◽  
Amp Activated Protein Kinase

Conventional (c) protein kinase C (PKC) activity has been shown to increase with skeletal muscle contraction, and numerous studies using primarily pharmacological inhibitors have implicated cPKCs in contraction-stimulated glucose uptake. Here, to confirm that cPKC activity is required for contraction-stimulated glucose uptake in mouse muscles, contraction-stimulated glucose uptake ex vivo was first evaluated in the presence of three commonly used cPKC inhibitors (calphostin C, Gö-6976, and Gö-6983) in incubated mouse soleus and extensor digitorum longus (EDL) muscles. All potently inhibited contraction-stimulated glucose uptake by 50–100%, whereas both Gö compounds, but not calphostin C, inhibited insulin-stimulated glucose uptake modestly. AMP-activated protein kinase (AMPK) and eukaryotic elongation factor 2 phosphorylation was unaffected by the blockers. PKCα was estimated to account for ∼97% of total cPKC protein expression in skeletal muscle. However, in muscles from PKCα knockout (KO) mice, neither contraction- nor phorbol ester-stimulated glucose uptake ex vivo differed compared with the wild type. Furthermore, the effects of calphostin C and Gö-6983 on contraction-induced glucose uptake were similar in muscles lacking PKCα and in the wild type. It can be concluded that PKCα, representing ∼97% of cPKC in skeletal muscle, is not required for contraction-stimulated glucose uptake. Thus the effect of the PKC blockers on glucose uptake is either nonspecific working on other parts of contraction-induced signaling or the remaining cPKC isoforms are sufficient for stimulating glucose uptake during contractions.

Sirtuin 1 is not required for contraction-stimulated glucose uptake in mouse skeletal muscle

2021 ◽  
Author(s):  
Ji Hyun Kang ◽  
Ji E. Park ◽  
Jason Dagoon ◽  
Stewart W.C. Masson ◽  
Troy L. Merry ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Glucose Uptake ◽  
Muscle Force ◽  
Ex Vivo ◽  
Sirtuin 1 ◽  
Muscle Type ◽  
Mouse Skeletal Muscle ◽  
Dependent Protein ◽  
Skeletal Muscle Glucose Uptake ◽  
Protein Deacetylase

While it has long been known that contraction robustly stimulates skeletal muscle glucose uptake, the molecular steps regulating this increase remain incompletely defined. The mammalian ortholog of Sir2, sirtuin 1 (SIRT1), is an NAD+-dependent protein deacetylase that is thought to link perturbations in energy flux associated with exercise to subsequent cellular adaptations. Nevertheless, its role in contraction-stimulated glucose uptake has not been described. The objective of this study was to determine the importance of SIRT1 to contraction-stimulated glucose uptake in mouse skeletal muscle. Using a radioactive 2-deoxyglucose uptake (2DOGU) approach, we measured ex vivo glucose uptake in unstimulated (rested) and electrically-stimulated (100 Hz contraction every 15s for 10 min; contracted) extensor digitorum longus (EDL) and soleus from ~15 week old male and female mice with muscle-specific knockout of SIRT1 deacetylase activity (mKO) and their wildtype (WT) littermates. Skeletal muscle force decreased over the contraction protocol, although there were no differences in the rate of fatigue between genotypes. In EDL and soleus, depletion of SIRT1 deacetylase activity did not affect contraction-induced increase in glucose uptake in either sex. Interestingly, the absolute rate of contraction-stimulated 2DOGU was ~1.4-fold higher in female compared to male mice, regardless of muscle type. Taken together, our findings demonstrate that SIRT1 is not required for contraction-stimulated glucose uptake in mouse skeletal muscle. Moreover, to our knowledge, this is the first demonstration of sex-based differences in contraction-stimulated glucose uptake in mouse skeletal muscle.

Skeletal muscle glucose uptake during contraction is regulated by nitric oxide and ROS independently of AMPK

2010 ◽  
Vol 298 (3) ◽  
pp. E577-E585 ◽  
Author(s):  
Troy L. Merry ◽  
Gregory R. Steinberg ◽  
Gordon S. Lynch ◽  
Glenn K. McConell
Keyword(s):  
Nitric Oxide ◽  
Skeletal Muscle ◽  
Glucose Uptake ◽  
Similar Extent ◽  
Independent Mechanism ◽  
Skeletal Muscle Glucose Uptake ◽  
Nos Inhibitor ◽  
Muscle Specific Kinase ◽  
Amp Activated Protein Kinase ◽  
Oxide Synthase

Reactive oxygen species (ROS) and nitric oxide (NO) have been implicated in the regulation of skeletal muscle glucose uptake during contraction, and there is evidence that they do so via interaction with AMP-activated protein kinase (AMPK). In this study, we tested the hypothesis that ROS and NO regulate skeletal muscle glucose uptake during contraction via an AMPK-independent mechanism. Isolated extensor digitorum longus (EDL) and soleus muscles from mice that expressed a muscle-specific kinase dead AMPKα2 isoform (AMPK-KD) and wild-type litter mates (WT) were stimulated to contract, and glucose uptake was measured in the presence or absence of the antioxidant N-acetyl-l-cysteine (NAC) or the nitric oxide synthase (NOS) inhibitor NG-monomethyl-l-arginine (l-NMMA). Contraction increased AMPKα2 activity in WT but not AMPK-KD EDL muscles. However, contraction increased glucose uptake in the EDL and soleus muscles of AMPK-KD and WT mice to a similar extent. In EDL muscles, NAC and l-NMMA prevented contraction-stimulated increases in oxidant levels (dichloroflourescein fluorescence) and NOS activity, respectively, and attenuated contraction-stimulated glucose uptake in both genotypes to a similar extent. In soleus muscles of AMPK-KD and WT mice, NAC prevented contraction-stimulated glucose uptake and l-NMMA had no effect. This is likely attributed to the relative lack of neuronal NOS in the soleus muscles compared with EDL muscles. Contraction increased AMPKα Thr172 phosphorylation in EDL and soleus muscles of WT but not AMPK-KD mice, and this was not affected by NAC or l-NMMA treatment. In conclusion, ROS and NO are involved in regulating skeletal muscle glucose uptake during contraction via an AMPK-independent mechanism.

CaMKK is an upstream signal of AMP-activated protein kinase in regulation of substrate metabolism in contracting skeletal muscle

2009 ◽  
Vol 297 (6) ◽  
pp. R1724-R1732 ◽  
Author(s):  
Marcia J. Abbott ◽  
Arthur M. Edelman ◽  
Lorraine P. Turcotte
Keyword(s):  
Skeletal Muscle ◽  
Protein Kinase ◽  
Muscle Contraction ◽  
Glucose Uptake ◽  
Protein Content ◽  
Moderate Intensity ◽  
Dependent Manner ◽  
Substrate Metabolism ◽  
Caffeine Treatment ◽  
Amp Activated Protein Kinase

Multiple signals have been shown to be involved in regulation of fatty acid (FA) and glucose metabolism in contracting skeletal muscle. This study aimed to determine whether a Ca2+-stimulated kinase, CaMKK, is involved in regulation of contraction-induced substrate metabolism and whether it does so in an AMP-activated protein kinase (AMPK)-dependent manner. Rat hindlimbs were perfused at rest ( n = 16), with 3 mM caffeine ( n = 15), with 2 mM 5-aminoimidazole-4-carboxamide 1-β-d-ribofuranoside (AICAR; n = 16), or during moderate-intensity muscle contraction (MC; n = 14) and with or without 5 μM STO-609, a CaMKK inhibitor. FA uptake and oxidation increased ( P < 0.05) 64% and 71% by caffeine, 42% and 93% by AICAR, and 65% and 143% by MC. STO-609 abolished ( P < 0.05) caffeine- and MC-induced FA uptake and oxidation but had no effect with AICAR treatment. Glucose uptake increased ( P < 0.05) 104% by caffeine, 85% by AICAR, and 130% by MC, and STO-609 prevented the increase in glucose uptake in caffeine and muscle contraction groups. CaMKKβ activity increased ( P < 0.05) 113% by caffeine treatment and 145% by MC but was not affected by AICAR treatment. STO-609 prevented the caffeine- and MC-induced increase in CaMKKβ activity. Caffeine, AICAR, and MC increased ( P < 0.05) AMPKα2 activity by 295%, 11-fold, and 7-fold but did not affect AMPKα1 activity. STO-609 decreased ( P < 0.05) AMPKα2 activity induced by caffeine treatment and MC by 60% and 61% but did not affect AICAR-induced activity. Plasma membrane transport protein content of CD36 and glucose transporter 4 (GLUT4) increased ( P < 0.05) with caffeine, AICAR, and MC, and STO-609 prevented caffeine- and MC-induced increases in protein content. These results show the importance of Ca2+-dependent signaling via CaMKK activation in the regulation of substrate uptake and FA oxidation in contracting rat skeletal muscle and agree with the notion that CaMKK is an upstream kinase of AMPK in the regulation of substrate metabolism in skeletal muscle.

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