scholarly journals Evidence that insulin and guanosine triphosphate regulate dephosphorylation of the β-subunit of the insulin receptor in sarcolemma membranes isolated from skeletal muscle

1986 ◽  
Vol 234 (3) ◽  
pp. 527-535 ◽  
Author(s):  
R S Horn ◽  
E Lystad ◽  
A Adler ◽  
O Walaas
Keyword(s):  
Skeletal Muscle ◽  
Membrane Protein ◽  
Insulin Receptor ◽  
Beta Subunit ◽  
Regulatory Proteins ◽  
Maximal Effect ◽  
Dependent Manner ◽  
Insulin Effect ◽  
Phosphoamino Acid Analysis ◽  
Dose Dependent

When sarcolemma membranes isolated from rat skeletal muscle were incubated with [gamma-32P]ATP, a membrane protein of apparent Mr 95,000 was rapidly phosphorylated, with the 32P content reaching a maximum within 2 s. On the basis of immunoprecipitation with anti-insulin-receptor antiserum, phosphoamino acid analysis and Mr, this protein probably represents the beta-subunit of the insulin receptor. Similarly, on incubation of the membrane with adenosine 5′-[gamma-[35S]thio] triphosphate the 95 kDa protein was thiophosphorylated, indicating thiophosphorylation of the beta-subunit of the insulin receptor on the basis of immunoprecipitation studies. The effect of insulin on the phosphorylation of this protein in the membrane was studied. Insulin induced a 20% decrease in the 32P labelling of the protein when the membranes were phosphorylated for 10 s. This insulin effect was dose-dependent, with half-maximal effect obtained at 2-3 nM-insulin. Addition of GTP, but not GDP or guanosine 5′-[beta, gamma-imido]triphosphate, enhanced the effect to 35% inhibition, with half-maximal effect of GTP obtained at 0.5 microM. GTP had no effect on the phosphorylation of the protein in the absence of insulin. Analysis of this insulin effect showed that insulin increased the rate of dephosphorylation of the 95 kDa protein in the membrane. In contrast, insulin had no effect on thiophosphorylation of the 95 kDa membrane protein after incubation with adenosine 5′-[gamma-[35S]thio]triphosphate. Since thiophosphorylated proteins are less sensitive to phosphatase action, these investigations suggest that insulin stimulated a protein phosphatase activity in a GTP-dependent manner. The possibility that GTP-regulatory proteins are involved in the action of insulin on the phosphorylation of the insulin receptor and other membrane proteins is discussed.

scholarly journals Effect of lemon peel flavonoids on anti-fatigue and anti-oxidation capacities of exhaustive exercise mice

2020 ◽  
Vol 63 (1) ◽  
Author(s):  
Guili Bao ◽  
Yinglong Zhang ◽  
Xiaoguang Yang
Keyword(s):  
Skeletal Muscle ◽  
Superoxide Dismutase ◽  
Manganese Superoxide Dismutase ◽  
Fatty Acid Content ◽  
Hepatic Tissue ◽  
Control Group ◽  
Dependent Manner ◽  
Lemon Peel ◽  
Tnf Α ◽  
Dose Dependent

AbstractIn this study, lemon peel flavonoids (LPF) were administered to investigate its effect on the anti-fatigue and antioxidant capacity of mice that undergo exercise until exhaustion. LPF (88.36 min in LPFH group mice) significantly increased the exhaustion swimming time compare to the untreated mice (40.36 min), increased the liver glycogen and free fatty acid content in mice and reduce lactic acid and BUN content in a dose-dependent manner. As the concentration of lemon peel flavonoids increased, the serum creatine kinase, aspartate aminotransferase, and alanine aminotransferase levels of mice gradually decreased. LPF increases superoxide dismutase (SOD) and catalase (CAT) levels in mice and reduces malondialdehyde levels in a dose-dependent manner. And LPF raises hepatic tissue SOD, CAT activities and reduces skeletal muscle tissue iNOS, TNF-α levels of mice compared to the control group. LPF also enhanced the expression of copper/zinc-superoxide dismutase (Cu/Zn-SOD), manganese-superoxide dismutase (Mn-SOD), and CAT mRNA in mouse liver tissue. LPF also enhanced the expression of alanine/serine/cysteine/threonine transporter 1 (ASCT1) mRNA and attenuate the expression of syncytin-1, inducible nitric oxide synthase (iNOS), and tumor necrosis factor (TNF)-α in mouse skeletal muscle. According to high-performance liquid chromatography (HPLC) analysis, it was found that LPF contains flavonoids such as rutin, astragalin, isomangiferin, naringin, and quercetin. Our experimental data show that LPF has good anti-fatigue effects and anti-oxidation ability. In summary, LPF has high prospects to be developed and added to nutritional supplements.

scholarly journals Ginsenoside Rb1 stimulates glucose uptake through insulin-like signaling pathway in 3T3-L1 adipocytes

2008 ◽  
Vol 198 (3) ◽  
pp. 561-569 ◽  
Author(s):  
Wenbin Shang ◽  
Ying Yang ◽  
Libin Zhou ◽  
Boren Jiang ◽  
Hua Jin ◽  
...  
Keyword(s):  
Insulin Receptor ◽  
Glucose Uptake ◽  
Signaling Pathway ◽  
Glucose Transport ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
C2c12 Myotubes ◽  
Maximal Effect ◽  
Dependent Manner ◽  
Active Constituent

A series of clinical trials and animal experiments have demonstrated that ginseng and its major active constituent, ginsenosides, possess glucose-lowering action. In our previous study, ginsenoside Rb1 has been shown to regulate peroxisome proliferator-activated receptor γ activity to facilitate adipogenesis of 3T3-L1 cells. However, the effect of Rb1 on glucose transport in insulin-sensitive cells and its molecular mechanism need further elucidation. In this study, Rb1 significantly stimulated basal and insulin-mediated glucose uptake in a time- and dose-dependent manner in 3T3-L1 adipocytes and C2C12 myotubes; the maximal effect was achieved at a concentration of 1 μM and a time of 3 h. In adipocytes, Rb1 promoted GLUT1 and GLUT4 translocations to the cell surface, which was examined by analyzing their distribution in subcellular membrane fractions, and enhanced translocation of GLUT4 was confirmed using the transfection of GLUT4-green fluorescence protein in Chinese Hamster Ovary cells. Meanwhile, Rb1 increased the phosphorylation of insulin receptor substrate-1 and protein kinase B (PKB), and stimulated phosphatidylinositol 3-kinase (PI3K) activity in the absence of the activation of the insulin receptor. Rb1-induced glucose uptake as well as GLUT1 and GLUT4 translocations was inhibited by the PI3K inhibitor. These results suggest that ginsenoside Rb1 stimulates glucose transport in insulin-sensitive cells by promoting translocations of GLUT1 and GLUT4 by partially activating the insulin signaling pathway. These findings are useful in understanding the hypoglycemic and anti-diabetic properties of ginseng and ginsenosides.

AMP-activated protein kinase activity and glucose uptake in rat skeletal muscle

2001 ◽  
Vol 280 (5) ◽  
pp. E677-E684 ◽  
Author(s):  
Nicolas Musi ◽  
Tatsuya Hayashi ◽  
Nobuharu Fujii ◽  
Michael F. Hirshman ◽  
Lee A. Witters ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Protein Kinase ◽  
Glucose Uptake ◽  
Muscle Contractions ◽  
Treadmill Running ◽  
Dependent Manner ◽  
Rat Skeletal Muscle ◽  
Amp Activated Protein Kinase ◽  
Dose Dependent

The AMP-activated protein kinase (AMPK) has been hypothesized to mediate contraction and 5-aminoimidazole-4-carboxamide 1-β-d-ribonucleoside (AICAR)-induced increases in glucose uptake in skeletal muscle. The purpose of the current study was to determine whether treadmill exercise and isolated muscle contractions in rat skeletal muscle increase the activity of the AMPKα1 and AMPKα2 catalytic subunits in a dose-dependent manner and to evaluate the effects of the putative AMPK inhibitors adenine 9-β-d-arabinofuranoside (ara-A), 8-bromo-AMP, and iodotubercidin on AMPK activity and 3- O-methyl-d-glucose (3-MG) uptake. There were dose-dependent increases in AMPKα2 activity and 3-MG uptake in rat epitrochlearis muscles with treadmill running exercise but no effect of exercise on AMPKα1 activity. Tetanic contractions of isolated epitrochlearis muscles in vitro significantly increased the activity of both AMPK isoforms in a dose-dependent manner and at a similar rate compared with increases in 3-MG uptake. In isolated muscles, the putative AMPK inhibitors ara-A, 8-bromo-AMP, and iodotubercidin fully inhibited AICAR-stimulated AMPKα2 activity and 3-MG uptake but had little effect on AMPKα1 activity. In contrast, these compounds had absent or minimal effects on contraction-stimulated AMPKα1 and -α2 activity and 3-MG uptake. Although the AMPKα1 and -α2 isoforms are activated during tetanic muscle contractions in vitro, in fast-glycolytic fibers, the activation of AMPKα2-containing complexes may be more important in regulating exercise-mediated skeletal muscle metabolism in vivo. Development of new compounds will be required to study contraction regulation of AMPK by pharmacological inhibition.

A new polymorphism in the type II deiodinase gene is associated with circulating thyroid hormone parameters

2005 ◽  
Vol 289 (1) ◽  
pp. E75-E81 ◽  
Author(s):  
Robin P. Peeters ◽  
Annewieke W. van den Beld ◽  
Hayat Attalki ◽  
Hans van Toor ◽  
Yolanda B. de Rijke ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Thyroid Hormone ◽  
Blood Donors ◽  
Muscle Size ◽  
Dependent Manner ◽  
Type Ii ◽  
Elderly Men ◽  
Similar Frequency ◽  
Dose Dependent ◽  
Tsh Levels

Type II deiodinase (D2) is important in the regulation of local thyroid hormone bioactivity in certain tissues. D2 in skeletal muscle may also play a role in serum triiodothyronine (T3) production. In this study, we identified a polymorphism in the 5′-UTR of the D2 gene (D2-ORFa-Gly3Asp). We investigated the association of D2-ORFa-Gly3Asp, and of the previously identified D2-Thr92Ala polymorphism, with serum iodothyronine levels. D2-ORFa-Gly3Asp was identified by sequencing the 5′-UTR of 15 randomly selected individuals. Genotypes for D2-ORFa-Gly3Asp were determined in 156 healthy blood donors (age 46.3 ± 12.2 yr) and 349 ambulant elderly men (age 77.7 ± 3.5 yr) and related to serum iodothyronine and TSH levels. D2-ORFa-Asp3had an allele frequency of 33.9% in blood bank donors and was associated with serum thyroxine (T4; Gly/Gly vs. Gly/Asp vs. Asp/Asp = 7.06 ± 0.14 vs. 6.74 ± 0.15 vs. 6.29 ± 0.27 μg/dl, P = 0.01), free T4(1.22 ± 0.02 vs. 1.16 ± 0.02 vs. 1.06 ± 0.04 ng/dl, P = 0.001), reverse T3( P = 0.01), and T3/T4ratio ( P = 0.002) in a dose-dependent manner, but not with serum T3( P = 0.59). In elderly men, D2-ORFa-Asp3had a similar frequency but was not associated with serum iodothyronine levels. This new polymorphism in the 5′-UTR of D2 is associated with iodothyronine levels in blood donors but not in elderly men. We hypothesize that this might be explained by the decline in skeletal muscle size during aging, resulting in a relative decrease in the contribution of D2 to serum T3production.

L(+)-Lactate Binding to a Protein in Rat Skeletal Muscle Plasma Membranes

1995 ◽  
Vol 20 (1) ◽  
pp. 112-124 ◽  
Author(s):  
Karl J. A. McCullagh ◽  
Arend Bonen
Keyword(s):  
Skeletal Muscle ◽  
Plasma Membranes ◽  
Mass Range ◽  
Dependent Manner ◽  
Rat Skeletal Muscle ◽  
Lactate Transport ◽  
Sds Page ◽  
Lactate And Pyruvate ◽  
Potential Inhibitors ◽  
Dose Dependent

Biochemical studies were conducted to determine the location of a putative lactate transport protein in rat skeletal muscle plasma membranes (PM). PM (50-100 μg protein) were incubated with [U-14C] L(+)-lactate, in the presence or absence of unlabeled monocarboxylates or potential inhibitors, after which proteins were separated by SDS-PAGE. Gel slices (2 mm) were cut and analyzed for14C. [U-14C] L(+)-lactate was bound to plasma membranes in the 30 to 40 kDa molecular mass range. Binding of [U-14C] L(+)-lactate was inhibited by N-ethylmaleimide, unlabeled L-lactate and pyruvate, and in a dose dependent manner by α-cyano-4-hydroxycinnamate (r = 0.995), but not by cytochalasin-B. The inhibition of [U-14C] L(+)-lactate binding was similar to the inhibition of lactate transport. Therefore the transport of L(+)-lactate across skeletal muscle plasma membranes involves a polypeptide of 30 to 40 kDa. Key words: transport, affinity labeling

scholarly journals Calcium-dependence of insulin receptor phosphorylation

1983 ◽  
Vol 214 (2) ◽  
pp. 361-366 ◽  
Author(s):  
W E Plehwe ◽  
P F Williams ◽  
I D Caterson ◽  
L C Harrison ◽  
J R Turtle
Keyword(s):  
Membrane Protein ◽  
Insulin Receptor ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Beta Subunit ◽  
Receptor Phosphorylation ◽  
32P Incorporation ◽  
Isolated Rat

Phosphorylation of the insulin receptor of isolated rat adipocytes in response to insulin has been studied. Immunoprecipitation of adipocyte membrane protein demonstrated increased incorporation of 32P after exposure to insulin for 15 min, but this was dependent on the presence of physiological concentrations of Ca2+ and Mg2+. Autoradiography of solubilized immunoprecipitated membrane protein after sodium dodecyl sulphate/polyacrylamide-gel electrophoresis revealed that most of the 32P incorporation occurred in a band corresponding to Mr 95 000, which has been identified previously as the beta-subunit of the insulin receptor. 32P incorporation was inhibited by 2,4-dinitrophenol and trifluoperazine. It is suggested that insulin-receptor phosphorylation is an energy-requiring process that is Ca2+-dependent and may be modulated by calmodulin. Phosphorylation may proceed independently of glucose transport.

Di(2-ethylhexyl)phthalate exposure impairs insulin receptor and glucose transporter 4 gene expression in L6 myotubes

2013 ◽  
Vol 33 (7) ◽  
pp. 685-700 ◽  
Author(s):  
P Rajesh ◽  
K Balasubramanian
Keyword(s):  
Gene Expression ◽  
Insulin Receptor ◽  
Glucose Uptake ◽  
Insulin Signaling ◽  
Glucose Transporter ◽  
Negative Influence ◽  
Receptor Protein ◽  
Dependent Manner ◽  
L6 Myotubes ◽  
Dose Dependent

Di(2-ethyl hexyl)-phthalate (DEHP) is an endocrine disrupter and is the most abundantly used phthalate derivative, which is suspected to be an inevitable environmental exposure contributing to the increasing incidence of type-2 diabetes in humans. Therefore, the present study was designed to address the dose-dependent effects of DEHP on insulin signaling molecules in L6 myotubes. L6 myotubes were exposed to different concentrations (25, 50, and 100 μM) of DEHP for 24 h. At the end of exposure, cells were utilized for assessing various parameters. Insulin receptor and glucose transporter4 (GLUT4) gene expression, insulin receptor protein concentration, glucose uptake and oxidation, and enzymatic and nonenzymatic antioxidants were significantly reduced, but glutamine fructose-6-phosphate amidotransferase, nitric oxide, lipid peroxidation, and reactive oxygen species levels were elevated in a dose-dependent manner in L6 myotubes exposed to DEHP. The present study in turn shows the direct adverse effect of DEHP on the expression of insulin receptor and GLUT4 gene, glucose uptake, and oxidation in L6 myotubes suggesting that DEHP exposure may have a negative influence on insulin signaling.

scholarly journals Modulation of electrical activity and ionic currents of isolated neurons by orexin A

2013 ◽  
Vol 11 (4) ◽  
pp. 54-60
Author(s):  
Petr Dmitriyevich Shabanov ◽  
Anatoliy Ivanovich Vislobokov
Keyword(s):  
Membrane Potential ◽  
Lymnaea Stagnalis ◽  
Ionic Currents ◽  
Maximal Effect ◽  
Dependent Manner ◽  
Isolated Neurons ◽  
Orexin A ◽  
High Concentrations ◽  
Kinetics Of ◽  
Dose Dependent

The changes in intracellular potential of resting (PR) and potential of action (PA) of the identified neurons of pedal and visceral ganglia of the CNS mollusk Planorbarius corneus registered by means of intracellular electrodes, and ionic currents of isolated neurons under fixed potential after administration of orexin A in concentrations 1, 10, 100 and 1000 µg/ml were studied by the method of fixation of membrane potential in isolated neurons of the Lymnaea stagnalis mollusk. Dibazol in concentrations of 1 and 10 µM effected slightly on the ionic currents. High concentrations of dibazol (100 and 1000 µM) inhibited all currents in dose dependent manner with maximal effect on potassium currents amplitude. ЕС50 were 7.4 мМ for INa, 4.0 мМ for ICa, 83.9 µM for IKs,1 (one group of neurons) and 2.9 мМ for IKs,2 (the another group of neurons). The voltage-amper membrane characteristics shift was not registered, but the kinetics of currents development was changed. Dibazol was more effective in inhibition of ionic currents compared to its structural analogs.

scholarly journals Bradykinin potentiates insulin-stimulated glucose uptake and enhances insulin signal through the bradykinin B2 receptor in dog skeletal muscle and rat L6 myoblasts

1998 ◽  
pp. 344-352 ◽  
Author(s):  
T Miyata ◽  
T Taguchi ◽  
M Uehara ◽  
S Isami ◽  
H Kishikawa ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Insulin Receptor ◽  
Glucose Uptake ◽  
Dependent Manner ◽  
Glut4 Translocation ◽  
Binding Studies ◽  
Insulin Receptor Tyrosine Kinase ◽  
Peripheral Tissues ◽  
Bradykinin B2 Receptor ◽  
Increase Glucose Uptake

Previously we demonstrated that bradykinin infusion could increase glucose uptake into dog peripheral tissues, and that bradykinin could potentiate insulin-induced glucose uptake through glucose transporter 4 (GLUT4) translocation in dog adipocytes. However, skeletal muscle is the predominant tissue for insulin-mediated glucose disposal. The aim of this study was to determine how bradykinin affected insulin-stimulated glucose uptake in dog skeletal muscle and myotubes transformed from rat L6 myoblasts. The bradykinin receptor binding studies revealed that dog skeletal muscle and rat L6 myoblasts possessed significant numbers of bradykinin receptors (Kd = 88 and 76 pmol/l, Bmax = 82.5 and 20 fmol/mg protein respectively). An RT-PCR (reverse transcriptase-polymerase chain reaction) amplification showed mRNA specific for bradykinin B2 receptor in both cells. Bradykinin significantly increased 2-deoxyglucose uptake in isolated muscle and L6 myoblasts in the presence of insulin (10(-7) mol/l) in a dose-dependent manner, but not in the absence of insulin. Bradykinin also enhanced insulin-stimulated GLUT4 translocation, and insulin-induced phosphorylation of insulin receptor beta subunit and insulin receptor substrate-1 (IRS-1) without affecting the binding affinities or numbers of cell surface insulin receptors in both cells. It is concluded that bradykinin could potentiate the insulin-induced glucose uptake through GLUT4 translocation in dog skeletal muscle and rat L6 myoblasts. This effect could be explained by the potency of bradykinin to upregulate the insulin receptor tyrosine kinase activity which stimulates phosphorylation of IRS-1, followed by an increase in GLUT4 translocation.

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