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

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.

scholarly journals Partial purification and reconstitution of the sarcolemmal l-lactate carrier from rat skeletal muscle

1994 ◽  
Vol 303 (1) ◽  
pp. 207-212 ◽  
Author(s):  
P J Allen ◽  
G A Brooks
Keyword(s):  
Skeletal Muscle ◽  
Sodium Phosphate ◽  
Partial Purification ◽  
Transport Activity ◽  
Rat Skeletal Muscle ◽  
Lactate Transport ◽  
Hindlimb Muscle ◽  
Sds Page ◽  
Transport Analysis ◽  
Lactate Exchange

Purified sarcolemmal membranes from mixed rat hindlimb muscle were solubilized with octylglucoside and the extract subjected to hydroxylapatite (HA) chromatography. Following protein elution with a sodium phosphate gradient and detergent removal by dialysis, the HA eluate was reconstituted into asolectin liposomes using a freeze-thaw procedure. Specific L-[14C]lactate transport activity eluting from the 0.2 M sodium phosphate fraction was 30-fold higher compared with native sarcolemmal vesicles (31.64 versus 1.06 nmol/min per mg). The reconstituted carrier exhibited Michaelis-Menten saturation kinetics with Km and Vmax. values of 46.2 +/- 6.6 mM and 498.7 +/- 17.2 nmol/15 s per mg respectively. L-Lactate transport activity was inhibited 57% by preincubation of proteoliposomes with 10 mM alpha-cyano-4-hydroxycinnamate, a known inhibitor of lactate transport. Analysis of the HA eluates by SDS/PAGE showed the presence of a 34 kDa band corresponding to lactate transport activity. Reconstitution of lactate transport activity eluting from the HA column, together with SDS/PAGE analysis suggests the presence of a 34 kDa polypeptide mediating sarcolemmal lactate exchange in rat skeletal muscle.

scholarly journals Reconstitution of the lactate carrier from rat skeletal-muscle sarcolemma

1994 ◽  
Vol 299 (2) ◽  
pp. 533-537 ◽  
Author(s):  
F Wibrand ◽  
C Juel
Keyword(s):  
Skeletal Muscle ◽  
Time Course ◽  
Rat Skeletal Muscle ◽  
Lactate Transport ◽  
Exchange Inhibitor ◽  
Transport Inhibitors ◽  
Lactate And Pyruvate ◽  
Lactate Uptake ◽  
Fold Excess ◽  
Sarcolemmal Vesicles

The lactate carrier was solubilized from purified rat skeletal-muscle sarcolemma with the detergent decanoyl-N-methyl-glucamide and the solubilized carrier was reconstituted into phospholipid vesicles. Reconstituted proteoliposomes showed a faster time course of L-lactate uptake than did protein-free liposomes. The rate of L-lactate uptake into the proteoliposomes was inhibited by the lactate-transport inhibitors p-chloromercuribenzenesulphonate, diethyl pyrocarbonate, alpha-cyano-4-hydroxycinnamate and quercetin. In contrast, the anion-exchange inhibitor DIDS (4,4′-di-isothiocyanostilbene-2,2′-disulphonate) had almost no effect on the uptake. The extent of L-lactate uptake at equilibrium was not affected by the presence of the transport inhibitors, but was sensitive to osmotic strength. L-Lactate and pyruvate, but not D-lactate, inhibited L-lactate uptake when present at 10-fold excess. The properties of L-lactate transport in reconstituted proteoliposomes were similar to those observed in native sarcolemmal vesicles, i.e. the lactate carrier seems to retain its transport characteristics during the solubilization and reconstitution steps.

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 Chlorotetracycline as a fluorescent Ca2+ probe in pancreatic islet cells.

1978 ◽  
Vol 76 (3) ◽  
pp. 652-674 ◽  
Author(s):  
I B Täljedal
Keyword(s):  
Plasma Membranes ◽  
Islet Cells ◽  
Dependent Manner ◽  
Cell Surfaces ◽  
Noticeable Effect ◽  
Cell Plasma ◽  
Glass Capillaries ◽  
Asymmetrically Distributed ◽  
Dose Dependent ◽  
Dispersed Cells

Pancreatic islets, or suspensions of islet cells, from noninbred ob/ob-mice were incubated with chlorotetracycline and analyzed for Ca2+-dependent fluorescence in a microscope. Unless logarithmically transformed, signals from islets were asymmetrically distributed with unstable variance. Signals from cells pelleted in glass capillaries were more homogeneous and depended linearly on the thickness of the sample. The effect of sample thickness and a significant enhancement of fluorescence by alloxan suggest that beta-cells were involved in producing the signal from whole islets. The signal from dispersed cells was probably diagnostic of Ca2+ in beta-cell plasma membranes because it was suppressed by La3+ and had a spectrum indicative of an apolar micromilieu; fluorescent staining of cell surfaces was directly seen at high magnification. Fluorescence from cells was enhanced by 0.5-10 mM Ca2+ in a dose-dependent manner, whereas less than 0.5 mM Ca2+ saturated the probe alone in methanol. The signal from islets or dispersed cells was suppressed by 5 mM theophylline; that from cells was also suppressed by 0.5 mM 3-isobutyl-1-methylxanthine, 1.2 or 15 mM Mg2+, 3-20 mM D-glucose, and, to a lesser extent, 20 mM 3-O-methyl-D-glucose. D-glucose was more inhibitory in the absence than in the presence of Mg2+, as if Mg2+ and D-glucose influenced the same Ca2+ pool. L-glucose, D-mannopheptulose, or diazoxide had no noticeable effect and 20 mM bicarbonate was stimulatory. The results suggest that microscopy of chlorotetracycline-stained cells can aid in characterizing calcium pools of importance for secretion. Initiation of insulin release may be associated with an increas

Insulin-degrading Activity of Plasma Membranes from Rat Skeletal Muscle: Its Isolation, Characterization, and Biologic Significance

Diabetes ◽  
1979 ◽  
Vol 28 (9) ◽  
pp. 810-817 ◽  
Author(s):  
K. Yokono ◽  
Y. Imamura ◽  
H. Sakai ◽  
S. Baba
Keyword(s):  
Skeletal Muscle ◽  
Plasma Membranes ◽  
Rat Skeletal Muscle

scholarly journals Antioxidant, Antibacterial, and Cytoprotective Activity of Agathi Leaf Protein

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
A. S. Zarena ◽  
Shubha Gopal ◽  
R. Vineeth
Keyword(s):  
Safety Assessment ◽  
Crude Protein ◽  
Leaf Protein ◽  
Dependent Manner ◽  
Cytoprotective Activity ◽  
Sesbania Grandiflora ◽  
Rp Hplc ◽  
Maximum Inhibition ◽  
Sds Page ◽  
Dose Dependent

In the present study a protein termed agathi leaf protein (ALP) fromSesbania grandiflora Linn. (agathi) leaves was isolated after successive precipitation with 65% ammonium sulphate followed by purification on Sephadex G 75. The column chromatography of the crude protein resulted in four peaks of which Peak I (P I) showed maximum inhibition activity against hydroxyl radical. SDS-PAGE analysis of P I indicated that the molecular weight of the protein is≈29 kDa. The purity of the protein was 98.4% as determined by RP-HPLC and showed a single peak with a retention time of 19.9 min. ALP was able to reduce oxidative damage by scavenging lipid peroxidation against erythrocyte ghost (85.50 ± 6.25%), linolenic acid (87.67 ± 3.14%) at 4.33 μM, ABTS anion (88 ± 3.22%), and DNA damage (83 ± 4.20%) at 3.44 μM in a dose-dependent manner. The purified protein offered significant protection to lymphocyte (72% at 30 min) induced damage by t-BOOH. In addition, ALP showed strong antibacterial activity againstPseudomonas aeruginosa(20 ± 3.64 mm) andStaphylococcus aureus(19 ± 1.53 mm) at 200 μg/mL. The safety assessment showed that ALP does not induce cytotoxicity towards human lymphocyte at the tested concentration of 0.8 mg/mL.

Extracellular ATP induces a nonspecific permeability of thymocyte plasma membranes

1989 ◽  
Vol 67 (9) ◽  
pp. 495-502 ◽  
Author(s):  
Chakib El-Moatassim ◽  
Nicole Bernad ◽  
Jean-Claude Mani ◽  
Jacques Dornand
Keyword(s):  
Purinergic Receptors ◽  
Plasma Membranes ◽  
Divalent Cations ◽  
Extracellular Atp ◽  
Active Species ◽  
Extracellular Nucleotides ◽  
Dependent Manner ◽  
Specific Receptors ◽  
Atp Analogs ◽  
Dose Dependent

We have previously demonstrated that extracellular ATP can give medullary thymocytes the calcium message required for the induction of their blastogenesis, without mobilization of intracellular calcium. We describe here the effects of extracellular nucleotides on membrane permeability to monovalent and divalent cations in mouse thymocytes. Among all nucleotides tested, under physiological conditions, only ATP and, to a lesser extent, 2-methylthio-ATP, adenosine 5′-O-(3-thio-triphosphate), and ADP were able to depolarize thymocyte plasma membranes and to induce Na+ and Ca2+ influxes into thymocytes; other nonhydrolysable ATP analogs were only effective in the absence of Mg2+. The ATP-induced effects were inhibited in a dose-dependent manner by Mg2+ and greatly potentiated in its absence, which suggests that the tetrabasic ATP4− is probably the active species and that a phosphotransferase activity is not involved in its effects. These ATP-mediated changes in ion fluxes result from an increase in nonspecific permeability of thymocyte membranes, probably by pore formation. These ion flux changes might be responsible for the mitogenic induction of phorbol 12-myristate 13-acetate treated medullary thymocytes. The potency order for the adenine derivatives to affect these fluxes (ATP>ADP> >AMP>adenosine) suggests the presence of ATP specific receptors (P2 purinergic receptors) on thymocyte plasma membranes.Key words: purinergic receptors, extracellular ATP, membrane potential, cation fluxes, thymocytes.

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.

Sign in / Sign up

Export Citation Format

Share Document