Regulation of mitochondrial respiration by inorganic phosphate; comparing permeabilized muscle fibers and isolated mitochondria prepared from type-1 and type-2 rat skeletal muscle

2008 ◽  
Vol 105 (2) ◽  
pp. 279-287 ◽  
Author(s):  
Morten Scheibye-Knudsen ◽  
Bjørn Quistorff
Keyword(s):  
Skeletal Muscle ◽  
Mitochondrial Respiration ◽  
Inorganic Phosphate ◽  
Muscle Fibers ◽  
Rat Skeletal Muscle ◽  
Isolated Mitochondria ◽  
Permeabilized Muscle

scholarly journals A comparative study of mitochondrial respiration in circulating blood cells and skeletal muscle fibers in women

2019 ◽  
Vol 317 (3) ◽  
pp. E503-E512 ◽  
Author(s):  
Shannon Rose ◽  
Eugenia Carvalho ◽  
Eva C. Diaz ◽  
Matthew Cotter ◽  
Sirish C. Bennuri ◽  
...  
Keyword(s):  
Insulin Resistance ◽  
Type 2 Diabetes ◽  
Skeletal Muscle ◽  
Mitochondrial Function ◽  
Mitochondrial Respiration ◽  
Blood Cells ◽  
Muscle Fibers ◽  
Circulating Cells ◽  
Permeabilized Muscle

Skeletal muscle mitochondrial respiration is thought to be altered in obesity, insulin resistance, and type 2 diabetes; however, the invasive nature of tissue biopsies is an important limiting factor for studying mitochondrial function. Recent findings suggest that bioenergetics profiling of circulating cells may inform on mitochondrial function in other tissues in lieu of biopsies. Thus, we sought to determine whether mitochondrial respiration in circulating cells [peripheral blood mononuclear cells (PBMCs) and platelets] reflects that of skeletal muscle fibers derived from the same subjects. PBMCs, platelets, and skeletal muscle (vastus lateralis) samples were obtained from 32 young (25–35 yr) women of varying body mass indexes. With the use of extracellular flux analysis and high-resolution respirometry, mitochondrial respiration was measured in intact blood cells as well as in permeabilized cells and permeabilized muscle fibers. Respiratory parameters were not correlated between permeabilized muscle fibers and intact PBMCs or platelets. In a subset of samples ( n = 12–13) with permeabilized blood cells available, raw measures of substrate (pyruvate, malate, glutamate, and succinate)-driven respiration did not correlate between permeabilized muscle (per mg tissue) and permeabilized PBMCs (per 106 cells); however, complex I leak and oxidative phosphorylation coupling efficiency correlated between permeabilized platelets and muscle (Spearman’s ρ = 0.64, P = 0.030; Spearman’s ρ = 0.72, P = 0.010, respectively). Our data indicate that bioenergetics phenotypes in circulating cells cannot recapitulate muscle mitochondrial function. Select circulating cell bioenergetics phenotypes may possibly inform on overall metabolic health, but this postulate awaits validation in cohorts spanning a larger range of insulin resistance and type 2 diabetes status.

Hindlimb Muscle Dissection, Permeabilization, and Respirometry v1

2021 ◽  
Author(s):  
Matthew D. Campbell ◽  
David J. Marcinek
Keyword(s):  
Skeletal Muscle ◽  
Muscle Biopsy ◽  
Mitochondrial Respiration ◽  
Gastrocnemius Muscle ◽  
Muscle Fibers ◽  
Mouse Muscle ◽  
Skeletal Muscle Biopsy ◽  
Cellular Environment ◽  
Permeabilized Muscle

The use of permeabilized muscle fibers (PMF) has emerged as a gold standard for assessing skeletal muscle mitochondrial function. PMF provide an intermediate approach between in vivo strategies and isolated mitochondria that allows the mitochondria to be maintained in close to their native morphology in the myofiber while allowing greater control of substrate and inhibitor concentrations. However, like mitochondrial isolation, the primary drawback to PMF is disruption of the cellular environment during the muscle biopsy and preparation. Despite all the benefits of permeabilized muscle fibers in evaluating mitochondrial respiration and dynamics one of the major drawbacks is increased variability introduced during a muscle biopsy as well as intrinsic variation that exists due to sex and age. This study was designed to evaluate how age, sex, and biopsy preparations affect mitochondrial respiration in extensor digitorum longus, soleus, and gastrocnemius muscle of mice. Here we detail a modified approach to skeletal muscle biopsy of the gastrocnemius muscle of mice focused on maintenance of intact fibers that results in greater overall respiration compared to cut fibers. The improved respiration of intact fibers is sex specific as are some of the changes in mitochondrial respiration with age. This study shows the need for standard practices when measuring mitochondrial respiration in permeabilized muscle and provides a protocol to control for variation introduced during a typical mouse muscle biopsy.

Mechanisms of reduced SR Ca2+ release induced by inorganic phosphate in rat skeletal muscle fibers

2001 ◽  
Vol 281 (2) ◽  
pp. C418-C429 ◽  
Author(s):  
Adrian M. Duke ◽  
Derek S. Steele
Keyword(s):  
Skeletal Muscle ◽  
Inorganic Phosphate ◽  
Inhibitory Action ◽  
Muscle Fibers ◽  
Inhibitory Effect ◽  
Rat Skeletal Muscle ◽  
Skeletal Muscle Fatigue ◽  
Skeletal Muscle Fibers ◽  
Atp Breakdown ◽  
Marked Dependence

The effects of inorganic phosphate (Pi) on Ca2+ release from the sarcoplasmic reticulum (SR) were studied in mechanically skinned rat skeletal muscle fibers. Application of caffeine or T-tubule depolarization was used to induce Ca2+ release from the SR, which was detected using fura 2 fluorescence. Addition of Pi (1–40 mM) caused a reversible and concentration-dependent reduction in the caffeine-induced Ca2+ transient. This effect was apparent at low Pi concentration (<5 mM), which did not result in detectable precipitation of calcium phosphate within the SR. The inhibitory effect of Pi exhibited a marked dependence on free Mg2+ concentration. At 0.5 mM free Mg2+, 5 mM Pi reduced the caffeine-induced transient by 25.1 ± 4.1% ( n = 13). However, at 1.5 mM free Mg2+, 5 mM Pi reduced the amplitude of caffeine-induced Ca2+ transients by 68.9 ± 3.1% ( n = 10). Depolarization-induced SR Ca2+release was similarly affected. These effects of Pi may be important in skeletal muscle fatigue, if an inhibitory action of Pi on SR Ca2+ release is augmented by the rise in cytosolic Mg2+ concentration, which accompanies ATP breakdown.

scholarly journals Human and Rodent Skeletal Muscles Express Angiotensin II Type 1 Receptors

Cells ◽  
2020 ◽  
Vol 9 (7) ◽  
pp. 1688
Author(s):  
Rafael Deminice ◽  
Hayden Hyatt ◽  
Toshinori Yoshihara ◽  
Mustafa Ozdemir ◽  
Branden Nguyen ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Angiotensin Ii ◽  
Muscle Atrophy ◽  
Muscle Fibers ◽  
Skeletal Muscle Atrophy ◽  
Type I ◽  
Single Muscle ◽  
Rat Skeletal Muscle ◽  
Skeletal Muscle Fibers

Abundant evidence reveals that activation of the renin-angiotensin system promotes skeletal muscle atrophy in several conditions including congestive heart failure, chronic kidney disease, and prolonged mechanical ventilation. However, controversy exists about whether circulating angiotensin II (AngII) promotes skeletal muscle atrophy by direct or indirect effects; the centerpiece of this debate is the issue of whether skeletal muscle fibers express AngII type 1 receptors (AT1Rs). While some investigators assert that skeletal muscle expresses AT1Rs, others argue that skeletal muscle fibers do not contain AT1Rs. These discordant findings in the literature are likely the result of study design flaws and additional research using a rigorous experimental approach is required to resolve this issue. We tested the hypothesis that AT1Rs are expressed in both human and rat skeletal muscle fibers. Our premise was tested using a rigorous, multi-technique experimental design. First, we established both the location and abundance of AT1Rs on human and rat skeletal muscle fibers by means of an AngII ligand-binding assay. Second, using a new and highly selective AT1R antibody, we carried out Western blotting and determined the abundance of AT1R protein within isolated single muscle fibers from humans and rats. Finally, we confirmed the presence of AT1R mRNA in isolated single muscle fibers from rats. Our results support the hypothesis that AT1Rs are present in both human and rat skeletal muscle fibers. Moreover, our experiments provide the first evidence that AT1Rs are more abundant in fast, type II muscle fibers as compared with slow, type I fibers. Together, these discoveries provide the foundation for an improved understanding of the mechanism(s) responsible for AngII-induced skeletal muscle atrophy.

Does impaired mitochondrial function affect insulin signaling and action in cultured human skeletal muscle cells?

2008 ◽  
Vol 294 (1) ◽  
pp. E97-E102 ◽  
Author(s):  
Audrey E. Brown ◽  
Matthias Elstner ◽  
Stephen J. Yeaman ◽  
Douglass M. Turnbull ◽  
Mark Walker
Keyword(s):  
Skeletal Muscle ◽  
Glucose Uptake ◽  
Insulin Signaling ◽  
Mitochondrial Respiration ◽  
Respiratory Function ◽  
Insulin Action ◽  
Human Skeletal Muscle ◽  
Human Skeletal Muscle Cells ◽  
Basal Glucose Uptake

Insulin-resistant type 2 diabetic patients have been reported to have impaired skeletal muscle mitochondrial respiratory function. A key question is whether decreased mitochondrial respiration contributes directly to the decreased insulin action. To address this, a model of impaired cellular respiratory function was established by incubating human skeletal muscle cell cultures with the mitochondrial inhibitor sodium azide and examining the effects on insulin action. Incubation of human skeletal muscle cells with 50 and 75 μM azide resulted in 48 ± 3% and 56 ± 1% decreases, respectively, in respiration compared with untreated cells mimicking the level of impairment seen in type 2 diabetes. Under conditions of decreased respiratory chain function, insulin-independent (basal) glucose uptake was significantly increased. Basal glucose uptake was 325 ± 39 pmol/min/mg (mean ± SE) in untreated cells. This increased to 669 ± 69 and 823 ± 83 pmol/min/mg in cells treated with 50 and 75 μM azide, respectively (vs. untreated, both P < 0.0001). Azide treatment was also accompanied by an increase in basal glycogen synthesis and phosphorylation of AMP-activated protein kinase. However, there was no decrease in glucose uptake following insulin exposure, and insulin-stimulated phosphorylation of Akt was normal under these conditions. GLUT1 mRNA expression remained unchanged, whereas GLUT4 mRNA expression increased following azide treatment. In conclusion, under conditions of impaired mitochondrial respiration there was no evidence of impaired insulin signaling or glucose uptake following insulin exposure in this model system.

Glycogen Depletion in Rat Skeletal Muscle Fibers During Exercise

1975 ◽  
pp. 397-401 ◽  
Author(s):  
R. B. Armstrong ◽  
C. W. Saubert ◽  
W. L. Sembrowich ◽  
R. E. Shepherd ◽  
P. D. Gollnick
Keyword(s):  
Skeletal Muscle ◽  
Muscle Fibers ◽  
Glycogen Depletion ◽  
Rat Skeletal Muscle ◽  
Skeletal Muscle Fibers

scholarly journals Sodium nitrate supplementation alters mitochondrial H2O2 emission but does not improve mitochondrial oxidative metabolism in the heart of healthy rats

2018 ◽  
Vol 315 (2) ◽  
pp. R191-R204 ◽  
Author(s):  
Cynthia M. F. Monaco ◽  
Paula M. Miotto ◽  
Jason S. Huber ◽  
Luc J. C. van Loon ◽  
Jeremy A. Simpson ◽  
...  
Keyword(s):  
Diastolic Pressure ◽  
Muscle Fibers ◽  
Coupling Efficiency ◽  
Left Ventricular ◽  
Lv Function ◽  
Mitochondrial Bioenergetics ◽  
Isolated Mitochondria ◽  
Nitrate Supplementation ◽  
Permeabilized Muscle ◽  
Invasive Hemodynamics

Supplementation with dietary inorganic nitrate ([Formula: see text]) is increasingly recognized to confer cardioprotective effects in both healthy and clinical populations. While the mechanism(s) remains ambiguous, in skeletal muscle oral consumption of NaNO3 has been shown to improve mitochondrial efficiency. Whether NaNO3 has similar effects on mitochondria within the heart is unknown. Therefore, we comprehensively investigated the effect of NaNO3 supplementation on in vivo left ventricular (LV) function and mitochondrial bioenergetics. Healthy male Sprague-Dawley rats were supplemented with NaNO3 (1 g/l) in their drinking water for 7 days. Echocardiography and invasive hemodynamics were used to assess LV morphology and function. Blood pressure (BP) was measured by tail-cuff and invasive hemodynamics. Mitochondrial bioenergetics were measured in LV isolated mitochondria and permeabilized muscle fibers by high-resolution respirometry and fluorometry. Nitrate decreased ( P < 0.05) BP, LV end-diastolic pressure, and maximal LV pressure. Rates of LV relaxation (when normalized to mean arterial pressure) tended ( P = 0.13) to be higher with nitrate supplementation. However, nitrate did not alter LV mitochondrial respiration, coupling efficiency, or oxygen affinity in isolated mitochondria or permeabilized muscle fibers. In contrast, nitrate increased ( P < 0.05) the propensity for mitochondrial H2O2 emission in the absence of changes in cellular redox state and decreased the sensitivity of mitochondria to ADP (apparent Km). These results add to the therapeutic potential of nitrate supplementation in cardiovascular diseases and suggest that nitrate may confer these beneficial effects via mitochondrial redox signaling.

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