Standardized bioenergetic profiling of adult mouse cardiomyocytes

Readnower, Ryan D.; Brainard, Robert E.; Hill, Bradford G.; Jones, Steven P.

doi:10.1152/physiolgenomics.00129.2012

Cited by 69 publications

(80 citation statements)

References 29 publications

(31 reference statements)

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“…More recently, bioenergetic analysis of cell suspensions such as hematopoietic cells and lymphocytes using XF analyzer has been published (31,46). A recent study used a similar protocol for bioenergetic profiling of adult mouse cardiac myocytes seeded on laminin-coated plates (33). Although immobilization is recommended for isolated cells, for large cells such as cardiac myocytes, we have performed the analysis in suspension as well as in immobilized cells and obtained similar results (26).…”

Section: Thloredoxin 2 Expressionsupporting

confidence: 53%

Ca²⁺entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance

Hoffman

Miller

Wang

et al. 2015

American Journal of Physiology-Heart and Circulatory Physiology

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Five days post-injection, gKO-GFP heart slices had higher reactive oxygen species (ROS) levels but lower oxygen consumption rate (OCR) than WT-GFP heart slices. Trpm2 but not E960D decreased ROS and restored OCR in gKO hearts back to normal levels. In gKO myocytes expressing Trpm2 or its mutants, Trpm2 but not E960D reduced the elevated mitochondrial superoxide (O 2 .Ϫ ) levels in gKO myocytes. After hypoxia-reoxygenation (H/R), Trpm2 but not E906D or P1018L (inactivates Trpm2 current) lowered O 2 .Ϫ levels in gKO myocytes and only in the presence of extracellular Ca 2ϩ , indicating sustained Ca 2ϩ entry is necessary for Trpm2-mediated preservation of mitochondrial function. After ischemic-reperfusion (I/R), cardiac-specific Trpm2 KO hearts exhibited lower maximal first time derivative of LV pressure rise (ϩdP/dt) than WT hearts in vivo. After doxorubicin treatment, Trpm2 KO mice had worse survival and lower ϩdP/dt. We conclude 1) cardiac Trpm2-mediated Ca 2ϩ influx is necessary to maintain mitochondrial function and protect against H/R injury; 2) Ca 2ϩ influx via cardiac Trpm2 confers protection against H/R and I/R injury by reducing mitochondrial oxidants; and 3) Trpm2 confers protection in doxorubicin cardiomyopathy. ischemic cardiomyopathy; doxorubicin cardiomyopathy; voltage-independent Ca 2ϩ channels; cardiac Trpm2 currents; mitochondrial superoxide; hypoxia-reoxygenation IN MAMMALIAN CELLS, THE TRANSIENT receptor potential (Trp) protein superfamily is a diverse group of voltage-independent, cation-permeable channels organized into six subfamilies based on amino acid sequence homology (9, 30). Monomeric Trp proteins have six putative transmembrane (TM) domains and intracellular NH 2 and COOH termini. To form a functional channel, Trp proteins assemble into either homo-or heterotetramers, with the putative pore formed by loops between the fifth and sixth TM domains (21, 23). The Trp-melastatin (Trpm) subfamily consists of eight mammalian members (Trpm1-Trpm8)(30), of which Trpm2 (27), -m4, -m5, and -m6 (49) are expressed in the heart. Although Trpm4 is associated with conduction abnormalities and cardiac arrhythmias (1, 36), there is little information on the physiological and pathophysiological function of Trpm2 in the heart.

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Section: Thloredoxin 2 Expressionsupporting

confidence: 53%

Ca²⁺entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance

Hoffman

Miller

Wang

et al. 2015

American Journal of Physiology-Heart and Circulatory Physiology

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“…It is of interest that interfibrillar mitochondria show a greater respiratory capacity than subsarcolem- mal mitochondria in healthy Spague-Dawley rat hearts but have greater depression of respiratory capacity and LV contractile dysfunction after 20 wk of pressure overload (35). The use of the Seahorse extracellular flux analyzer provides the ability to localize the metabolic changes to intact cardiomyocytes excluding contamination from multiple cells types, maintaining mitochondrial localization in the cell, and obviating damage to mitochondria during isolation (14,16,32). However, because the cells are not contracting, the maximum OCR does not provide information regarding ATP production, but the lack of sensitivity to oligomycin suggests that the capacity of the mitochondria in the unstressed condition to make ATP is not impaired by VO.…”

Section: Discussionmentioning

confidence: 99%

“…However, the efficacy of XO inhibition has not been tested in the pathogenesis of a pure VO. Here, we studied chronic VO of ACF in the rat and attempted to determine whether XO inhibition has a long-term beneficial effect on LV remodeling/function and cardiomyocyte bioenergetics using the extracellular flux (XF24) analyzer from Seahorse Bioscience, which allows for quantitative real-time measurements of O 2 consumption and pH in cardiomyocyte culture (14,24,25,32).…”

mentioning

confidence: 99%

Xanthine oxidase inhibition preserves left ventricular systolic but not diastolic function in cardiac volume overload

Gladden

Zelickson

Guichard

et al. 2013

American Journal of Physiology-Heart and Circulatory Physiology

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In the setting of increased ATP demand, XO-mediated ROS can decrease mitochondrial respiration and contractile function. Thus, we tested the hypothesis that XO inhibition improves cardiomyocyte bioenergetics and LV function in chronic ACF in the rat. Sprague-Dawley rats were randomized to either sham or ACF Ϯ allopurinol (100 mg·kg Ϫ1 ·day Ϫ1 , n Ն7 rats/group). Echocardiography at 8 wk demonstrated a similar 37% increase in LV end-diastolic dimension (P Ͻ 0.001), a twofold increase in LV end-diastolic pressure/wall stress (P Ͻ 0.05), and a twofold increase in lung weight (P Ͻ 0.05) in treated and untreated ACF groups versus the sham group. LV ejection fraction, velocity of circumferential shortening, maximal systolic elastance, and contractile efficiency were significantly depressed in ACF and significantly improved in ACF ϩ allopurinol rats, all of which occurred in the absence of changes in the maximum O 2 consumption rate measured in isolated cardiomyocytes using the extracellular flux analyzer. However, the improvement in contractile function is not paralleled by any attenuation in LV dilatation, LV end-diastolic pressure/wall stress, and lung weight. In conclusion, allopurinol improves LV contractile function and efficiency possibly by diminishing the known XO-mediated ROS effects on myofilament Ca 2ϩ sensitivity. However, LV remodeling and diastolic properties are not improved, which may explain the failure of XO inhibition to improve symptoms and hospitalizations in patients with severe heart failure. heart failure; volume overload; xanthine oxidase; mitochondria A PURE VOLUME OVERLOAD (VO), without an increase in systolic pressure, increases diastolic load and results in an adverse eccentric left ventricular (LV) hypertrophy and remodeling manifested by wall thinning, cardiomyocyte elongation, and a decrease in the LV mass to volume.Currently, there is no medical therapy to attenuate this remodeling process and progression to heart failure (10). Recently, our laboratory has demonstrated increased cardiomyocyte oxidative stress in patients with VO of isolated chronic mitral regurgitation (1) and in rats with VO of aortocaval fistula (ACF) (18, 39). Specifically, xanthine oxidase (XO) is elevated in cardiomyocytes along with myofibrillar degeneration and mitochondrial dysfunction and damage, despite a preserved LV ejection fraction (LVEF). The preservation of LV shortening has been shown to belie underlying cardiomyocyte dysfunction due to ejection into the low pressure left atrium and, in this animal model, ejection into the aortovenous fistula. XO is a critical mediator of ATP and purine degradation and produces ROS when O 2 is used as an electron acceptor during purine metabolism (30). We have postulated that increased myocardial O 2 demand in the volume-overloaded heart with increased XO activity and ATP turnover sets up a vicious cycle of ROS production that can negatively impact multiple processes in the cardiomyocyte by altering mitochondrial function and production of ROS that impact other sensiti...

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“…Cardiac metabolism was analyzed using the Seahorse XFe 96 analyzer (Seahorse Bioscience, North Billerica, MA) using the protocol of Readnower et al 23 with minor changes. Cell metabolism was analyzed with the XF glycolysis stress test kit for analysis of glycolytic function and the XF Mito stress test kit for analysis of mitochondrial function according to the protocol provided by the manufacturer.…”

Section: Bioenergetic Profilingmentioning

confidence: 99%

Myostatin Regulates Energy Homeostasis in the Heart and Prevents Heart Failure

Biesemann

Mendler

Wietelmann

et al. 2014

Circulation Research

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Rationale: Myostatin is a major negative regulator of skeletal muscle mass and initiates multiple metabolic changes, including enhanced insulin sensitivity. However, the function of myostatin in the heart is barely understood, although it is upregulated in the myocardium under several pathological conditions.Objective: Here, we aimed to decipher the role of myostatin and myostatin-dependent signaling pathways for cardiac function and cardiac metabolism in adult mice. To avoid potential counterregulatory mechanisms occurring in constitutive and germ-line-based myostatin mutants, we generated a mouse model that allows myostatin inactivation in adult cardiomyocytes. Methods and Results:Cardiac MRI revealed that genetic inactivation of myostatin signaling in the adult murine heart caused cardiac hypertrophy and heart failure, partially recapitulating effects of the age-dependent decline of the myostatin paralog growth and differentiation factor 11. We found that myostatin represses AMP-activated kinase activation in the heart via transforming growth factor-β-activated kinase 1, thereby preventing a metabolic switch toward glycolysis and glycogen accumulation. Furthermore, myostatin stimulated expression of regulator of G-protein signaling 2, a GTPase-activating protein that restricts Gaq and Gas signaling and thereby protects against cardiac failure. Inhibition of AMP-activated kinase in vivo rescued cardiac hypertrophy and prevented enhanced glycolytic flow and glycogen accumulation after inactivation of myostatin in cardiomyocytes. Conclusions:

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Standardized bioenergetic profiling of adult mouse cardiomyocytes

Abstract: Readnower RD, Brainard RE, Hill BG, Jones SP. Standardized bioenergetic profiling of adult mouse cardiomyocytes.

Cited by 69 publications

References 29 publications

Ca²⁺entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance

Ca²⁺entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance

Xanthine oxidase inhibition preserves left ventricular systolic but not diastolic function in cardiac volume overload

Myostatin Regulates Energy Homeostasis in the Heart and Prevents Heart Failure

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Standardized bioenergetic profiling of adult mouse cardiomyocytes

Abstract: Readnower RD, Brainard RE, Hill BG, Jones SP. Standardized bioenergetic profiling of adult mouse cardiomyocytes.

Cited by 69 publications

References 29 publications

Ca2+entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance

Ca2+entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance

Xanthine oxidase inhibition preserves left ventricular systolic but not diastolic function in cardiac volume overload

Myostatin Regulates Energy Homeostasis in the Heart and Prevents Heart Failure

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Ca²⁺entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance

Ca²⁺entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance