scholarly journals Fluctuations in mitochondrial membrane potential caused by repetitive gating of the permeability transition pore

1999 ◽  
Vol 343 (2) ◽  
pp. 311-317 ◽  
Author(s):  
Jörg HüSER ◽  
Lothar A. BLATTER
Keyword(s):  
Oxidative Stress ◽  
Living Cell ◽  
Structural Damage ◽  
Laser Scanning ◽  
Mitochondrial Permeability Transition ◽  
Electrical Potential ◽  
Permeability Transition Pore ◽  
Permeability Transition ◽  
Laser Scanning Microscopy ◽  
Confocal Laser

Confocal laser scanning microscopy and the potentiometric fluorescence probe tetramethylrhodamine ethyl ester were used to measure changes in membrane electrical potential (δΨm) in individual mitochondria after isolation or in the living cell. Recordings averaged over small mitochondrial populations revealed a gradual decline in δΨmcaused by the light-induced generation of free radicals. Depolarization was attenuated by dithiothreitol or acidification. In contrast, individual organelles displayed rapid spontaneous depolarizations caused by openings of the mitochondrial permeability transition pore (MTP). Repetitive openings and closings of the pore gave rise to marked fluctuations in δΨm between the fully charged and completely depolarized state. Rapid spontaneous fluctuations in δΨm were observed in mitochondria isolated from rat heart and in mitochondria in living endothelial cells. The loss of δΨm of mitochondria in the living cell coincided with swelling of the organelle and the breakdown of long mitochondrial filaments. In the individual mitochondrion, oxidative stress initially triggered pore openings of shorter duration, before prolonged openings caused the complete dissipation of δΨm and a measurable efflux of larger solutes. Generalizing this scheme, we suggest that under conditions of prolonged oxidative stress and/or cellular Ca2+ overload, short openings of MTP might serve as an emergency mechanism allowing the partial dissipation of δΨm, the fast release of accumulated Ca2+ ions and the decreased generation of endogenous oxygen radicals. In contrast, loss of matrix metabolites, swelling and other structural damage of the organelle render prolonged openings of the transition pore deleterious to mitochondria and to the cell.

scholarly journals Curcumin Downregulates Phosphate Carrier and Protects against Doxorubicin Induced Cardiomyocyte Apoptosis

2016 ◽  
Vol 2016 ◽  
pp. 1-6 ◽  
Author(s):  
Lu Junkun ◽  
Chu Erfu ◽  
Hasahya Tony ◽  
Li Xin ◽  
K. C. Sudeep ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Laser Scanning ◽  
Mitochondrial Permeability Transition ◽  
Permeability Transition Pore ◽  
Permeability Transition ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
H9c2 Cell ◽  
Malondialdehyde Content ◽  
Phosphate Carrier

Aim. To explore the effects of curcumin on phosphate carrier (PiC) and its role in protection against doxorubicin induced myocyte toxicity.Methods. Using H9c2 cell line, the cardiotoxic effect of doxorubicin and its mitigation by curcumin were studied. H9c2 cells were cultured with doxorubicin and/or curcumin at various concentrations. Analysis for apoptosis of H9c2 was done using flow cytometry. Confocal laser scanning microscopy was used to record the fluorescence intensity ratios and to determine the mitochondrial permeability transition pore (MPTP) opening state. Oxidative stress was measured using glutathione level, superoxide dismutase activities, and malondialdehyde content. The effect of doxorubicin and curcumin on PiC gene expression was measured by real-time PCR.Results. Curcumin decreased mRNA of PiC and was partly protective against oxidative stress, loss of mitochondrial transmembrane potential, and apoptosis induced by doxorubicin. Interestingly, the effect was not clearly dose dependent and the concentration of 12 mg/L was more efficient than 15 and 10 mg/L.Conclusion. Curcumin downregulates PiC and partly protects against doxorubicin induced oxidative stress and myocyte apoptosis.

Abstract 305: Nebivolol Confers Mitochondria-Targeted Cardioprotection in Isoproterenol-Induced Acute Stressor State

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
Sumeet S Vaikunth ◽  
Karl T Weber ◽  
Syamal K Bhattacharya
Keyword(s):  
Oxidative Stress ◽  
Cardiac Tissue ◽  
Mitochondrial Permeability Transition ◽  
Therapeutic Potential ◽  
Permeability Transition Pore ◽  
Permeability Transition ◽  
Antioxidant Defenses ◽  
Sprague Dawley ◽  
Acute Stressor ◽  
And Oxidative Stress

Introduction: Isoproterenol-induced acute stressor state simulates injury from burns or trauma, and results in Ca 2+ overloading and oxidative stress in diverse tissues, including cardiac myocytes and their subsarcolemmal mitochondria (SSM), overwhelming endogenous Zn 2+ -based antioxidant defenses. We hypothesized that pretreatment with nebivolol (Nebi), having dual beta-1 antagonistic and novel beta-3 receptor agonistic properties, would prevent Ca 2+ overloading and oxidative stress and upregulate Zn 2+ -based antioxidant defenses, thus enhancing its overall cardioprotective potential in acute stressor state. Methods: Eight-week-old male Sprague-Dawley rats received a single subcutaneous dose of isoproterenol (1 mg/kg) and compared to those treated with Nebi (10 mg/kg by gavage) for 10 days prior to isoproterenol. SSM were harvested from cardiac tissue at sacrifice. Total Ca 2+ , Zn 2+ and 8-isoprostane levels in tissue, and mitochondrial permeability transition pore (mPTP) opening, free [Ca 2+ ] m and H 2 O 2 production in SSM were monitored. Untreated, age-/sex-matched rats served as controls; each group had six rats and data shown as mean±SEM. Results: Compared to controls, isoproterenol rats revealed: (1) Significantly (*p<0.05) increased cardiac tissue Ca 2+ (8.2±0.8 vs. 13.7±1.0*, nEq/mg fat-free dry tissue (FFDT)), which was abrogated ( # p<0.05) by Nebi (8.9±0.4 # ); (2) Reduced cardiac Zn 2+ (82.8±2.4 vs. 78.5±1.0*, ng/mg FFDT), but restored by Nebi (82.4±0.6 # ); (3) Two-fold rise in cardiac 8-isoprostane (111.4±13.7 vs. 232.1±17.2*, pmoles/mg protein), and negated by Nebi (122.3+14.5 # ); (4) Greater opening propensity for mPTP that diminished by Nebi; (5) Elevated [Ca 2+ ] m (88.8±2.5 vs. 161.5±1.0*, nM), but normalized by Nebi (93.3±2.7 # ); and (6) Increased H 2 O 2 production by SSM (97.4±5.3 vs. 142.8±7.0*, pmoles/mg protein/min), and nullified by Nebi (106.8±9.0 # ). Conclusions : Cardioprotection conferred by Nebi, a unique beta-blocker, prevented Ca 2+ overloading and oxidative stress in cardiac tissue and SSM, while simultaneously augmenting antioxidant capacity and promoting mPTP stability. Therapeutic potential of Nebi in patients with acute stressor states remains a provocative possibility that deserves to be explored.

scholarly journals Potential role of nuclear translocation of glyceraldehyde-3-phosphate dehydrogenase in apoptosis and oxidative stress

2001 ◽  
Vol 114 (9) ◽  
pp. 1643-1653 ◽  
Author(s):  
Z. Dastoor ◽  
J.L. Dreyer
Keyword(s):  
Oxidative Stress ◽  
Laser Scanning ◽  
Fluorescent Protein ◽  
Nuclear Translocation ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Apoptotic Cells ◽  
Transfected Cells ◽  
And Oxidative Stress

Recent studies indicating a role of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in apoptosis or oxidative stress has been reported. Using confocal laser-scanning microscopy, we have investigated the cellular distribution of GAPDH in central nervous system (CNS)-derived cells (neuroblastoma mNB41A3), in non-CNS derived cells (R6 fibroblast) and in an apoptosis-resistant Bcl2 overexpressing cell line (R6-Bcl2). Induction of apoptosis by staurosporine or MG132 and oxidative stress by H(2)O(2) or FeCN enhanced the nuclear translocation of endogenous GAPDH in all cell types, as detected by immunocytochemistry. In apoptotic cells, GAPDH expression is three times higher than in non-apoptotic cells. Consistent with a role for GAPDH in apoptosis, overexpression of a GAPDH-green fluorescent protein (GAPDH-GFP) hybrid increased nuclear import of GAPDH-GFP into transfected cells and the number of apoptotic cells, and made them more sensitive to agents that induce apoptosis. Bcl2 overexpression prevents nuclear translocation of GAPDH and apoptosis in untransfected cells, but not in transfected cells that overexpress GAPDH-GFP. Our observations indicate that nuclear translocation of GAPDH may play a role in apoptosis and oxidative stress, probably related to the activity of GAPDH as a DNA repair enzyme or as a nuclear carrier for pro-apoptotic molecules.

Chronic Tempol treatment restores pharmacological preconditioning in the senescent rat heart

2013 ◽  
Vol 304 (5) ◽  
pp. H649-H659 ◽  
Author(s):  
Jiang Zhu ◽  
Mario J. Rebecchi ◽  
Qiang Wang ◽  
Peter S. A. Glass ◽  
Peter R. Brink ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Mitochondrial Permeability Transition Pore ◽  
Mitochondrial Permeability Transition ◽  
Permeability Transition Pore ◽  
Permeability Transition ◽  
Cyclophilin D ◽  
Mitochondrial Permeability ◽  
Anesthetic Preconditioning ◽  
Permeability Transition Pore Opening ◽  
Pore Opening

Cardioprotective effects of anesthetic preconditioning and cyclosporine A (CsA) are lost with aging. To extend our previous work and address a possible mechanism underlying age-related differences, we investigated the role of oxidative stress in the aging heart by treating senescent animals with the oxygen free radical scavenger Tempol. Old male Fischer 344 rats (22–24 mo) were randomly assigned to control or Tempol treatment groups for 2 or 4 wk (T×2wk and T×4wk, respectively). Rats received isoflurane 30 min before ischemia-reperfusion injury or CsA just before reperfusion. Myocardial infarction sizes were significantly reduced by isoflurane or CsA in the aged rats treated with Tempol (T×4wk) compared with old control rats. In other experiments, young (4–6 mo) and old rats underwent either chronic Tempol or vehicle treatment, and the levels of myocardial protein oxidative damage, antioxidant enzymes, mitochondrial Ca2+ uptake, cyclophilin D protein, and mitochondrial permeability transition pore opening times were measured. T×4wk significantly increased MnSOD enzyme activity, GSH-to-GSSH ratios, MnSOD protein level, mitochondrial Ca2+ uptake capacity, reduced protein nitrotyrosine levels, and normalized cyclophilin D protein expression in the aged rat heart. T×4wk also significantly prolonged mitochondrial permeability transition pore opening times induced by reactive oxygen species in old cardiomyocytes. Our studies demonstrate that 4 wk of Tempol pretreatment restores anesthetic preconditioning and cardioprotection by CsA in the old rat and that this is associated with decreased oxidative stress and improved mitochondrial function. Our results point to a new protective strategy for the ischemic myocardium in the high-risk older population.

Abstract MP124: De-palmitoylation of Cysteine 202 of Cyclophilin-D by Calcium is Important for the Activation of the Permeability Transition Pore

2020 ◽  
Vol 127 (Suppl_1) ◽  
Author(s):  
Georgios Amanakis ◽  
Junhui Sun ◽  
Maria Fergusson ◽  
Chengyu Liu ◽  
Jeff D Molkentin ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Mitochondrial Permeability Transition ◽  
Ischemia Reperfusion ◽  
Permeability Transition Pore ◽  
Permeability Transition ◽  
Calcium Overload ◽  
Mitochondrial Calcium ◽  
Cyclophilin D ◽  
Perfused Heart ◽  
Knock Out

Cyclophilin-D (CypD) is a well-known regulator of the mitochondrial permeability transition pore (PTP), the main effector of cardiac ischemia/reperfusion (I/R) injury characterized by oxidative stress and calcium overload. However, the mechanism by which CypD activates PTP is poorly understood. Cysteine 202 of CypD (C202) is highly conserved across species and can undergo redox-sensitive post-translational modifications, such as S-nitrosylation and oxidation. To study the importance of C202, we developed a knock-in mouse model using CRISPR where CypD-C202 was mutated to a serine (C202S). Hearts from these mice are protected against I/R injury. We found C202 to be abundantly S-palmitoylated under baseline conditions while C202 was de-palmitoylated during ischemia in WT hearts. To further investigate the mechanism of de-palmitoylation during ischemia, we considered the increase of matrix calcium, oxidative stress and uncoupling of ATP synthesis from the electron transport chain. We tested the effects of these conditions on the palmitoylation of CypD in isolated cardiac mitochondria. The palmitoylation of CypD was assessed using a resin-assisted capture (Acyl-RAC). We report that oxidative stress (phenylarsenide) and uncoupling (CCCP) had no effect on CypD palmitoylation (p>0.05, n=3 and n=7 respectively). However, calcium overload led to de-palmitoylation of CypD to the level observed at the end ischemia (1±0.10 vs 0.63±0.09, p=0.012, n=9). To further test the hypothesis that calcium regulates S-palmitoylation of CypD we measured S-palmitoylation of CypD in non-perfused heart lysates from global germline mitochondrial calcium uniporter knock-out mice (MCU-KO), which have reduced mitochondrial calcium and we found an increase in S-palmitoylation of CypD (WT 1±0.04 vs MCU-KO 1.603±0.11, p<0.001, n=6). The data are consistent with the hypothesis that C202 is important for the CypD mediated activation of PTP. Ischemia leads to increased matrix calcium which in turn promotes the de-palmitoylation of CypD on C202. The now free C202 can further be oxidized during reperfusion leading to the activation of PTP. Thus, S-palmitoylation and oxidation of CypD-C202 possibly target CypD to the PTP, making them potent regulators of cardiac I/R injury.

scholarly journals The P66Shc/Mitochondrial Permeability Transition Pore Pathway Determines Neurodegeneration

2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
Costanza Savino ◽  
PierGiuseppe Pelicci ◽  
Marco Giorgio
Keyword(s):  
Oxidative Stress ◽  
Knockout Mice ◽  
Mitochondrial Permeability Transition ◽  
Neuronal Damage ◽  
Permeability Transition Pore ◽  
Permeability Transition ◽  
Mitochondrial Swelling ◽  
Cyclophilin D ◽  
Knock Out ◽  
Mitochondrial Permeability

Mitochondrial-mediated oxidative stress and apoptosis play a crucial role in neurodegenerative disease and aging. Both mitochondrial permeability transition (PT) and swelling of mitochondria have been involved in neurodegeneration. Indeed, knockout mice for cyclophilin-D (Cyc-D), a key regulatory component of the PT pore (PTP) that triggers mitochondrial swelling, resulted to be protected in preclinical models of multiple sclerosis (MS), Parkinson’s disease (PD), and amyotrophic lateral sclerosis (ALS). However, how neuronal stress is transduced into mitochondrial oxidative stress and swelling is unclear. Recently, the aging determinant p66Shc that generates H2O2reacting with cytochrome c and induces oxidation of PTP and mitochondrial swelling was found to be involved in MS and ALS. To investigate the role of p66Shc/PTP pathway in neurodegeneration, we performed experimental autoimmune encephalomyelitis (EAE) experiments in p66Shc knockout mice (p66Shc−/−), knock out mice for cyclophilin-D (Cyc-D−/−), and p66Shc Cyc-D double knock out (p66Shc/Cyc-D−/−) mice. Results confirm that deletion of p66Shc protects from EAE without affecting immune response, whereas it is not epistatic to the Cyc-D mutation. These findings demonstrate that p66Shc contributes to EAE induced neuronal damage most likely through the opening of PTP suggesting that p66Shc/PTP pathway transduces neurodegenerative stresses.

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