scholarly journals The very low number of calcium-induced permeability transition pores in the single mitochondrion

2020 ◽  
Vol 152 (10) ◽  
Author(s):  
Maria A. Neginskaya ◽  
Jasiel O. Strubbe ◽  
Giuseppe F. Amodeo ◽  
Benjamin A. West ◽  
Shoshana Yakar ◽  
...  

Mitochondrial permeability transition (PT) is a phenomenon of stress-induced increase in nonspecific permeability of the mitochondrial inner membrane that leads to disruption of oxidative phosphorylation and cell death. Quantitative measurement of the membrane permeability increase during PT is critically important for understanding the PT’s impact on mitochondrial function. The elementary unit of PT is a PT pore (PTP), a single channel presumably formed by either ATP synthase or adenine nucleotide translocator (ANT). It is not known how many channels are open in a single mitochondrion during PT, which makes it difficult to quantitatively estimate the overall degree of membrane permeability. Here, we used wide-field microscopy to record mitochondrial swelling and quantitatively measure rates of single-mitochondrion volume increase during PT-induced high-amplitude swelling. PT was quantified by calculating the rates of water flux responsible for measured volume changes. The total water flux through the mitochondrial membrane of a single mitochondrion during PT was in the range of (2.5 ± 0.4) × 10−17 kg/s for swelling in 2 mM Ca2+ and (1.1 ± 0.2) × 10−17 kg/s for swelling in 200 µM Ca2+. Under these experimental conditions, a single PTP channel with ionic conductance of 1.5 nS could allow passage of water at the rate of 0.65 × 10−17 kg/s. Thus, we estimate the integral ionic conductance of the whole mitochondrion during PT to be 5.9 ± 0.9 nS for 2 mM concentration of Ca2+ and 2.6 ± 0.4 nS for 200 µM of Ca2+. The number of PTPs per mitochondrion ranged from one to nine. Due to the uncertainties in PTP structure and model parameters, PTP count results may be slightly underestimated. However, taking into account that each mitochondrion has ∼15,000 copies of ATP synthases and ANTs, our data imply that PTP activation is a rare event that occurs only in a small subpopulation of these proteins.

2014 ◽  
Vol 2014 ◽  
pp. 1-14 ◽  
Author(s):  
Elena A. Belyaeva ◽  
Larisa V. Emelyanova ◽  
Sergey M. Korotkov ◽  
Irina V. Brailovskaya ◽  
Margarita V. Savina

Previously we have shown that opening of the mitochondrial permeability transition pore in its low conductance state is the case in hepatocytes of the Baltic lamprey (Lampetra fluviatilis L.) during reversible metabolic depression taking place in the period of its prespawning migration when the exogenous feeding is switched off. The depression is observed in the last year of the lamprey life cycle and is conditioned by reversible mitochondrial dysfunction (mitochondrial uncoupling in winter and coupling in spring). To further elucidate the mechanism(s) of induction of the mitochondrial permeability transition pore in the lamprey liver, we used Cd2+and Ca2+plus Pias the pore inducers. We found that Ca2+plus Piinduced the high-amplitude swelling of the isolated “winter” mitochondria both in isotonic sucrose and ammonium nitrate medium while both low and high Cd2+did not produce the mitochondrial swelling in these media. Low Cd2+enhanced the inhibition of basal respiration rate of the “winter” mitochondria energized by NAD-dependent substrates whereas the same concentrations of the heavy metal evoked its partial stimulation on FAD-dependent substrates. The above changes produced by Cd2+or Ca2+plus Piin the “winter” mitochondria were only weakly (if so) sensitive to cyclosporine A (a potent pharmacological desensitizer of the nonselective pore) added alone and they were not sensitive to dithiothreitol (a dithiol reducing agent). Under monitoring of the transmembrane potential of the “spring” lamprey liver mitochondria, we revealed that Cd2+produced its decrease on both types of the respiratory substrates used that was strongly hampered by cyclosporine A, and the membrane potential was partially restored by dithiothreitol. The effects of different membrane permeability modulators on the lamprey liver mitochondria function and the seasonal changes in their action are discussed.


2013 ◽  
Vol 33 (6) ◽  
pp. 928-934 ◽  
Author(s):  
Ping Gong ◽  
Rong Hua ◽  
Yu Zhang ◽  
Hong Zhao ◽  
Ziren Tang ◽  
...  

Increasing evidence has shown that mild hypothermia is neuroprotective for comatose patients resuscitated from cardiac arrest, but the mechanism of this protection is not fully understood. The aim of this study was to determine whether prolonged whole-body mild hypothermia inhibits mitochondrial membrane permeability (MMP) in the cerebral cortex after return of spontaneous circulation (ROSC). Thirty-seven inbred Chinese Wuzhishan minipigs were successfully resuscitated after 8 minutes of untreated ventricular fibrillation (VF) and underwent recovery under normothermic (NT) or prolonged whole-body mild hypothermic (HT; 33°C) conditions for 24 or 72 hours. Cerebral samples from the frontal cortex were collected at 24 and 72 hours after ROSC. Mitochondria were isolated by differential centrifugation. At 24 hours, relative to NT, HT was associated with reductions in opening of the mitochondrial permeability transition pore, release of pro-apoptotic substances from mitochondria, caspase 3 cleavage, apoptosis, and neurologic deficit scores, as well as increases in mitochondrial membrane potential and mitochondrial respiration. Together, these findings suggest that mild hypothermia inhibits ischemia-induced increases in MMP, which may provide neuroprotection against cerebral injury after cardiac arrest.


2015 ◽  
Vol 117 (suppl_1) ◽  
Author(s):  
Jason Karch ◽  
Tobias G Schips ◽  
Matthew J Brody ◽  
Onur Kanisicak ◽  
Michelle A Sargent ◽  
...  

In times of energy depletion, a cell will attempt to maintain metabolic homeostasis and viability by degrading and recycling organelles and intracellular components and proteins in a process referred to as autophagy. However, if the energy depletion persists, the cell will be overwhelmed by the autophagic process and will succumb to autophagic cell death. This form of cell death has been implicated in cardiac remodeling during heart failure and damage during ischemic injury. Two proteins that have been previously shown to play a role in virtually every form of regulated cell death, including autophagy, are Bax and Bak. These effectors are responsible for cytochrome-c release during apoptosis and effect mitochondrial permeability transition pore opening during regulated necrosis. Although the expression of either Bax or Bak is required for autophagic cell death to occur, the role of Bax/Bak in this type of cell death is poorly understood, although the lysosome appears to be centrally involved. Here we show that Bax/Bak DKO MEFs subjected to several days of serum starvation contain intact lysosomes compared to WT MEFs. Furthermore, the acidity of the lysosomes in starved DKO MEFs is preserved compared with starved WT MEFs. Bax and Bak are both found in isolated lysosomal preparations and Bax targeted to the lysosome can completely restore autophagic cell death in DKO MEFs. Finally, although Bax oligomerization is required for apoptosis, it is not necessary for autophagic cell death, as DKO MEFs expressing an oligomerization defective mutant of Bax are still susceptible to this form of death, as monomeric Bax can still increase membrane permeability. In conclusion our results suggest that lysosomal membrane permeability through Bax or Bak is required for autophagic cell death to occur and without Bax or Bak the lysosomes remain intact where they can function as an energy source during times of nutrient deprivation.


2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii99-ii100
Author(s):  
Martyn Sharpe ◽  
David Baskin ◽  
Santosh Helekar

Abstract Magnetic fields in the mT range influence spin state pairing in redox-active radical pairs generating spin-forbidden quantum states which are kinetically inert. Studies examining the effects of static magnetic fields on mitochondrial electron transfer kinetics have demonstrated only modest effects. When neodymium super-magnets are securely attached to and precision-balanced on the shafts of electronically-controlled motors it is possible to generate rotating magnetic fields of desirable strengths and frequencies. Unlike a static magnetic field or an alternating field in a static electromagnetic coil, oscillating magnetic field (OMF) produced by rotating lines of force of a spinning permanent magnet can dynamically couple the electron spins of radical pairs within proteins whose orientations are ‘fixed’. The frequencies of rotation of magnets can also be tuned to appropriate electron cycling resonances within the proteins. Using OMF of appropriate field strength, frequency and on/off acceleration/deceleration profiles we can completely arrest electron transport in isolated respiring rat liver mitochondria. Parallel to this inhibition of electron flux, we also independently observe an increase in superoxide and hydrogen peroxide. Under certain OMF exposure regimes, we observe membrane permeability transition in these mitochondria when using succinate as substrate, and show that the mitochondrial membrane permeability transition effect can be blocked by bongkrekic acid. We have examined the effect of OMF on oxygen consumption in cultured primary cancer cells with a rotating magnet (oncoscillator) that is an integral component of a new anti-cancer Oncomagnetic device. We observe three main effects in addition to the inhibition of respiratory flux in cancer cells – damage to the respiratory complex, uncoupling and generation of superoxide/hydrogen peroxide. OMF generated by oncoscillators can induce mitochondrial permeability transition in primary cultured malignant meningioma, diffuse intrinsic pontine glioma and GBM cells. Parallel experiments with normal human astrocytes show only minor changes in cellular/mitochondrial function under these conditions.


2021 ◽  
Vol 12 (2) ◽  
pp. 419-439
Author(s):  
Elena P. Isakova ◽  
Olga I. Klein ◽  
Yulia I. Deryabina

In this study, the mechanism of non-specific membrane permeability (yPTP) in the Endomyces magnusii yeast mitochondria under oxidative stress due to blocking the key antioxidant enzymes has been investigated. We used monitoring the membrane potential at the cellular (potential-dependent staining) and mitochondrial levels and mitochondria ultra-structural images with transmission electron microscopy (TEM) to demonstrate the mitochondrial permeability transition induction due to the pore opening. Analysis of the yPTP opening upon respiring different substrates showed that NAD(P)H completely blocked the development of the yPTP. The yPTP opening was inhibited by 5–20 mM Pi, 5 mM Mg2+, adenine nucleotides (AN), 5 mM GSH, the inhibitor of the Pi transporter (PiC), 100 μM mersalyl, the blockers of the adenine nucleotide transporter (ANT) carboxyatractyloside (CATR), and bongkrekic acid (BA). We concluded that the non-specific membrane permeability pore opens in the E. magnusii mitochondria under oxidative stress, and the ANT and PiC are involved in its formation. The crucial role of the Ca2+ ions in the process has not been confirmed. We showed that the Ca2+ ions affected the yPTP both with and without the Ca2+ ionophore ETH129 application insignificantly. This phenomenon in the E. magnusii yeast unites both mitochondrial unselective channel (ScMUC) features in the Saccharomyces cerevisiae mitochondria and the classical membrane pore in the mammalian ones (mPTP).


2003 ◽  
Vol 2 (1) ◽  
pp. 167 ◽  
Author(s):  
L ARGAUD ◽  
O GATEAUROESCH ◽  
D MUNTEAN ◽  
L GOMEZ ◽  
L CHALABREYSSE ◽  
...  

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