Measurement of membrane permeability and the mitochondrial permeability transition

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.

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EXTH-58. ONCOSCILLATORS: INDUCTION OF THE MITOCHONDRIAL PERMEABILITY TRANSITION IN CANCER CELLS

Neuro-Oncology ◽

10.1093/neuonc/noaa215.412 ◽

2020 ◽

Vol 22 (Supplement_2) ◽

pp. ii99-ii100

Author(s):

Martyn Sharpe ◽

David Baskin ◽

Santosh Helekar

Keyword(s):

Magnetic Field ◽

Hydrogen Peroxide ◽

Magnetic Fields ◽

Cancer Cells ◽

Membrane Permeability ◽

Mitochondrial Permeability Transition ◽

Permeability Transition ◽

Radical Pairs ◽

Membrane Permeability Transition ◽

Mitochondrial Permeability

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.

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