Fatty acid‐induced uncoupling of oxidative phosphorylation is partly due to opening of the mitochondrial permeability transition pore

Wiêckowski, Mariusz R.; Wojtczak, Lech

doi:10.1016/s0014-5793(98)00118-5

Cited by 117 publications

(65 citation statements)

References 35 publications

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“…Determination of ⌬⌿ m -Mitochondrial membrane potential was measured by using the fluorescence signal of the cationic dye safranine O, which is accumulated and quenched inside energized mitochondria (45). Mitochondria (0.2 mg protein/ml) were incubated in the standard respiration buffer supplemented with 10 M safranine.…”

Section: Methodsmentioning

confidence: 99%

Mitochondrial Bound Hexokinase Activity as a Preventive Antioxidant Defense

da‐Silva

Gómez‐Puyou

et al. 2004

Journal of Biological Chemistry

252

112

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Brain hexokinase is associated with the outer membrane of mitochondria, and its activity has been implicated in the regulation of ATP synthesis and apoptosis. Reactive oxygen species (ROS) are by-products of the electron transport chain in mitochondria. Here we show that the ADP produced by hexokinase activity in rat brain mitochondria (mt-hexokinase) controls both membrane potential (⌬⌿ m ) and ROS generation. Exposing control mitochondria to glucose increased the rate of oxygen consumption and reduced the rate of hydrogen peroxide generation. Mitochondrial associated hexokinase activity also regulated ⌬⌿ m , because glucose stabilized low ⌬⌿ m values in state 3. Interestingly, the addition of glucose 6-phosphate significantly reduced the time of state 3 persistence, leading to an increase in the ⌬⌿ m and in H 2 O 2 generation. The glucose analogue 2-deoxyglucose completely impaired H 2 O 2 formation in state 3-state 4 transition. In sharp contrast, the mt-hexokinase-depleted mitochondria were, in all the above mentioned experiments, insensitive to glucose addition, indicating that the mt-hexokinase activity is pivotal in the homeostasis of the physiological functions of mitochondria. When mt-hexokinase-depleted mitochondria were incubated with exogenous yeast hexokinase, which is not able to bind to mitochondria, the rate of H 2 O 2 generation reached levels similar to those exhibited by control mitochondria only when an excess of 10-fold more enzyme activity was supplemented. Hyperglycemia induced in embryonic rat brain cortical neurons increased ROS production due to a rise in the intracellular glucose 6-phosphate levels, which were decreased by the inclusion of 2-deoxyglucose, N-acetyl cysteine, or carbonyl cyanide p-trifluoromethoxyphenylhydrazone. Taken together, the results presented here indicate for the first time that mt-hexokinase activity performed a key role as a preventive antioxidant against oxidative stress, reducing mitochondrial ROS generation through an ADP-recycling mechanism.

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Section: Methodsmentioning

confidence: 99%

Mitochondrial Bound Hexokinase Activity as a Preventive Antioxidant Defense

da‐Silva

Gómez‐Puyou

et al. 2004

Journal of Biological Chemistry

252

112

View full text Add to dashboard Cite

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“…Determination of ⌬⌿-Mitochondrial membrane potential was measured by using the fluorescence signal of the cationic dye safranine O, which is accumulated and quenched inside energized mitochondria (17). Fluorescence was detected with an excitation wavelength of 495 nm (slit 5 nm) and an emission wavelength of 586 nm.…”

Section: Isolation Of Mitochondria From Rat Bat and Liver-adult Malementioning

confidence: 99%

Identification of a Ca2+-ATPase in Brown Adipose Tissue Mitochondria

Meis

Arruda

Costa

et al. 2006

Journal of Biological Chemistry

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In brown adipose tissue (BAT) adrenaline promotes a rise of the cytosolic Ca 2؉ concentration from 0.05 up to 0.70 M. It is not known how the rise of Ca 2؉ concentration activates BAT thermogenesis. In this report we compared the effects of Ca 2؉ in BAT and liver mitochondria. Using electron microscopy and immunolabeling we identified a sarco/endoplasmic reticulum (ER) Ca 2؉ -ATPase bound to the inner membrane of BAT mitochondria. A Ca 2؉ -dependent ATPase activity was detected in BAT mitochondria when the respiratory substrates malate and pyruvate were included in the medium. ATP and Ca 2؉ enhanced the amount of heat produced by BAT mitochondria during respiration. The Ca 2؉ concentration needed for half-maximal activation of the ATPase activity and rate of heat production were the same and varied between 0.1 and 0.2 M. Heat production was partially inhibited by the proton ionophore carbonyl cyanide p-trifluoromethoxyphenylhydrazone and abolished by thapsigargin, a specific ER Ca 2؉ -ATPase inhibitor, and by both rotenone and KCN, two substances that inhibit the electron transfer trough the mitochondrial cytochrome chain. In liver mitochondria Ca 2؉ did not stimulate the ATPase activity nor increase the rate of heat production. Thapsigargin had no effect on liver mitochondria. In conclusion, this is the first report of a Ca 2؉ -ATPase in mitochondria that is BAT-specific and can generate heat in the presence of Ca 2؉ concentrations similar to those noted in the cell during adrenergic stimulation. BAT3 is capable of rapidly converting fat stores to heat and has been used as a model system for the understanding of nonshivering heat production and mechanisms of energy wasting to control obesity (1-9). The signal that activates the rate of heat production in BAT cells is the rise of the cytosolic Ca 2ϩ concentration from a basal level of 0.05 M up to the range of 0.2-0.7 M. This is promoted by ␣ 1 -and ␤ 3 -adrenergic receptors located in the cell membrane. Activation of ␣ 1 -adrenoreceptors leads to the release of Ca 2ϩ from intracellular stores into the cytosol, whereas ␤ 3 -adrenergic receptors promote the release of free fatty acids and increase the effect of Ca 2ϩ release induced by ␣ 1 -adrenoreceptors (10, 11). At present we do not know how the rise of the cytosolic Ca 2ϩ concentration activates the rate of heat production in BAT cells.In a previous report (12) we identified a sarco/ER Ca 2ϩ ATPase (SERCA 1) in vesicles derived from BAT ER. In this report we show that the rate of heat produced by BAT mitochondria is enhanced when the Ca 2ϩ concentration in the medium is raised to a level similar to that observed in BAT cells during adrenergic stimulation. This effect is not observed in liver mitochondria, a tissue that is not specialized in heat production. EXPERIMENTAL PROCEDURES Isolation of Mitochondria from Rat BAT and Liver-Adult maleWistar rats were killed by decapitation. Mitochondria were isolated as previously described (13). Briefly, interscapular BAT and liver were removed and homogenized in a mix...

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“…В ізопротереноловій моделі пошкодження міокарда важливе значення відводиться погіршенню функціонування дихального ланцюга мітохондрій кардіоміоцитів, дії вільних радикалів, які утворюються в резуль-таті процесів перекисного окиснення ліпідів (ПОЛ) [2]. Важливу роль у порушенні різних клітинних функцій відіграє кальційзалежна циклоспоринчутлива мітохондріальна пора (МП) [1,3], яка вважається потужним фар-макологічним об'єктом при корекції хвороб, пов'язаних з мітохондріальною дисфункцією та індукцією клітинної смерті [2,[4][5][6].…”

Section: вступunclassified

“…Так, показано, що ССМ мають вищий вміст кардіоліпіну, а ІФМ -вищу експресію ключових компонентів пори, зокрема, потенціалзалежного аніонного каналу і циклофіліну D, крім того, вищий вміст цитохрому с [4]. Серед можливих механізмів негативної дії ізопротеренолу на мітохондрії може бути також те, що катехоламіни спричиняють вивільнення вільних жирних кислот, які в свою чергу діють як роз'єднувачі окисного фосфорилювання та індукують відкриття мітохондріальної пори, знижуючи мембран-ний потенціал нижче від порогового рівня чи прямо взаємодіючи з компонентами пори, зокрема з аденіннуклеотидтранслоказою [6]. Водночас ω-3 ПНЖК знижують вміст вільних жирних кислот [8].…”

Section: результати та їх обговоренняunclassified