A study of the effects of flocalin on respiration and potassium transport of rat-heart and liver mitochondria

The effects of the drug flocalin, which possesses cardioprotective properties, on the respiration rates of rat-heart and liver mitochondria in different functional states, the efficiency of oxidative phosphorylation, as well as the transport of potassium ions in these organelles, were studied. It was found that flocalin at concentrations of 7–30 μm stimulated respiration of rat-heart and liver mitochondria in V ₂ and V ₄ states in the presence of succinic add as a respiration substrate in a potassium-containing medium. In the absence of potassium ions in the incubation medium, flocalin had no effect on mitochondrial respiration in these states. Studying the functioning of the potassium transport system revealed that flocalin at these concentrations dose-dependently activated the ATP-dependent transport of potassium ions in rat-heart and liver mitochondria. The data we obtained indicate that the cardioprotective effect of flocalin can be associated with activation of the ATP-dependent potassium channel of the inner mitochondrial membrane.     

Effect of bedaquiline on the functions of rat liver mitochondria

The paper considers the effects of bedaquiline (BDQ), an antituberculous preparation of the new generation, on rat liver mitochondria. It was shown that 50?μM BDQ inhibited mitochondrial respiration measured with substrates of complexes I and II (glutamate/malate and succinate/rotenone systems respectively) in the states V₃ and V_DNP. At the same time, at concentrations below 50?μM, BDQ slightly stimulated respiration with substrates of complex I in the state V₂. BDQ was also found to suppress, in a dose-dependent manner, the activity of complex II and the total activity of complexes II?+?III of the mitochondrial transport chain. It was discovered that at concentrations up to 10?μM, BDQ inhibited H₂O₂ production in mitochondria. BDQ (10–50?μM) suppressed the opening of Ca²⁺-dependent CsA-sensitive mitochondrial permeability transition pore. The latter was revealed experimentally as the inhibition of Ca²⁺/P_i-dependent swelling of mitochondria, suppression of cytochrome c release, and an increase in the Ca²⁺ capacity of the organelles. BDQ also decreased the rate of mitochondrial energy-dependent K⁺ transport, which was evaluated by the energy-dependent swelling of mitochondria in a K⁺ buffer and DNP-induced K⁺ efflux from the organelles. The possible mechanisms of BDQ effect of rat liver mitochondria are discussed.     

Possible Mechanism for Formation and Regulation of the Palmitate-Induced Cyclosporin A-Insensitive Mitochondrial Pore

The mechanism of the palmitate-induced opening of the mitochondrial Ca2+-dependent cyclosporin A (CsA)-insensitive pore was studied, as well as the influence on this process of well-known modulators of the CsA-sensitive Ca2+-dependent pore. Palmitic acid, which can bind Ca2+ with high affinity, induced the cyclosporin A-insensitive swelling of mitochondria, whereas palmitoleic and 2-bromopalmitic acids, which have no such affinity for Ca2+, failed to induce the pore opening. The palmitate-induced Ca2+-dependent swelling of mitochondria was not affected by a well-known inhibitor of the CsA-sensitive pore (ADP) and an activator of this pore (inorganic phosphate, P(i)). However, this swelling was inhibited by physiological concentrations of ATP ([I]50 = 1.3 mM), but 100 microM ATP increased by 30% the rate of mitochondria swelling if Ca2+ had been added earlier. The effects of ATP (inhibition and activation) manifested themselves from different sides of the inner mitochondrial membrane. Mg2+ inhibited the palmitate-induced Ca2+-dependent swelling of mitochondria with [I]50 = 0.8 mM. It is concluded that palmitic acid induces the opening of the CsA-insensitive pore due to its ability for complexing with Ca2+. A possible mechanism of the pore formation and the influence of some modulators on this process are discussed.     

On the mechanism of palmitic acid-induced apoptosis: the role of a pore induced by palmitic acid and Ca2+ in mitochondria

Palmitic acid (Pal) is known to promote apoptosis (Sparagna G et al (2000) Am J Physiol Heart Circ Physiol 279: H2124–H2132) and its amount in blood and mitochondria increases under some pathological conditions. Yet, the mechanism of the proapoptotic action of Pal has not been elucidated. We present evidence for the involvement of the mitochondrial cyclosporin A-insensitive pore induced by Pal/Ca²⁺ complexes in the apoptotic process. Opening of this pore led to a fall of the mitochondrial membrane potential and the release of the proapoptotic signal cytochrome c. The addition of cytochrome c prevented these effects and recovered membrane potential, which is in contrast to the cyclosporin A-sensitive mitochondrial permeability transition pore. Oleic and linoleic acids prevented the Pal/Ca²⁺-induced pore opening in the intact mitochondria, this directly and significantly correlating with the effect of these fatty acids on Pal-induced apoptosis in cells (Hardy S et al (2003) J Biol Chem 278: 31861–31870). The specific probe for cardiolipin, 10-N-nonyl acridine orange, inhibited formation of this pore.     

Influence of cholesterol on the formation of palmitate/Ca<Superscript>2+</Superscript>-activated pores in mitochondria and liposomes

The influence of cholesterol on the formation of a mitochondrial cyclosporin A-insensitive palmitate/Ca²⁺-activated pore has been studied. Loading of mitochondrial membranes with cholesterol increases the rate of mitochondrial swelling induced by palmitic acid (≥20 μM) and Ca²⁺ (30 μM). This effect is not related to changes in the functional activity of organelles, since cholesterol does not influence the mitochondrial respiration in different metabolic states. At the same time, palmitate/Ca²⁺-induced permeabilization of azolectin/cholesterol liposomes is more pronounced than that of azolectin liposomes. In the liposomal membrane, Ca²⁺ induces phase separation of palmitic acid into distinct membrane domains; the presence of cholesterol in membranes enhances this effect.     

Energetic,oxidative and ionic exchange in rat brain and liver mitochondria at experimental audiogenic epilepsy (Krushinsky–Molodkina model)

The role of brain and liver mitochondria at epileptic seizure was studied on Krushinsky-Molodkina (KM) rats which respond to sound with an intensive epileptic seizure (audiogenic epilepsy). We didn't find significant changes in respiration rats of brain and liver mitochondria of KM and control rats; however the efficiency of АТР synthesis in the KM rat mitochondria was 10% lower. In rats with audiogenic epilepsy the concentration of oxidative stress marker malondialdehyde in mitochondria of the brain (but not liver) was 2-fold higher than that in the control rats. The rate of H₂O₂ generation in brain mitochondria of КМ rats was twofold higher than in the control animals when using NAD-dependent substrates. This difference was less pronounced in liver mitochondria. In KM rats, the activity of mitochondrial ATP-dependent potassium channel was lower than in liver mitochondria of control rats. The comparative study of the mitochondria ability to retain calcium ions revealed that in the case of using the complex I and complex II substrates, permeability transition pore is easier to trigger in brain and liver mitochondria of KM and КМs rats than in the control ones. The role of the changes in the energetic, oxidative, and ionic exchange in the mechanism of audiogenic epilepsy generation in rats and the possible correction of the epilepsy seizures are discussed.     

Role of mitochondria in hepatotoxicity of ethanol

The current understanding of the effects of alcohol intoxication on the basic mitochondrial functions has been presented. Both, the direct toxic effect of ethanol on biological membranes and various cellular systems and the toxicity of acetaldehyde and reactive oxygen species (the products of ethanol oxidation) are discussed, with emphasis on the effect of ethanol on the basic functions of mitochondria and Ca²⁺-dependent mitochondrial permeability transition. Based on the available experimental data, it is demonstrated that acute alcohol intoxication causes a global mitochondrial dysfunction in the liver, resulting in considerable disturbance of the whole cellular metabolism. Alcohol poisoning of the liver leads to a decreased ability of cells to withstand oxidative stress, to support the synthesis of vital metabolic intermediates (e.g., methyl groups), as well as to produce urea from ammonia, due to a decreased permeability of the outer membrane and impaired exchange of substrates between the cytoplasm and the mitochondrial matrix. This review emphasizes the role of porin channels of the outer mitochondrial membrane in ethanol-mediated disturbances of basic mitochondrial functions and its consequences for the entire cell metabolism in the liver.     

Ca<Superscript>2+</Superscript>-Induced Phase Separation in the Membrane of Palmitate-Containing Liposomes and Its Possible Relation to Membrane Permeabilization

A Ca(2+)-induced phase separation of palmitic acid (PA) in the membrane of azolectin unilamellar liposomes has been demonstrated with the fluorescent membrane probe nonyl acridine orange (NAO). It has been shown that NAO, whose fluorescence in liposomal membranes is quenched in a concentration-dependent way, can be used to monitor changes in the volume of lipid phase. The incorporation of PA into NAO-labeled liposomes increased fluorescence corresponding to the expansion of membrane. After subsequent addition of Ca(2+), fluorescence decreased, which indicated separation of PA/Ca(2+) complexes into distinct membrane domains. The Ca(2+)-induced phase separation of PA was further studied in relation to membrane permeabilization caused by Ca(2+) in the PA-containing liposomes. A supposition was made that the mechanism of PA/Ca(2+)-induced membrane permeabilization relates to the initial stage of Ca(2+)-induced phase separation of PA and can be considered as formation of fast-tightening lipid pores due to chemotropic phase transition in the lipid bilayer.     

Ultrastructural and Functional Changes in Liver Mitochondria in a Rat Model of Type I Diabetes Mellitus

Biophysics - Abstract—The characteristics of the ultrastructure and functioning of mitochondria in the liver of Sprague Dawley rats in the experimental model of type I diabetes mellitus have...     

Mitochondrial lipid pore in the mechanism of glutamate-induced calcium deregulation of brain neurons

The work examines the mechanism of central nerve cell death upon stimulation of brain NMDA receptors with the stimulatory mediator glutamate. A prolonged stimulation of neurons with glutamate is known to result in the disorder of Ca²⁺ homeostasis and severe mitochondrial depolarization followed by cell death. It has been shown that the overload of mitochondria with Sr²⁺ leads to the release of the cation, medium alkalization, decrease of membrane potential and mitochondrial swelling, indicating a nonspecific permeabilization of the mitochondrial membrane. The permeabilization, in our opinion, is caused by the activation of Ca²⁺/Sr²⁺-dependent phospholipase A₂ (PLA₂), resulting in the formation of free palmitic and stearic acids in the mitochondrial membrane. These fatty acids bind Ca²⁺ with high affinity and the process of binding is accompanied by the formation of a transient lipid pore—a phenomenon demonstrated earlier on both artificial and mitochondrial membranes. The inhibitors of PLA₂ have been shown to suppress permeabilization of mitochondrial membranes. In the culture of granular cerebellum neurons, the PLA₂ inhibitors prolonged the lag of the delayed Sr²⁺ deregulation and membrane depolarization. On the basis of data obtained on isolated mitochondria and neurons we suppose that the initial stages of glutamate-induced Ca²⁺ deregulation of neurons are underlain by the opening of lipid pores in brain mitochondria.