Inhibition Of Mitochondrial Electron Transfer Research Articles

AKD-2023: A novel miticide. Biological activity and mode of action

AKD-2023, 3-dodecyl-1,4-dihydro-1,4-dioxo-2-naphthyl acetate (proposed common name acequinocyl) is a novel miticide now under commercial development. It has outstanding miticidal activity against economically important spider mites with the new mode of action of inhibition of mitochondrial electron transfer at complex III.

Nitric Oxide, Superoxide, and Hydrogen Peroxide Production in Brain Mitochondria after Haloperidol Treatment

Inhibition of mitochondrial respiration and free radical induction have been suggested to be involved in haloperidol neurotoxicity. In this study, mice were injected i.p. with haloperidol, according to two different treatments: (a) a single injection (1 mg/kg), sacrificed 1 h after the injection (single-dose model); and (b) two injections (1 mg/kg each), sacrificed 24 h after the first dose (double-dose model). Determinations of oxygen consumption and hydrogen peroxide (H2O2) production rate were carried out in isolated brain mitochondria. Nitric oxide (NO) and superoxide (O−2) production rates were measured in submitochondrial particles (SMP). Single-dose haloperidol treatment produced a 33% inhibition in malate–glutamate-dependent respiration, while no significant changes were found after double-dose treatment. NO production was inhibited by 39 and 54% in SMP from haloperidol-treated mice (single- and double-dose treatments, respectively) (control value: 1.6 ± 0.2 nmol/min mg protein). NO steady-state concentration was estimated at about 16.5 nM and was decreased by 40% by haloperidol treatment. Increases of 105 and 54% were found in succinate-supported O−2 and H2O2 production rates, respectively, after haloperidol single-dose treatment. Haloperidol treatment generated a 248% increase in SMP O−2 production rate when measured in the presence of NADH plus rotenone. Our results suggest that haloperidol neurotoxicity would be mediated by a decreased mitochondrial NO production, a decreased intramitochondrial NO steady-state concentration, and by an inhibition of mitochondrial electron transfer with enhancement of O−2 and H2O2 production. This inhibition does not seem to be caused by increased NO or ONOO− formation.

Nitric Oxide Inhibits Electron Transfer and Increases Superoxide Radical Production in Rat Heart Mitochondria and Submitochondrial Particles

Nitric oxide (·NO) released byS-nitrosoglutathione (GSNO) inhibited enzymatic activities of rat heart mitochondrial membranes. Cytochrome oxidase activity was inhibited to one-half at an effective ·NO concentration of 0.1 μM, while succinate– and NADH–cytochrome-creductase activities were half-maximally inhibited at 0.3 μM·NO. Submitochondrial particles treated with ·NO (either from GSNO or from a pure solution) showed increased[formula]and H2O2production when supplemented with succinate alone, at rates that were comparable to those of control particles with added succinate and antimycin. Rat heart mitochondria treated with ·NO also showed increased H2O2production. Cytochrome spectra and decreased enzymatic activities in the presence of ·NO are consistent with a multiple inhibition of mitochondrial electron transfer at cytochrome oxidase and at the ubiquinone–cytochromebregion of the respiratory chain, the latter leading to the increased[formula]production. Electrochemical detection showed that the buildup of a ·NO concentration from GSNO was interrupted by submitochondrial particles supplemented with succinate and antimycin and was restored by addition of superoxide dismutase. The inhibitory effect of ·NO on cytochrome oxidase was also prevented under the same conditions. Apparently, mitochondrial[formula]reacts with ·NO to form peroxynitrite and, by removing ·NO, reactivates the previously inhibited cytochrome oxidase. It is suggested that, at physiological concentrations of ·NO, inhibition of electron transfer, ·NO-induced[formula]production, and ONOO−formation participate in the regulatory control of mitochondrial oxygen uptake.

Monitoring of mitochondrial NADH levels by surface fluorimetry as an indication of ischaemia during hepatic and renal transplantation.

One of the major causes of dysfunction in transplanted organs is ischaemia-reperfusion (IR) injury. Impairment of mitochondrial function is likely to be central to many of the known consequences of ischaemia; these include loss of cellular homeostasis involving a fall in intracellular pH (Fuller et al., 1988), mitochondrial calcium loading and cellular swelling (Caiman et al., 1973), accumulation of reduced pyridine nucleotides, inhibition of mitochondrial electron transfer, and a fall in ATP levels (Hardy et al., 1991). In irreversibly damaged cells, respiratory control is lost and is accompanied by oxidation of cytochromes a and a3 and NADH (Taegtmeyer et al., 1985). The latter was attributed originally to substrate deficiency (Chance and Williams, 1955) but more recent studies indicate that an enzymological defect develops resulting in an inability to metabolise NADH-linked substrates (Taegtmeyer et al., 1985 and Hardy et al., 1991). In vitro studies of the respiratory chain (RC) complexes have been made in several tissues including cardiac and renal tissue, subjected to ischaemia-reperfusion injury and it was found that complexes I and IV are major defective sites (Hardy et al., 1991 and Veitch et al., 1992). Return of function may, therefore, relate to preservation of inner mitochondrial membrane integrity, and the structure and activities of the RC complexes. The integrity of oxidative metabolic pathways and capacity to resynthesise ATP rather than the immediate post-ischaemic ATP levels appears to determine the return of function (Taegtmeyer et al., 1985).

Time course and mechanism of oxidative stress and tissue damage in rat liver subjected to in vivo ischemia-reperfusion.

The time course of oxidative stress and tissue damage in zonal liver ischemia-reperfusion in rat liver in vivo was evaluated. After 180 min of ischemia, surface chemiluminescence decreased to zero, state 3 mitochondrial respiration decreased by 70-80%, and xanthine oxidase activity increased by 26% without change in the water content and in the activities of superoxide dismutase, catalase, and glutathione peroxidase. After reperfusion, marked increases in oxyradical production and tissue damage were detected. Mitochondrial oxygen uptake in state 3 and respiratory control as well as the activities of superoxide dismutase, catalase, and glutathione peroxidase and the level of nonenzymatic antioxidants (evaluated by the hydroperoxide-initiated chemiluminescence) were decreased. The severity of the post-reperfusion changes correlated with the time of ischemia. Morphologically, hepatocytes appeared swollen with zonal cord disarrangement which ranged from mild to severe for the tissue reperfused after 60-180 min of ischemia. Neutrophil infiltration was observed after 180 min of ischemia and 30 min of reperfusion. Mitochondria appear as the major source of hydrogen peroxide in control and in reperfused liver, as indicated by the almost complete inhibition of hydrogen peroxide production exerted by the uncoupler carbonylcyanide p-(trifluoromethoxy) phenylhydrazone. Additionally, inhibition of mitochondrial electron transfer by antimycin in liver slices reproduced the inhibition of state 3 mitochondrial respiration and the increase in hydrogen peroxide steady-state concentration found in reperfused liver. Increased rates of oxyradical production by inhibited mitochondria appear as the initial cause of oxidative stress and liver damage during early reperfusion in rat liver.

Effect of photosynthesis on dark mitochondrial respiration in green cells

Green plant cells can generate ATP in both chloroplasts and mitochondria. Hence the effect of photosynthesis on dark mitochondrial respiration can be considered at a variety of levels. Turnover of ceitric acid cycle dehydrogenases, which is essential for supply of carbon skeletons for amino acid synthesis, seems to be largely unaffected during photosynthesis. The source of carbon for the anaplerotic function of the citric acid cycle in light is however, not known with certainty. NADH generated in these reactions is probably not oxidised via the mitochondrial electron transfer chain coupled to ATP synthesis. However, it may be oxidised by the alternative cyanide-insensitive pathway, exported to the cytosol via the oxaloacetate-malate dicarboxylate shuttle or directly utilised for cytosolic nitrate reduction. Oxidation of succinate via cytochrome oxidase may also be similarly inhibited in light. Whether increase in the cytosolic ATP/ADP ratio in light is responsible for the inhibition of mitochondrial electron transfer to O 2 is not clearly established, because the ATP/ADP ratio is reported to be already quite high in the dark. Effective collaboration between photophosphorylation and oxidative phosphorylation in order to maintain the cytosolic energy charge at a present high level is discussed.

Effects of dibromothymoquinone on mung bean mitochondrial electron transfer and membrane fluidity

The effects of the quinone analog dibromothymoquinone on electron transfer in isolated mung bean mitochondria are described. Both the main, cyanidesensitive and the alternate, cyanide-insensitive pathways are inhibited by dibromothymoquinone but in markedly different fashions. Half-maximal inhibition appeared at 40 μM and 20 μM dibromothymoquinone for the cyanide-sensitive and alternate pathways, respectively. With succinate as the electron donor, dibromothymoquinone inhibited the alternate pathway at a single site; showing a mixed, non-competitive type inhibition. On the succinate, cyanide-sensitive pathway dibromothymoquinone showed two sites of inhibition and neither coincides with the site of inhibition associated with the alternate pathway. With malate as the electron donor, two sites of inhibition by dibromothymoquinone were observed regardless of the pathway measured. Dibromothymoquinone also inhibited the rate of valinomycin-induced swelling of isolated mung bean mitochondria. Steady-state kinetics showed the inhibition to be non-competitive with respect to valinomycin. Additionally dibromothymoquinone was observed to increase the fluorescence polarization associated with the hydrophobic probe 1,6-diphenylhexatriene. The results indicated that dibromothymoquinone decreased the fluidity of the inner mitochondrial membrane and suggested that the inhibition of mitochondrial electron transfer by dibromothymoquinone may be associated with this decrease in membrane fluidity. The relationship of the multisite nature of the inhibition of electron transfer by dibromothymoquinone and the possible role of mobile electron carriers such as ubiquinone on the main and alternate respiratory pathways of higher plants is discussed.

Diminished inhibition of mitochondrial electron transfer from succinate to cytochrome c by thenoyltrifluoroacetone induced by antimycin.

Diminished inhibition of mitochondrial electron transfer from succinate to cytochrome c by thenoyltrifluoroacetone induced by antimycin.

Inhibition of Mitochondrial Electron Transfer in Rats by Ethanethiol and Methanethiol

1. We have investigated the effects of ethanethiol, methanethiol and dimethyl sulphide on some metabolic processes of isolated rat hepatocytes, isolated mitochondria from liver and brain and ox-heart submitochondrial particles. 2. Ethanethiol, but not dimethyl sulphide, inhibited both gluconeogenesis and ureogenesis from various substrates in rat hepatocytes, depressed cellular ATP content and caused an increased reduction of the mitochondria. 3. Ethanethiol inhibited respiration in isolated rat-liver mitochondria with several substrates, both in the presence of ADP and phosphate or in the presence of an uncoupling agent. Ethanethiol also inhibited respiration in isolated rat-brain mitochondria. Dimethyl sulphide was much less effective in inhibiting mitochondrial respiration. 4. In ox-heart submitochondrial particles ethanethiol inhibited electron transfer between cytochrome c and oxygen. 5. Purified cytochrome c oxidase was inhibited by ethanethiol in a non-competitive manner. 6. Methanethiol inhibited cytochrome c oxidase and was an effective inhibitor of mitochondrial electron transfer, both in liver and brain. 7. The difference in inhibitory properties between ethanethiol, methanethiol and dimethyl sulphide observed in our experiments coincides with the difference in potency to elicit coma in rats. We suggest that inhibition of mitochondrial electron transfer by mercaptans may be relevant to the mechanism by which energy production in brain is depressed during hepatic coma.

The site of inhibition of mitochondrial electron transfer by coenzyme Q analogues

The effects of 33 quinone derivatives on mitochondrial electron transfer in yeast were examined. Twenty-two of the compounds were also tested for their effects on the growth of yeast cells. Four strong inhibitors of electron transfer were identified: 5- n-undecyl-6-hydroxy-4, 7-dioxobenzothiazole, 7-ω-cyclohexyloctyl-6-hydroxy-5,8-quinolinequinone, 7- n-hexadecyl-mercapto-6-hydroxy-5, 8-quinolinequinone, and 3- n-dodecylmercapto-2-hydroxy-1, 4-naphthoquinone. They inhibit the growth of yeast with ethanol as an energy source, but not when glucose is the energy source. The NADH oxidase activity of isolated mitochondria is 50% inhibited by these quinone derivatives at about 10 −8 m, or 0.5 μmol/g mitochondrial protein; 1000-fold higher concentrations do not affect electron transfer from NADH or succinate to coenzyme Q 2. The effects of the inhibitors on cytochrome spectra indicate that they block electron transfer between cytochromes b and c 1. A possible antagonism between these compounds and coenzyme Q at a site between cytochromes b and C 1 is discussed in terms of Mitchell's “protonmotive Q cycle” hypothesis (Mitchell, P. (1976) J. Theor. Biol. 62, 327–367). 6-β-naphthylmercapto-5-chloro-2,3-dimethoxy-1,4-benzoquinone inhibits electron transfer between succinate and coenzyme Q 2 or phenazine methosulfate, suggesting a site in the succinate-coenzyme Q reductase complex with a different quinone specificity from that of the site in the cytochrome bc 1 complex. Seven of the quinone derivatives inhibit growth on both glucose and ethanol media, indicating that their effect is not the result of inhibition of respiration.