Mode of action of oxathiin systemic fungicides. V. Effect on electron transport system of Ustilago maydis and Saccharomyces cerevisiae

The systemic fungicide carboxin (5,6-dihydro-2-methyl-1,4-oxathiin-3-carboxanilide) at 100 mum inhibited succinate cytochrome c reductase in mitochondria from Ustilago maydis and Saccharomyces cerevisiae. It did not have any effect on reduced nicotinamide adenine dinucleotide (NADH) cytochrome c reductase. Succinate coenzyme Q reductase was also inhibited, but NADH coenzyme Q reductase was not. When dichlorophenolindophenol (DCIP) was used as the terminal acceptor of electrons from the oxidation of succinate, carboxin was very effective in inhibiting succinate-DCIP reductase. Carboxin was inhibitory to succinic dehydrogenase assayed with phenazine methosulfate plus DCIP when intact mitochondria were used as the enzyme source but not when solubilized enzyme was used. The main site of action of carboxin, therefore, appears to lie between succinate and coenzyme Q. The dioxide analogue of carboxin was also effective in inhibiting succinate-cytochrome c reductase, succinate-coenzyme Q reductase, or succinate-DCIP reductase, whereas the monoxide analogue was less effective in inhibiting these enzymes.     

Decline in cytochrome c oxidase activity in rat-brain mitochondria with aging. Role of peroxidized cardiolipin and beneficial effect of melatonin

Reactive oxygen species (ROS) are considered a key factor in mitochondrial dysfunction associated with brain aging process. Mitochondrial respiration is an important source of ROS and hence a potential contributor to brain functional changes with aging. In this study, we examined the effect of aging on cytochrome c oxidase activity and other bioenergetic processes such as oxygen consumption, membrane potential and ROS production in rat brain mitochondria. We found a significant age-dependent decline in the cytochrome c oxidase activity which was associated with parallel changes in state 3 respiration, membrane potential and with an increase in H₂O₂ generation. The cytochrome aa₃ content was practically unchanged in mitochondria from young and aged animals. The age-dependent decline of cytochrome c oxidase activity could be restored, in situ, to the level of young animals, by exogenously added cardiolipin. In addition, exposure of brain mitochondria to peroxidized cardiolipin resulted in an inactivation of this enzyme complex. It is suggested that oxidation/depletion of cardiolipin could be responsible, at least in part, for the decline of cytochrome c oxidase and mitochondrial dysfunction in brain aging. Melatonin treatment of old animals largely prevented the age-associated alterations of mitochondrial bioenergetic parameters. These results may prove useful in elucidating the molecular mechanisms underlying mitochondrial dysfunction associated with brain aging process, and may have implications in etiopathology of age-associated neurodegenerative disorders and in the development of potential treatment strategies.     

An antimycin-insensitive succinate-cytochrome c reductase activity in pure reconstitutively active succinate dehydrogenase

Antimycin-insensitive succinate-cytochrome c reductase activity has been detected in pure, reconstitutively active succinate dehydrogenase. The enzyme catalyzes electron transfer from succinate to cytochrome c at a rate of 0.7 mumole succinate oxidized per min per mg protein, in the presence of 100 microM cytochrome c. This activity, which is about 2% of that of reconstitutive (the ability of succinate dehydrogenase to reconstitute with coenzyme ubiquinone-binding proteins (QPs) to form succinate-ubiquinone reductase) or succinate-phenazine methosulfate activity in the preparation, differs from antimycin-insensitive succinate-cytochrome c reductase activity detected in submitochondrial particles or isolated succinate-cytochrome c reductase. The Km for cytochrome c for the former is too high to be measured. The Km for the latter is about 4.4 microM, similar to that of antimycin-sensitive succinate-cytochrome c activity in isolated succinate-cytochrome c reductase, suggesting that antimycin-insensitive succinate-cytochrome c activity of succinate-cytochrome c reductase probably results from incomplete inhibition by antimycin. Like reconstitutive activity of succinate dehydrogenase, the antimycin-insensitive succinate-cytochrome c activity of succinate dehydrogenase is sensitive to oxygen; the half-life is about 20 min at 0 degrees C at a protein concentration of 23 mg/ml. In the presence of QPs, the antimycin-insensitive succinate-cytochrome c activity of succinate dehydrogenase disappears and at the same time a thenoyltrifluoroacetone-sensitive succinate-ubiquinone reductase activity appears. This suggests that antimycin-insensitive succinate-cytochrome c reductase activity of succinate dehydrogenase appears when succinate dehydrogenase is detached from the membrane or from QPs. Reconstitutively active succinate dehydrogenase oxidizes succinate using succinylated cytochrome c as electron acceptor, suggesting that a low potential intermediate (radical) may be involved. This suggestion is confirmed by the detection of an unknown radical by spin trapping techniques. When a spin trap, alpha-phenyl-N-tert-butylnitrone (PBN), is added to a succinate oxidizing system containing reconstitutively active succinate dehydrogenase, a PBN spin adduct is generated. Although this PBN spin adduct is identical to that generated by xanthine oxidase, indicating that a perhydroxy radical might be involved, the insensitivity of this antimycin-insensitive succinate-cytochrome c reductase activity to superoxide dismutase and oxygen questions the nature of this observed radical.     

Effects of L-malate on mitochondrial oxidoreductases in liver of aged rats

Accumulation of oxidative damage has been implicated to be a major causative factor in the decline in physiological functions that occur during the aging process. The mitochondrial respiratory chain is a powerful source of reactive oxygen species (ROS), considered as the pathogenic agent of many diseases and aging. L-malate, a tricarboxylic acid cycle intermediate, plays an important role in transporting NADH from cytosol to mitochondria for energy production. Previous studies in our laboratory reported L-malate as a free radical scavenger in aged rats. In the present study we focused on the effect of L-malate on the activities of electron transport chain in young and aged rats. We found that mitochondrial membrane potential (MMP) and the activities of succinate dehydrogenase, NADH-cytochrome c oxidoreductase and cytochrome c oxidase in liver of aged rats were significantly decreased when compared to young control rats. Supplementation of L-malate to aged rats for 30 days slightly increased MMP and improved the activities of NADH-dehydrogenase, NADH-cytochrome c oxidoreductase and cytochrome c oxidase in liver of aged rats when compared with aged control rats. In young rats, L-malate administration increased only the activity of NADH-dehydrogenase. Our result suggested that L-malate could improve the activities of electron transport chain enzymes in aged rats.     

Effect of Aging on the Metabolism of Pyruvate and 3-Hydroxybutyrate in Nonsynaptic and Synaptic Mitochondria from Rat Brain

Abstract: Age-dependent changes in the oxidative metabolism in nonsynaptic and synaptic mitochondria from brains of 3, 12, and 24-month-old rats were investigated. When pyruvate and malate were used in conjunction as substrates, a significant reduction in State 3 respiration was observed in both mitochondrial populations from 12-and 24-month-old rats compared with 3-month-old animals. A similar age-dependent reduction in the oxidation of [1-¹¹C]pyruvate was also observed in nonsynaptic and synaptic mitochondria from senescent rats. Pyruvate dehydrogenase complex activity (both active and total) was, however, not decreased in the two mitochondrial populations from brains of 3, 12, and 24-month-old rats. When DL-3-hydroxybutyrate plus malate were used as substrates, a decrease in State 3 respiration was observed only in synaptic mitochondria from 24-month-old rats compared with 3- month-old animals. Similarly, an age-dependent reduction in the oxidation of 3-hydroxy[3-¹¹C]butyrate was also observed only in synaptic mitochondria from 12-and 24-month-old rats. However, a significant reduction in the activities of ketone body-metabolizing enzymes, namely, 3-hydroxybutyrate dehydrogenase, 3-ketoacid CoA transferase, and acetoacetyl-CoA thiolase was observed in both mitochondrlal populations from 12- and 24-month-old rats compared with 3 month-old animals. These findings show that specific alterations in oxidative metabolism occur in nonsynaptic and synaptic mitochondria from aging rats. The data also suggest that in addition to alterations in enzyme activities, permeability of anions (e.g. pyruvate) across the inner mitochondrial membrane may be altered in nonsynaptic and synaptic mitochondria from senescent animals.     

Action ofl-acetylcarnitine on different cerebral mitochondrial populations from hippocampus and striatum during aging

The maximum rates (V_max) of some mitochondrial enzyme activities related to energy transduction (citrate synthase, malate dehydrogenase, NADH cytochrome c reductase, cytochrome oxidase) and amino acid metabolism (glutamate dehydrogenase) were evaluated in non-synaptic (free) and synaptic mitochondria from rat hippocampus and striatum. Three types of mitochondria were isolated from control rats aged 4, 8, 12, 16, 20 and 24 months and treated ones withl-acetylcarnitine (100 mg·kg^–1, i.p., 60 min). Enzyme activities of non-synaptic and synaptic mitochondria are different in hippocampus and striatum., confirming that a different metabolic machinery exists in various types of brain mitochondria. During aging, enzyme activities behave quite similarly in both areas. In vivo administration ofl-acetylcarnitine decreased the enzyme activities related to Krebs' cycle mainly of synaptic mitochondria, suggesting a specific subcellular trigger site of action. The drug increased cytochrome oxidase activity of synaptic and non-synaptic mitochondria, indicating the specificity of molecular interaction with this enzyme.     

Biochemical effects of PR toxin on rat liver mitochondrial respiration and oxidative phosphorylation

The in vitro effects of PR toxin, a toxic secondary metabolite produced by certain strains of Penicillium roqueforti, on the membrane structure and function of rat liver mitochondria were investigated. It was found that the respiratory control and oxidative phosphorylation of the isolated mitochondria decreased concomitantly when the toxin was added to the assay system. The respiratory control ratio decreased about 60% and the ADP/O ratio decreased about 40% upon addition of 3.1 X 10(-5) M PR toxin to the highly coupled mitochondria. These findings suggest that PR toxin impairs the structural integrity of mitochondrial membranes. On the other hand, the toxin inhibited mitochondrial respiratory functions. It exhibited noncompetitive inhibitions to succinate oxidase, succinate-cytochrome c reductase, and succinate dehydrogenase activities of the mitochondrial respiratory chain. The inhibitory constants of PR toxin to these three enzyme systems were estimated to be 5.1 X 10(-6), 2.4 X 10(-5), and 5.2 X 10(-5) M, respectively. Moreover, PR toxin was found to change the spectral features of succinate-reduced cytochrome b and cytochrome c1 in succinate-cytochrome c reductase and inhibited the electron transfer between the two cytochromes. These observations indicate that the electron transfer function of succinate-cytochrome c reductase was perturbed by the toxin. However, PR toxin did not show significant inhibition of either cytochrome oxidase or NADH dehydrogenase activity of the mitochondria. It is thus concluded that PR toxin exerts its effect on the mitochondrial respiration and oxidative phosphorylation through action on the membrane and the succinate-cytochrome c reductase complex of the mitochondria.     

Oxidation of succinate in heart, brain, and kidney mitochondria in hypobaria and hypoxia.

Exposure of rats to hypobaric stress for periods of up to 36 h caused a consistent change in the succinate-NT reductase activity of the heart mitochondria whereas there was no significant change in the activities of either succinate dehydrogenase and succinate-NT reductase of the brain and the kidney. Mitochondrial succinate dehydrogenase of the heart, the brain and the kidney was activated 2- to 7-fold with the substrate and malonate. The activations obtained with oxalate, citrate and dinitrophenol were relatively lower in comparison to succinate and malonate. Benzohydroquinone and 2-nitrophenol had no stimulatory effect on the heart, the brain and the kidney mitochondria. THE ACTIVATIONS OBTAINED WITH THE VARIOUS EFFECTORS PARTIALLY (OR COMPLETELY IN THE CASE OF SUCCINATE) REVERSED ON WASHING THE MITOCHONDRIAL SAMPLES WITH THE SUCROSE HOMOGENIZING MEDIUM. The effect of ubiquinol, which also activated the enzyme, was only partially reversed after the second preincubation with succinate in the brain and the kidney whereas in the heart the activity was fully reversed. The increased activity of succinate dehydrogenase obtained with ATP and ADP was further enhanced by Mg2+ exclusively in the brain mitochondria, suggesting the possibility of Mg2+-AIP complex as the active species. Succinate-NT reductase of the heart, the brain and the kidney mitochondria showed a high activation with ubiquinone whereas its reduced form had no stimulatory effect.     

Enzymatic and immunologic study of the role of the thyroid hormone in the formation of the internal mitochondrial membrane during postnatal development of the rat

Hypothyroidism induces an increase of liver D-beta-hydroxybutyrate dehydrogenase activity. Injection of thyroid hormone reverses the phenomena. The use of monospecific antibody raised against the purified enzyme indicates that there was not an increase of apoenzyme biosynthesis. The thyroid hormone negative control is due to a metabolism alteration of the membrane phospholipids which are directly involved in the apoenzyme activity. The highest difference is observed with 20 days old rats. Opposite effects were obtained on succinate cytochrome c reductase.     

Effect of organophosphorus insecticides on the oxidative processes in rat brain synaptosomes

Abstract: This paper describes the effect of four organophosphorus insecticides: Dipterex, DDVP, Ronnel and its oxygen analogue on the respiration of rat brain synaptosomes. Dipterex and DDVP in the concentrations used, 5, 50, or 500 μM, did not change the rate of oxygen uptake and oxidative phosphorylation in rat brain synaptosomes. Ronnel in the highest concentration (500 μM) inhibited respiration in state 3 conditions and abolished respiratory control by ADP. This inhibition was correlated with a change of cytochrome c oxidase activity. The oxygen analogue of Ronnel (OAR) in micromolar concentrations (50 μM) increased the rate of respiration of synaptosomes utilizing glutamate plus malate as substrate. Higher concentrations of OAR produced inhibition of respiration, cytochrome c oxidase and NADH: cytochrome c reductase activities. These observations are typical for uncouplers of oxidative phosphorylation. Noteworthy is the fact that the uncoupling activity of OAR was observed at concentrations which did not inhibit acetylcholinesterase activity. These findings seem to suggest that disturbances in oxidative processes could play an important role in the toxicity of organophosphorus insecticides. The relation between chemical structure and the ability of insecticides to affect oxidative phosphorylation is discussed.     

Influence of increased environmental temperature on oxidation processes in rat liver mitochondria

Exposure of rats to elevated temperature of 28 degrees C or 35 degrees C for 3 days six hours daily resulted in a decreased rate of oxidation with succinate or glutamate + malate as substrates, by the mitochondria of liver. The higher decrease was observed in environment temperature of 35 degrees C. There was no change in ADP/O ratio. The activities of NADH: cytochrome c reductase and cytochrome oxidase were stimulated but activities of succinate dehydrogenase and succinate cytochrome reductase were decreased.     

Mitochondria alterations in Cd2+-treated rats: general regression of inner membrane cristae and electron transport impairment

Young adult rats absorbed 50 p.p.m. Cd2+ added to drinking water. After 6 weeks, 3, 6 and 9 months of treatment, the ultrastructural condition of liver, kidney and muscle was observed by electron microscopy. The choice of these tissues was determined by their differences in the capacity to accumulate Cd2+: the liver is able to concentrate a considerable amount of metal, but redistributes it throughout the entire organism, while the kidney collects it in view of its elimination. Muscle contains the least Cd2+. A general regression in mitochondria cristae accompanied by a vesiculation and a fragmentation of endoplasmic reticulum appeared simultaneously in the three tissues, at as early as 6 weeks of treatment, and extended progressively with its continuation supporting evidence of a general attack of the intracellular membrane systems. Cd2+ stimulation of membrane-degrading enzymes such as phospholipases and proteases was suggested. A concomitant diminution in glycogen stores was noted. Active synthesis of neutral lipids, especially cholesterol esters, took place in liver mitochondria of treated rats in collaboration with rough endoplasmic reticulum, and progressively generated a multiplication of electron-transparent inclusions in cytoplasm. Isolated mitochondria from liver, kidney and muscle of Cd2+-treated rats maintained partial energy coupling, but displayed a rapid early fall in cytochrome oxidase followed by a partial restoration after 6 months of treatment, and a progressively slackening of succinate dehydrogenase. Isolated vesicles of liver mitochondria inner membrane of treated rats behaved as intact mitochondria, indicating changes inside the membrane itself. Addition in vitro of the metal ion to mitochondria and also to inner membrane vesicles isolated from control rats revealed that Cd2+ was able to stop completely succinate dehydrogenase, but was totally ineffective on cytochrome oxidase. Membrane fixation of Cd2+ on the flavoprotein or SH associated with succinate dehydrogenase is proposed. Considering the close parallelism of the extensive depression of microsomal NADPH cytochrome c reductase and the rapid fall in mitochondrial cytochrome oxidase, it is suggested that an indirect inhibition process occurs, through Cd2+-induced diminution of a constituent common to all cytochromes in the cell.     

The regulation of glycerol 3-phosphate oxidase of rat brownadipose tissue mitochondria by long-chain free fatty acids

Added free fatty acids inhibit oxidation of glycerol 3-phosphate, succinate and NADH in brown-adipose tissue mitochondria from 10-day-old rats. The most pronounced is the inhibitory effect of glycerol 3-phosphate cytochrome c reductase (GP-cyto. c reductase). Contrary to other reductases, GP-cyto. c reductase activity of freshly isolated mitochondria is already inhibited by the fraction of endogenous free fatty acids. Both added and endogenous free fatty acids inhibition of GP-cyto. c reductase is fully reversible by the removal of free fatty acids by bovine serum albumine treatment. The inhibition of GP-cyto. c reductase is of strictly non-competitive type. The most inhibitory are unsaturated long-chain free fatty acids-oleic and linoleic acid. Results are discussed with regards to the regulatory importance of free fatty acids in brown-adiposetissue during intensive non-shivering thermogenesis.     

Development of glial cells in primary cultures: Energy metabolism and lactate dehydrogenase isoenzymes

Primary cultures of glial cells prepared from brains of newborn rats were grown for periods of 1–5 weeks. After a proliferative phase of between 2 and 3 weeks, the cultures were maintained in stationary phase, during which a significant increase of oxygen consumption and of the activities of lactate dehydrogenase, succinate dehydrogenase, and mitochondrial glycerolphosphate dehydrogenase could be observed. Furthermore, qualitative changes in the lactate dehydrogenase isoenzyme pattern were found with time, characterized by a shift toward an enhanced synthesis of H subunits. A similar development was found in comparing the LDH isoenzyme pattern in the brain of 15-day-old rat embryo with those of newborn and adult rat brains. It is suggested that some aspects of maturation of glial cells in culture are comparable to those occurring in whole brain in vivo, namely a shift towards an enhanced aerobic metabolism.     

The influence of storage temperature on the transition, activation enthalpy, and activity of enzymes associated with inner mitochondrial membranes

The effects of storage at low temperature on the transition in enzyme function, Tf*, and the Arrhenius activation energy, Ea, were determined for several enzymes associated with the inner membrane of rat liver mitochondria. The enzymes studied were succinate:cytochrome c reductase, cytochrome c oxidase, beta-hydroxybutyrate dehydrogenase, and oligomycin-sensitive, Mg2+-activated ATPase. For freshly isolated mitochondria the Tf*, for succinate:cytochrome c reductase and cytochrome c oxidase, occurred at approximately 23 degrees C and was coincident with a transition in structure, Ts*, determined as the change in temperature coefficient of motion for a spin label intercalated with the membrane lipids. This suggest that the change in thermal response of the membrane-associated enzymes is related to a change in molecular ordering of the membrane lipids. When mitochondria were stored at -12 degrees C, the specific activities of succinate:cytochrome c reductase and cytochrome c oxidase decreased. Concomitant with these changes the Ea, above Tf*, increased. After 100 days storage at -12 degrees C, Ea above Tf* approached the value for Ea below Tf* such that the transition in thermal response could no longer be detected. In contrast, for mitochondria stored at -196 degrees C, although the specific activity declined over the 100 days storage, no changes in either Ea or Tf* were evident. The results indicate a need for caution in evaluating comparative studies of Tf and Ea, for membrane-associated enzymes, using mitochondria which have been frozen and stored.     

Enzyme, electron microscopic and polarographic characteristics of isolated rat brain mitochondria. III. Quantitative assessment of their distribution in fractions of the homogenate]

Activities of succinate dehydrogenase, succinate- and NAD-H-cytochrome c--reductases, and cytochrome c--oxidase was compared in 1 g tissue homogenate and homogenate fractions made from 1 g brain tissue using various solutions. Fractionation resulted in the increased activities of NADH- and succinate cytochrome reductases, and in the loss of succinate dehydrogenase activity, cytochrome oxidase was less influenced. These phenomena are regarded as signs of the interrelation between mitochondria and other constituents of brain cell within homogenates. Maximal quantity of mitochondria isolated from homogenates is no more than 20% of all the mitochondrial homogenates (according enzyme data). The electronogram of the brain mitochondrial preparation isolated in the Krebs--Ringer solution without glucose pointed out to a high homogeneity of mitochondria in the residue.     

The effect of ring substituents on the mechanism of interaction of exogenous quinones with the mitochondrial respiratory chain

In uncoupled pig-heart mitochondria the rate of the reduction of duroquinone by succinate in the presence of cyanide is inhibited by about 50% by antimycin. This inhibition approaches completion when myxothiazol is also added or British anti-Lewisite-treated (BAL-treated) mitochondria are used. If mitochondria are replaced by isolated succinate:cytochrome c oxidoreductase, the inhibition by antimycin alone is complete. The reduction of a plastoquinone homologue with an isoprenoid side chain (plastoquinone-2) is strongly inhibited by antimycin with either mitochondria or succinate:cytochrome c reductase. The reduction by succinate of plastoquinone analogues with an n-alkyl side chain in the presence of mitochondria is inhibited neither by antimycin nor by myxothiazol, but is sensitive to the combined use of these two inhibitors. On the other hand, the reduction of the ubiquinone homologues Q2, Q4, Q6 and Q10 and an analogue, 2,3-dimethoxyl-5-n-decyl-6-methyl-1,4-benzoquinone, is not sensitive to any inhibitor of QH2:cytochrome c reductase tested or their combined use, either in normal or BAL-treated mitochondria or in isolated succinate:cytochrome c reductase. It is concluded that quinones with a ubiquinone ring can be reduced directly by succinate:Q reductase, whereas those with a plastoquinone ring can not. Reduction of the latter compounds requires participation of either center i or center o (Mitchell, P. (1975) FEBS Lett. 56, 1-6) or both, of QH2:cytochrome c oxidoreductase. It is proposed that a saturated side chain promotes, while an isoprenoid side chain prevents reduction of these compounds at center o.     

Oxidation of acetoacetate and palmitylcarnitine by brain and liver mitochondria from suckling and adult rats

Experiments in which we investigated the possible oxidative utilization of lipoid substrates by brain and liver mitochondria were carried out with rats aged 5 and 90 days, kept under completely standardized conditions. Brain mitochondria were isolated on a Ficoll gradient after Clark and Nicklas (1970). Respiratory activity (or the respiratory control index-R.C.) was determined in the manner described in an earlier paper (Dobesová and Mourek 1980). Na succinate or Na malate was used as the testing substrate; palmityl carnitine, acetyl carnitine and acetoacetate were used as lipoid substrates. Oxygen consumption was measured with a Clark's oxygen electrode and respiration was expressed in nAt oxygen per min per mg protein, which was measured by the method of Lowry et al. (1951). When using succinate or malate, in agreement with our previous results we did not find any development changes in the respiratory activity of the brain mitochondria. The oxidation of acetoacetate by the brain mitochondria of 5-day-old rats was about five times greater, and of acetyl carnitine over two times greater, compared with the CNS mitochondria of adult rats. The oxidative utilization of lipoid substrates by the liver mitochondria of 5-day-old rats was significantly greater than their utilization by CNS mitochondria (in the case of palmityl carnitine three times greater, for example) and was always significantly greater than in the liver mitochondria of adult rats. We demonstrated that mitochondria isolated from the brain of 5-day-old rats are equipped with an enzymatic apparatus which allows them to utilize lipoid substrates on a significantly greater scale than in adulthood.     

Reduction of cytochrome b in mitochondria from yeast lacking coenzyme Q

Mitochondria isolated from coenzyme Q deficient yeast cells had no detectable NADH:cytochrome c reductase or succinate:cytochrome c reductase but had comparable amounts of cytochromes b and c1 as wild-type mitochondria. Addition of succinate to the mutant mitochondria resulted in a slight reduction of cytochrome b; however, the subsequent addition of antimycin resulted in a biphasic reduction of cytochrome b, leading to reduction of 68% of the total dithionite-reducible cytochrome b. No "red" shift in the absorption maximum was observed, and no cytochrome c1 was reduced. The addition of either myxothiazol or alkylhydroxynaphthoquinone blocked the reduction of cytochrome b observed with succinate and antimycin, suggesting that the reduction of cytochrome b-562 in the mitochondria lacking coenzyme Q may proceed by a pathway involving cytochrome b at center o where these inhibitors block. Cyanide did not prevent the reduction of cytochrome b by succinate and antimycin the the mutant mitochondria. These results suggest that the succinate dehydrogenase complex can transfer electrons directly to cytochrome b in the absence of coenzyme Q in a reaction that is enhanced by antimycin. Reduced dichlorophenolindophenol (DCIP) acted as an effective bypass of the antimycin block in complex III, resulting in oxygen uptake with succinate in antimycin-treated mitochondria. By contrast, reduced DCIP did not restore oxygen uptake in the mutant mitochondria, suggesting that coenzyme Q is necessary for the bypass. The addition of low concentrations of DCIP to both wild-type and mutant mitochondria reduced with succinate in the presence of antimycin resulted in a rapid oxidation of cytochrome b perhaps by the pathway involving center o, which does not require coenzyme Q.     

Age-Related Changes in Aldehyde Dehydrogenase Activity of Rat Brain, Liver, and Heart

Abstract: Aldehyde dehydrogenase (ALDH) activity was measured in brains, livers, and hearts of 23–26-month-old and 3-month-old rats. A significant increase of ALDH activity was found in whole brain of old rats with both acetaldehyde (39%) and propionylaldehyde (15%) used as substrates. In different brain areas of old rats, with acetaldehyde used as substrate, a significant increase of ALDH activity was found in striatum (30–50%) and cerebral cortex (37%). However, no significant difference in ALDH activity was found in livers and hearts of young and old rats. Preliminary experiments showed a significant increase of aldehyde reductase activity (52%) with p -nitrobenzaldehyde used as substrate in whole brain of old rats compared with young rats. The present work indicates that an increase of ALDH activity in brain of old rats may be an adaptive phenomenon.