Reinstate hydrogen peroxide as the product of alternative oxidase of plant mitochondria

Chill treatment of potato tubers for 8 days induced mitochondrial O-2 consumption by cyanide-insensitive alternative oxidase (AOX). About half of the total O-2 consumption in such mitochondria was found to be sensitive to salicylhydroxamate (SHAM), a known inhibitor of AOX activity. Addition of catalase to the reaction mixture of AOX during the reaction decreased the rate of SHAM-sensitive O-2 consumption by nearly half, and addition at the end of the reaction released half of the O-2 consumed by AOX, both typical of catalase action on H2O2. This reaffirmed that the product of reduction of O-2 by plant AOX was H2O2 as found earlier and not H2O as reported in some recent reviews.

Ferrous-iron induces lipid peroxidation with little damage to energy transduction in mitochondria

Addition of ferrous sulfate, but not ferric chloride, in micromolar concentrations to rat liver mitochondria induced high rates of consumption of oxygen. The oxygen consumed was several times in excess of the reducing capacity of ferrous-iron (O: Fe ratios 5�8). This occurred in the absence of NADPH or any exogenous oxidizable substrate. The reaction terminated on oxidation of ferrous ions. Malondialdehyde (MDA), measured as thiobarbituric acid-reacting material, was produced indicating peroxidation of lipids. The ratio of O2: MDA was about 4: 1. Pretreatment of mitochondria with ferrous sulfate decreased the rate of oxidation (state 3) with glutamate (+malate) as the substrate by about 40% but caused little damage to energy tranduction process as represented by ratios of ADP: O and respiratory control, as well as calcium-stimulated oxygen uptake and energy-dependent uptake of [45Ca]-calcium. Addition of succinate or ubiquinone decreased ferrous iron-induced lipid peroxidation in intact mitochondria. In frozen-thawed mitochondria, addition of succinate enhanced lipid peroxidation whereas ubiquinone had little effect. These results suggest that ferrous-iron can cause peroxidation of mitochondrial lipids without affecting the energy transduction systems, and that succinate and ubiquinone can offer protection from damage due to such ferrous-iron released from the stores within the cells.

High level stable expression of rat brain type IIA sodium channel ?-subunit in CHO cells

The neuronal sodium channels are responsible for the rising phase of action potential and are composed of three subunits, of which the alpha-subunit has been shown to be adequate for most of its functional properties. We have stably expressed the rat brain type IIA sodium channel alpha-subunit in CHO cell tine using a CMV promoter-based vector. The expression was confirmed by detecting a 6.5 kb RNA corresponding to sodium channel alpha-subunit using Northern hybridization. The cells stably expressing the alpha-subunit, yield isolated sodium currents of amplitudes greater than 4nA when studied in whole-cell configuration of the patch-clamp technique. The sodium currents are characterized by activation and inactivation properties similar to neuronal sodium channels, and are blocked by the voltage gated sodium channel blocker tetrodotoxin (TTX).

Regulation of diacylglycerol kinase in rat brain membranes by docosahexaenoic acid

The effect of docosahexaenoic acid (DHA) on the diacylglycerol kinase (DG kinase) activity in rat brain membranes was investigated. DHA at 500 mu M concentration, stimulated the enzyme activity by about 2 fold. This effect was concentration-and time-dependent and was observed after very short periods of incubation (one min). DHA stimulation of DG kinase was observed only with rat brain membranes, and not with rat brain cytosol or rat liver membranes. Treating the rat brain membranes with phospholipase A(2) which released free fatty acids including DHA, significantly stimulated the DG kinase activity. It is concluded that DHA through its stimulatory effect on DG kinase may regulate the signalling events in growth-related situations in the brain such as synaptogenesis.

Understanding How the Brain Changes Its Mind: Microstimulation in the Macaque Frontal Eye Field Reveals How Saccade Plans Are Changed

Accumulator models that integrate incoming sensory information into motor plans provide a robust framework to understand decision making. However, their applicability to situations that demand a change of plan raises an interesting problem for the brain. This is because interruption of the current motor plan must occur by a competing motor plan, which is necessarily weaker in strength. To understand how changes of mind get expressed in behavior, we used a version of the double-step task called the redirect task, in which monkeys were trained to modify a saccade plan. We microstimulated the frontal eye fields during redirect behavior and systematically measured the deviation of the evoked saccade from the response field to causally track the changing saccade plan. Further, to identify the underlying mechanisms, eight different computational models of redirect behavior were assessed. It was observed that the model that included an independent, spatially specific inhibitory process, in addition to the two accumulators representing the preparatory processes of initial and final motor plans, best predicted the performance and the pattern of saccade deviation profile in the task. Such an inhibitory process suppressed the preparation of the initial motor plan, allowing the final motor plan to proceed unhindered. Thus, changes of mind are consistent with the notion of a spatially specific, inhibitory process that inhibits the current inappropriate plan, allowing expression of the new plan.

Phase synchronization in brain networks derived from correlation between probabilities of recurrences in functional MRI data

It is increasingly being recognized that resting state brain connectivity derived from functional magnetic resonance imaging (fMRI) data is an important marker of brain function both in healthy and clinical populations. Though linear correlation has been extensively used to characterize brain connectivity, it is limited to detecting first order dependencies. In this study, we propose a framework where in phase synchronization (PS) between brain regions is characterized using a new metric ``correlation between probabilities of recurrence'' (CPR) and subsequent graph-theoretic analysis of the ensuing networks. We applied this method to resting state fMRI data obtained from human subjects with and without administration of propofol anesthetic. Our results showed decreased PS during anesthesia and a biologically more plausible community structure using CPR rather than linear correlation. We conclude that CPR provides an attractive nonparametric method for modeling interactions in brain networks as compared to standard correlation for obtaining physiologically meaningful insights about brain function.

Cytochrome P450-mediated metabolism in brain: functional roles and their implications

Introduction: Cytochromes P450 (P450) and associated monooxygenases are a family of heme proteins involved in metabolism of endogenous compounds (arachidonic acid, eicosanoids and prostaglandins) as also xenobiotics including drugs and environmental chemicals. Liver is the major organ involved in P450-mediated metabolism and hepatic enzymes have been characterized. Extrahepatic organs, such as lung, kidney and brain have the capability for biotransformation through P450 enzymes. Brain, including human brain, expresses P450 enzymes that metabolize xenobiotics and endogenous compounds. Areas covered: An overview of P450-mediated metabolism in brain is presented focusing on distinct differences seen in expression of P450 enzymes, generation of unique P450 enzymes in brain through alternate splicing and their consequences in terms of metabolism of psychoactive drugs and inflammatory prompts, such as leukotrienes, thus modulating inflammatory response. Expert opinion: The brain possesses unique P450s that metabolize drugs and endogenous compounds through pathways that are markedly different from that seen in liver indicating that extrapolation directly from liver to brain is not appropriate. It is therefore necessary to characterize the unique brain P450s and their ability to metabolize xenobiotics and endogenous compounds to better understand the functions of this important class of enzymes in brain, especially human brain.

Antioxidant and oxidative stress parameters in brain of Heteropneustes fossilis under air exposure condition; role of mitochondrial electron transport chain

Many fishes are exposed to air in their natural habitat or during their commercial handling. In natural habitat or during commercial handling, the cat fish Heteropneustes fossilis is exposed to air for > 24 h. Data on its oxidative metabolism in the above condition are not available. Oxidative stress (OS) indices (lipid and protein oxidation), toxic reactive oxygen species (ROS: H2O2) generation, antioxidative status (levels of superoxide dismutase, catalase, glutathione peroxidase and reductase, ascorbic acid and nonprotein sulfhydryl) and activities of electron transport chain (ETC) enzymes (complex I-IV) were investigated in brain tissue of H. fossilis under air exposure condition (0, 3, 6, 12 and 18 h at 25 degrees C). Decreased activities of antioxidant (except catalase) and ETC enzymes (except complex II) with increased H2O2 and OS levels were observed in the tissue under water deprivation condition. Positive correlation was observed for complex II activity and non-protein thiol groups with time period of air exposure. The critical time period to induce OS and to reduce most of the studied antioxidant level in brain was found to be 3-6 h air exposure. The data can be useful to minimize the stress generated during commercial handling of the live fishes those exposed to air in general and H. fossilis in particular. (C) 2013 Elsevier Inc. All rights reserved.

Mitochondria-Targeting Oxidovanadium(IV) Complex as a Near-IR Light Photocytotoxic Agent

Oxidovanadium(IV) complexes VO(L-1)(phen)]Cl (1) and VO(L-2)(L-3)]Cl (2), in which HL1 is 2-{(benzimidazol-2-yl)methylimino]-methyl}phenol (sal-ambmz), HL2 is 2-({1-(anthracen-9-yl)methyl]-benzimidazol-2-yl}methylimino)-met hyl]phenol (sal-an-ambmz), phen is 1,10-phenanthroline and L-3 is dipyrido3,2-a:2,3-c]phenazine (dppz) conjugated to a Gly-Gly-OMe dipeptide moiety, were prepared, characterized, and their DNA binding, photoinduced DNA-cleavage, and photocytotoxic properties were studied. Fluorescence microscopy studies were performed by using complex 2 in HeLa and HaCaT cells. Complex 1, structurally characterized by X-ray crystallography, has a vanadyl group in VO2N4 core with the VO2+ moiety bonded to N,N-donor phen and a N,N,O-donor Schiff base. Complex 2, having an anthracenyl fluorophore, showed fluorescence emission bands at 397, 419, and 443nm. The complexes are redox-active exhibiting the V(IV)/V(III) redox couple near -0.85V versus SCE in DMF 0.1M tetrabutylammonium perchlorate (TBAP). Complex 2, having a dipeptide moiety, showed specific binding towards poly(dAdT)(2) sequence. The dppz-Gly-Gly-OMe complex showed significant DNA photocleavage activity in red light of 705nm through a hydroxyl radical ((OH)-O-.) pathway. Complex 2 showed photocytotoxicity in HaCaT and HeLa cells in visible light (400-700nm) and red light (620-700nm), however, the complex was less toxic in the dark. Fluorescence microscopy revealed the localization of complex 2 primarily in mitochondria. Apoptosis was found to occur inside mitochondria (intrinsic pathway) caused by ROS generation.

Effects of temperature on complexes I and II mediated respiration, ROS generation and oxidative stress status in isolated gill mitochondria of the mud crab Scylla serrata

Effects of fluctuations in habitat temperature (18-30 degrees) on mitochondrial respiratory behavior and oxidative metabolic responses in the euryhaline ectotherm Scylla serrate are not fully understood. In the present study, effects of different temperatures ranging from 12 to 40 degrees C on glutamate and succinate mediated mitochondrial respiration, respiratory control ratio (RCR), ATP generation rate, ratio for the utilization of phosphate molecules per atomic oxygen consumption (P/O), levels of lipid peroxidation and H2O2 in isolated gill mitochondria of S. serrata are reported. The pattern of variation in the studied parameters was similar for the two substrates at different temperatures. The values recorded for RCR ( >= 3) and P/O ratio (1.4-2.7) at the temperature range of 15-25 degrees C were within the normal range reported for other animals (3-10 for RCR and 1.5-3 for P/O). Values for P/O ratio, ATP generation rate and RCR were highest at 18 degrees C when compared to the other assay temperatures. However, at low and high extreme temperatures, i.e. at 12 and 40 degrees C, states III and IV respiration rates were not clearly distinguishable from each other indicating that mitochondria were completely uncoupled. Positive correlations were noticed between temperature and the levels of both lipid peroxidation and H2O2. It is inferred that fluctuations on either side of ambient habitat temperature may adversely influence mitochondria respiration and oxidative metabolism in S. serrata. The results provide baseline data to understand the impacts of acute changes in temperature on ectotherms inhabiting estuarine or marine environments. (C) 2014 Elsevier Ltd. All rights reserved.

Mitochondria-Targeted Photoinduced Anticancer Activity of Oxidovanadium( IV) Complexes of Curcumin in Visible Light

Oxidovanadium(IV) complexes VO(py-aebmz)(B)]Cl (1, 2) and VO(napth-py-aebmz)(cur)]Cl 3; py-aebmz = 2-(1H-benzimidazol-2-yl)-N-(pyridin-2-ylmethylene)ethanamine, HB = acetylacetone (Hacac, 1) and curcumin (Hcur, 2), napth-py-aebmz = naphthalimide conjugated to py-aebmz ] have been prepared, characterized and their photoinduced DNA cleavage activities and photocytotoxicities studied. Complexes 1-3 each exhibited an irreversible cyclic voltammetric response of the V-IV/V-III redox couple at around -0.85 V versus SCE in dmf/0.1 M tbap. The complexes showed DNA photocleavage activity in visible light of 454, 530 and 647 nm through hydroxyl radical and singlet oxygen pathways. Fluorescence microscopy data suggest mitochondrial localization of complex 3 bearing a naphthalimide with a two-fold increase in photocytotoxicity in HaCaT cells with an IC50 value of 6.3 M and a three-fold increase in MCF-7 cells with an IC50 of 5.4 M compared with complex 2. Both 2 and 3 were non-toxic in the dark.