Biomedical subjects

Virginia Haynes

Publications and source records attributed to Virginia Haynes.

4 recordsLinked to original sources

PubMed2005

Differential requirements of calcium for oxoglutarate dehydrogenase and mitochondrial nitric-oxide synthase under hypoxia: impact on the regulation of mitochondrial oxygen consumption.

Studies with isolated mitochondria are performed at artificially high pO(2) (220 to 250 microM oxygen), although this condition is hyperoxic for these organelles. It was the aim of this study to evaluate the effect of hypoxia (20-30 microM) on the calcium-dependent activation of 2-oxoglutarate dehydrogenase (or 2-ketoglutarate dehydrogenase; OGDH) and mitochondrial nitric-oxide synthase (mtNOS). Mitochondria had a P/O value 15% higher in hypoxia than that in normoxia, indicating that oxidative phosphorylation and electron transfer were more efficiently coupled, whereas the intramitochondrial free calcium concentrations were higher (2-3-fold) at lower pO(2). These increases were abrogated by ruthenium red indicating that the higher uptake via the calcium uniporter was involved in this process. Mitochondria at high calcium concentration microdomains may produce nitric oxide, given the K(0.5) of calcium for OGDH (0.16 microM) and mtNOS (approximately 1 microM). Nitric oxide, by binding to cytochrome oxidase in competition with oxygen, decreases the rate of oxygen consumption. This condition is highly beneficial for the following reasons: i, these mitochondria are still able to produce ATP and support calcium clearance; ii, it prevents the accumulation of ROS by slowing the rate of oxygen consumption (hence ROS production); iii, the onset of anoxia is delayed, allowing oxygen to diffuse back to these sites, thereby ameliorating the oxygen gradient between regions of high and low calcium concentration. In this way, oxygen depletion at the latter sites is prevented. This, in turn, assures continued aerobic metabolism which may involve the activated dehydrogenases.

Animals↗

PubMed2004

Role of calcium signaling in the activation of mitochondrial nitric oxide synthase and citric acid cycle.

An apparent discrepancy arises about the role of calcium on the rates of oxygen consumption by mitochondria: mitochondrial calcium increases the rate of oxygen consumption because of the activation of calcium-activated dehydrogenases, and by activating mitochondrial nitric oxide synthase (mtNOS), decreases the rates of oxygen consumption because nitric oxide is a competitive inhibitor of cytochrome oxidase. To this end, the rates of oxygen consumption and nitric oxide production were followed in isolated rat liver mitochondria in the presence of either L-Arg (to sustain a mtNOS activity) or N(G)-monomethyl-L-Arg (NMMA, a competitive inhibitor of mtNOS) under State 3 conditions. In the presence of NMMA, the rates of State 3 oxygen consumption exhibited a K(0.5) of 0.16 microM intramitochondrial free calcium, agreeing with those required for the activation of the Krebs cycle. By plotting the difference between the rates of oxygen consumption in State 3 with L-Arg and with NMMA at various calcium concentrations, a K(0.5) of 1.2 microM intramitochondrial free calcium was obtained, similar to the K(0.5) (0.9 microM) of the dependence of the rate of nitric oxide production on calcium concentrations. The activation of dehydrogenases, followed by the activation of mtNOS, would lead to the modulation of the Krebs cycle activity by the modulation of nitric oxide on the respiratory rates. This would ensue in changes in the NADH/NAD and ATP/ADP ratios, which would influence the rate of the cycle and the oxygen diffusion.

Animals↗

PubMed2004

Mitochondrial nitric-oxide synthase: enzyme expression, characterization, and regulation.

Nitric oxide is generated in vivo by nitric-oxide synthase (NOS) during the conversion of L-Arg to citrulline. Using a variety of biological systems and approaches emerging evidence has been accumulated for the occurrence of a mitochondrial NOS (mtNOS), identified as the alpha isoform of neuronal or NOS-1. Under physiological conditions, the production of nitric oxide by mitochondria has an important implication for the maintenance of the cellular metabolism, i.e. modulates the oxygen consumption of the organelles through the competitive (with oxygen) and reversible inhibition of cytochrome c oxidase. The transient inhibition suits the continuously changing energy and oxygen requirements of the tissue; it is a short-term regulation with profound pathophysiological consequences. This review describes the identification of mtNOS and the role of posttranslational modifications on mtNOS' activity and regulation.

Adaptation, Physiological↗

PubMedDate not supplied

Mitochondrial nitric-oxide synthase: role in pathophysiology.

The biochemistry of the mitochondrial production of nitric oxide is reviewed to gain insight into the basic role of this radical in mitochondrial and cellular oxidative metabolism. The mitochondrial production of nitric oxide is catalyzed by a nitric-oxide synthase (mtNOS). This enzyme has the same cofactor and substrate requirements as other constitutive nitric-oxide synthases. Its occurrence was demonstrated in various mitochondrial preparations from different organs and species using diverse approaches (oxidation of oxymyoglobin, electron paramagnetic resonance in conjunction with spin trap, radiolabeled L-arginine, immunohistochemistry, nitric-oxide electrode). MtNOS has been identified as the alpha isoform of nNOS, acylated at a Thr or Ser residue, and phosphorylated at the C-terminal end. Endogenous nitric oxide reversibly inhibits oxygen consumption and ATP synthesis by competitive inhibition of cytochrome oxidase. Nitric oxide is the first molecule that fulfills the requirement for a cytochrome oxidase activity modulator: it is a competitive inhibitor, produced endogenously at a fair rate near the target site, at concentrations high enough to exhibit an inhibitory effect on cytochrome oxidase. The role of the mitochondrial nitric oxide production is discussed in terms of the physiological (modulating oxygen gradients into tissues) and pathological (abrogation of oxygen gradient modification, apoptosis, protein nitrative/oxidative stress) implications.

Animals↗