Characterization of Superoxide-producing Sites in Isolated Brain Mitochondria

Because short-term incubation of rat brain mitochondria with H2O2 induced increased H2O2 production at this site we propose that reactive oxygen species can activate a selfaccelerating vicious cycle causing mitochondrial damage and neuronal cell death

Alexei P. Kudin; Nana Yaw-B. Bimpong-Buta; Stefan Vielhaber; Christian E. Elger; Wolfram S. Kunz

2004

Scholarcy highlights

  • Idence that superoxide and H2O2 contribute to the pathogenesis of certain neurodegenerative diseases
  • Because it is well documented that mitochondrial superoxide production is strongly dependent on oxygen concentration we determined the oxygen concentration dependence of H2O2 production of rat brain mitochondria in the presence of the respiratory substrates glutamateϩmalate, glutamateϩ malateϩrotenone, and succinate
  • In the present study we have characterized the H2O2 generation of mitochondria isolated from rat brain and human parahippocampal cortex
  • In accordance with several previous reports, but in clear contrast to others, we observed that the main quantity of H2O2 is produced at a site within complex I of the respiratory chain whereas only rather small quantities are produced at the bc1 complex in the presence of antimycin A
  • In accordance with previous work we detected a clear oxygen dependence of H2O2 generation with the rather high Km value of 0.92 Ϯ 0.01 mM oxygen for all conditions studied. In line with these results, at 100% oxygen saturation the superoxide production of brain mitochondria leads to a diminished rotenone sensitivity of NADH/coenzyme Q1 oxidoreductase
  • The different total H2O2 generation rates observed for isolated rat and human brain mitochondria are most likely related to the different purity of the individual mitochondrial preparations
  • In the present study we avoided the use of ketoacids as mitochondrial substrates and used p-hydroxyphenylacetate, which did not show considerable endogenous fluorescence changes as obtained with the more sensitive dye Amplex Red. To obtain reliable data for maximal H2O2 and superoxide generation rates we used oxygen saturated media

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