Membrane potential-related effect of calcium on reactive oxygen species generation in isolated brain mitochondria

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The effect of Ca2+ applied in high concentrations (50 and 300μM) was addressed on the generation of reactive oxygen species in isolated mitochondria from guinea-pig brain. The experiments were performed in the presence of ADP, a very effective inhibitor of mitochondrial permeability transition. Moderate increase in H2O2 release from mitochondria was induced by Ca2+ applied in 50μM, but not in 300μM concentration as measured with Amplex red fluorescent assay starting with a delay of 100-150sec after exposure to Ca2+. Parallel measurements of membrane potential (δψm) by safranine fluorescence showed a transient depolarization by Ca2+ followed by the recovery of δψm to a value, which was more negative than that observed before addition of Ca2+ indicating a relative hyperpolarization. NAD(P)H fluorescence was also increased by Ca2+ given in 50μM concentration. In mitochondria having high δψm in the presence of oligomycin or ATP, the basal rate of release of H2O2 was significantly higher than that observed in a medium containing ADP and Ca2+ no longer increased but rather decreased the rate of H2O2 release. With 300μM Ca2+ only a loss but no tendency of a recovery of δψm was detected and H2O2 release was unchanged. It is suggested that in the presence of nucleotides the effect of Ca2+ on mitochondrial ROS release is related to changes in δψm; in depolarized mitochondria, in the presence of ADP, moderate increase in H2O2 release is induced by calcium, but only in ≤ 100μM concentration, when after a transient Ca2+-induced depolarization mitochondria became more polarized. In highly polarized mitochondria, in the presence of ATP or oligomycin, where no hyperpolarization follows the Ca2+-induced depolarization, Ca2+ fails to stimulate mitochondrial ROS generation. These effects of calcium (≤ 300μM) are unrelated to mitochondrial permeability transition.

Original languageEnglish
Pages (from-to)922-928
Number of pages7
JournalBiochimica et Biophysica Acta - Bioenergetics
Issue number6-7
Publication statusPublished - Jun 1 2010


  • Brain
  • Calcium
  • Membrane potential
  • Mitochondria
  • Oxidative stress
  • Reactive oxygen species

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Cell Biology

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