High Ca2+ load promotes Hydrogen peroxide generation via activation of α-glycerophosphate dehydrogenase in brain mitochondria

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Abstract

H2O2 generation associated with α- glycerophosphate (α-GP) oxidation was addressed in guinea pig brain mitochondria challenged with high Ca2+ load (10 μM). Exposure to 10 μM Ca2+ induced an abrupt 2.5-fold increase in H 2O2 release compared to that measured in the presence of a physiological cytosolic Ca2+ concentration (100 nM) from mitochondria respiring on 5 mM α-GP in the presence of ADP (2 mM). The Ca2+-induced stimulation of H2O2 generation was reversible and unaltered by the uniporter blocker Ru 360, indicating that it did not require Ca2+ uptake into mitochondria. Enhanced H 2O2 generation by Ca2+ was also observed in the absence of ADP when mitochondria exhibited permeability transition pore opening with a decrease in the NAD(P)H level, dissipation of membrane potential, and mitochondrial swelling. Furthermore, mitochondria treated with the pore-forming peptide alamethicin also responded with an elevated H2O2 generation to a challenge with 10 μM Ca2+. Ca2+-induced promotion of H2O2 formation was further enhanced by the complex III inhibitor myxothiazol. With 20 mM α-GP concentration, stimulation of H2O2 formation by Ca2+ was detected only in the presence, not in the absence, of ADP. It is concluded that α-glycerophosphate dehydrogenase, which is accessible to and could be activated by a rise in the level of cytosolic Ca2+, makes a major contribution to Ca2+-stimulated H2O2 generation. This work highlights a unique high-Ca2+-stimulated reactive oxygen species-forming mechanism in association with oxidation of α-GP, which is largely independent of the bioenergetic state and can proceed even in damaged, functionally incompetent mitochondria.

Original languageEnglish
Pages (from-to)2119-2130
Number of pages12
JournalFree Radical Biology and Medicine
Volume53
Issue number11
DOIs
Publication statusPublished - Dec 1 2012

Fingerprint

Glycerolphosphate Dehydrogenase
Mitochondria
Hydrogen Peroxide
Brain
Chemical activation
Adenosine Diphosphate
Alamethicin
Mitochondrial Swelling
Glycerophosphates
Oxidation
Electron Transport Complex III
NAD
Membrane Potentials
Energy Metabolism
Swelling
Permeability
Reactive Oxygen Species
Guinea Pigs
Association reactions
Membranes

Keywords

  • α-Glycerophosphate dehydrogenase
  • α-Glycerophosphate shuttle
  • Calcium
  • Free radicals
  • Hydrogen peroxide
  • Mitochondria
  • Myxothiazol
  • Permeability transition pore
  • Reactive oxygen species
  • Reverse electron transport

ASJC Scopus subject areas

  • Biochemistry
  • Physiology (medical)
  • Medicine(all)

Cite this

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title = "High Ca2+ load promotes Hydrogen peroxide generation via activation of α-glycerophosphate dehydrogenase in brain mitochondria",
abstract = "H2O2 generation associated with α- glycerophosphate (α-GP) oxidation was addressed in guinea pig brain mitochondria challenged with high Ca2+ load (10 μM). Exposure to 10 μM Ca2+ induced an abrupt 2.5-fold increase in H 2O2 release compared to that measured in the presence of a physiological cytosolic Ca2+ concentration (100 nM) from mitochondria respiring on 5 mM α-GP in the presence of ADP (2 mM). The Ca2+-induced stimulation of H2O2 generation was reversible and unaltered by the uniporter blocker Ru 360, indicating that it did not require Ca2+ uptake into mitochondria. Enhanced H 2O2 generation by Ca2+ was also observed in the absence of ADP when mitochondria exhibited permeability transition pore opening with a decrease in the NAD(P)H level, dissipation of membrane potential, and mitochondrial swelling. Furthermore, mitochondria treated with the pore-forming peptide alamethicin also responded with an elevated H2O2 generation to a challenge with 10 μM Ca2+. Ca2+-induced promotion of H2O2 formation was further enhanced by the complex III inhibitor myxothiazol. With 20 mM α-GP concentration, stimulation of H2O2 formation by Ca2+ was detected only in the presence, not in the absence, of ADP. It is concluded that α-glycerophosphate dehydrogenase, which is accessible to and could be activated by a rise in the level of cytosolic Ca2+, makes a major contribution to Ca2+-stimulated H2O2 generation. This work highlights a unique high-Ca2+-stimulated reactive oxygen species-forming mechanism in association with oxidation of α-GP, which is largely independent of the bioenergetic state and can proceed even in damaged, functionally incompetent mitochondria.",
keywords = "α-Glycerophosphate dehydrogenase, α-Glycerophosphate shuttle, Calcium, Free radicals, Hydrogen peroxide, Mitochondria, Myxothiazol, Permeability transition pore, Reactive oxygen species, Reverse electron transport",
author = "L. Tretter and V. {\'A}d{\'a}m-Vizi",
year = "2012",
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T1 - High Ca2+ load promotes Hydrogen peroxide generation via activation of α-glycerophosphate dehydrogenase in brain mitochondria

AU - Tretter, L.

AU - Ádám-Vizi, V.

PY - 2012/12/1

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N2 - H2O2 generation associated with α- glycerophosphate (α-GP) oxidation was addressed in guinea pig brain mitochondria challenged with high Ca2+ load (10 μM). Exposure to 10 μM Ca2+ induced an abrupt 2.5-fold increase in H 2O2 release compared to that measured in the presence of a physiological cytosolic Ca2+ concentration (100 nM) from mitochondria respiring on 5 mM α-GP in the presence of ADP (2 mM). The Ca2+-induced stimulation of H2O2 generation was reversible and unaltered by the uniporter blocker Ru 360, indicating that it did not require Ca2+ uptake into mitochondria. Enhanced H 2O2 generation by Ca2+ was also observed in the absence of ADP when mitochondria exhibited permeability transition pore opening with a decrease in the NAD(P)H level, dissipation of membrane potential, and mitochondrial swelling. Furthermore, mitochondria treated with the pore-forming peptide alamethicin also responded with an elevated H2O2 generation to a challenge with 10 μM Ca2+. Ca2+-induced promotion of H2O2 formation was further enhanced by the complex III inhibitor myxothiazol. With 20 mM α-GP concentration, stimulation of H2O2 formation by Ca2+ was detected only in the presence, not in the absence, of ADP. It is concluded that α-glycerophosphate dehydrogenase, which is accessible to and could be activated by a rise in the level of cytosolic Ca2+, makes a major contribution to Ca2+-stimulated H2O2 generation. This work highlights a unique high-Ca2+-stimulated reactive oxygen species-forming mechanism in association with oxidation of α-GP, which is largely independent of the bioenergetic state and can proceed even in damaged, functionally incompetent mitochondria.

AB - H2O2 generation associated with α- glycerophosphate (α-GP) oxidation was addressed in guinea pig brain mitochondria challenged with high Ca2+ load (10 μM). Exposure to 10 μM Ca2+ induced an abrupt 2.5-fold increase in H 2O2 release compared to that measured in the presence of a physiological cytosolic Ca2+ concentration (100 nM) from mitochondria respiring on 5 mM α-GP in the presence of ADP (2 mM). The Ca2+-induced stimulation of H2O2 generation was reversible and unaltered by the uniporter blocker Ru 360, indicating that it did not require Ca2+ uptake into mitochondria. Enhanced H 2O2 generation by Ca2+ was also observed in the absence of ADP when mitochondria exhibited permeability transition pore opening with a decrease in the NAD(P)H level, dissipation of membrane potential, and mitochondrial swelling. Furthermore, mitochondria treated with the pore-forming peptide alamethicin also responded with an elevated H2O2 generation to a challenge with 10 μM Ca2+. Ca2+-induced promotion of H2O2 formation was further enhanced by the complex III inhibitor myxothiazol. With 20 mM α-GP concentration, stimulation of H2O2 formation by Ca2+ was detected only in the presence, not in the absence, of ADP. It is concluded that α-glycerophosphate dehydrogenase, which is accessible to and could be activated by a rise in the level of cytosolic Ca2+, makes a major contribution to Ca2+-stimulated H2O2 generation. This work highlights a unique high-Ca2+-stimulated reactive oxygen species-forming mechanism in association with oxidation of α-GP, which is largely independent of the bioenergetic state and can proceed even in damaged, functionally incompetent mitochondria.

KW - α-Glycerophosphate dehydrogenase

KW - α-Glycerophosphate shuttle

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KW - Free radicals

KW - Hydrogen peroxide

KW - Mitochondria

KW - Myxothiazol

KW - Permeability transition pore

KW - Reactive oxygen species

KW - Reverse electron transport

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