Effects of Partial Inhibition of Respiratory Complex I on H2O2 Production by Isolated Brain Mitochondria in Different Respiratory States

Author(s)

Michelini, Luiz G. B. ^[1] ; Benevento, Carlos E. ^[1] ; Rossato, Franco A. ^[1] ; Siqueira-Santos, Edilene S. ^[1] ; Castilho, Roger F. ^[1]

Affiliation

^[1] Univ Estadual Campinas, Dept Patol Clin, Fac Ciencias Med, BR-13083887 Campinas, SP - Brazil

• Document type: Journal article
• Source: Neurochemical Research; v. 39, n. 12, p. 2419-2430, DEC 2014.
• Web of Science Citations: 5

Abstract

The aim of this work was to characterize the effects of partial inhibition of respiratory complex I by rotenone on H2O2 production by isolated rat brain mitochondria in different respiratory states. Flow cytometric analysis of membrane potential in isolated mitochondria indicated that rotenone leads to uniform respiratory inhibition when added to a suspension of mitochondria. When mitochondria were incubated in the presence of a low concentration of rotenone (10 nm) and NADH-linked substrates, oxygen consumption was reduced from 45.9 +/- A 1.0 to 26.4 +/- A 2.6 nmol O-2 mg(-1) min(-1) and from 7.8 +/- A 0.3 to 6.3 +/- A 0.3 nmol O-2 mg(-1) min(-1) in respiratory states 3 (ADP-stimulated respiration) and 4 (resting respiration), respectively. Under these conditions, mitochondrial H2O2 production was stimulated from 12.2 +/- A 1.1 to 21.0 +/- A 1.2 pmol H2O2 mg(-1) min(-1) and 56.5 +/- A 4.7 to 95.0 +/- A 11.1 pmol H2O2 mg(-1) min(-1) in respiratory states 3 and 4, respectively. Similar results were observed when comparing mitochondrial preparations enriched with synaptic or nonsynaptic mitochondria or when 1-methyl-4-phenylpyridinium ion (MPP+) was used as a respiratory complex I inhibitor. Rotenone-stimulated H2O2 production in respiratory states 3 and 4 was associated with a high reduction state of endogenous nicotinamide nucleotides. In succinate-supported mitochondrial respiration, where most of the mitochondrial H2O2 production relies on electron backflow from complex II to complex I, low rotenone concentrations inhibited H2O2 production. Rotenone had no effect on mitochondrial elimination of micromolar concentrations of H2O2. The present results support the conclusion that partial complex I inhibition may result in mitochondrial energy crisis and oxidative stress, the former being predominant under oxidative phosphorylation and the latter under resting respiration conditions. (AU)

• FAPESP's process: 11/50400-0 - Mitochondrial energy metabolism, redox state and functionality in cell death and cardiometabolic and neurodegenerative disorders
• Grantee: Aníbal Eugênio Vercesi
• Support Opportunities: Research Projects - Thematic Grants
• FAPESP's process: 11/14229-4 - Mitochondrial generation of reactive oxygen species in the presence of a partial complex I inhibition: Effect of different metabolic states
• Grantee: Luiz Guilherme Michelini Bueno
• Support Opportunities: Scholarships in Brazil - Master