Synergistic inhibition of respiration in brain mitochondria by nitric oxide and dihydroxyphenylacetic acid (DOPAC)

The inhibition of mitochondrial respiration by nitric oxide (NO) at cytochrome c oxidase level has been established as a physiological regulatory mechanism of mitochondrial function. Given, on the one hand, the potential involvement of NO and dopamine metabolism in mitochondrial dysfunction associated with neurodegeneration and, on the other hand, the reported interaction of NO with dihydroxyphenylacetic acid (DOPAC), a major mitochondrial-associated dopamine metabolite, we examined the combined effects of NO and DOPAC on the respiratory chain of isolated rat brain mitochondria. Whereas dopamine or DOPAC induced no measurable effects on the mitochondrial respiration rate, a mixture of NO with DOPAC inhibited the rate in a way stronger than that exerted by NO. This effect was noticed with actively respiring (state 3) and resting (state 4) mitochondria. At variance with DOPAC, dopamine failed to potentiate NO inhibitory effects. The inhibition was dependent on the concentration of both compounds, NO and DOPAC, and exhibited characteristics similar to those exerted by NO, namely: it was reversible and dependent on the concentration of oxygen. Analysis of respiratory enzymatic activities demonstrated a selective inhibition at the level of cytochrome c oxidase (complex IV). Insights into the chemical mechanisms underlying the inhibitory effect were inferred from experiments using metmyoglobin (a ligand for NO and derived species, such as nitroxyl anion) and ferrocyanide (a reductant of NO, producing nitroxyl anion). Whereas metmyoglobin decreased the inhibition, ferrocyanide potentiated the inhibition. Moreover, a mixture of ferrocyanide with NO reproduced the effects exerted by the mixture of NO with DOPAC. The results are consistent with the notion of a reaction of NO with DOPAC producing a nitric oxide-derived compound(s), which inhibit O2 uptake at the cytochrome oxidase level. Although the mechanism in question remains to be clearly elucidated it is suggested that the NO/DOPAC-dependent inhibition of cytochrome oxidase may involve nitroxyl anion. The significance of these observations for mitochondrial dysfunction inherent in Parkinson's disease is discussed.