Reactive oxygen species production in synaptosomes is independent of ΔΨm

This study addressed whether dissipation of ΔΨm had any influence on ROS generation of in situ neuronal mitochondria. The ROS formation was measured as a release of H2O2 by Amplex Red assay. Complete blockage of Complex I by rotenone or Complex III by antimycin caused enhanced H2O2 release. Dissipation of ΔΨm by FCCP or DNP had no effect on the H2O2 production induced by rotenone or antimycin. Antimycin substantially diminishes the ΔΨm, but part of the membrane potential still maintained by the reverse function of mitochondrial ATP synthase. When antimycin was applied together with oligomycin, ΔΨm was totally dissipated, but the ROS production decreased only by 15%. Rotenone inhibited the antimycin‐induced H2O2 release by 55%, but not eliminated completely. These experiments suggest that in in situ mitochondria of synaptosomes (i) ROS generation because of inhibition of Complex I or III is not dependent on ΔΨm and (ii) when Complex I is completely inhibited electrons entering the respiratory chain distal from rotenone site could fuel the ROS formation.