Putative roles of Ca(2+) -independent phospholipase A2 in respiratory chain-associated ROS production in brain mitochondria: influence of docosahexaenoic acid and bromoenol lactone.

Abstract

Ca(2+) -independent phospholipase A2 (iPLA2 ) is hypothesized to control mitochondrial reactive oxygen species (ROS) generation. Here, we modulated the influence of iPLA2 -induced liberation of non-esterified free fatty acids on ROS generation associated with the electron transport chain. We demonstrate enzymatic activity of membrane-associated iPLA2 in native, energized rat brain mitochondria (RBM). Theoretically, enhanced liberation of free fatty acids by iPLA2 modulates mitochondrial ROS generation, either attenuating the reversed electron transport (RET) or deregulating the forward electron transport of electron transport chain. For mimicking such conditions, we probed the effect of docosahexaenoic acid (DHA), a major iPLA2 product on ROS generation. We demonstrate that the adenine nucleotide translocase partly mediates DHA-induced uncoupling, and that low micromolar DHA concentrations diminish RET-dependent ROS generation. Uncoupling proteins have no effect, but the adenine nucleotide translocase inhibitor carboxyatractyloside attenuates DHA-linked uncoupling effect on RET-dependent ROS generation. Under physiological conditions of forward electron transport, low micromolar DHA stimulates ROS generation. Finally, exposure of RBM to the iPLA2 inhibitor bromoenol lactone (BEL) enhanced ROS generation. BEL diminished RBM glutathione content. BEL-treated RBM exhibits reduced Ca(2+) retention capacity and partial depolarization. Thus, we rebut the view that iPLA2 attenuates oxidative stress in brain mitochondria. However, the iPLA2 inhibitor BEL has detrimental activities on energy-dependent mitochondrial functions. The Ca(2+) -independent phospholipase A2 (iPLA2 ), a FFA (free fatty acids)-generating membrane-attached mitochondrial phospholipase, is potential to regulate ROS (reactive oxygen species) generation by mitochondria. FFA can either decrease reversed electron transport (RET)-linked or enhance forward electron transport (FET)-linked ROS generation. In the physiological mode of FET, iPLA2 activity increases ROS generation. The iPLA2 inhibitor BEL exerts detrimental effects on energy-dependent mitochondrial functions.