Mitochondria-Derived Reactive Oxygen Species Dilate Cerebral Arteries by Activating Ca 2+ Sparks

Abstract

Mitochondria regulate intracellular calcium (Ca 2+ ) signals in smooth muscle cells, but mechanisms mediating these effects, and the functional relevance, are poorly understood. Similarly, antihypertensive ATP-sensitive potassium (K ATP ) channel openers (KCOs) activate plasma membrane K ATP channels and depolarize mitochondria in several cell types, but the contribution of each of these mechanisms to vasodilation is unclear. Here, we show that cerebral artery smooth muscle cell mitochondria are most effectively depolarized by diazoxide (−15%, tetramethylrhodamine [TMRM]), less so by levcromakalim, and not depolarized by pinacidil. KCO-induced mitochondrial depolarization increased the generation of mitochondria-derived reactive oxygen species (ROS) that stimulated Ca 2+ sparks and large-conductance Ca 2+ -activated potassium (K Ca ) channels, leading to transient K Ca current activation. KCO-induced mitochondrial depolarization and transient K Ca current activation were attenuated by 5-HD and glibenclamide, K ATP channel blockers. MnTMPyP, an antioxidant, and Ca 2+ spark and K Ca channel blockers reduced diazoxide-induced vasodilations by >60%, but did not alter dilations induced by pinacidil, which did not elevate ROS. Data suggest diazoxide drives ROS generation by inducing a small mitochondrial depolarization, because nanomolar CCCP, a protonophore, similarly depolarized mitochondria, elevated ROS, and activated transient K Ca currents. In contrast, micromolar CCCP, or rotenone, an electron transport chain blocker, induced a large mitochondrial depolarization (−84%, TMRM), reduced ROS, and inhibited transient K Ca currents. In summary, data demonstrate that mitochondria-derived ROS dilate cerebral arteries by activating Ca 2+ sparks, that some antihypertensive KCOs dilate by stimulating this pathway, and that small and large mitochondrial depolarizations lead to differential regulation of ROS and Ca 2+ sparks.