A Mechanistic Link Between Complex II and the mitoKATP Channel in Ischemic Preconditioning

Thesis (Ph.D.)--University of Rochester. School of Medicine and Dentistry. Dept. of Pharmacology and Physiology, 2010.The preservation of mitochondrial function is a major protective strategy against damage induced by cardiac ischemia reperfusion (IR) injury. An endogenous cardioprotective mechanism is ischemic preconditioning (IPC) and a critical component of this mechanism is the mitochondrial ATP-sensitive potassium channel (mitoKATP). Agonists of the mitoKATP mimic IPC while inhibitors prevent the protection evoked by either channel agonists or IPC. Despite the pivotal role the mitoKATP plays in protecting the heart, its molecular identity remains controversial, and the validity of current methods to assay mitoKATP activity is disputed. Pharmacologic overlap between the mitoKATP and complex II of the mitochondrial respiratory chain implicates complex II in the regulation of the channel. The work herein investigates a mechanistic link between complex II and the mitoKATP in IPC, using isolated mitochondria, cardiomyocytes and perfused heart models of IR injury. These studies demonstrate the endogenous formation of a complex II inhibitor under conditions that mimic IPC; this inhibitor can activate the mitoKATP. Furthermore, complex II inhibition protects against IR injury via a mitoKATP dependent mechanism, suggesting that complex II regulates mitoKATP function. A new method to measure mitoKATP activity was developed and utilized to investigate the properties of the K+ transporting component of the mitoKATP and its role in IPC. Using this method, novel regulatory compounds and a mechanism of recovering lost channel activity were elucidated. The results support the hypothesis that complex II acts as a regulator of the mitoKATP and that the K+ transporting component of the mitoKATP is an inward rectifying K+ channel (Kir), likely the Kir6.2 isoform. This unique complex II-mediated control of mitoKATP activity has led to the discovery of the most potent mitoKATP agonist and cardioprotective agent to date, and may facilitate the development of improved therapeutics