Oxidation and Pharmacologic Activation of IK/SK Channels

中（IK）和小（SK）电导Ca激活的K+通道在血管平滑肌内皮依赖性松弛中的作用已得到明确证明。在激动剂和血流诱导的血管舒张以及活性氧（ROS）增加的情况下，IK和SK通道都被激活，这几乎与所有心血管疾病有关。内皮中这些通道的表达也被证明在球囊血管成形术后受损。最后，已知这些通道对edhf介导的血管舒张至关重要，而edhf介导的血管舒张在许多心血管疾病中受到损害。这些结果表明，内皮细胞IK和SK通道的药理激活将在广泛的心血管疾病中具有临床益处。我们的实验室首先确定了一系列结构相似的IK和SK通道的药理学打开剂。进一步明确IK和SK通道在内皮细胞中的作用

The role of intermediate (IK) and small (SK) conductance, Ca -activated K+ channels have been unequivocally demonstrated in the endothelial-dependent relaxation of vascular smooth muscle. Both IK and SK channels are activated during agonist- and flow-induced vasodilation as well as in the presence of increased reactive oxygen species(ROS), which are associated with virtually all cardiovascular disease. The expression of these channels in endothelia has also been shown to be compromised following balloon angioplasty. Finally, these channels are known to be critical to EDHF-mediated vasodilation, which is compromised in a host of cardiovascular diseases. These results have led to the proposal that the pharmacological activation of endothelial IK and SK channels would be of clinical benefit in a wide array of cardiovascular diseases. Our laboratory was the first to identify a series of structurally similar pharmacological openers of IK and SK channels. To further define the role of IK and SK channels in endothelial function and how they may be pharmacologically manipulated for clinical benefit requires us to answer two critical unknowns. First, how do ROS alter IK and SK channel function and therefore endothelial function? Second, what is the molecular mechanism of action for the known openers of IK and SK channels? Thus, we propose the following aims: (i) Define the mechanisms involved in the reactive oxygen species-dependent regulation of IK and SK channels. We will utilize a combination of patch-clamp and mutagenesis techniques to define the mechanisms whereby oxidizing agents activate IK and SK channels. These studies will be carried out on both heterologously expressed channels as well as on primary cultures of endothelial cells, (ii) We will define the molecular mechanism whereby pharmacological activators of IK and SK channels increase channel activity. These studies will be carried out utilizing a combination of patch-clamp and mutagenesis techniques, (iii) We will utilize FRET to define inter- and intra-subunit domain interactions in IK and SK channels and how physiological and pharmacological regulators of channel function modify these interactions. Defining how these interactions are altered is critical to our understanding of how these channels are regulated during the inflammatory process and how they may be manipulated pharmacologically. The results of these studies will clearly define the mechanism whereby ROS activate endothelial IK and SK channels, and thus alter vascular tone, as well as define the molecular mechanism underlying pharmacological activation of these channels; thereby furthering our understanding of how these channels may be manipulated for therapeutic benefit.