Large conductance K+ channels activated by elevated levels of intracellular Na+ were first described in the heart in 1984. At that time it was postulated that the current (IKNa) conducted by these channels was most likely only activated during severe ischemia and has since been largely ignored. Six years ago, the channel that conducts IKNa was cloned and named Slo2.1. However, little is known regarding the structural basis of Slo2.1 channel function, including the molecular basis of the weak voltage dependence of channel activation, the location of the activation gate, or the mechanisms that couple intracellular Na+ binding to channel opening. Our preliminary findings indicate two novel features of Slo2.1. First, the usual voltage-sensor (S1-S4 segments) has no role in sensing transmembrane voltage. Second, the selectivity filter, not the S6 bundle crossing, may function as the activation gate.
In Aim1 we will use site-directed mutagenesis, heterologous expression in Xenopus oocytes and voltage clamp techniques to substantiate these initial findings and definitively describe the molecular basis of Slo2.1 channel activation. Cardiac arrest is often associated with a combination of ventricular fibrillation (VF) and ischemia (VF/ischemia), both conditions promoting a rise in [Na+]i. A potential and yet unexplored role for IKNa during VF/ischemia is its contribution to depression and loss of excitability via an increase in the net outward current and extracellular K+ accumulation. During prolonged VF/ischemia, large gradients of activation rate and excitability develop between the enodcardium and the epicardium and between the right and the left ventricle (RV and LV), which are highly relevant to the outcomes of defibrillation and post-shock resuscitation. Yet the exact mechanism of these gradients remains a puzzle. Our preliminary results reveal regional differences in Slo 2.1 expression which may explain the electrophysiological gradients. There is also direct and indirect evidence supporting an important role of IKNa in outward K+ leak during ischemia. Unfortunately, knowledge of the role of IKNa in ischemia or VF/ischemia is very limited.
In Aim 2 we will test the hypothesis that in the dog heart, the LV epicardium is the first region to exhibit inexcitability during VF/ischemia because myocytes in this region either accumulate cytoplasmic Na+ faster or express a greater density of IKNa channels compared to the LV endocardium or RV/septum. KNa channels are activated during ischemia, reperfusion and rapid heart rates when [Na+]i is transiently increased.
In Aim 3, activation of KNa channels under these conditions will be examined in isolated canine ventricular myocytes. Finally, the relative contributions of IKNa and IKATP in these phenomena will also be investigated in the whole heart (Aim 2) and in isolated myocytes (Aim 3) of dogs. Together these studies will define 1) the molecular basis of KNa channel activation, 2) the role of these channels under normal and pathophysiological conditions and 3) the potential role of these channels as modulators of defibrillation and post-shock resuscitation.

Public Health Relevance

Ion channels are membrane-bound proteins that selectively conduct specific ions in and out of cells. A plethora of potassium-selective ion channels are expressed in the human heart and are important components of the electrical activity that is responsible for the normal pump function of this organ. One type of potassium ion channel is called Slo2.1 and these channels are only activated if the intracellular concentration of sodium is elevated to abnormal levels as can occur during ischemia. Myocardial ischemia is both a frequent cause and a consequence of ventricular tachycardia and fibrillation, two potentially lethal forms of cardiac arrhythmia. The goals of this project are to understand the molecular details of Slo2.1 channel activation and define their role in the gradient of electrical excitability that develops between various layers and regions of the ventricle in the heart during ischemia. These electrical gradients are highly relevant to the outcomes of cardiac defibrillation and resuscitation of individuals who have experienced sudden cardiac arrest.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Project (R01)
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Electrical Signaling, Ion Transport, and Arrhythmias Study Section (ESTA)
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Lathrop, David A
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University of Utah
Schools of Medicine
Salt Lake City
United States
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Warren, Mark; Sciuto, Katie J; Taylor, Tyson G et al. (2017) Blockade of CaMKII depresses conduction preferentially in the right ventricular outflow tract and promotes ischemic ventricular fibrillation in the rabbit heart. Am J Physiol Heart Circ Physiol 312:H752-H767
Warren, Mark; Zaitsev, Alexey V (2017) CaMKII blockade, cardiac conduction, and arrhythmia. Cardiovasc Res 113:1798-1799
Suzuki, Tomoyuki; Hansen, Angela; Sanguinetti, Michael C (2016) Hydrophobic interactions between the S5 segment and the pore helix stabilizes the closed state of Slo2.1 potassium channels. Biochim Biophys Acta 1858:783-92
Thomson, Steven J; Hansen, Angela; Sanguinetti, Michael C (2015) Identification of the Intracellular Na+ Sensor in Slo2.1 Potassium Channels. J Biol Chem 290:14528-35
Garg, Vivek; Taylor, Tyson; Warren, Mark et al. (2015) ?-Adrenergic stimulation and rapid pacing mutually promote heterogeneous electrical failure and ventricular fibrillation in the globally ischemic heart. Am J Physiol Heart Circ Physiol 308:H1155-70
Venable, Paul W; Sciuto, Katie J; Warren, Mark et al. (2015) Mitochondrial depolarization and asystole in the globally ischemic rabbit heart: coordinated response to interventions affecting energy balance. Am J Physiol Heart Circ Physiol 308:H485-99
Shibayama, Junko; Yuzyuk, Tatiana N; Cox, James et al. (2015) Metabolic remodeling in moderate synchronous versus dyssynchronous pacing-induced heart failure: integrated metabolomics and proteomics study. PLoS One 10:e0118974
Diakos, Nikolaos A; Selzman, Craig H; Sachse, Frank B et al. (2014) Myocardial atrophy and chronic mechanical unloading of the failing human heart: implications for cardiac assist device-induced myocardial recovery. J Am Coll Cardiol 64:1602-12
Garg, Priyanka; Sanguinetti, Michael C (2014) Intracellular ATP does not inhibit Slo2.1 K+ channels. Physiol Rep 2:
Taylor, Tyson G; Venable, Paul W; Booth, Alicja et al. (2013) Does the combination of hyperkalemia and KATP activation determine excitation rate gradient and electrical failure in the globally ischemic fibrillating heart? Am J Physiol Heart Circ Physiol 305:H903-12

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