kn-93 and Ventricular-Fibrillation

Hypokalemia is known to promote ventricular arrhythmias, especially in combination with class III antiarrhythmic drugs like dofetilide. Here, we evaluated the underlying molecular mechanisms.. Arrhythmias were recorded in isolated rabbit and rat hearts or patch-clamped ventricular myocytes exposed to hypokalemia (1.0-3.5 mmol/L) in the absence or presence of dofetilide (1 μmol/L). Spontaneous early afterdepolarizations (EADs) and ventricular tachycardia/fibrillation occurred in 50% of hearts at 2.7 mmol/L [K] in the absence of dofetilide and 3.3 mmol/L [K] in its presence. Pretreatment with the Ca-calmodulin kinase II (CaMKII) inhibitor KN-93, but not its inactive analogue KN-92, abolished EADs and hypokalemia-induced ventricular tachycardia/fibrillation, as did the selective late Na current (INa) blocker GS-967. In intact hearts, moderate hypokalemia (2.7 mmol/L) significantly increased tissue CaMKII activity. Computer modeling revealed that EAD generation by hypokalemia (with or without dofetilide) required Na-K pump inhibition to induce intracellular Na and Ca overload with consequent CaMKII activation enhancing late INa and the L-type Ca current. K current suppression by hypokalemia and dofetilide alone in the absence of CaMKII activation were ineffective at causing EADs.. We conclude that Na-K pump inhibition by even moderate hypokalemia plays a critical role in promoting EAD-mediated arrhythmias by inducing a positive feedback cycle activating CaMKII and enhancing late INa. Class III antiarrhythmic drugs like dofetilide sensitize the heart to this positive feedback loop.

Topics: Action Potentials; Animals; Benzylamines; Computer Simulation; Hypokalemia; Male; Phenethylamines; Pyridines; Rabbits; Rats; Rats, Inbred F344; Sodium-Potassium-Exchanging ATPase; Sulfonamides; Triazoles; Ventricular Fibrillation

Action potential duration alternans (APD-ALT), defined as long-short-long repetitive pattern of APD, potentially leads to lethal ventricular arrhythmia. However, the mechanisms of APD-ALT in the arrhythmogenesis of cardiac hypertrophy remain undetermined. Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) is known to modulate the function of cardiac sarcoplasmic reticulum and play an important role in Ca(2+) cycling. We thus aimed to determine the role of CaMKII in the increased susceptibility to APD-ALT and arrhythmogenesis in the hypertrophied heart. APD was measured by high-resolution optical mapping in left ventricular (LV) anterior wall from normotensive Wistar-Kyoto (WKY; n = 10) and spontaneously hypertensive rats (SHR; n = 10) during rapid ventricular pacing. APD-ALT was evoked at significantly lower pacing rate in SHR compared with WKY (382 ± 43 vs. 465 ± 45 beats/min, P < 0.01). These changes in APD-ALT in SHR were completely reversed by KN-93 (1 μmol/l; n = 5), an inhibitor of CaMKII, but not its inactive analog, KN-92 (1 μmol/l; n = 5). The magnitude of APD-ALT was also significantly greater in SHR than WKY and was completely normalized by KN-93. Ventricular fibrillation (VF) was induced by rapid pacing more frequently in SHR than in WKY (60 vs. 10%; P < 0.05), which was also abolished by KN-93 (0%, P < 0.05). Western blot analyses indicated that the CaMKII autophosphorylation at Thr287 was significantly increased in SHR compared with WKY. The increased susceptibility to APD-ALT and VF during rapid pacing in hypertrophied heart was prevented by KN-93. CaMKII could be an important mechanism of arrhythmogenesis in cardiac hypertrophy.

Topics: Action Potentials; Animals; Anti-Arrhythmia Agents; Benzylamines; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cardiac Pacing, Artificial; Cardiomegaly; Disease Models, Animal; Heart Ventricles; Hypertension; Male; Phosphorylation; Protein Kinase Inhibitors; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Sulfonamides; Threonine; Time Factors; Ventricular Fibrillation

Diastolic spontaneous Ca(2+) waves (DCWs) are recognized as important contributors to triggered arrhythmias. DCWs are thought to arise when [Ca(2+)] in sarcoplasmic reticulum ([Ca(2+)](SR)) reaches a certain threshold level, which might be reduced in cardiac disease as a consequence of sensitization of ryanodine receptors (RyR2s) to luminal Ca(2+).. We investigated the mechanisms of DCW generation in myocytes from normal and diseased hearts, using a canine model of post-myocardial infarction ventricular fibrillation (VF).. The frequency of DCWs, recorded during periodic pacing in the presence of a β-adrenergic receptor agonist isoproterenol, was significantly higher in VF myocytes than in normal controls. Rather than occurring immediately on reaching a final [Ca(2+)](SR), DCWs arose with a distinct time delay after attaining steady [Ca(2+)](SR) in both experimental groups. Although the rate of [Ca(2+)](SR) recovery after the SR Ca(2+) release was similar between the groups, in VF myocytes the latency to DCWs was shorter, and the [Ca(2+)](SR) at DCW initiation was lower. The restitution of depolarization-induced Ca(2+) transients, assessed by a 2-pulse protocol, was significantly faster in VF myocytes than in controls. The VF-related alterations in myocyte Ca(2+) cycling were mimicked by the RyR2 agonist, caffeine. The reducing agent, mercaptopropionylglycine, or the CaMKII inhibitor, KN93, decreased DCW frequency and normalized restitution of Ca(2+) release in VF myocytes.. The attainment of a certain threshold [Ca(2+)](SR) is not sufficient for the generation of DCWs. Postrelease Ca(2+) signaling refractoriness critically influences the occurrence of DCWs. Shortened Ca(2+) signaling refractoriness due to RyR2 phosphorylation and oxidation is responsible for the increased rate of DCWs observed in VF myocytes and could provide a substrate for synchronization of arrhythmogenic events at the tissue level in hearts prone to VF.

Topics: Adrenergic beta-Agonists; Animals; Benzylamines; Caffeine; Calcium Channel Agonists; Calcium Signaling; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cardiac Pacing, Artificial; Death, Sudden, Cardiac; Disease Models, Animal; Dogs; Excitation Contraction Coupling; Female; Isoproterenol; Male; Myocardial Infarction; Myocytes, Cardiac; Oxidation-Reduction; Phosphorylation; Protein Kinase Inhibitors; Reaction Time; Reducing Agents; Ryanodine Receptor Calcium Release Channel; Sarcoplasmic Reticulum; Sulfonamides; Time Factors; Tiopronin; Ventricular Fibrillation

Oxidative stress with hydrogen peroxide (H(2)O(2)) readily promotes early afterdepolarizations (EADs) and triggered activity (TA) in isolated rat and rabbit ventricular myocytes. Here we examined the effects of H(2)O(2) on arrhythmias in intact Langendorff rat and rabbit hearts using dual-membrane voltage and intracellular calcium optical mapping and glass microelectrode recordings. Young adult rat (3-5 mo, N = 25) and rabbit (3-5 mo, N = 6) hearts exhibited no arrhythmias when perfused with H(2)O(2) (0.1-2 mM) for up to 3 h. However, in 33 out of 35 (94%) aged (24-26 mo) rat hearts, 0.1 mM H(2)O(2) caused EAD-mediated TA, leading to ventricular tachycardia (VT) and fibrillation (VF). Aged rabbits (life span, 8-12 yr) were not available, but 4 of 10 middle-aged rabbits (3-5 yr) developed EADs, TA, VT, and VF. These arrhythmias were suppressed by the reducing agent N-acetylcysteine (2 mM) and CaMKII inhibitor KN-93 (1 microM) but not by its inactive form (KN-92, 1 microM). There were no significant differences between action potential duration (APD) or APD restitution slope before or after H(2)O(2) in aged or young adult rat hearts. In histological sections, however, trichrome staining revealed that aged rat hearts exhibited extensive fibrosis, ranging from 10-90%; middle-aged rabbit hearts had less fibrosis (5-35%), whereas young adult rat and rabbit hearts had <4% fibrosis. In aged rat hearts, EADs and TA arose most frequently (70%) from the left ventricular base where fibrosis was intermediate ( approximately 30%). Computer simulations in two-dimensional tissue incorporating variable degrees of fibrosis showed that intermediate (but not mild or severe) fibrosis promoted EADs and TA. We conclude that in aged ventricles exposed to oxidative stress, fibrosis facilitates the ability of cellular EADs to emerge and generate TA, VT, and VF at the tissue level.

Topics: Acetylcysteine; Action Potentials; Age Factors; Aging; Animals; Antioxidants; Benzylamines; Calcium Signaling; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cell Communication; Computer Simulation; Disease Models, Animal; Electrocardiography; Enzyme Activation; Fibrosis; Heart Ventricles; Hydrogen Peroxide; Male; Models, Cardiovascular; Myocytes, Cardiac; Oxidants; Oxidative Stress; Perfusion; Protein Kinase Inhibitors; Rabbits; Rats; Rats, Inbred F344; Sulfonamides; Tachycardia, Ventricular; Time Factors; Ventricular Fibrillation