toxin-ii-(anemonia-sulcata) and Tachycardia--Ventricular

Brugada syndrome (BrS) is associated with a pronounced risk to develop sudden cardiac death (SCD). Up to 21% of patients are related to mutations in SCN5A. Studies identified SCN10A as a contributor of BrS. However, the investigation of the human cellular phenotype of BrS in the presence of SCN10A mutations remains lacking. The objective of this study was to establish a cellular model of BrS in presence of SCN10A mutations using human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs).. Dermal fibroblasts obtained from a BrS patient suffering from SCD harbouring the SCN10A double variants (c.3803G>A and c.3749G>A) and three independent healthy control subjects were reprogrammed to hiPSCs. Human-induced pluripotent stem cells were differentiated into cardiomyocytes (hiPSC-CMs).The hiPSC-CMs from the BrS patient showed a significantly reduced peak sodium channel current (INa) and a significantly reduced ATX II (sea anemone toxin, an enhancer of late INa) sensitive as well as A-887826 (a blocker of SCN10A channel) sensitive late sodium channel current (INa) when compared with the healthy control hiPSC-CMs, indicating loss-of-function of sodium channels. Consistent with reduced INa the action potential amplitude and upstroke velocity (Vmax) were significantly reduced, which may contribute to arrhythmogenesis of BrS. Moreover, Ajmaline effects on action potentials were stronger in BrS-hiPSC-CMs than in healthy control cells. This is in agreement with the higher susceptibility of patients to sodium channel blocking drugs in unmasking BrS.. Patient-specific hiPSC-CMs are able to recapitulate single-cell phenotype features of BrS with SCN10A mutations and may provide novel opportunities to further elucidate the cellular disease mechanism.

Topics: Action Potentials; Ajmaline; Brugada Syndrome; Cardiotonic Agents; Case-Control Studies; Cellular Reprogramming Techniques; Cnidarian Venoms; Death, Sudden, Cardiac; Humans; Induced Pluripotent Stem Cells; Loss of Function Mutation; Male; Middle Aged; Morpholines; Mutation; Myocytes, Cardiac; NAV1.8 Voltage-Gated Sodium Channel; Niacinamide; Patch-Clamp Techniques; Phenotype; Tachycardia, Ventricular; Voltage-Gated Sodium Channel Blockers

Female gender is a risk factor for long QT-related arrhythmias, but the underlying mechanisms remain uncertain. Here, we tested the hypothesis that gender-dependent function of the post-depolarization 'late' sodium current (I(Na-L)) contributes.. Studies were conducted in mice in which the canonical cardiac sodium channel Scn5a locus was disrupted, and expression of human wild-type SCN5A cDNA substituted. Baseline QT intervals were similar in male and female mice, but exposure to the sodium channel opener anemone toxin ATX-II elicited polymorphic ventricular tachycardia in 0/9 males vs. 6/9 females. Ventricular I(Na-L) and action potential durations were increased in myocytes isolated from female mice compared with those from males before and especially after treatment with ATX-II. Further, ATX-II elicited potentially arrhythmogenic early afterdepolarizations in myocytes from 0/5 male mice and 3/5 female mice.. These data identify variable late I(Na) as a modulator of gender-dependent arrhythmia susceptibility.

Topics: Acetanilides; Action Potentials; Animals; Cnidarian Venoms; Disease Models, Animal; Electrocardiography; Female; Genetic Predisposition to Disease; Humans; Long QT Syndrome; Male; Mice; Mice, 129 Strain; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; NAV1.5 Voltage-Gated Sodium Channel; Piperazines; Ranolazine; Risk Factors; Sex Factors; Tachycardia, Ventricular; Time Factors

Late Na(+) current (I(NaL)) and Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) are both increased in the diseased heart. Recently, CaMKII was found to phosphorylate the Na(+) channel 1.5 (Na(v)1.5), resulting in enhanced I(NaL). Conversely, an increase of I(NaL) would be expected to cause elevation of intracellular Ca(2+) and activation of CaMKII. However, a relationship between enhancement of I(NaL) and activation of CaMKII has yet to be demonstrated. We investigated whether Na(+) influx via Na(v)1.5 leads to CaMKII activation and explored the functional significance of this pathway. In neonatal rat ventricular myocytes (NRVM), treatment with the I(NaL) activators anemone toxin II (ATX-II) or veratridine increased CaMKII autophosphorylation and increased phosphorylation of CaMKII substrates phospholamban and ryanodine receptor 2. Knockdown of Na(v)1.5 (but not Na(v)1.1 or Na(v)1.2) prevented ATX-II-induced CaMKII phosphorylation, providing evidence for a specific role of Na(v)1.5 in CaMKII activation. In support of this view, CaMKII activity was also increased in hearts of transgenic mice overexpressing a gain-of-function Na(v)1.5 mutant (N(1325)S). The effects of both ATX-II and the N(1325)S mutation were reversed by either I(NaL) inhibition (with ranolazine or tetrodotoxin) or CaMKII inhibition (with KN93 or autocamtide 2-related inhibitory peptide). Furthermore, ATX-II treatment also induced CaMKII-Na(v)1.5 coimmunoprecipitation. The same association between CaMKII and Na(v)1.5 was also found in N(1325)S mice, suggesting a direct protein-protein interaction. Pharmacological inhibitions of either CaMKII or I(NaL) also prevented ATX-II-induced cell death in NRVM and reduced the incidence of polymorphic ventricular tachycardia induced by ATX-II in rat perfused hearts. Taken together, these results suggest that a Na(v)1.5-dependent increase in Na(+) influx leads to activation of CaMKII, which in turn phosphorylates Na(v)1.5, further promoting Na(+) influx. Pharmacological inhibition of either CaMKII or Na(v)1.5 can ameliorate cardiac dysfunction caused by excessive Na(+) influx.

Topics: Acetanilides; Amino Acid Substitution; Animals; Animals, Newborn; Calcium; Calcium Signaling; Calcium-Binding Proteins; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Caspase 3; Cell Death; Cell Survival; Cnidarian Venoms; Dose-Response Relationship, Drug; Electrophysiological Phenomena; Female; Gene Expression; Heart Ventricles; Humans; Mice; Mice, Inbred Strains; Mice, Transgenic; Myocytes, Cardiac; NAV1.5 Voltage-Gated Sodium Channel; Peptides; Perfusion; Phosphorylation; Piperazines; Protein Binding; Rabbits; Ranolazine; Rats; Rats, Sprague-Dawley; RNA, Small Interfering; Ryanodine Receptor Calcium Release Channel; Sodium; Sodium Channels; Sodium-Calcium Exchanger; Tachycardia, Ventricular; Tetrodotoxin; Veratridine

Although long QT syndrome (LQTS) and coronary occlusion-reperfusion (O/R) are arrhythmogenic, they affect ventricular action potential duration (APD) differently. In contrast to the prolonged APD in LQTS, ischemia abbreviates APD after a transient prolongation. Thus we hypothesized that the dynamic interactive effects of ischemia and LQTS on APD and its dispersion would affect ventricular arrhythmogenicity. We mapped transmural distribution of action potentials in 6 groups of 10 isolated wedges of canine ventricular walls: LQTS-O/R, LQTS only, and O/R only, with separate groups for pacing cycle lengths (PCL) of 1,000 and 2,000 ms. We created type 3 LQTS with anemone toxin (ATX) II followed >30 min later by arterial occlusion (40 min) and reperfusion (>100 min). Arterial occlusion initially (first 4 min) prolonged and then shortened APD. Early afterdepolarizations (EADs) occurred during the initial 4 min of occlusion in 4 of the 10 LQTS-O/R wedges at PCL of 2,000 ms but not in the other groups. Reperfusion restored APD in the O/R-only groups but caused APD to overshoot its original duration, indicating depressed repolarization reserve, in the LQTS-O/R group. Reperfusion increased the dispersion of APDs and initiated ventricular tachycardia-fibrillation in 7 of 10 and 6 of 10 LQTS-O/R wedges and in 2 of 10 and 1 of 10 O/R-only wedges at PCLs of 1,000 and 2,000 ms, respectively. The LQTS-only wedges exhibited neither EADs nor ventricular tachycardia. We conclude that coronary O/R increased the arrhythmogenicity of LQTS via cumulative prolongation of APD, increase in repolarization dispersion, and suppression of repolarization reserve.

Topics: Action Potentials; Animals; Cnidarian Venoms; Coronary Disease; Disease Models, Animal; Dogs; In Vitro Techniques; Long QT Syndrome; Myocardial Reperfusion Injury; Reaction Time; Tachycardia, Ventricular

To explore the mechanism, we tested the hypothesis that premature epicardial stimulation transiently increased the dispersion of repolarization leading to VT.. Premature stimulation initiated ventricular tachycardia (VT) when applied to the epicardium but not to the endocardium in a canine model of long QT syndrome (LOTS).. We optically mapped action potentials (APs) on the cut-exposed transmural surfaces of isolated wedges of canine ventricular walls perfused with anemone toxin II (ATX-II), which produced type 3 LQTS with an asymmetrical transmural profile of repolarization that was earliest in the epicardium and latest in deep subendocardium.. Earliest excitable epicardial stimulation triggered VT in 5 of 18 wedges receiving > or =5 nmol/L ATX-II by direct activation of epicardium, which delayed repolarization in the still refractory midmyocardium and further enhanced the dispersion of repolarization. These VTs were initiated 197 +/- 72 ms (n = 10) after the premature stimulation, from focal regions of earliest repolarization downstream to the steepest local spatial gradients of repolarization, and maintained by new focal activation and reentry. Transmural differences in the cycle lengths of activations altered conduction pathways and resulted in torsades de pointes-like polymorphic VT. In contrast, VTs were not initiated by endocardial stimulation at the same premature intervals or when ATX-II was < or =2.5 nmol/L. Failed VT initiation was associated with significantly lower maximum local gradient of repolarization.. Heterogeneic repolarization in LQTS provides a transmural asymmetrical substrate for the earliest excitable epicardial, but not endocardial, stimulation to further delay midmyocardial repolarization and produce a steep spatial gradient of repolarization potential initiating torsades de pointes-like polymorphic VT.

Topics: Action Potentials; Animals; Cardiotonic Agents; Cnidarian Venoms; Dogs; Electric Stimulation; Endocardium; In Vitro Techniques; Long QT Syndrome; Models, Animal; Perfusion; Pericardium; Tachycardia, Ventricular