Prolonged Refractoriness Research Articles

Mechanisms of torsades de pointes: an update.

Torsades de Pointes (TdP) refers to a polymorphic ventricular tachycardia (VT) with undulating QRS axis that occurs in long QT syndrome (LQTS), although the term has been used to describe polymorphic ventricular tachyarrhythmias in which QT intervals are not prolonged, such as short-coupled variant of TdP currently known as short-coupled ventricular fibrillation (VF) and Brugada syndrome. Extensive works on LQTS-related TdP over more than 50 years since it was first recognized by Dessertennes who coined the French term meaning "twisting of the points", have led to current understanding of the electrophysiological mechanism that TdP is initiated by triggered activity due to early afterdepolarization (EAD) and maintained by reentry within a substrate of inhomogeneous repolarization. While a recently emerging notion that steep voltage gradients rather than EADs are crucial to generate premature ventricular contractions provides additions to the initiation mode, the research to elucidate the maintenance mechanism hasn't made much progress. The reentrant activity that produces the specific form of VT is not well characterized. We have conducted optical mapping in a rabbit model of electrical storm by electrical remodeling (QT prolongation) due to chronic complete atrioventricular block and demonstrated that a tissue-island with prolonged refractoriness due to enhanced late Na+ current (INa-L) contributes to the generation of drifting rotors in a unique manner, which may explain the ECG characteristic of TdP. Moreover, we have proposed that the neural Na+ channel NaV1.8-mediated INa-L may be a new player to form the substrate for TdP. Here we discuss TdP mechanisms by comparing the findings in electrical storm rabbits with recently published studies by others in simulation models and human and animal models of LQTS.

ERG potassium channels and T-type calcium channels contribute to the pacemaker and atrioventricular conduction in zebrafish larvae.

Bradyarrhythmias result from inhibition of automaticity, prolonged repolarization, or slow conduction in the heart. The ERG channels mediate the repolarizing current IKr in the cardiac action potential, whereas T-type calcium channels (TTCC) are involved in the sinoatrial pacemaker and atrioventricular conduction in mammals. Zebrafish have become a valuable research model for human cardiac electrophysiology and disease. Here, we investigate the contribution of ERG channels and TTCCs to the pacemaker and atrioventricular conduction in zebrafish larvae and determine the mechanisms causing atrioventricular block. Zebrafish larvae expressing ratiometric fluorescent Ca2+ biosensors in the heart were used to measure Ca2+ levels and rhythm in beating hearts invivo, concurrently with contraction and hemodynamics. The atrioventricular delay (the time between the start of atrial and ventricular Ca2+ transients) was used to measure impulse conduction velocity and distinguished between slow conduction and prolonged refractoriness as the cause of the conduction block. ERG blockers caused bradycardia and atrioventricular block by prolonging the refractory period in the atrioventricular canal and in working ventricular myocytes. In contrast, inhibition of TTCCs caused bradycardia and second-degree block (Mobitz type I) by slowing atrioventricular conduction. TTCC block did not affect ventricular contractility, despite being highly expressed in cardiomyocytes. Concomitant measurement of Ca2+ levels and ventricular size showed mechano-mechanical coupling: increased preload resulted in a stronger heart contraction invivo. ERG channels and TTCCs influence the heart rate and atrioventricular conduction in zebrafish larvae. The zebrafish lines expressing Ca2+ biosensors in the heart allow us to investigate physiological feedback mechanisms and complex arrhythmias.

In-silico drug trials for precision medicine in atrial fibrillation: From ionic mechanisms to electrocardiogram-based predictions in structurally-healthy human atria.

Atrial fibrillation (AF) inducibility, sustainability and response to pharmacological treatment of individual patients are expected to be determined by their ionic current properties, especially in structurally-healthy atria. Mechanisms underlying AF and optimal cardioversion are however still unclear. In this study, in-silico drug trials were conducted using a population of human structurally-healthy atria models to 1) identify key ionic current properties determining AF inducibility, maintenance and pharmacological cardioversion, and 2) compare the prognostic value for predicting individual AF cardioversion of ionic current properties and electrocardiogram (ECG) metrics. In the population of structurally-healthy atria, 477 AF episodes were induced in ionic current profiles with both steep action potential duration (APD) restitution (eliciting APD alternans), and high excitability (enabling propagation at fast rates that transformed alternans into discordant). High excitability also favored 211 sustained AF episodes, so its decrease, through prolonged refractoriness, explained pharmacological cardioversion. In-silico trials over 200 AF episodes, 100 ionic profiles and 10 antiarrhythmic compounds were consistent with previous clinical trials, and identified optimal treatments for individual electrophysiological properties of the atria. Algorithms trained on 211 simulated AF episodes exhibited >70% accuracy in predictions of cardioversion for individual treatments using either ionic current profiles or ECG metrics. In structurally-healthy atria, AF inducibility and sustainability are enabled by discordant alternans, under high excitability and steep restitution conditions. Successful pharmacological cardioversion is predicted with 70% accuracy from either ionic or ECG properties, and it is optimal for treatments maximizing refractoriness (thus reducing excitability) for the given ionic current profile of the atria.

Selective epicardial pulsed field ablation of atrial ganglionated plexi causes anti-arrhythmic prolongation of refractoriness: demonstration of feasibility in cardiac surgery patients

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – EU funding. Main funding source(s): EU Horizon 2020 SME Instrument. Background Epicardial ganglionated plexi (GP) play a significant role in the initiation and maintenance of atrial fibrillation. However, modulation of this effect, through GP ablation, has had limited success; outcomes being confounded by unnecessary atrial tissue ablation and inability to access and ablate all of the atrial GPs. Selective pulsed field ablation (PFA) of GPs, using epicardial access, provides the opportunity to better identify the role of GP ablation in the treatment of atrial fibrillation. Purpose This study aimed to assess the safety and feasibility of selective GP ablation in patients undergoing elective coronary artery bypass grafting (CABG). It was hypothesized that GP ablation would provide an acute extension of atrial tissue refractoriness, which constitutes its antiarrhythmic effect. Methods Using a monopolar, monophasic PFA system, atrial GPs were ablated in nineteen patients with or without atrial fibrillation, undergoing CABG. The Oblique Sinus GP, Right Superior GP, Transverse Sinus GP, Left Superior GP and Ligament of Marshall GP were each ablated with up to sixty PFA pulses of 1000 V amplitude and 100 µs pulse width. Atrial Effective Refractory Period (AERP) was measured before and after all GP ablations, at the left atrial appendage (LAA) and on the right atrium (RA). Patients were monitored through to discharge for post-operative atrial fibrillation (POAF). Results Complete ablation of the GPs was performed in nineteen patients (aged 63.4 ± 6.6 years, 63.1% male) immediately after sternotomy. Electric field pulses were ECG-gated, with energy delivery during the ventricular refractory period. All GP sites were successfully accessed and ablated; all patients progressed immediately to their planned elective surgery and were discharged on schedule. Procedure time, for all GP ablations and AERP measurements was in the range 35-45 minutes. Thirteen valid pre- and post-ablation datasets were obtained. AERP (LAA and RA combined) increased upon GP ablation on average by 23% (220 ± 46 ms pre-ablation versus 269 ± 59 ms post-ablation, p = 0.002). Four patients experienced POAF; there was no evident correlation between POAF and AERP data. Only three of the enrolled patients had a prior history of AF; none of these exhibited AF on 24-hour Holter monitoring at 3-month follow-up. Conclusions Selective epicardial PFA of GPs is feasible and safe. An acute increase in atrial tissue refractoriness is promising but further studies are required to see how this translates to longer term outcomes in symptomatic AF patients and in a percutaneous epicardial access setting.

Axonal Excitability Findings in Acute Inflammatory Demyelinating Polyneuropathy Related to SARS-CoV-2

Guillain–Barré syndrome (GBS) is a disorder of the peripheral nervous system characterized by acute-onset ascendance paresis. We present a patient who was diagnosed as having facial-onset acute inflammatory demyelinating polyneuropathy after being infected with SARS-CoV-2. A 51-year-old man presented to the emergency department with facial diplegia. He then developed bilateral ascendance paralysis. He had noticed that for 1 month, he had smell and taste disturbances. SARS-CoV-2 infection was suspected. Nasopharyngeal swab polymerase chain reaction test was negative, but anti-SARS-CoV-2 antibody was found to be positive. A nerve conduction study showed prolonged motor distal and F wave latencies with decreased motor and sensory compound muscle action potential amplitudes. Lumbar puncture revealed albuminocytologic dissociation. According to the neurologic examination and laboratory findings, the patient was diagnosed as having acute inflammatory demyelinating polyneuropathy. An axonal excitability study revealed fanning in pattern with prolonged refractoriness, which indicates nodal sodium channel disturbances. Facial-onset SARS-CoV-2–related GBS has been rarely reported; however, facial involvement seems to be one of the features of the neurologic findings.

CaMKII inhibition has dual effects on spontaneous Ca2+ release and Ca2+ alternans in ventricular cardiomyocytes from mice with a gain-of-function RyR2 mutation.

In conditions with abnormally increased activity of the cardiac ryanodine receptor (RyR2), Ca2+/calmodulin-dependent protein kinase II (CaMKII) can contribute to a further destabilization of RyR2 that results in triggered arrhythmias. Therefore, inhibition of CaMKII in such conditions has been suggested as a strategy to suppress RyR2 activity and arrhythmias. However, suppression of RyR2 activity can lead to the development of arrhythmogenic Ca2+ alternans. The aim of this study was to test whether the suppression of RyR2 activity caused by inhibition of CaMKII increases propensity for Ca2+ alternans. We studied spontaneous Ca2+ release events and Ca2+ alternans in isolated left ventricular cardiomyocytes from mice carrying the gain-of-function RyR2 mutation RyR2-R2474S and from wild-type mice. CaMKII inhibition by KN-93 effectively decreased the frequency of spontaneous Ca2+ release events in RyR2-R2474S cardiomyocytes exposed to the β-adrenoceptor agonist isoprenaline. However, KN-93-treated RyR2-R2474S cardiomyocytes also showed increased propensity for Ca2+ alternans and increased Ca2+ alternans ratio compared with both an inactive analog of KN-93 and with vehicle-treated controls. This increased propensity for Ca2+ alternans was explained by prolongation of Ca2+ release refractoriness. Importantly, the increased propensity for Ca2+ alternans in KN-93-treated RyR2-R2474S cardiomyocytes did not surpass that of wild type. In conclusion, inhibition of CaMKII efficiently reduces spontaneous Ca2+ release but promotes Ca2+ alternans in RyR2-R2474S cardiomyocytes with a gain-of-function RyR2 mutation. The dominant effect in RyR2-R2474S is to reduce spontaneous Ca2+ release, which supports this intervention as a therapeutic strategy in this specific condition. However, future studies on CaMKII inhibition in conditions with increased propensity for Ca2+ alternans should include investigation of both phenomena.NEW & NOTEWORTHY Genetically increased RyR2 activity promotes arrhythmogenic Ca2+ release. Inhibition of CaMKII suppresses RyR2 activity and arrhythmogenic Ca2+ release. Suppression of RyR2 activity prolongs refractoriness of Ca2+ release. Prolonged refractoriness of Ca2+ release leads to arrhythmogenic Ca2+ alternans. CaMKII inhibition promotes Ca2+ alternans by prolonging Ca2+ release refractoriness.

CaMKII as a therapeutic target in catecholaminergic polymorphic ventricular tachycardia type 1

Abstract Funding Acknowledgements Type of funding sources: Public Institution(s). Main funding source(s): South-Eastern Norway Regional Health Authority Background In catecholaminergic polymorphic ventricular tachycardia type 1 (CPVT1) adrenergic activation of the cardiac ryanodine receptor (RyR2) causes spontaneous calcium release and triggers arrhythmias. Calcium/calmodulin-dependent protein kinase II (CaMKII) can contribute to such arrhythmogenic calcium release and has been proposed as a therapeutic target in CPVT1. To predict the efficacy and safety of this strategy, it is necessary to know whether the mechanism for CaMKII activation is important for its arrhythmogenic effects, and if inhibition has proarrhythmic effects. We tested (1) if oxidation of CaMKII contributes to spontaneous calcium release in CPVT1 and (2) if inhibition of CaMKII in this condition can induce calcium alternans. Methods Mice with the CPVT1-causative mutation RyR2-R2474S (RyR2-RS) were crossed with mice with the CaMKII M281/282V (MMVV) mutation that prevents CaMKII M281/282 oxidation, to create double mutants (RyR2-RSxMMVV). Telemetric ECG surveillance was used to study in vivo arrhythmias following an adrenergic challenge by i.p. administration of the beta-adrenoceptor agonist isoprenaline. Confocal line-scan imaging and whole-cell calcium imaging were used to study arrhythmogenic calcium release in isolated left ventricular cardiomyocytes during stimulation with isoprenaline. Results As expected, RyR2-RS mice exhibited more arrhythmic events and spontaneous calcium release (i.e. calcium sparks and calcium waves) compared to wild-type mice. Treatment of RyR2-RS cardiomyocytes with either the CaMKII inhibitor KN-93 or the antioxidant n-acetyl-cysteine reduced spontaneous calcium release (i.e. calcium sparks and calcium waves, for KN-93 and n-acetyl-cysteine, respectively). Interestingly, CaMKII inhibition by KN-93 also increased both incidence and degree of arrhythmogenic calcium alternans in RyR2-RS cardiomyocytes. This adverse effect was a result of prolonged refractoriness of calcium release. Furthermore, to test whether the protective effect of antioxidant treatment in RyR2-RS was mediated via CaMKII oxidation, we compared arrhythmias and spontaneous calcium release (i.e. calcium waves) in RyR2-RSxMMVV with RyR2-RS. However, these two genotypes did not differ in either incidence or severity of arrhythmias, and showed similar degree of spontaneous calcium release. Conclusions Inhibition of CaMKII protects against spontaneous calcium release in CPVT1, and is a promising therapeutic strategy. However, the fact that such inhibition also induces calcium alternans needs further exploration. Antioxidative agents also attenuate arrhythmogenic calcium release in CPVT1 cardiomyocytes, but this effect does not seem to involve the M281/282 CaMKII oxidation site. Future studies should explore other oxidation sites.

A case of infra-nodal Wenckebach conduction block with alternating bundle branch block

BackgroundAtrioventricular (AV) node normally has decremental conduction property and a longer refractory period than His-Purkinje system (HPS). This results in AV conduction delay or block at the level of AV node in response to short-coupled atrial premature beats. Prolonged refractoriness in HPS can produce unusual physiological patterns of AV conduction such as conduction delay or infra-nodal block in the distal elements of HPS.Case presentationWe present a case in which atrial premature stimulation produces infra-nodal Wenckebach conduction block which initiates long-short cycle sequence within the bundle branches resulted in alternating bundle branch block and atypical pattern of Ashman phenomenon.ConclusionsThis case highlights the importance of recognizing the unusual physiological AV conduction patterns of HPS. The long-short cycle sequence in the bundle branches of distal HPS and linking phenomenon can result in alternating bundle branch block without the presence of HPS disease.

Парадоксальная рефрактерность, возникающая при аллогенном переносе животным с аллоксановым диабетом суммарной РНК лимфоцитов селезенки вылеченных особей

Ранее нами было показано, что препараты аллогенной суммарной РНК, выделенной из лимфоидных и стволовых клеток здоровых животных, вызывают нормализацию уровня глюкозы крови у крыс со стойким аллоксановым сахарным диабетом. Цель исследования -- выяснение влияния суммарной РНК лимфоцитов селезенки крыс, ранее перенесших стойкий аллоксановый диабет, на особенности течения эксперименального аллоксанового диабета у животных. Методика. Эксперимент выполнен на 35 белых беспородных крысах-самках массой 250-280 г: 5 интактных животных, 20 - с экспериментальным аллоксановым сахарным диабетом и 10 животных, ранее перенесших стойкий аллоксановый диабет, у которых уровень глюкозы крови был полностью нормализован введением суммарных РНК клеток костного мозга, селезенки и поджелудочной железы. Диабет у этих животных моделировали однократным подкожным введением полного адъюванта Фрейнда (0,5 мл на крысу) и последующим подкожным введением аллоксана тригидрата в дозе 200 мг/кг. Достигнутая нормогликемия у животных подтверждалась в течение последующих 60 сут. Из селезенок и костного мозга этих животных методом фенол-хлороформной экстракции была выделена суммарная РНК. На 20 крысах с экспериментальным аллоксановым сахарным диабетом изучали эффекты однократного внутрибрюшинного введения полученной суммарной РНК (15 мкг/100 г массы тела животного). Результаты. Обнаружено, что при введении суммарной РНК селезенки крыс, ранее перенесших аллоксановый диабет, животным с аллоксановым диабетом у последних развивалась стойкая и длительная рефрактерность к лечению терапевтическими препаратами РНК. Заключение. 1. Выявлен феномен повышенной чувствительности крыс с аллоксановым диабетом к действию суммарной РНК селезенки животных, ранее перенесших аллоксановый диабет. 2. Предполагаемой причиной рефрактерности является клональный компонент суммарной РНК CD8+ Т-лимфоцитов памяти из селезенки животных, ранее перенесших аллоксановый диабет. 3. Предполагается наличие механизма адресного взаимодействия отдельных компонентов суммарной РНК селезенки крыс, ранее перенесших аллоксановый диабет, с их лимфоцитами-мишенями в организме реципиентов с аллоксановым диабетом. Introduction. Earlier we have shown that preparations of allogenic total RNA from lymphoid and stem cells of healthy animals contribute to normalization of blood glucose levels in white outbred rats with persistent alloxan-induced diabetes mellitus. The aim of this study was to find out whether allogenic total RNA isolated from the spleen of rats treated for alloxan diabetes affects its course, as determined by changes in blood glucose, in animals with persistent alloxan diabetes. Methods. Experiments were performed on 35 white outbred female rats weighing 250-280 g. Rats were divided into intact animals (n=5), rats with experimental alloxan diabetes mellitus (n=20), and rats after persistent alloxan diabetes (n=10) whose blood glucose level had been completely normalized by administrating total RNA of bone marrow, splenic, and pancreatic cells (post-diabetes group). Diabetes mellitus was modeled with a single subcutaneous (s.c.) injection of complete Freund’s adjuvant (0.5 ml) followed by a s.c. injection of alloxan trihydrate (200 mg/kg). Achievement of normoglycemia in animals of the post-diabetes group was confirmed over the next 60 days. Then total RNA was isolated from their spleen and bone marrow by phenol-chloroform extraction, and the effect of a single intraperitoneal injection of total RNA (15 μg/100 g body weight) was studied. Results. Administration of splenic total RNA from rats previously treated for alloxan diabetes to animals with alloxan diabetes resulted in development of stable and prolonged refractoriness of diabetic rats to the treatment with therapeutic RNA preparations [11]. Conclusions. This study discovered a phenomenon of hypersensitivity of rats with alloxan diabetes to the diabetogenic effect of total RNA from animals that had previously had alloxan diabetes. Apparently, this refractoriness was caused by the clonal component of the total RNA of CD8 + T memory lymphocytes from the spleen of post-diabetes animals. A mechanism is proposed for the interaction between individual components of splenic total RNA from post-diabetes rats and their target lymphocytes in recipients with alloxan diabetes.

P699Mechanisms of action of the small conductance Ca2+-activated K+-channel modulator AP30663, a novel compound being developed for treatment of atrial fibrillation in man

Abstract Background Small conductance Ca2+-activated K+-channels (SK-channels) are a promising new atrial selective target for treatment of atrial fibrillation (AF). AP30663 is a small molecule inhibitor of SK-channels that is effective in converting vernakalant-resistant AF in tachy-paced pigs. Detailed understanding of the molecular mechanism of AP30663 is important for the development of SK channel inhibition for use in man. Purpose To establish the electrophysiological profile, mechanism of action and efficacy in prolonging atrial refractoriness ex vivo of AP30663. Methods AP30663 potency and mechanism of action were established by whole cell and inside-out patch clamp recordings of expressed SK channels. The ion channel selectivity profile of AP30663 was investigated on heterologous expressed channels. Effects of AP30663 or vehicle (DMSO) on atrial refractoriness (AERP) and ventricular repolarization (QTcB) were investigated on isolated perfused guinea pig hearts. Results AP30663 was found to be a selective negative allosteric modulator of SK channels (IC50=0.77±x0.13 μM) with no or minor effects on a panel of other cardiac ion channels, including hERG/KV11.1, (IKr), KV7.1/KCNE1 (IKs), KV4.3/KChiP2 (Ito), Kir2.1 (IK1), Kir3.1/Kir3.4 (IKACh), KV1.5 (IKur), NaV1.5 (INa) and CaV1.2 (ICa). AP30663 inhibited the SK-channel by right-shifting the calcium activation curve of the SK-channel (the EC50 of Ca2+ increased from 0.43±0.02 μM (control, n=6) to 1.37±0.05 μM (in the presence of 7μM AP30663, n=6). In isolated guinea pig hearts, administration of vehicle had no effect on AERP or QTcB. AP30663 significantly prolonged the AERP in 3 μM (to 131±6% of baseline) and 10 μM (to 165±3% of baseline) without any effects on the QTcB. Conclusion AP30663 is a selective negative allosteric modulator of SK channels, acting by means of shifting the calcium dependence of SK-channel activation. AP30663 prolonged atrial refractoriness without affecting the QT-interval in isolated perfused heart preparations. These properties support continued development of AP30663 for treatment of AF in man. Acknowledgement/Funding Innovation Fund Denmark, Wellcome Trust

Electrocardiographic Changes and Arrhythmia in Fabry Disease.

Fabry disease is an X-chromosome-linked lysosomal storage disease characterized by a deficient activity or, in most males, absence of the enzyme α-galactosidase A (a-Gal A) leading to systemic, primary lysosomal accumulation of globotriaosylceramide (Gb3) (1). Recent literature refers to an overall birth prevalence of 1:40,000–170,000; however, such data do not allow an estimation on an actual patient number suffering from Fabry disease (2). Multisystem morbidity commonly develops in childhood and, with progression of the disease, life-threatening complications often occur in adulthood, including renal failure, cardiovascular dysfunction, neuropathy, and stroke (3–6). Life expectancy is reduced by an average of 15 years in female patients and 20 years in male patients (7, 8). The pathognomonic Gb3 accumulation has been repeatedly observed over the past decades by many groups in vascular endothelial and smooth muscle cells, cardiomyocytes, cardiac conduction tissue, and valvular fibroblasts (3). Although incompletely described, it is likely that inflammatory and neurohormonal mechanisms are involved in subsequent cellular and vascular dysfunction, leading to tissue ischemia, hypertrophy, and fibrosis (9). Furthermore, recently published works on cardiomyocyte dysfunction and conduction tissue involvement have suggested that cardiac dysfunction may reflect increased myocardial nitric oxide production with oxidative damage of cardiomyocyte myofilaments and DNA, causing cell dysfunction and death, and accelerated conduction with prolonged refractoriness and electric instability (10, 11).

Early afterdepolarizations promote transmural reentry in ischemic human ventricles with reduced repolarization reserve

AimsAcute ischemia is a major cause of sudden arrhythmic death, further promoted by potassium current blockers. Macro-reentry around the ischemic region and early afterdepolarizations (EADs) caused by electrotonic current have been suggested as potential mechanisms in animal and isolated cell studies. However, ventricular and human-specific arrhythmia mechanisms and their modulation by repolarization reserve remain unclear. The goal of this paper is to unravel multiscale mechanisms underlying the modulation of arrhythmic risk by potassium current (IKr) block in human ventricles with acute regional ischemia. Methods and resultsA human ventricular biophysically-detailed model, with acute regional ischemia is constructed by integrating experimental knowledge on the electrophysiological ionic alterations caused by coronary occlusion. Arrhythmic risk is evaluated by determining the vulnerable window (VW) for reentry following ectopy at the ischemic border zone. Macro-reentry around the ischemic region is the main reentrant mechanism in the ischemic human ventricle with increased repolarization reserve due to the ATP-sensitive potassium current (IK(ATP)) activation. Prolongation of refractoriness by 4% caused by 30% IKr reduction counteracts the establishment of macro-reentry and reduces the VW for reentry (by 23.5%). However, a further decrease in repolarization reserve (50% IKr reduction) is less anti-arrhythmic despite further prolongation of refractoriness. This is due to the establishment of transmural reentry enabled by electrotonically-triggered EADs in the ischemic border zone. EADs are produced by L-type calcium current (ICaL) reactivation due to prolonged low amplitude electrotonic current injected during the repolarization phase. ConclusionsElectrotonically-triggered EADs are identified as a potential mechanism facilitating intramural reentry in a regionally-ischemic human ventricles model with reduced repolarization reserve.

Pathology and Function of Conduction Tissue in Fabry Disease Cardiomyopathy

Cardiac arrhythmias are common in Fabry disease (FD) and may occur in prehypertrophic cardiomyopathy suggesting an early compromise of conduction tissue (CT). Therefore, FD X-linked and CT may be variously involved in male and female patients with FD cardiomyopathy, affecting CT function. Among 74 patients with endomyocardial biopsy diagnosis of FD cardiomyopathy, 13 (6 men; 7 women; mean age, 50.1±13.5 years; maximal wall thickness, 16.7±3.7 mm) had CT included in histological specimens and 6 also at electron microscopy. CT glycolipid infiltration was defined as focal, moderate, extensive, or massive, if involved ≤30%, ≤50%, >50%, or 100% of cells; identified as loosely arranged small myocytes positive to HCN4 immunostaining, supplied by a centrally placed thick-walled arteriole. CT involvement was correlated with age, sex, and α-Gal gene mutation. CT function was evaluated by electrophysiological study and arrhythmias at Holter registration. CT infiltration was focal/moderate in 4 women with no arrhythmias and normal electrophysiological study, extensive in 3 women with atrial or ventricular arrhythmias and short HV interval, and massive in 6 men with atrial fibrillation or ventricular arrhythmias and short HV. Short PR/AH with increased refractoriness was additionally found in 3 patients with extensive/massive CT infiltration. A male patient with the shortest HV presented infra-Hissian block during decremental atrial stimulation. There was no correlation with age, maximal wall thickness, and type of gene mutation. CT infiltration in FD cardiomyopathy is constant in men and variable in women because of skewed X-chromosome inactivation; its extensive/massive involvement causes accelerated conduction with prolonged refractoriness and electric instability.

A computational modelling approach combined with cellular electrophysiology data provides insights into the therapeutic benefit of targeting the late Na+ current.

The ventricular action potential plateau is a phase of high resistance, which makes ventricular myocytes vulnerable to small electrical perturbations. We developed a computationally based model of GS-458967 interaction with the cardiac Na+ channel, informed by experimental data recorded from guinea pig isolated single ventricular myocytes. The model predicts that the therapeutic potential of GS-458967 derives largely from the designed property of significant potent selectivity for INaL. Selective inhibition of the slowly inactivating or late Na(+) current (INaL) in patients with inherited or acquired arrhythmia syndrome may confer therapeutic benefit by reducing the incidence of triggers for arrhythmia and suppressing one component of arrhythmia-promoting cardiac substrates (e.g. prolonged refractoriness and spatiotemporal dispersion of action potential duration). Recently, a novel compound that preferentially and potently reduces INaL, GS-458967 (IC50 for block of INaL = 130 nM) has been studied. Experimental measurements of the effects of GS-458967 on endogenous INaL in guinea pig ventricular myocytes demonstrate a robust concentration-dependent reduction in action potential duration (APD). Using experimental data to calibrate INaL and the rapidly activating delayed rectifier K(+) current, IKr, in the Faber-Rudy computationally based model of the guinea pig ventricular action potential, we simulated effects of GS-458967 on guinea pig ventricular APD. GS-458967 (0.1 μM) caused a 28.67% block of INaL and 12.57% APD shortening in experiments, while the model predicted 10.06% APD shortening with 29.33% block of INaL. An additional effect of INaL block is to reduce the time during which the membrane potential is in a high resistance state (i.e. the action potential plateau). To test the hypothesis that targeted block of INaL would make ventricular myocytes less susceptible to small electrical perturbations, we used the computational model to test the degree of APD prolongation induced by small electrical perturbations in normal cells and in cells with simulated long QT syndrome. The model predicted a substantial dose-dependent reduction in sensitivity to small electrical perturbations as evidenced by action potential duration at 90% repolarization variability in the presence of GS-458967-induced INaL block. This effect was especially potent in the 'disease setting' of inherited long QT syndrome. Using a combined experimental and theoretical approach, our results suggest that INaL block is a potent therapeutic strategy. This is because reduction of INaL stabilizes the action potential waveform by reducing depolarizing current during the plateau phase of the action potential. This reduces the most vulnerable phase of the action potential with high membrane resistance. In summary, by reducing the sensitivity of the myocardial substrate to small electrical perturbations that promote arrhythmia triggers, agents such as GS-458967 may constitute an effective antiarrhythmic pharmacological strategy.

Role for MicroRNA-21 in Atrial Profibrillatory Fibrotic Remodeling Associated With Experimental Postinfarction Heart Failure

Atrial tissue fibrosis is often an important component of the atrial fibrillation (AF) substrate. Small noncoding microRNAs are important mediators in many cardiac remodeling paradigms. MicroRNA-21 (miR-21) has been suggested to be important in ventricular fibrotic remodeling by downregulating Sprouty-1, a protein that suppresses fibroblast proliferation. The present study examined the potential role of miR-21 in the atrial AF substrate resulting from experimental heart failure after myocardial infarction (MI). Large MIs (based on echocardiographic left ventricular wall motion score index) were created by left anterior descending coronary artery ligation in rats. Changes induced by MI versus sham controls were first characterized with echocardiography, histology, biochemistry, and in vivo electrophysiology. Additional MI rats were then randomized to receive anti-miR-21 (KD21) or scrambled control sequence (Scr21) injections into the left atrial myocardium. Progressive left ventricular enlargement, hypocontractility, left atrial dilation, fibrosis, refractoriness prolongation, and AF promotion occurred in MI rats versus sham controls. Atrial tissues of MI rats showed upregulation of miR-21, along with dysregulation of the target genes Sprouty-1, collagen-1, and collagen-3. KD21 treatment reduced atrial miR-21 expression levels in MI rats to values in sham rats, decreased AF duration from 417 (69-1595; median [Q1-Q3]) seconds to 3 (2-16) seconds (8 weeks after MI; P<0.05), and reduced atrial fibrous tissue content from 14.4 ± 1.8% (mean ± SEM) to 4.9 ± 1.2% (8 weeks after MI; P<0.05) versus Scr21 controls. MI-induced heart failure leads to AF-promoting atrial remodeling in rats. Atrial miR-21 knockdown suppresses atrial fibrosis and AF promotion, implicating miR-21 as an important signaling molecule for the AF substrate and pointing to miR-21 as a potential target for molecular interventions designed to prevent AF.

Supernormal conduction and linking in an accessory AV pathway

1 IntroductionSpontaneous intermittent block over an accessory pathwaymanifested as alternating QRS complexes of pre-excitedand normally conducted beats suggests a prolonged ante-grade refractory period [1]. Similarly, sudden loss of thedelta wave during exercise [2] is also considered evidenceof prolonged refractoriness over an accessory A–Vpathway. However, the antegrade refractory period of anaccessory A–V pathway is not the only determinant ofconduction over bypass tracts. Two other important butfrequently overlooked electrophysiologic mechanismsmodulate conduction over accessory A–V pathways:supernormal conduction [3]andlinking[4] between thenormal conduction system and the accessory A–Vpathway.We present a patient that, despite intermittent preexcita-tion at rest and loss of preexcitation during exercise, had anaccessory A–V pathway able to conduct at fast rates asdemonstrated during the electrophysiology study. Bothsupernormal conduction and the linking phenomenonaccounted for this unexpected behavior.2 Case reportAn 18-year-old asymptomatic male with a structurallynormal heart was found to have intermittent ventricular

Estradiol and Progesterone Exert Opposite Effects on Cardiac Repolarization and Arrhythmogenesis in Transgenic Long QT Syndrome 2 Rabbits

Introduction: Adult women with LQT2 are at higher risk for sudden cardiac death (SCD) than men with an increased risk during the postpartum. We have created transgenic rabbits over-expressing a pore mutant of the human ERG channel in the heart (LQT2) and showing the human long QT phenotype (Brunner et al. JCI, 2008). 4/4 LQT2 females used for breeding died of SCD during the postpartum. We hypothesize that sex hormones modulate cardiac repolarization and arrhythmogenesis in LQT2 females.Methods: Prepubertal ovariectomized LQT2 females were implanted with 90day release-pellets of estradiol (EST), progesterone (Prog), dihydrotestosterone (DHT), or placebo (OVX) (n=6 each). All groups underwent telemetric ECG monitoring and in vivo electrophysiological studies (EPS) after 8 weeks and first optical mapping experiments were performed in OVX, EST- and DHT-treated rabbits.Results: EST treatment steepened the QT/RR slope of prepubertal rabbits (p<0.05), whereas DHT or OVX decreased the QT/RR slope steepness (p<0.05). Prog did not alter the QT/RR slope. In vivo EPS revealed a longer ventricular refractory period (VERP) in EST- than in DHT- or Prog-treated rabbits (DHT: p<0.05, Prog: p<0.01). Within 8 weeks of hormone-treatment, 4 of 6 EST-treated rabbits died of polymorphic VT, while no SCD occurred in 6 DHT- and 6 Prog-treated LQT2 females (p<0.05). Preliminary optical mapping experiments revealed heterogeneous APD dispersion with islands of prolonged refractoriness in EST- rabbits contrasting with smoother APD dispersion in OVX and DHT-treated rabbits.Conclusions: EST increases the QT/RR steepness and prolongs cardiac refractoriness, whereas DHT and Prog shorten cardiac refractoriness. EST predisposes prepubertal LQT2 rabbits to polymorphic VT. Heterogeneous ADP dispersion might underlie this proarrhythmic effect of EST. We are currently using high-throughput molecular approaches to elucidate the underlying mechanisms.

Increased cycle length during long-duration ventricular fibrillation is caused by decreased upstroke velocity as well as prolonged refractoriness

Cycle length (CL) increases as ventricular fibrillation (VF) progresses. The purpose of this study was to test the hypotheses that increased CL is due to increased diastolic interval (DI), not increased action potential duration (APD), and that the DI increase is not solely due to increased postrepolarization refractoriness. In 10 swine, VF was recorded for 20 minutes using a floating microelectrode through a hole in a 504-electrode epicardial plaque. Mean APD, DI, action potential amplitude (APA), maximum change in voltage during the AP upstroke (V(max)), and CL were calculated from the floating microelectrode recordings each minute of VF. The refractory period was estimated from the minimum DI (DI(min)). In two animals, rapid pacing was performed to gauge refractoriness. As VF progressed, CL, DI, and DI(min) increased (P <.05), whereas APD, V(max), and APA decreased (P <.05). At 20 minutes, DI(min) was not different from mean DI at VF onset. Pacing captured, but 53% of paced wavefronts blocked within the plaque. Increasing CL in VF is due to increased DI and not APD, which shortens. The increase in DI(min) over time is much less than the increase in mean DI, indicating that the myocardium is excitable during much of the DI. This finding, along with the ability to pace at a CL shorter than the native VF CL and the poor paced wavefront propagation, suggests that the increase in DI is due not only to increased postrepolarization refractoriness but also to poor wavefront propagation because of decreased APA and V(max) secondary to global ischemia caused by VF.

Short-term pacing in the mouse alters cardiac expression of connexin43

BackgroundCardiac insults such as ischemia, infarction, hypertrophy and dilatation are often accompanied by altered abundance and/or localization of the connexin43 gap junction protein, which may predispose towards arrhythmic complications. Models of chronic dyssynchronous cardiac activation have also been shown to result in redistribution of connexin43 in cardiomyocytes. We hypothesized that alterations in connexin43 expression and localization in the mouse heart might be induced by ventricular pacing over a short period of time.ResultsThe subdiaphragmatic approach was used to pace a series of wild type mice for six hours before the hearts were removed for analysis. Mice were paced at 10–15% above their average anesthetized sinus rate and monitored to ensure 1:1 capture. Short-term pacing resulted in a significant reduction in connexin43 mRNA abundance, a partial redistribution of connexin43 from the sarcolemma to a non-sarcolemmal fraction, and accumulation of ubiquitinated connexin43 without a significant change in overall connexin43 protein levels. These early pacing-induced changes in connexin43 expression were not accompanied by decreased cardiac function, prolonged refractoriness or increased inducibility into sustained arrhythmias.ConclusionOur data suggest that short-term pacing is associated with incipient changes in the expression of the connexin43 gap junction, possibly including decreased production and a slowed rate of degradation. This murine model may facilitate the study of early molecular changes induced by pacing and may ultimately assist in the development of strategies to prevent gap junction remodeling and the associated arrhythmic complications of cardiac disease.

Electrophysiological Effects of Carvedilol Administration in Patients with Dilated Cardiomyopathy

Several studies suggest the clinical efficacy of carvedilol in reducing atrial and ventricular arrhythmias in patients with left ventricular dysfunction (LVD) due to congestive heart failure (CHF) or following myocardial infarction. However, the mechanisms supporting its antiarrhythmic efficacy have been derived from experimental studies. In this prospective, placebo-controlled trial we examined the electrophysiological effects of a high oral dose of carvedilol in patients with CHF and LVD due to non-ischemic dilated cardiomyopathy. Thirty-one patients with stable CHF underwent electrophysiological study and were randomly assigned to treatment with carvedilol or placebo. After 2 months of treatment the study was repeated. Carvedilol prolonged almost all conduction times. In the same group atrial and ventricular effective refractory periods were significantly prolonged, while the parameters of repolarization remained virtually unchanged. The prolongation of refractoriness was most pronounced in the atrium. The change in ventricular refractoriness was correlated with ejection fraction (r = 0.94, p < 0.01) suggesting that patients with more preserved left ventricular function responded to treatment with greater prolongation. Even after a short period of administration carvedilol has marked and diffused electrophysiological effects that would be beneficial for patients with CHF and may contribute to the positive outcome of clinical trials.