Arrhythmogenic Right Ventricular Cardiomyopathy: Considerations from in Silico Experiments

Objective: Arrhythmogenic right ventricular cardiomyopathy (ARVC) is associated with remodeling of gap junctions and also, although less well-defined, down-regulation of the fast sodium current. The gap junction remodeling and down-regulation of sodium current have been proposed as contributors to arrhythmogenesis in ARVC by slowing conduction. The objective of the present study was to assess the amount of conduction slowing due to the observed gap junction remodeling and down-regulation of sodium current. Methods: The effects of (changes in) gap junctional conductance, cell dimensions, and sodium current on both longitudinal and transversal conduction velocity were tested by simulating action potential propagation in linear strands of human ventricular cells that were either arranged end-to-end or side-by-side. Results: A 50% reduction in gap junction content, as commonly observed in ARVC, gives rise to an 11% decrease in longitudinal conduction velocity and a 29% decrease in transverse conduction velocity. A down-regulation of the sodium current through a 50% decrease in peak current density as well as a −15 mV shift in steady-state inactivation, as observed in an experimental model of ARVC, decreases conduction velocity in either direction by 32%. In combination, the gap junction remodeling and down-regulation of sodium current result in a 40% decrease in longitudinal conduction velocity and a 52% decrease in transverse conduction velocity. Conclusion: The gap junction remodeling and down-regulation of sodium current do result in conduction slowing, but heterogeneity of gap junction remodeling, in combination with down-regulation of sodium current, rather than gap junction remodeling per se may be a critical factor in arrhythmogenesis in ARVC

Cellular Mechanisms of Sinus Node Dysfunction in Carriers of the SCN5A-E161K Mutation and Role of the H558R Polymorphism

Background: Carriers of the E161K mutation in the SCN5A gene, encoding the NaV1.5 pore-forming α-subunit of the ion channel carrying the fast sodium current (INa), show sinus bradycardia and occasional exit block. Voltage clamp experiments in mammalian expression systems revealed a mutation-induced 2.5- to 4-fold reduction in INa peak current density as well as a +19 mV shift and reduced steepness of the steady-state activation curve. The highly common H558R polymorphism in NaV1.5 limits this shift to +13 mV, but also introduces a -10 mV shift in steady-state inactivation.Aim: We assessed the cellular mechanism by which the E161K mutation causes sinus node dysfunction in heterozygous mutation carriers as well as the potential role of the H558R polymorphism.Methods: We incorporated the mutation-induced changes in INa into the Fabbri-Severi model of a single human sinoatrial node cell and the Maleckar et al. human atrial cell model, and carried out simulations under control conditions and over a wide range of acetylcholine levels.Results: In absence of the H558R polymorphism, the E161K mutation increased the basic cycle length of the sinoatrial node cell from 813 to 866 ms. In the simulated presence of 10 and 25 nM acetylcholine, basic cycle length increased from 1027 to 1131 and from 1448 to 1795 ms, respectively. The increase in cycle length was the result of a significant slowing of diastolic depolarization. The mutation-induced reduction in INa window current had reduced the contribution of the mutant component of INa to the net membrane current during diastolic depolarization to effectively zero. Highly similar results were obtained in presence of the H558R polymorphism. Atrial excitability was reduced, both in absence and presence of the H558R polymorphism, as reflected by an increase in threshold stimulus current and a concomitant decrease in capacitive current of the atrial cell.Conclusion: We conclude that the experimentally identified mutation-induced changes in INa can explain the clinically observed sinus bradycardia and potentially the occasional exit block. Furthermore, we conclude that the common H558R polymorphism does not significantly alter the effects of the E161K mutation and can thus not explain the reduced penetrance of the E161K mutation

Effects of Acetylcholine and Noradrenalin on Action Potentials of Isolated Rabbit Sinoatrial and Atrial Myocytes

The autonomic nervous system controls heart rate and contractility through sympathetic and parasympathetic inputs to the cardiac tissue, with acetylcholine (ACh) and noradrenalin (NA) as the chemical transmitters. In recent years, it has become clear that specific Regulators of G protein Signaling proteins (RGS proteins) suppress muscarinic sensitivity and parasympathetic tone, identifying RGS proteins as intriguing potential therapeutic targets. In the present study, we have identified the effects of 1 μM ACh and 1 μM NA on the intrinsic action potentials of sinoatrial (SA) nodal and atrial myocytes. Single cells were enzymatically isolated from the SA node or from the left atrium of rabbit hearts. Action potentials were recorded using the amphotericin-perforated patch-clamp technique in the absence and presence of ACh, NA, or a combination of both. In SA nodal myocytes, ACh increased cycle length and decreased diastolic depolarization rate, whereas NA decreased cycle length and increased diastolic depolarization rate. Both ACh and NA increased maximum upstroke velocity. Furthermore, ACh hyperpolarized the maximum diastolic potential. In atrial myocytes stimulated at 2 Hz, both ACh and NA hyperpolarized the maximum diastolic potential, increased the action potential amplitude, and increased the maximum upstroke velocity. Action potential duration at 50 and 90% repolarization was decreased by ACh, but increased by NA. The effects of both ACh and NA on action potential duration showed a dose dependence in the range of 1–1000 nM, while a clear-cut frequency dependence in the range of 1–4 Hz was absent. Intermediate results were obtained in the combined presence of ACh and NA in both SA nodal and atrial myocytes. Our data uncover the extent to which SA nodal and atrial action potentials are intrinsically dependent on ACh, NA, or a combination of both and may thus guide further experiments with RGS proteins

Is sodium current present in human sinoatrial node cells?

Pacemaker activity of the sinoatrial node has been studied extensively in various animal species, but is virtually unexplored in man. As such, it is unknown whether the fast sodium current (INa) plays a role in the pacemaker activity of the human sinoatrial node. Recently, we had the unique opportunity to perform patch-clamp experiments on single pacemaker cells isolated from a human sinoatrial node. In 2 out of the 3 cells measured, we observed large inward currents with characteristics of INa. Although we were unable to analyze the current in detail, our findings provide strong evidence that INa is present in human sinoatrial node pacemaker cells, and that this INa is functionally available at potentials negative to -60 mV

Long QT Syndrome and Sinus Bradycardia–A Mini Review

Congenital long-QT syndrome (LQTS) is an inherited cardiac disorder characterized by the prolongation of ventricular repolarization, susceptibility to Torsades de Pointes (TdP), and a risk for sudden death. Various types of congenital LQTS exist, all due to specific defects in ion channel-related genes. Interestingly, almost all of the ion channels affected by the various types of LQTS gene mutations are also expressed in the human sinoatrial node (SAN). It is therefore not surprising that LQTS is frequently associated with a change in basal heart rate (HR). However, current data on how the LQTS-associated ion channel defects result in impaired human SAN pacemaker activity are limited. In this mini-review, we provide an overview of known LQTS mutations with effects on HR and the underlying changes in expression and kinetics of ion channels. Sinus bradycardia has been reported in relation to a large number of LQTS mutations. However, the occurrence of both QT prolongation and sinus bradycardia on a family basis is almost completely limited to LQTS types 3 and 4 (LQT3 and Ankyrin-B syndrome, respectively). Furthermore, a clear causative role of this sinus bradycardia in cardiac events seems reserved to mutations underlying LQT3

Acetylcholine Reduces L-Type Calcium Current without Major Changes in Repolarization of Canine and Human Purkinje and Ventricular Tissue

Vagal nerve stimulation (VNS) holds a strong basis as a potentially effective treatment modality for chronic heart failure, which explains why a multicenter VNS study in heart failure with reduced ejection fraction is ongoing. However, more detailed information is required on the effect of acetylcholine (ACh) on repolarization in Purkinje and ventricular cardiac preparations to identify the advantages, risks, and underlying cellular mechanisms of VNS. Here, we studied the effect of ACh on the action potential (AP) of canine Purkinje fibers (PFs) and several human ventricular preparations. In addition, we characterized the effects of ACh on the L-type Ca2+ current (I-CaL) and AP of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) and performed computer simulations to explain the observed effects. Using microelectrode recordings, we found a small but significant AP prolongation in canine PFs. In the human myocardium, ACh slightly prolonged the AP in the midmyocardium but resulted in minor AP shortening in subepicardial tissue. Perforated patch-clamp experiments on hiPSC-CMs demonstrated that 5 mu M ACh caused an approximate to 15% decrease in I-CaL density without changes in gating properties. Using dynamic clamp, we found that under blocked K+ currents, 5 mu M ACh resulted in an approximate to 23% decrease in AP duration at 90% of repolarization in hiPSC-CMs. Computer simulations using the O'Hara-Rudy human ventricular cell model revealed that the overall effect of ACh on AP duration is a tight interplay between the ACh-induced reduction in I-CaL and ACh-induced changes in K+ currents. In conclusion, ACh results in minor changes in AP repolarization and duration of canine PFs and human ventricular myocardium due to the concomitant inhibition of inward I-CaL and outward K+ currents, which limits changes in net repolarizing current and thus prevents major changes in AP repolarization

Early detection and intervention using neutrophil gelatinase-associated lipocalin (NGAL) may improve renal outcome of acute contrast media induced nephropathy: A randomized controlled trial in patients undergoing intra-arterial angiography (ANTI-CIN Study)

<p>Abstract</p> <p>Background</p> <p>Patients with pre-existing impaired renal function are prone to develop acute contrast media induced nephropathy (CIN). Neutrophil gelatinase-associated lipocalin (NGAL), a new biomarker predictive for acute kidney injury (AKI), has been shown to be useful for earlier diagnosis of CIN; however, urinary NGAL values may be markedly increased in chronic renal failure at baseline. Results from those studies suggested that urinary NGAL values may not be helpful for the clinician. An intravenous volume load is a widely accepted prophylactic measure and possibly a reasonable intervention to prevent deterioration of renal function. The aim of our study is to evaluate NGAL as an early predictor of CIN and to investigate the clinical benefit of early post-procedural i.v. hydration.</p> <p>Methods/Design</p> <p>The study will follow a prospective, open-label, randomized controlled design. Patients requiring intra-arterial contrast media (CM) application will be included and receive standardized, weight-based, intravenous hydration before investigation. Subjects with markedly increased urinary NGAL values after CM application will be randomized into one of two study groups. Group A will receive 3-4 ml/kg BW/h 0.9% saline intravenously for 6 hours. Group B will undergo only standard treatment consisting of unrestricted oral fluid intake. The primary outcome measure will be CIN defined by an increase greater than 25% of baseline serum creatinine. Secondary outcomes will include urinary NGAL values, cystatin C values, contrast media associated changes in cardiac parameters such as NT-pro-BNP/troponin T, changes in urinary cytology, need for renal replacement treatment, length of stay in hospital and death.</p> <p>We assume that 20% of the included patients will show a definite rise in urinary NGAL. Prospective statistical power calculations indicate that the study will have 80% statistical power to detect a clinically significant decrease of CIN of 40% in the treatment arm if 1200 patients are recruited into the study.</p> <p>Discussion</p> <p>A volume expansion strategy showing a benefit from earlier intervention for patients with markedly elevated urinary NGAL values, indicating a CIN, might arise from data from this study.</p> <p>Trial registration</p> <p>ClinicalTrials.gov <a href="http://www.clinicaltrials.gov/ct2/show/NCT01292317">NCT01292317</a></p

Self-restoration of cardiac excitation rhythm by anti-arrhythmic ion channel gating

Homeostatic regulation protects organisms against hazardous physiological changes. However, such regulation is limited in certain organs and associated biological processes. For example, the heart fails to self-restore its normal electrical activity once disturbed, as with sustained arrhythmias. Here we present proof-of-concept of a biological self-restoring system that allows automatic detection and correction of such abnormal excitation rhythms. For the heart, its realization involves the integration of ion channels with newly designed gating properties into cardiomyocytes. This allows cardiac tissue to i) discriminate between normal rhythm and arrhythmia based on frequency-dependent gating and ii) generate an ionic current for termination of the detected arrhythmia. We show in silico, that for both human atrial and ventricular arrhythmias, activation of these channels leads to rapid and repeated restoration of normal excitation rhythm. Experimental validation is provided by injecting the designed channel current for arrhythmia termination in human atrial myocytes using dynamic clamp

Acetylcholine Reduces IKr and Prolongs Action Potentials in Human Ventricular Cardiomyocytes

Vagal nerve stimulation (VNS) has a meaningful basis as a potentially effective treatment for heart failure with reduced ejection fraction. There is an ongoing VNS randomized study, and four studies are completed. However, relatively little is known about the effect of acetylcholine (ACh) on repolarization in human ventricular cardiomyocytes, as well as the effect of ACh on the rapid component of the delayed rectifier K(+) current (I(Kr)). Here, we investigated the effect of ACh on the action potential parameters in human ventricular preparations and on I(Kr) in human induced pluripotent stem-cell-derived cardiomyocytes (hiPSC-CMs). Using standard microelectrode technique, we demonstrated that ACh (5 µM) significantly increased the action potential duration in human left ventricular myocardial slices. ACh (5 µM) also prolonged repolarization in a human Purkinje fiber and a papillary muscle. Optical mapping revealed that ACh increased the action potential duration in human left ventricular myocardial slices and that the effect was dose-dependent. Perforated patch clamp experiments demonstrated action potential prolongation and a significant decrease in I(Kr) by ACh (5 µM) in hiPSC-CMs. Computer simulations of the electrical activity of a human ventricular cardiomyocyte showed an increase in action potential duration upon implementation of the experimentally observed ACh-induced changes in the fully activated conductance and steady-state activation of I(Kr). Our findings support the hypothesis that ACh can influence the repolarization in human ventricular cardiomyocytes by at least changes in I(Kr)

Effects of muscarinic receptor stimulation on Ca2+ transient, cAMP production and pacemaker frequency of rabbit sinoatrial node cells

We investigated the contribution of the intracellular calcium (Cai2+) transient to acetylcholine (ACh)-mediated reduction of pacemaker frequency and cAMP content in rabbit sinoatrial nodal (SAN) cells. Action potentials (whole cell perforated patch clamp) and Cai2+ transients (Indo-1 fluorescence) were recorded from single isolated rabbit SAN cells, whereas intracellular cAMP content was measured in SAN cell suspensions using a cAMP assay (LANCE®). Our data show that the Cai2+ transient, like the hyperpolarization-activated “funny current” (If) and the ACh-sensitive potassium current (IK,ACh), is an important determinant of ACh-mediated pacemaker slowing. When If and IK,ACh were both inhibited, by cesium (2 mM) and tertiapin (100 nM), respectively, 1 μM ACh was still able to reduce pacemaker frequency by 72%. In these If and IK,ACh-inhibited SAN cells, good correlations were found between the ACh-mediated change in interbeat interval and the ACh-mediated change in Cai2+ transient decay (r2 = 0.98) and slow diastolic Cai2+ rise (r2 = 0.73). Inhibition of the Cai2+ transient by ryanodine (3 μM) or BAPTA-AM (5 μM) facilitated ACh-mediated pacemaker slowing. Furthermore, ACh depressed the Cai2+ transient and reduced the sarcoplasmic reticulum (SR) Ca2+ content, all in a concentration-dependent fashion. At 1 μM ACh, the spontaneous activity and Cai2+ transient were abolished, but completely recovered when cAMP production was stimulated by forskolin (10 μM) and IK,ACh was inhibited by tertiapin (100 nM). Also, inhibition of the Cai2+ transient by ryanodine (3 μM) or BAPTA-AM (25 μM) exaggerated the ACh-mediated inhibition of cAMP content, indicating that Cai2+ affects cAMP production in SAN cells. In conclusion, muscarinic receptor stimulation inhibits the Cai2+ transient via a cAMP-dependent signaling pathway. Inhibition of the Cai2+ transient contributes to pacemaker slowing and inhibits Cai2+-stimulated cAMP production. Thus, we provide functional evidence for the contribution of the Cai2+ transient to ACh-induced inhibition of pacemaker activity and cAMP content in rabbit SAN cells