SUMMARY Organ-specific functions of tissue-resident macrophages in the steady-state heart are unknown. Here we show that cardiac macrophages facilitate electrical conduction through the distal atrioventricular node, where conducting cells densely intersperse with elongated macrophages expressing connexin 43. When coupled to spontaneously beating cardiomyocytes via connexin 43-containing gap junctions, cardiac macrophages have a negative resting membrane potential and depolarize in synchrony with cardiomyocytes. Conversely, macrophages render the resting membrane potential of cardiomyocytes more positive and, according to computational modeling, accelerate their repolarization. Photostimulation of channelrhodopsin 2-expressing macrophages improves atrioventricular conduction, while conditional deletion of connexin 43 in macrophages and congenital lack of macrophages delay atrioventricular conduction. In the Cd11bDTR mouse, macrophage ablation induces progressive atrioventricular block. These observations implicate macrophages in normal and aberrant cardiac conduction.

Atrial fibrillation is a highly prevalent arrhythmia and a major risk factor for stroke, heart failure and death1. We conducted a genome-wide association study (GWAS) in individuals of European ancestry, including 6,707 with and 52,426 without atrial fibrillation. Six new atrial fibrillation susceptibility loci were identified and replicated in an additional sample of individuals of European ancestry, including 5,381 subjects with and 1 0,030 subjects without atrial fibrillation (P < 5 × 10−8). Four of the loci identified in Europeans were further replicated in silico in a GWAS of Japanese individuals, including 843 individuals with and 3,350 individuals without atrial fibrillation. The identified loci implicate candidate genes that encode transcription factors related to cardiopulmonary development, cardiac-expressed ion channels and cell signaling molecules.

A new facet of calcium signalling involves the nuclear import of the NF-AT transcription factors from their dormant position in the cytoplasm. The protein phosphatase calcineurin appears to play an essential role in activating NF-AT nuclear import, as the calcineurin inhibitors cyclosporin A and FK506 block dephosphorylation and nuclear import of NF-AT (refs 4-7). Here we show that calcium signalling induces an association between NF-AT4 and calcineurin, and that these molecules are transported, as a complex, to the nucleus, where calcineurin continues to dephosphorylate NF-AT4. We propose that a nuclear complex of NF-AT4 and calcineurin maintains calcium signalling by counteracting a vigorous nuclear NF-AT kinase.

Atrial fibrillation (AF) is the most common sustained arrhythmia. A subset of patients with lone AF have no overt heart disease and an increased heritability of AF. We sought to identify common genetic variants underlying lone AF. Cases were from the German AF Network, Heart and Vascular Health Study, Atherosclerosis Risk in Communities Study, Cleveland Clinic, and Massachusetts General Hospital. Subjects were genotyped, HapMap SNPs imputed, and age- sex- and hypertension-adjusted analyses performed. A meta-analysis was conducted using 1,335 cases of lone AF and 12,844 referents. A novel locus on chromosome 1q21 was identified, and the most significant SNP, rs13376333, had an adjusted odds ratio of 1.56 (P=6.3×10−12). This association was replicated in two cohorts with lone AF for an overall odds ratio of 1.52 (P=1.83×10−21). Rs13376333 is intronic to KCNN3, a potassium channel involved in atrial repolarization. KCNN3 represents a novel potential therapeutic target in the treatment of AF.

Background-Drug-induced QT prolongation and torsades de pointes remain significant and often unpredictable clinical problems. Current in vitro preclinical assays are limited by biological simplicity, and in vivo models suffer from expense and low throughput. Methods and Results-During a screen for the effects of 100 small molecules on the heart rate of the zebrafish, Danio rerio, we found that drugs that cause QT prolongation in humans consistently caused bradycardia and AV block in the zebrafish. Of 23 such drugs tested, 18 were positive in this initial screen. Poor absorption explained 4 of 5 false-negative results, as demonstrated by microinjection. Overall, 22 of 23 compounds that cause repolarization abnormalities were positive in this assay. Antisense "knockdown" of the zebrafish KCNH2 ortholog yielded bradycardia in a dose dependent manner confirming the effects of reduction of repolarizing potassium current in this model. Classical drug-drug interactions between erythromycin and cisapride, as well as cimetidine and terfenadine, were also reproduced. Conclusion-This simple high-throughput assay is a promising addition to the repertoire of preclinical tests for drug-induced repolarization abnormalities. The genetic tractability of the zebrafish will allow the exploration of heritable modifiers of such drug effects. Key Words: drugs Ⅲ electrophysiology Ⅲ arrhythmia Ⅲ genes L ife-threatening arrhythmia in the setting of QT prolongation occurs as a result of inherited mutations in ion channel genes or, more commonly, as a consequence of drugs that affect cardiac repolarization. 1,2 This latter mechanism is the focus of increasing regulatory attention as several pharmaceuticals have been withdrawn from the US market due to torsades de pointes (TdP). Despite their medical importance, drug-related repolarization abnormalities and related arrhythmias remain difficult to predict. 3 Repolarization is complex, depending on individual channels, receptors, cytoskeletal elements, and the membrane. Additional complexity results from regional heterogeneity within the heart. 4 Further, some drugs, although safe in isolation, cause repolarization abnormalities when given with other medications, 1 through pharmacokinetic or pharmacodynamic interactions. Genetic variation may contribute to individual susceptibility to drug-induced arrhythmias. 3 A tractable model system enabling the identification of genes responsible for such variation would represent a significant advance.Virtually all QT-prolonging drugs identified to date inhibit the rapid component of the repolarizing potassium current (IKr). A channel composed of at least two subunits, KCNH2 and KCNE2, is responsible for this current. In vitro assays focusing on IKr are limited by biological simplicity, low throughput, and inability to detect drug-drug interactions. 5,6 Animal models, although more physiological, have an even lower throughput, restricting their ability to screen systematically for drug-drug interactions. 6 The zebrafish has a beating heart with both a ...

To increase the facility and throughput of scoring phenotypic traits in embryonic zebrafish, we developed an automated micro-well assay for heart rate using automated fluorescence microscopy of transgenic embryos expressing green fluorescent protein in myocardium. The assay measures heart rates efficiently and accurately over a large linear dynamic range, and it rapidly characterizes dose dependence and kinetics of small molecule-induced changes in heart rate. This is the first high-throughput micro-well assay for organ function in an intact vertebrate.

Rationale More than 25 million individuals suffer from heart failure worldwide, with nearly 4,000 patients currently awaiting heart transplantation in the United States. Donor organ shortage and allograft rejection remain major limitations with only about 2,500 hearts transplanted each year. As a theoretical alternative to allotransplantation, patient-derived bioartificial myocardium could provide functional support and ultimately impact the treatment of heart failure. Objective The objective of this study is to translate previous work to human scale and clinically relevant cells, for the bioengineering of functional myocardial tissue based on the combination of human cardiac matrix and human iPS-derived cardiac myocytes. Methods and Results To provide a clinically relevant tissue scaffold, we translated perfusion-decellularization to human scale and obtained biocompatible human acellular cardiac scaffolds with preserved extracellular matrix composition, architecture, and perfusable coronary vasculature. We then repopulated this native human cardiac matrix with cardiac myocytes derived from non-transgenic human induced pluripotent stem cells (iPSCs) and generated tissues of increasing three-dimensional complexity. We maintained such cardiac tissue constructs in culture for 120 days to demonstrate definitive sarcomeric structure, cell and matrix deformation, contractile force, and electrical conduction. To show that functional myocardial tissue of human scale can be built on this platform, we then partially recellularized human whole heart scaffolds with human iPSC-derived cardiac myocytes. Under biomimetic culture, the seeded constructs developed force-generating human myocardial tissue, showed electrical conductivity, left ventricular pressure development, and metabolic function. Conclusions Native cardiac extracellular matrix scaffolds maintain matrix components and structure to support the seeding and engraftment of human iPS-derived cardiac myocytes, and enable the bioengineering of functional human myocardial-like tissue of multiple complexities.

Elevated resting heart rate is associated with greater risk of cardiovascular disease and mortality. In a 2-stage meta-analysis of genome-wide association studies in up to 181,171 individuals, we identified 14 new loci associated with heart rate and confirmed associations with all 7 previously established loci. Experimental downregulation of gene expression in Drosophila melanogaster and Danio rerio identified 20 genes at 11 loci that are relevant for heart rate regulation and highlight a role for genes involved in signal transmission, embryonic cardiac development and the pathophysiology of dilated cardiomyopathy, congenital heart failure and/or sudden cardiac death. In addition, genetic susceptibility to increased heart rate is associated with altered cardiac conduction and reduced risk of sick sinus syndrome, and both heart rate–increasing and heart rate–decreasing variants associate with risk of atrial fibrillation. Our findings provide fresh insights into the mechanisms regulating heart rate and identify new therapeutic targets.