Fibroblast-Specific Proteotranscriptomes Reveal Distinct Fibrotic Signatures of Human Sinoatrial Node in Nonfailing and Failing Hearts.

BACKGROUND: Up to 50% of the adult human sinoatrial node (SAN) is composed of dense connective tissue. Cardiac diseases including heart failure (HF) may increase fibrosis within the SAN pacemaker complex, leading to impaired automaticity and conduction of electric activity to the atria. Unlike the role of cardiac fibroblasts in pathologic fibrotic remodeling and tissue repair, nothing is known about fibroblasts that maintain the inherently fibrotic SAN environment. METHODS: Intact SAN pacemaker complex was dissected from cardioplegically arrested explanted nonfailing hearts (non-HF; n=22; 48.7±3.1 years of age) and human failing hearts (n=16; 54.9±2.6 years of age). Connective tissue content was quantified from Masson trichrome-stained head-center and center-tail SAN sections. Expression of extracellular matrix proteins, including collagens 1 and 3A1, CILP1 (cartilage intermediate layer protein 1), and POSTN (periostin), and fibroblast and myofibroblast numbers were quantified by in situ and in vitro immunolabeling. Fibroblasts from the central intramural SAN pacemaker compartment (≈10×5×2 mm3) and right atria were isolated, cultured, passaged once, and treated ± transforming growth factor ß1 and subjected to comprehensive high-throughput next-generation sequencing of whole transcriptome, microRNA, and proteomic analyses. RESULTS: Intranodal fibrotic content was significantly higher in SAN pacemaker complex from HF versus non-HF hearts (57.7±2.6% versus 44.0±1.2%; P<0.0001). Proliferating phosphorylated histone 3+/vimentin+/CD31- (cluster of differentiation 31) fibroblasts were higher in HF SAN. Vimentin+/α-smooth muscle actin+/CD31- myofibroblasts along with increased interstitial POSTN expression were found only in HF SAN. RNA sequencing and proteomic analyses identified unique differences in mRNA, long noncoding RNA, microRNA, and proteomic profiles between non-HF and HF SAN and right atria fibroblasts and transforming growth factor ß1-induced myofibroblasts. Specifically, proteins and signaling pathways associated with extracellular matrix flexibility, stiffness, focal adhesion, and metabolism were altered in HF SAN fibroblasts compared with non-HF SAN. CONCLUSIONS: This study revealed increased SAN-specific fibrosis with presence of myofibroblasts, CILP1, and POSTN-positive interstitial fibrosis only in HF versus non-HF human hearts. Comprehensive proteotranscriptomic profiles of SAN fibroblasts identified upregulation of genes and proteins promoting stiffer SAN extracellular matrix in HF hearts. Fibroblast-specific profiles generated by our proteotranscriptomic analyses of the human SAN provide a comprehensive framework for future studies to investigate the role of SAN-specific fibrosis in cardiac rhythm regulation and arrhythmias.

Comprehensive evaluation of electrophysiological and 3D structural features of human atrial myocardium with insights on atrial fibrillation maintenance mechanisms.

Atrial fibrillation (AF) occurrence and maintenance is associated with progressive remodeling of electrophysiological (repolarization and conduction) and 3D structural (fibrosis, fiber orientations, and wall thickness) features of the human atria. Significant diversity in AF etiology leads to heterogeneous arrhythmogenic electrophysiological and structural substrates within the 3D structure of the human atria. Since current clinical methods have yet to fully resolve the patient-specific arrhythmogenic substrates, mechanism-based AF treatments remain underdeveloped. Here, we review current knowledge from in-vivo, ex-vivo, and in-vitro human heart studies, and discuss how these studies may provide new insights on the synergy of atrial electrophysiological and 3D structural features in AF maintenance. In-vitro studies on surgically acquired human atrial samples provide a great opportunity to study a wide spectrum of AF pathology, including functional changes in single-cell action potentials, ion channels, and gene/protein expression. However, limited size of the samples prevents evaluation of heterogeneous AF substrates and reentrant mechanisms. In contrast, coronary-perfused ex-vivo human hearts can be studied with state-of-the-art functional and structural technologies, such as high-resolution near-infrared optical mapping and contrast-enhanced MRI. These imaging modalities can resolve atrial arrhythmogenic substrates and their role in reentrant mechanisms maintaining AF and validate clinical approaches. Nonetheless, longitudinal studies are not feasible in explanted human hearts. As no approach is perfect, we suggest that combining the strengths of direct human atrial studies with high fidelity approaches available in the laboratory and in realistic patient-specific computer models would elucidate deeper knowledge of AF mechanisms. We propose that a comprehensive translational pipeline from ex-vivo human heart studies to longitudinal clinically relevant AF animal studies and finally to clinical trials is necessary to identify patient-specific arrhythmogenic substrates and develop novel AF treatments.

Adenosine-Induced Atrial Fibrillation: Localized Reentrant Drivers in Lateral Right Atria due to Heterogeneous Expression of Adenosine A1 Receptors and GIRK4 Subunits in the Human Heart.

BACKGROUND: Adenosine provokes atrial fibrillation (AF) with a higher activation frequency in right atria (RA) versus left atria (LA) in patients, but the underlying molecular and functional substrates are unclear. We tested the hypothesis that adenosine-induced AF is driven by localized reentry in RA areas with highest expression of adenosine A1 receptor and its downstream GIRK (G protein-coupled inwardly rectifying potassium channels) channels (IK,Ado). METHODS: We applied biatrial optical mapping and immunoblot mapping of various atrial regions to reveal the mechanism of adenosine-induced AF in explanted failing and nonfailing human hearts (n=37). RESULTS: Optical mapping of coronary-perfused atria (n=24) revealed that adenosine perfusion (10-100 µmol/L) produced more significant shortening of action potential durations in RA (from 290±45 to 239±41 ms, 17.3±10.4%; P<0.01) than LA (from 307±24 to 286±23 ms, 6.7±6.6%; P<0.01). In 10 hearts, adenosine induced AF (317±116 s) that, when sustained (≥2 minutes), was primarily maintained by 1 to 2 localized reentrant drivers in lateral RA. Tertiapin (10-100 nmol/L), a selective GIRK channel blocker, counteracted adenosine-induced action potential duration shortening and prevented AF induction. Immunoblotting showed that the superior/middle lateral RA had significantly higher adenosine A1 receptor (2.7±1.7-fold; P<0.01) and GIRK4 (1.7±0.8-fold; P<0.05) protein expression than lateral/posterior LA. CONCLUSIONS: This study revealed a 3-fold RA-to-LA adenosine A1 receptor protein expression gradient in the human heart, leading to significantly greater RA versus LA repolarization sensitivity in response to adenosine. Sustained adenosine-induced AF is maintained by reentrant drivers localized in lateral RA regions with the highest adenosine A1 receptor/GIRK4 expression. Selective atrial GIRK channel blockade may effectively treat AF during conditions with increased endogenous adenosine.

Calsequestrin 2 deletion causes sinoatrial node dysfunction and atrial arrhythmias associated with altered sarcoplasmic reticulum calcium cycling and degenerative fibrosis within the mouse atrial pacemaker complex1.

AIMS: Loss-of-function mutations in Calsequestrin 2 (CASQ2) are associated with catecholaminergic polymorphic ventricular tachycardia (CPVT). CPVT patients also exhibit bradycardia and atrial arrhythmias for which the underlying mechanism remains unknown. We aimed to study the sinoatrial node (SAN) dysfunction due to loss of CASQ2. METHODS AND RESULTS: In vivo electrocardiogram (ECG) monitoring, in vitro high-resolution optical mapping, confocal imaging of intracellular Ca(2+) cycling, and 3D atrial immunohistology were performed in wild-type (WT) and Casq2 null (Casq2(-/-)) mice. Casq2(-/-) mice exhibited bradycardia, SAN conduction abnormalities, and beat-to-beat heart rate variability due to enhanced atrial ectopic activity both at baseline and with autonomic stimulation. Loss of CASQ2 increased fibrosis within the pacemaker complex, depressed primary SAN activity, and conduction, but enhanced atrial ectopic activity and atrial fibrillation (AF) associated with macro- and micro-reentry during autonomic stimulation. In SAN myocytes, CASQ2 deficiency induced perturbations in intracellular Ca(2+) cycling, including abnormal Ca(2+) release, periods of significantly elevated diastolic Ca(2+) levels leading to pauses and unstable pacemaker rate. Importantly, Ca(2+) cycling dysfunction occurred not only at the SAN cellular level but was also globally manifested as an increased delay between action potential (AP) and Ca(2+) transient upstrokes throughout the atrial pacemaker complex. CONCLUSIONS: Loss of CASQ2 causes abnormal sarcoplasmic reticulum Ca(2+) release and selective interstitial fibrosis in the atrial pacemaker complex, which disrupt SAN pacemaking but enhance latent pacemaker activity, create conduction abnormalities and increase susceptibility to AF. These functional and extensive structural alterations could contribute to SAN dysfunction as well as AF in CPVT patients.

Atrial fibrillation driven by micro-anatomic intramural re-entry revealed by simultaneous sub-epicardial and sub-endocardial optical mapping in explanted human hearts.

AIMS: The complex architecture of the human atria may create physical substrates for sustained re-entry to drive atrial fibrillation (AF). The existence of sustained, anatomically defined AF drivers in humans has been challenged partly due to the lack of simultaneous endocardial-epicardial (Endo-Epi) mapping coupled with high-resolution 3D structural imaging. METHODS AND RESULTS: Coronary-perfused human right atria from explanted diseased hearts (n = 8, 43-72 years old) were optically mapped simultaneously by three high-resolution CMOS cameras (two aligned Endo-Epi views (330 µm2 resolution) and one panoramic view). 3D gadolinium-enhanced magnetic resonance imaging (GE-MRI, 80 µm3 resolution) revealed the atrial wall structure varied in thickness (1.0 ± 0.7-6.8 ± 2.4 mm), transmural fiber angle differences, and interstitial fibrosis causing transmural activation delay from 23 ± 11 to 43 ± 22 ms at increased pacing rates. Sustained AF (>90 min) was induced by burst pacing during pinacidil (30-100 µM) perfusion. Dual-sided sub-Endo-sub-Epi optical mapping revealed that AF was driven by spatially and temporally stable intramural re-entry with 107 ± 50 ms cycle length and transmural activation delay of 67 ± 31 ms. Intramural re-entrant drivers were captured primarily by sub-Endo mapping, while sub-Epi mapping visualized re-entry or 'breakthrough' patterns. Re-entrant drivers were anchored on 3D micro-anatomic tracks (15.4 ± 2.2 × 6.0 ± 2.3 mm2, 2.9 ± 0.9 mm depth) formed by atrial musculature characterized by increased transmural fiber angle differences and interstitial fibrosis. Targeted radiofrequency ablation of the tracks verified these re-entries as drivers of AF. CONCLUSIONS: Integrated 3D structural-functional mapping of diseased human right atria ex vivo revealed that the complex atrial microstructure caused significant differences between Endo vs. Epi activation during pacing and sustained AF driven by intramural re-entry anchored to fibrosis-insulated atrial bundles.

Upregulation of adenosine A1 receptors facilitates sinoatrial node dysfunction in chronic canine heart failure by exacerbating nodal conduction abnormalities revealed by novel dual-sided intramural optical mapping.

BACKGROUND: Although sinoatrial node (SAN) dysfunction is a hallmark of human heart failure (HF), the underlying mechanisms remain poorly understood. We aimed to examine the role of adenosine in SAN dysfunction and tachy-brady arrhythmias in chronic HF. METHODS AND RESULTS: We applied multiple approaches to characterize SAN structure, SAN function, and adenosine A1 receptor expression in control (n=17) and 4-month tachypacing-induced chronic HF (n=18) dogs. Novel intramural optical mapping of coronary-perfused right atrial preparations revealed that adenosine (10 µmol/L) markedly prolonged postpacing SAN conduction time in HF by 206 ± 99 milliseconds (versus 66 ± 21 milliseconds in controls; P=0.02). Adenosine induced SAN intranodal conduction block or microreentry in 6 of 8 dogs with HF versus 0 of 7 controls (P=0.007). Adenosine-induced SAN conduction abnormalities and automaticity depression caused postpacing atrial pauses in HF versus control dogs (17.1 ± 28.9 versus 1.5 ± 1.3 seconds; P<0.001). Furthermore, 10 µmol/L adenosine shortened atrial repolarization and led to pacing-induced atrial fibrillation in 6 of 7 HF versus 0 of 7 control dogs (P=0.002). Adenosine-induced SAN dysfunction and atrial fibrillation were abolished or prevented by adenosine A1 receptor antagonists (50 µmol/L theophylline/1 µmol/L 8-cyclopentyl-1,3-dipropylxanthine). Adenosine A1 receptor protein expression was significantly upregulated during HF in the SAN (by 47 ± 19%) and surrounding atrial myocardium (by 90 ± 40%). Interstitial fibrosis was significantly increased within the SAN in HF versus control dogs (38 ± 4% versus 23 ± 4%; P<0.001). CONCLUSIONS: In chronic HF, adenosine A1 receptor upregulation in SAN pacemaker and atrial cardiomyocytes may increase cardiac sensitivity to adenosine. This effect may exacerbate conduction abnormalities in the structurally impaired SAN, leading to SAN dysfunction, and potentiate atrial repolarization shortening, thereby facilitating atrial fibrillation. Atrial fibrillation may further depress SAN function and lead to tachy-brady arrhythmias in HF.

Optimization of catheter ablation of atrial fibrillation: insights gained from clinically-derived computer models.

Atrial fibrillation (AF) is the most common heart rhythm disturbance, and its treatment is an increasing economic burden on the health care system. Despite recent intense clinical, experimental and basic research activity, the treatment of AF with current antiarrhythmic drugs and catheter/surgical therapies remains limited. Radiofrequency catheter ablation (RFCA) is widely used to treat patients with AF. Current clinical ablation strategies are largely based on atrial anatomy and/or substrate detected using different approaches, and they vary from one clinical center to another. The nature of clinical ablation leads to ambiguity regarding the optimal patient personalization of the therapy partly due to the fact that each empirical configuration of ablation lines made in a patient is irreversible during one ablation procedure. To investigate optimized ablation lesion line sets, in silico experimentation is an ideal solution. 3D computer models give us a unique advantage to plan and assess the effectiveness of different ablation strategies before and during RFCA. Reliability of in silico assessment is ensured by inclusion of accurate 3D atrial geometry, realistic fiber orientation, accurate fibrosis distribution and cellular kinetics; however, most of this detailed information in the current computer models is extrapolated from animal models and not from the human heart. The predictive power of computer models will increase as they are validated with human experimental and clinical data. To make the most from a computer model, one needs to develop 3D computer models based on the same functionally and structurally mapped intact human atria with high spatial resolution. The purpose of this review paper is to summarize recent developments in clinically-derived computer models and the clinical insights they provide for catheter ablation.

Altered microRNA and mRNA profiles during heart failure in the human sinoatrial node.

Heart failure (HF) is frequently accompanied with the sinoatrial node (SAN) dysfunction, which causes tachy-brady arrhythmias and increased mortality. MicroRNA (miR) alterations are associated with HF progression. However, the transcriptome of HF human SAN, and its role in HF-associated remodeling of ion channels, transporters, and receptors responsible for SAN automaticity and conduction impairments is unknown. We conducted comprehensive high-throughput transcriptomic analysis of pure human SAN primary pacemaker tissue and neighboring right atrial tissue from human transplanted HF hearts (n = 10) and non-failing (nHF) donor hearts (n = 9), using next-generation sequencing. Overall, 47 miRs and 832 mRNAs related to multiple signaling pathways, including cardiac diseases, tachy-brady arrhythmias and fibrosis, were significantly altered in HF SAN. Of the altered miRs, 27 are predicted to regulate mRNAs of major ion channels and neurotransmitter receptors which are involved in SAN automaticity (e.g. HCN1, HCN4, SLC8A1) and intranodal conduction (e.g. SCN5A, SCN8A) or both (e.g. KCNJ3, KCNJ5). Luciferase reporter assays were used to validate interactions of miRs with predicted mRNA targets. In conclusion, our study provides a profile of altered miRs in HF human SAN, and a novel transcriptome blueprint to identify molecular targets for SAN dysfunction and arrhythmia treatments in HF.

Optical Mapping-Validated Machine Learning Improves Atrial Fibrillation Driver Detection by Multi-Electrode Mapping.

BACKGROUND: Atrial fibrillation (AF) can be maintained by localized intramural reentrant drivers. However, AF driver detection by clinical surface-only multielectrode mapping (MEM) has relied on subjective interpretation of activation maps. We hypothesized that application of machine learning to electrogram frequency spectra may accurately automate driver detection by MEM and add some objectivity to the interpretation of MEM findings. METHODS: Temporally and spatially stable single AF drivers were mapped simultaneously in explanted human atria (n=11) by subsurface near-infrared optical mapping (NIOM; 0.3 mm2 resolution) and 64-electrode MEM (higher density or lower density with 3 and 9 mm2 resolution, respectively). Unipolar MEM and NIOM recordings were processed by Fourier transform analysis into 28 407 total Fourier spectra. Thirty-five features for machine learning were extracted from each Fourier spectrum. RESULTS: Targeted driver ablation and NIOM activation maps efficiently defined the center and periphery of AF driver preferential tracks and provided validated annotations for driver versus nondriver electrodes in MEM arrays. Compared with analysis of single electrogram frequency features, averaging the features from each of the 8 neighboring electrodes, significantly improved classification of AF driver electrograms. The classification metrics increased when less strict annotation, including driver periphery electrodes, were added to driver center annotation. Notably, f1-score for the binary classification of higher-density catheter data set was significantly higher than that of lower-density catheter (0.81±0.02 versus 0.66±0.04, P<0.05). The trained algorithm correctly highlighted 86% of driver regions with higher density but only 80% with lower-density MEM arrays (81% for lower-density+higher-density arrays together). CONCLUSIONS: The machine learning model pretrained on Fourier spectrum features allows efficient classification of electrograms recordings as AF driver or nondriver compared with the NIOM gold-standard. Future application of NIOM-validated machine learning approach may improve the accuracy of AF driver detection for targeted ablation treatment in patients.

Impaired neuronal sodium channels cause intranodal conduction failure and reentrant arrhythmias in human sinoatrial node.

Mechanisms for human sinoatrial node (SAN) dysfunction are poorly understood and whether human SAN excitability requires voltage-gated sodium channels (Nav) remains controversial. Here, we report that neuronal (n)Nav blockade and selective nNav1.6 blockade during high-resolution optical mapping in explanted human hearts depress intranodal SAN conduction, which worsens during autonomic stimulation and overdrive suppression to conduction failure. Partial cardiac (c)Nav blockade further impairs automaticity and intranodal conduction, leading to beat-to-beat variability and reentry. Multiple nNav transcripts are higher in SAN vs atria; heterogeneous alterations of several isoforms, specifically nNav1.6, are associated with heart failure and chronic alcohol consumption. In silico simulations of Nav distributions suggest that INa is essential for SAN conduction, especially in fibrotic failing hearts. Our results reveal that not only cNav but nNav are also integral for preventing disease-induced failure in human SAN intranodal conduction. Disease-impaired nNav may underlie patient-specific SAN dysfunctions and should be considered to treat arrhythmias.

Silencing miR-370-3p rescues funny current and sinus node function in heart failure.

Bradyarrhythmias are an important cause of mortality in heart failure and previous studies indicate a mechanistic role for electrical remodelling of the key pacemaking ion channel HCN4 in this process. Here we show that, in a mouse model of heart failure in which there is sinus bradycardia, there is upregulation of a microRNA (miR-370-3p), downregulation of the pacemaker ion channel, HCN4, and downregulation of the corresponding ionic current, If, in the sinus node. In vitro, exogenous miR-370-3p inhibits HCN4 mRNA and causes downregulation of HCN4 protein, downregulation of If, and bradycardia in the isolated sinus node. In vivo, intraperitoneal injection of an antimiR to miR-370-3p into heart failure mice silences miR-370-3p and restores HCN4 mRNA and protein and If in the sinus node and blunts the sinus bradycardia. In addition, it partially restores ventricular function and reduces mortality. This represents a novel approach to heart failure treatment.

Unmasking Arrhythmogenic Hubs of Reentry Driving Persistent Atrial Fibrillation for Patient-Specific Treatment.

Background Atrial fibrillation (AF) driver mechanisms are obscured to clinical multielectrode mapping approaches that provide partial, surface-only visualization of unstable 3-dimensional atrial conduction. We hypothesized that transient modulation of refractoriness by pharmacologic challenge during multielectrode mapping improves visualization of hidden paths of reentrant AF drivers for targeted ablation. Methods and Results Pharmacologic challenge with adenosine was tested in ex vivo human hearts with a history of AF and cardiac diseases by multielectrode and high-resolution subsurface near-infrared optical mapping, integrated with 3-dimensional structural imaging and heart-specific computational simulations. Adenosine challenge was also studied on acutely terminated AF drivers in 10 patients with persistent AF. Ex vivo, adenosine stabilized reentrant driver paths within arrhythmogenic fibrotic hubs and improved visualization of reentrant paths, previously seen as focal or unstable breakthrough activation pattern, for targeted AF ablation. Computational simulations suggested that shortening of atrial refractoriness by adenosine may (1) improve driver stability by annihilating spatially unstable functional blocks and tightening reentrant circuits around fibrotic substrates, thus unmasking the common reentrant path; and (2) destabilize already stable reentrant drivers along fibrotic substrates by accelerating competing fibrillatory wavelets or secondary drivers. In patients with persistent AF, adenosine challenge unmasked hidden common reentry paths (9/15 AF drivers, 41±26% to 68±25% visualization), but worsened visualization of previously visible reentry paths (6/15, 74±14% to 34±12%). AF driver ablation led to acute termination of AF. Conclusions Our ex vivo to in vivo human translational study suggests that transiently altering atrial refractoriness can stabilize reentrant paths and unmask arrhythmogenic hubs to guide targeted AF driver ablation treatment.

A robust computational framework for estimating 3D Bi-Atrial chamber wall thickness.

Atrial fibrillation (AF) is the most prevalent form of cardiac arrhythmia. The atrial wall thickness (AWT) can potentially improve our understanding of the mechanism underlying atrial structure that drives AF and provides important clinical information. However, most existing studies for estimating AWT rely on ruler-based measurements performed on only a few selected locations in 2D or 3D using digital calipers. Only a few studies have developed automatic approaches to estimate the AWT in the left atrium, and there are currently no methods to robustly estimate the AWT of both atrial chambers. Therefore, we have developed a computational pipeline to automatically calculate the 3D AWT across bi-atrial chambers and extensively validated our pipeline on both ex vivo and in vivo human atria data. The atrial geometry was first obtained by segmenting the atrial wall from the MRIs using a novel machine learning approach. The epicardial and endocardial surfaces were then separated using a multi-planar convex hull approach to define boundary conditions, from which, a Laplace equation was solved numerically to automatically separate bi-atrial chambers. To robustly estimate the AWT in each atrial chamber, coupled partial differential equations by coupling the Laplace solution with two surface trajectory functions were formulated and solved. Our pipeline enabled the reconstruction and visualization of the 3D AWT for bi-atrial chambers with a relative error of 8% and outperformed existing algorithms by >7%. Our approach can potentially lead to improved clinical diagnosis, patient stratification, and clinical guidance during ablation treatment for patients with AF.

Human Atrial Fibrillation Drivers Resolved With Integrated Functional and Structural Imaging to Benefit Clinical Mapping.

OBJECTIVES: This study sought to improve atrial fibrillation (AF) driver identification by integrating clinical multielectrode mapping with driver fingerprints defined by high-resolution ex vivo 3-dimensional (3D) functional and structural imaging. BACKGROUND: Clinical multielectrode mapping of AF drivers suffers from variable contact, signal processing, and structural complexity within the 3D human atrial wall, raising questions on the validity of such drivers. METHODS: Sustained AF was mapped in coronary-perfused explanted human hearts (n = 11) with transmural near-infrared optical mapping (∼0.3 mm2 resolution). Simultaneously, custom FIRMap catheters (∼9 × 9 mm2 resolution) mapped endocardial and epicardial surfaces, which were analyzed by Focal Impulse and Rotor Mapping activation and Rotational Activity Profile (Abbott Labs, Chicago, Illinois). Functional maps were integrated with contrast-enhanced cardiac magnetic resonance imaging (∼0.1 mm3 resolution) analysis of 3D fibrosis architecture. RESULTS: During sustained AF, near-infrared optical mapping identified 1 to 2 intramural, spatially stable re-entrant AF drivers per heart. Driver targeted ablation affecting 2.2 ± 1.1% of the atrial surface terminated and prevented AF. Driver regions had significantly higher phase singularity density and dominant frequency than neighboring nondriver regions. Focal Impulse and Rotor Mapping had 80% sensitivity to near-infrared optical mapping-defined driver locations (16 of 20), and matched 14 of 20 driver visualizations: 10 of 14 re-entries seen with Rotational Activity Profile; and 4 of 6 breakthrough/focal patterns. Focal Impulse and Rotor Mapping detected 1.1 ± 0.9 false-positive rotational activity profiles per recording, but these regions had lower intramural contrast-enhanced cardiac magnetic resonance imaging fibrosis than did driver regions (14.9 ± 7.9% vs. 23.2 ± 10.5%; p < 0.005). CONCLUSIONS: The study revealed that both re-entrant and breakthrough/focal AF driver patterns visualized by surface-only clinical multielectrodes can represent projections of 3D intramural microanatomic re-entries. Integration of multielectrode mapping and 3D fibrosis analysis may enhance AF driver detection, thereby improving the efficacy of driver-targeted ablation.

Fibrosis and Atrial Fibrillation: Computerized and Optical Mapping; A View into the Human Atria at Submillimeter Resolution.

Recent studies strongly suggest that the majority of atrial fibrillation (AF) patients with diagnosed or subclinical cardiac diseases have established or even pre-existing fibrotic structural remodeling, which may lead to conduction abnormalities and reentrant activity that sustain AF. As conventional treatments fail to treat AF in far too many cases, an urgent need exists to identify specific structural arrhythmogenic fibrosis patterns, which may maintain AF, in order to identify effective ablation targets for AF treatment. However, the existing challenge is to define what exact structural remodeling within the complex 3D human atrial wall is arrhythmogenic, as well as linking arrhythmogenic fibrosis to an underlying mechanism of AF maintenance in the clinical setting. This review is focused on the role of 3D fibrosis architecture in the mechanisms of AF maintenance revealed by submillimeter, high-resolution ex-vivo imaging modalities directly of human atria, as well as from in-silico 3D computational techniques that can be able to overcome in-vivo clinical limitations. The systematic integration of functional and structural imaging ex-vivo may inform the necessary integration of electrode and structural mapping in-vivo. A holistic view of AF driver mechanisms may begin to identify the defining characteristics or "fingerprints" of reentrant AF drivers, such as 3D fibrotic architecture, in order to design optimal patient-specific ablation strategies.

Atrial fibrillation driver mechanisms: Insight from the isolated human heart.

Although there have been great technological advances in the treatment of atrial fibrillation (AF), current therapies remain limited due to a narrow understanding of AF mechanisms in the human heart. This review will highlight our recent studies on explanted human hearts where we developed and employed a novel functional-structural mapping approach by integrating high-resolution simultaneous endo-epicardial and panoramic optical mapping with 3D gadolinium-enhanced MRI to define the spatiotemporal characteristics of AF drivers and their structural substrates. The results allow us to postulate that the primary mechanism of AF maintenance in human hearts is a limited number of localized intramural microanatomic reentrant AF drivers anchored to heart-specific 3D fibrotically insulated myobundle tracks, which may remain hidden to clinical single-surface electrode mapping. We suggest that ex vivo human heart studies, by using an integrated 3D functional and structural mapping approach, will help to reveal defining features of AF drivers as well as validate and improve clinical approaches to detect and target these AF drivers in patients with cardiac diseases.

Redundant and diverse intranodal pacemakers and conduction pathways protect the human sinoatrial node from failure.

The human sinoatrial node (SAN) efficiently maintains heart rhythm even under adverse conditions. However, the specific mechanisms involved in the human SAN's ability to prevent rhythm failure, also referred to as its robustness, are unknown. Challenges exist because the three-dimensional (3D) intramural structure of the human SAN differs from well-studied animal models, and clinical electrode recordings are limited to only surface atrial activation. Hence, to innovate the translational study of human SAN structural and functional robustness, we integrated intramural optical mapping, 3D histology reconstruction, and molecular mapping of the ex vivo human heart. When challenged with adenosine or atrial pacing, redundant intranodal pacemakers within the human SAN maintained automaticity and delivered electrical impulses to the atria through sinoatrial conduction pathways (SACPs), thereby ensuring a fail-safe mechanism for robust maintenance of sinus rhythm. During adenosine perturbation, the primary central SAN pacemaker was suppressed, whereas previously inactive superior or inferior intranodal pacemakers took over automaticity maintenance. Sinus rhythm was also rescued by activation of another SACP when the preferential SACP was suppressed, suggesting two independent fail-safe mechanisms for automaticity and conduction. The fail-safe mechanism in response to adenosine challenge is orchestrated by heterogeneous differences in adenosine A1 receptors and downstream GIRK4 channel protein expressions across the SAN complex. Only failure of all pacemakers and/or SACPs resulted in SAN arrest or conduction block. Our results unmasked reserve mechanisms that protect the human SAN pacemaker and conduction complex from rhythm failure, which may contribute to treatment of SAN arrhythmias.

Three-dimensional Integrated Functional, Structural, and Computational Mapping to Define the Structural "Fingerprints" of Heart-Specific Atrial Fibrillation Drivers in Human Heart Ex Vivo.

BACKGROUND: Structural remodeling of human atria plays a key role in sustaining atrial fibrillation (AF), but insufficient quantitative analysis of human atrial structure impedes the treatment of AF. We aimed to develop a novel 3-dimensional (3D) structural and computational simulation analysis tool that could reveal the structural contributors to human reentrant AF drivers. METHODS AND RESULTS: High-resolution panoramic epicardial optical mapping of the coronary-perfused explanted intact human atria (63-year-old woman, chronic hypertension, heart weight 608 g) was conducted during sinus rhythm and sustained AF maintained by spatially stable reentrant AF drivers in the left and right atrium. The whole atria (107×61×85 mm3) were then imaged with contrast-enhancement MRI (9.4 T, 180×180×360-µm3 resolution). The entire 3D human atria were analyzed for wall thickness (0.4-11.7 mm), myofiber orientations, and transmural fibrosis (36.9% subendocardium; 14.2% midwall; 3.4% subepicardium). The 3D computational analysis revealed that a specific combination of wall thickness and fibrosis ranges were primarily present in the optically defined AF driver regions versus nondriver tissue. Finally, a 3D human heart-specific atrial computer model was developed by integrating 3D structural and functional mapping data to test AF induction, maintenance, and ablation strategies. This 3D model reproduced the optically defined reentrant AF drivers, which were uninducible when fibrosis and myofiber anisotropy were removed from the model. CONCLUSIONS: Our novel 3D computational high-resolution framework may be used to quantitatively analyze structural substrates, such as wall thickness, myofiber orientation, and fibrosis, underlying localized AF drivers, and aid the development of new patient-specific treatments.