Background—: Delayed afterdepolarizations (DADs) carried by Na ⁺ -Ca ²⁺ -exchange current (I _NCX ) in response to sarcoplasmic reticulum (SR) Ca ²⁺ leak can promote atrial fibrillation (AF). The mechanisms leading to delayed afterdepolarizations in AF patients have not been defined. Methods and Results—: Protein levels (Western blot), membrane currents and action potentials (patch clamp), and [Ca ²⁺ ] _i (Fluo-3) were measured in right atrial samples from 76 sinus rhythm (control) and 72 chronic AF (cAF) patients. Diastolic [Ca ²⁺ ] _i and SR Ca ²⁺ content (integrated I _NCX during caffeine-induced Ca ²⁺ transient) were unchanged, whereas diastolic SR Ca ²⁺ leak, estimated by blocking ryanodine receptors (RyR2) with tetracaine, was ≈50% higher in cAF versus control. Single-channel recordings from atrial RyR2 reconstituted into lipid bilayers revealed enhanced open probability in cAF samples, providing a molecular basis for increased SR Ca ²⁺ leak. Calmodulin expression (60%), Ca ²⁺ /calmodulin-dependent protein kinase-II (CaMKII) autophosphorylation at Thr287 (87%), and RyR2 phosphorylation at Ser2808 (protein kinase A/CaMKII site, 236%) and Ser2814 (CaMKII site, 77%) were increased in cAF. The selective CaMKII blocker KN-93 decreased SR Ca ²⁺ leak, the frequency of spontaneous Ca ²⁺ release events, and RyR2 open probability in cAF, whereas protein kinase A inhibition with H-89 was ineffective. Knock-in mice with constitutively phosphorylated RyR2 at Ser2814 showed a higher incidence of Ca ²⁺ sparks and increased susceptibility to pacing-induced AF compared with controls. The relationship between [Ca ²⁺ ] _i and I _NCX density revealed I _NCX upregulation in cAF. Spontaneous Ca ²⁺ release events accompanied by inward I _NCX currents and delayed afterdepolarizations/triggered activity occurred more often and the sensitivity of resting membrane voltage to elevated [Ca ²⁺ ] _i (diastolic [Ca ²⁺ ] _i –voltage coupling gain) was higher in cAF compared with control. Conclusions—: Enhanced SR Ca ²⁺ leak through CaMKII-hyperphosphorylated RyR2, in combination with larger I _NCX for a given SR Ca ²⁺ release and increased diastolic [Ca ²⁺ ] _i -voltage coupling gain, causes AF-promoting atrial delayed afterdepolarizations/triggered activity in cAF patients.

Background: Atrial fibrillation (AF) is frequently associated with enhanced inflammatory response. The NLRP3 (NACHT, LRR, and PYD domain containing protein 3) inflammasome mediates caspase-1 activation and interleukin-1β release in immune cells but is not known to play a role in cardiomyocytes (CMs). Here, we assessed the role of CM NLRP3 inflammasome in AF. Methods: NLRP3 inflammasome activation was assessed by immunoblot in atrial whole-tissue lysates and CMs from patients with paroxysmal AF or long-standing persistent (chronic) AF. To determine whether CM-specific activation of NLPR3 is sufficient to promote AF, a CM-specific knockin mouse model expressing constitutively active NLRP3 (CM-KI) was established. In vivo electrophysiology was used to assess atrial arrhythmia vulnerability. To evaluate the mechanism of AF, electric activation pattern, Ca ²⁺ spark frequency, atrial effective refractory period, and morphology of atria were evaluated in CM-KI mice and wild-type littermates. Results: NLRP3 inflammasome activity was increased in the atrial CMs of patients with paroxysmal AF and chronic AF. CM-KI mice developed spontaneous premature atrial contractions and inducible AF, which was attenuated by a specific NLRP3 inflammasome inhibitor, MCC950. CM-KI mice exhibited ectopic activity, abnormal sarcoplasmic reticulum Ca ²⁺ release, atrial effective refractory period shortening, and atrial hypertrophy. Adeno-associated virus subtype-9–mediated CM-specific knockdown of Nlrp3 suppressed AF development in CM-KI mice. Finally, genetic inhibition of Nlrp3 prevented AF development in CREM transgenic mice, a well-characterized mouse model of spontaneous AF. Conclusions: Our study establishes a novel pathophysiological role for CM NLRP3 inflammasome signaling, with a mechanistic link to the pathogenesis of AF, and establishes the inhibition of NLRP3 as a potential novel AF therapy approach.

The cardiac ryanodine receptor (RyR2)/calcium release channel on the sarcoplasmic reticulum is required for muscle excitation-contraction coupling. Using site-directed mutagenesis, we identified the specific Ca ²⁺ /calmodulin-dependent protein kinase II (CaMKII) phosphorylation site on recombinant RyR2, distinct from the site for protein kinase A (PKA) that mediates the “fight-or-flight” stress response. CaMKII phosphorylation increased RyR2 Ca ²⁺ sensitivity and open probability. CaMKII was activated at increased heart rates, which may contribute to enhanced Ca ²⁺ -induced Ca ²⁺ release. Moreover, rate-dependent CaMKII phosphorylation of RyR2 was defective in heart failure. CaMKII-mediated phosphorylation of RyR2 may contribute to the enhanced contractility observed at higher heart rates. The full text of this article is available online at http://circres.ahajournals.org .

There has been a significant progress in our understanding of the molecular mechanisms by which calcium (Ca ²⁺ ) ions mediate various types of cardiac arrhythmias. A growing list of inherited gene defects can cause potentially lethal cardiac arrhythmia syndromes, including catecholaminergic polymorphic ventricular tachycardia, congenital long QT syndrome, and hypertrophic cardiomyopathy. In addition, acquired deficits of multiple Ca ²⁺ -handling proteins can contribute to the pathogenesis of arrhythmias in patients with various types of heart disease. In this review article, we will first review the key role of Ca ²⁺ in normal cardiac function—in particular, excitation–contraction coupling and normal electric rhythms. The functional involvement of Ca ²⁺ in distinct arrhythmia mechanisms will be discussed, followed by various inherited arrhythmia syndromes caused by mutations in Ca ²⁺ -handling proteins. Finally, we will discuss how changes in the expression of regulation of Ca ²⁺ channels and transporters can cause acquired arrhythmias, and how these mechanisms might be targeted for therapeutic purposes.