Serine-threonine protein phosphatase regulation of Cx43 dephosphorylation in arrhythmogenic disorders.

Regulation of cell-to-cell communication in the heart by the gap junction protein Connexin43 (Cx43) involves modulation of Cx43 phosphorylation state by protein kinases, and dephosphorylation by protein phosphatases. Dephosphorylation of Cx43 has been associated with impaired intercellular coupling and enhanced arrhythmogenesis in various pathologic states. While there has been extensive study of the protein kinases acting on Cx43, there has been limited studies of the protein phosphatases that may underlie Cx43 dephosphorylation. The focus of this review is to introduce serine-threonine protein phosphatase regulation of Cx43 phosphorylation state and cell-to-cell communication, and its impact on arrhythmogenesis in the setting of chronic heart failure and myocardial ischemia, as well as on atrial fibrillation. We also discuss the therapeutic potential of modulating protein phosphatases to treat arrhythmias in these clinical settings.

Identification of Nrf2-responsive microRNA networks as putative mediators of myocardial reductive stress.

Although recent advances in the treatment of acute coronary heart disease have reduced mortality rates, few therapeutic strategies exist to mitigate the progressive loss of cardiac function that manifests as heart failure. Nuclear factor, erythroid 2 like 2 (Nfe2l2, Nrf2) is a transcriptional regulator that is known to confer transient myocardial cytoprotection following acute ischemic insult; however, its sustained activation paradoxically causes a reductive environment characterized by excessive antioxidant activity. We previously identified a subset of 16 microRNAs (miRNA) significantly diminished in Nrf2-ablated (Nrf2-/-) mouse hearts, leading to the hypothesis that increasing levels of Nrf2 activation augments miRNA induction and post-transcriptional dysregulation. Here, we report the identification of distinct miRNA signatures (i.e. "reductomiRs") associated with Nrf2 overexpression in a cardiac-specific and constitutively active Nrf2 transgenic (caNrf2-Tg) mice expressing low (TgL) and high (TgH) levels. We also found several Nrf2 dose-responsive miRNAs harboring proximal antioxidant response elements (AREs), implicating these "reductomiRs" as putative meditators of Nrf2-dependent post-transcriptional regulation. Analysis of mRNA-sequencing identified a complex network of miRNAs and effector mRNAs encoding known pathological hallmarks of cardiac stress-response. Altogether, these data support Nrf2 as a putative regulator of cardiac miRNA expression and provide novel candidates for future mechanistic investigation to understand the relationship between myocardial reductive stress and cardiac pathophysiology.

Molecular remodeling of Cx43, but not structural remodeling, promotes arrhythmias in an arrhythmogenic canine model of nonischemic heart failure.

BACKGROUND: Both gap junctional remodeling and interstitial fibrosis have been linked to impaired electrical conduction velocity (CV) and fatal ventricular arrhythmias in nonischemic heart failure (HF). However, the arrhythmogenic role of the ventricular gap junctional Cx43 in nonischemic HF remains in debate. Here, we assessed this in a newly developed arrhythmogenic canine model of nonischemic HF. METHODS AND RESULTS: Nonischemic HF was induced in canines by combined aortic valve insufficiency and aortic constriction. Left ventricular (LV) myocardium from HF dogs showed similar pathological changes to that of humans. HF dogs had reduced LV function, widened QRS complexes, and spontaneous nonsustained ventricular tachycardia. CV was measured in intact LV epicardium with high-density grid mapping. Total (Cx43-T) and nonphosphorylated Cx43 (Cx43-NP) and histological interstitial fibrosis were assessed from these mapped LV tissues. Longitudinal CV, which was slowed in HF (49 ± 1 vs. 65 ± 2 cm/s in Ctl), was positively correlated with reduced total junctional Cx43 and negatively correlated with markedly increased junctional Cx43-NP (2-fold) in HF. Cx43 dephosphorylation in HF was associated with enhanced colocalization of PP2A at the level of Cx43. Unchanged action potential upstroke and transverse CV were associated with unaltered Cx43 lateralization and interstitial fibrosis in the nonischemic HF canine LV. CONCLUSION: Our unique arrhythmogenic canine model of HF resembles human nonischemic HF (prior to the end stage). Cx43 remodeling occurs prior to the structural remodeling (with lack of fibrosis) in HF and it is crucial in slowed CV and ventricular arrhythmia development. Our findings suggest that altered Cx43 alone is arrhythmogenic and modulation of Cx43 has the anti-arrhythmic therapeutic potential for HF patients.

Atrial arrhythmia related outcomes in critically ill COVID-19 patients.

RATIONALE: Coronavirus disease 2019 (COVID-19) is associated with many clinical manifestations including respiratory failure and cardiovascular compromise. OBJECTIVES: We examine outcomes in critically ill individuals with COVID-19 who develop atrial tachyarrhythmias. METHODS: We collected data from electrocardiograms and the electronic medical record of COVID-19 positive (COVID+ ) and negative (COVID- ) individuals admitted to our medical intensive care unit between February 29 and June 28, 2020. We compared clinical and demographic characteristics, new onset atrial tachyarrhythmia, hemodynamic compromise following atrial tachyarrhythmia, and in-hospital mortality in COVID+ versus COVID- . Hemodynamic compromise was defined as having a new or increased vasopressor requirement or the need for direct current cardioversion for hemodynamic instability within 1 hour of atrial tachyarrhythmia onset. RESULTS: Of 300 individuals included, 200 were COVID+ and 100 were COVID- . Mean age was 60 ± 16 years, 180 (60%) were males, and 170 (57%) were African American. New onset atrial tachyarrhythmia occurred in 16% of COVID+ and 19% of COVID- individuals (P = .51). When compared to COVID- participants without atrial tachyarrhythmia, COVID+ individuals with new onset atrial tachyarrhythmia had higher mortality after multivariable adjustment (OR 5.0, 95% CI 1.9-13.5). New onset atrial tachyarrhythmia was followed by hemodynamic compromise in 18 COVID+ but no COVID- participants (P = .0001). COVID+ individuals with hemodynamic compromise after atrial tachyarrhythmia required increased ventilatory support at the time of atrial tachyarrhythmia onset. CONCLUSIONS: Atrial tachyarrhythmia is associated with increased mortality in critically ill individuals with COVID-19, especially those mechanically ventilated. Recognition of this could assist with clinical care for individuals with COVID-19.

Racial and socioeconomic disparity associates with differences in cardiac DNA methylation among men with end-stage heart failure.

Heart failure (HF) is a multifactorial syndrome that remains a leading cause of worldwide morbidity. Despite its high prevalence, only half of patients with HF respond to guideline-directed medical management, prompting therapeutic efforts to confront the molecular underpinnings of its heterogeneity. In the current study, we examined epigenetics as a yet unexplored source of heterogeneity among patients with end-stage HF. Specifically, a multicohort-based study was designed to quantify cardiac genome-wide cytosine-p-guanine (CpG) methylation of cardiac biopsies from male patients undergoing left ventricular assist device (LVAD) implantation. In both pilot (n = 11) and testing (n = 31) cohorts, unsupervised multidimensional scaling of genome-wide myocardial DNA methylation exhibited a bimodal distribution of CpG methylation found largely to occur in the promoter regions of metabolic genes. Among the available patient attributes, only categorical self-identified patient race could delineate this methylation signature, with African American (AA) and Caucasian American (CA) samples clustering separately. Because race is a social construct, and thus a poor proxy of human physiology, extensive review of medical records was conducted, but ultimately failed to identify covariates of race at the time of LVAD surgery. By contrast, retrospective analysis exposed a higher all-cause mortality among AA (56.3%) relative to CA (16.7%) patients at 2 yr following LVAD placement (P = 0.03). Geocoding-based approximation of patient demographics uncovered disparities in income levels among AA relative to CA patients. Although additional studies are needed, the current analysis implicates cardiac DNA methylation as a previously unrecognized indicator of socioeconomic disparity in human heart failure outcomes.NEW & NOTEWORTHY A bimodal signature of cardiac DNA methylation in heart failure corresponds with racial differences in all-cause mortality following mechanical circulatory support. Racial differences in promoter methylation disproportionately affect metabolic signaling pathways. Socioeconomic factors are associated with racial differences in the cardiac methylome among men with end-stage heart failure.

Alterations of housekeeping proteins in human aged and diseased hearts.

Pathological remodeling includes alterations of ion channel function and calcium homeostasis and ultimately cardiac maladaptive function during the process of disease development. Biochemical assays are important approaches for assessing protein abundance and post-translational modification of ion channels. Several housekeeping proteins are commonly used as internal controls to minimize loading variabilities in immunoblotting protein assays. Yet, emerging evidence suggests that some housekeeping proteins may be abnormally altered under certain pathological conditions. However, alterations of housekeeping proteins in aged and diseased human hearts remain unclear. In the current study, immunoblotting was applied to measure three commonly used housekeeping proteins (ß-actin, calsequestrin, and GAPDH) in well-procured human right atria (RA) and left ventricles (LV) from diabetic, heart failure, and aged human organ donors. Linear regression analysis suggested that the amounts of linearly loaded total proteins and quantified intensity of total proteins from either Ponceau S (PS) blot-stained or Coomassie Blue (CB) gel-stained images were highly correlated. Thus, all immunoblotting data were normalized with quantitative CB or PS data to calibrate potential loading variabilities. In the human heart, ß-actin was reduced in diabetic RA and LV, while GAPDH was altered in aged and diabetic RA but not LV. Calsequestrin, an important Ca2+ regulatory protein, was significantly changed in aged, diabetic, and ischemic failing hearts. Intriguingly, expression levels of all three proteins were unchanged in non-ischemic failing human LV. Overall, alterations of human housekeeping proteins are heart chamber specific and disease context dependent. The choice of immunoblotting loading controls should be carefully evaluated. Usage of CB or PS total protein analysis could be a viable alternative approach for some complicated pathological specimens.

A novel case of reverse Takotsubo cardiomyopathy following brain biopsy.

Reverse Takotsubo cardiomyopathy (rTTC) is a variant of Takotsubo cardiomyopathy (TTC) or stress-induced cardiomyopathy. TTC is a transient cardiomyopathy resulting in a heart failure syndrome, triggered by emotional and/or physical stressors, that is usually self-limited. rTTC is characterized by basal wall hypokinesis and apical wall hyperkinesis, the opposite of TTC. rTTC is more commonly associated with neurologic conditions, most notably intracranial hemorrhage. We present the first case in the literature of rTTC specifically following brain biopsy.

Heart Failure with Preserved Ejection Fraction in a Postpartum Patient with Superimposed Preeclampsia and COVID-19.

Our understanding of COVID-19 in pregnant and postpartum women is rapidly evolving. We present a case from March 2020 of a 25-year-old G2P2002 whose delivery was complicated by preeclampsia with severe features who presented to the emergency department 9 days after cesarean delivery with chest tightness and dyspnea on exertion. On presentation she had severe hypertension, pulmonary edema, elevated brain natriuretic peptide, and high-sensitivity troponin-I, suggesting a diagnosis of hypertensive emergency leading to heart failure with a preserved ejection fraction resulting in pulmonary edema and abnormal cardiac screening tests. However, bilateral opacities were seen on a computed tomography of the chest, and COVID-19 testing was positive. A high index of suspicion for both COVID-19 and cardiovascular complications are critical for optimal patient outcomes and protection of health care workers.

Genome-wide DNA methylation encodes cardiac transcriptional reprogramming in human ischemic heart failure.

Ischemic cardiomyopathy (ICM) is the clinical endpoint of coronary heart disease and a leading cause of heart failure. Despite growing demands to develop personalized approaches to treat ICM, progress is limited by inadequate knowledge of its pathogenesis. Since epigenetics has been implicated in the development of other chronic diseases, the current study was designed to determine whether transcriptional and/or epigenetic changes are sufficient to distinguish ICM from other etiologies of heart failure. Specifically, we hypothesize that genome-wide DNA methylation encodes transcriptional reprogramming in ICM. RNA-sequencing analysis was performed on human ischemic left ventricular tissue obtained from patients with end-stage heart failure, which enriched known targets of the polycomb methyltransferase EZH2 compared to non-ischemic hearts. Combined RNA sequencing and genome-wide DNA methylation analysis revealed a robust gene expression pattern consistent with suppression of oxidative metabolism, induced anaerobic glycolysis, and altered cellular remodeling. Lastly, KLF15 was identified as a putative upstream regulator of metabolic gene expression that was itself regulated by EZH2 in a SET domain-dependent manner. Our observations therefore define a novel role of DNA methylation in the metabolic reprogramming of ICM. Furthermore, we identify EZH2 as an epigenetic regulator of KLF15 along with DNA hypermethylation, and we propose a novel mechanism through which coronary heart disease reprograms the expression of both intermediate enzymes and upstream regulators of cardiac metabolism such as KLF15.

Noninvasive Activation Imaging of Ventricular Arrhythmias by Spatial Gradient Sparse in Frequency Domain-Application to Mapping Reentrant Ventricular Tachycardia.

The aim of this paper is to develop and evaluate a novel imaging method [spatial gradient sparse in frequency domain (SSF)] for the reconstruction of activation sequences of ventricular arrhythmia from noninvasive body surface potential map (BSPM) measurements. We formulated and solved the electrocardiographic inverse problem in the frequency domain, and the activation time was encoded in the phase information of the imaging solution. A cellular automaton heart model was used to generate focal ventricular tachycardia (VT). Different levels of Gaussian white noise were added to simulate noise-contaminated BSPM. The performance of SSF was compared with that of weighted minimum norm inverse solution. We also evaluated the method in a swine model with simultaneous intracardiac and body surface recordings. Four reentrant VTs were observed in pigs with myocardial infarction generated by left anterior descending artery occlusion. The imaged activation sequences of reentrant VTs were compared with those obtained from intracardiac electrograms. In focal VT simulation, SSF has increased the correlation coefficient (CC) by 5% and decreased localization errors (LEs) by 2.7 mm on average under different noise levels. In the animal validation with reentrant VT, SSF has achieved an average CC of 88% and an average LE of 6.3 mm in localizing the earliest and latest activation site in the reentry circuit. Our promising results suggest that the SSF provides noninvasive imaging capability of detecting and mapping macro-reentrant circuits in 3-D ventricular space. The SSF may become a useful imaging tool of identifying and localizing the potential targets for ablation of focal and reentrant VT.

Evidence for a hyper-reductive redox in a sub-set of heart failure patients.

BACKGROUND: Oxidative stress has been linked to heart failure (HF) in humans. Antioxidant-based treatments are often ineffective. Therefore, we hypothesize that some of the HF patients might have a reductive stress (RS) condition. Investigating RS-related mechanisms will aid in personalized optimization of redox homeostasis for better outcomes among HF patients. METHODS: Blood samples were collected from HF patients (n = 54) and healthy controls (n = 42) and serum was immediately preserved in - 80 °C for redox analysis. Malondialdehyde (MDA; lipid peroxidation) levels by HPLC, reduced glutathione (GSH) and its redox ratio (GSH/GSSG) using enzymatic-recycling assay in the serum of HF patients were measured. Further, the activities of key antioxidant enzymes were analyzed by UV-Vis spectrophotometry. Non-invasive echocardiography was used to relate circulating redox status with cardiac function and remodeling. RESULTS: The circulatory redox state (GSH/MDA ratio) was used to stratify the HF patients into normal redox (NR), hyper-oxidative (HO), and hyper-reductive (HR) groups. While the majority of the HF patients exhibited the HO (42%), 41% of them had a normal redox (NR) state. Surprisingly, a subset of HF patients (17%) belonged to the hyper-reductive group, suggesting a strong implication for RS in the progression of HF. In all the groups of HF patients, SOD, GPx and catalase were significantly increased while GR activity was significantly reduced relative to healthy controls. Furthermore, echocardiography analyses revealed that 55% of HO patients had higher systolic dysfunction while 62.5% of the hyper-reductive patients had higher diastolic dysfunction. CONCLUSION: These results suggest that RS may be associated with HF pathogenesis for a subset of cardiac patients. Thus, stratification of HF patients based on their circulating redox status may serve as a useful prognostic tool to guide clinicians designing personalized antioxidant therapies.

Altered Repolarization Reserve in Failing Rabbit Ventricular Myocytes: Calcium and ß-Adrenergic Effects on Delayed- and Inward-Rectifier Potassium Currents.

BACKGROUND: Electrophysiological remodeling and increased susceptibility for cardiac arrhythmias are hallmarks of heart failure (HF). Ventricular action potential duration (APD) is typically prolonged in HF, with reduced repolarization reserve. However, underlying K+ current changes are often measured in nonphysiological conditions (voltage clamp, low pacing rates, cytosolic Ca2+ buffers). METHODS AND RESULTS: We measured the major K+ currents (IKr, IKs, and IK1) and their Ca2+- and ß-adrenergic dependence in rabbit ventricular myocytes in chronic pressure/volume overload-induced HF (versus age-matched controls). APD was significantly prolonged only at lower pacing rates (0.2-1 Hz) in HF under physiological ionic conditions and temperature. However, when cytosolic Ca2+ was buffered, APD prolongation in HF was also significant at higher pacing rates. Beat-to-beat variability of APD was also significantly increased in HF. Both IKr and IKs were significantly upregulated in HF under action potential clamp, but only when cytosolic Ca2+ was not buffered. CaMKII (Ca2+/calmodulin-dependent protein kinase II) inhibition abolished IKs upregulation in HF, but it did not affect IKr. IKs response to ß-adrenergic stimulation was also significantly diminished in HF. IK1 was also decreased in HF regardless of Ca2+ buffering, CaMKII inhibition, or ß-adrenergic stimulation. CONCLUSIONS: At baseline Ca2+-dependent upregulation of IKr and IKs in HF counterbalances the reduced IK1, maintaining repolarization reserve (especially at higher heart rates) in physiological conditions, unlike conditions of strong cytosolic Ca2+ buffering. However, under ß-adrenergic stimulation, reduced IKs responsiveness severely limits integrated repolarizing K+ current and repolarization reserve in HF. This would increase arrhythmia propensity in HF, especially during adrenergic stress.

Knockout of the ATPase inhibitory factor 1 protects the heart from pressure overload-induced cardiac hypertrophy.

Mitochondrial ATP synthase catalyzes the coupling of oxidative phosphorylation. Under pathological conditions, ATP synthase hydrolyzes ATP to replenish protons from the matrix into the intermembrane space, sustaining mitochondrial membrane potential. ATPase inhibitory factor 1 (IF1) is a nuclear-encoded, ATP synthase-interacting protein that selectively inhibits the hydrolysis activity of ATP synthase, which may render the protective role of IF1 in ischemic hearts. However, the in vivo cardiac function of IF1 and the potential therapeutic application targeting IF1 remain obscure. In the present study, we uncovered that IF1 is upregulated in mouse hearts with pressure overload-induced hypertrophy and in human hearts with dilated cardiomyopathy. IF1 knockout (KO) mice were protected against cardiac dysfunction and pathological development induced by transverse aortic constriction (TAC) or isoproterenol infusion. The reduced ATP hydrolysis activated AMPK activity in IF1 KO hearts, which together facilitated autophagy. These results suggest that IF1 upregulation in the failing heart may be a maladaptive response. Inhibiting IF1 in the hypertrophied heart not only prevents cell death from excessive mitochondrial depolarization but also activates AMPK signaling and increases autophagy. Therefore, IF1 inhibition may serve as a potential therapeutic target in treating pathological cardiac hypertrophy and heart failure.

Regulation of cardiac gap junctions by protein phosphatases.

Sufficient connexin-mediated intercellular coupling is critical to maintain gap junctional communication for proper cardiac function. Alterations in connexin phosphorylation state, particularly dephosphorylation of connexin 43 (Cx43), may impact cell coupling and conduction in disease states. Cx43 dephosphorylation may be carried out by protein phosphatase activity. Here, we present an overview of the key phosphatases known to interact with Cx43 or modulators of Cx43, as well as some possible therapeutic targets to regulate phosphatase activity in the heart.

Reduced Arrhythmia Inducibility With Calcium/Calmodulin-dependent Protein Kinase II Inhibition in Heart Failure Rabbits.

RATIONALE: Calcium/calmodulin-dependent protein kinase II (CaMKII) is activated in heart failure (HF) and can contribute to arrhythmias induced by ß-adrenergic receptor-mediated sarcoplasmic reticulum calcium leak. OBJECTIVE: To evaluate the effect of CaMKII inhibition on ventricular tachycardia (VT) induction in conscious HF and naive rabbits. METHODS AND RESULTS: Nonischemic HF was induced by aortic insufficiency and constriction. Electrocardiograms were recorded in rabbits pretreated with vehicle (saline) or the CaMKII inhibitor KN-93 (300 µg/kg); VT was induced by infusion of increasing doses of norepinephrine (1.56-25 µg·kg⁻¹·min⁻¹) in naive (n = 8) and HF (n = 7) rabbits. With saline, median VT dose threshold in HF was 6.25 versus 12.5 µg·kg⁻¹·min⁻¹ norepinephrine in naive rabbits (P = 0.06). Pretreatment with KN-93 significantly increased VT threshold in HF and naive rabbits (median = 25 µg·kg⁻¹·min⁻¹, P < 0.05 vs. saline for both groups). Mean cycle length of VT initiation was shorter in HF (221 ± 20 milliseconds) than naive (296 ± 23 milliseconds, P < 0.05) rabbits with saline; this difference was not significant after treatment with KN-93. CONCLUSIONS: KN-93 significantly reduced arrhythmia inducibility and slowed initiation of VT, suggesting that CaMKII inhibition may have antiarrhythmic effects in the failing human heart.

Fibro-porous poliglecaprone/polycaprolactone conduits: synergistic effect of composition and in vitro degradation on mechanical properties.

Blends of poliglecaprone (PGC) and polycaprolactone (PCL) of varying compositions were electrospun into tubular conduits and their mechanical, morphological, thermal and in vitro degradation properties were evaluated under simulated physiological conditions. Generally, mechanical strength, modulus and hydrophilic nature were enhanced by the addition of PGC to PCL. An in vitro degradation study in phosphate-buffered saline (pH 7.3) was carried out for up to 1 month to understand the hydrolytic degradation effect on the mechanical properties in both the longitudinal and circumferential directions. Pure PCL and 4:1 PCL/PGC blend scaffolds exhibited considerable elastic stiffening after a 1 month in vitro degradation. Fourier transform infrared spectroscopic and DSC techniques were used to understand the degradation behavior and the changes in structure and crystallinity of the polymeric blends. A 3:1 PCL/PGC blend was concluded to be a judicious blend composition for tubular grafts based on overall results on the mechanical properties and performance after a 1 month in vitro degradation study.

Imaging cardiac activation sequence during ventricular tachycardia in a canine model of nonischemic heart failure.

Noninvasive cardiac activation imaging of ventricular tachycardia (VT) is important in the clinical diagnosis and treatment of arrhythmias in heart failure (HF) patients. This study investigated the ability of the three-dimensional cardiac electrical imaging (3DCEI) technique for characterizing the activation patterns of spontaneously occurring and norepinephrine (NE)-induced VTs in a newly developed arrhythmogenic canine model of nonischemic HF. HF was induced by aortic insufficiency followed by aortic constriction in three canines. Up to 128 body-surface ECGs were measured simultaneously with bipolar recordings from up to 232 intramural sites in a closed-chest condition. Data analysis was performed on the spontaneously occurring VTs (n=4) and the NE-induced nonsustained VTs (n=8) in HF canines. Both spontaneously occurring and NE-induced nonsustained VTs initiated by a focal mechanism primarily from the subendocardium, but occasionally from the subepicardium of left ventricle. Most focal initiation sites were located at apex, right ventricular outflow tract, and left lateral wall. The NE-induced VTs were longer, more rapid, and had more focal sites than the spontaneously occurring VTs. Good correlation was obtained between imaged activation sequence and direct measurements (averaged correlation coefficient of ∼0.70 over 135 VT beats). The reconstructed initiation sites were ∼10 mm from measured initiation sites, suggesting good localization in such a large animal model with cardiac size similar to a human. Both spontaneously occurring and NE-induced nonsustained VTs had focal initiation in this canine model of nonischemic HF. 3DCEI is feasible to image the activation sequence and help define arrhythmia mechanism of nonischemic HF-associated VTs.

Mitochondria-derived ROS bursts disturb Ca²âº cycling and induce abnormal automaticity in guinea pig cardiomyocytes: a theoretical study.

Mitochondria are in close proximity to the redox-sensitive sarcoplasmic reticulum (SR) Ca(2+) release [ryanodine receptors (RyRs)] and uptake [Ca(2+)-ATPase (SERCA)] channels. Thus mitochondria-derived reactive oxygen species (mdROS) could play a crucial role in modulating Ca(2+) cycling in the cardiomyocytes. However, whether mdROS-mediated Ca(2+) dysregulation translates to abnormal electrical activities under pathological conditions, and if yes what are the underlying ionic mechanisms, have not been fully elucidated. We hypothesize that pathological mdROS induce Ca(2+) elevation by modulating SR Ca(2+) handling, which activates other Ca(2+) channels and further exacerbates Ca(2+) dysregulation, leading to abnormal action potential (AP). We also propose that the morphologies of elicited AP abnormality rely on the time of mdROS induction, interaction between mitochondria and SR, and intensity of mitochondrial oxidative stress. To test the hypotheses, we developed a multiscale guinea pig cardiomyocyte model that incorporates excitation-contraction coupling, local Ca(2+) control, mitochondrial energetics, and ROS-induced ROS release. This model, for the first time, includes mitochondria-SR microdomain and modulations of mdROS on RyR and SERCA activities. Simulations show that mdROS bursts increase cytosolic Ca(2+) by stimulating RyRs and inhibiting SERCA, which activates the Na(+)/Ca(2+) exchanger, Ca(2+)-sensitive nonspecific cationic channels, and Ca(2+)-induced Ca(2+) release, eliciting abnormal AP. The morphologies of AP abnormality are largely influenced by the time interval among mdROS burst induction and AP firing, dosage and diffusion of mdROS, and SR-mitochondria distance. This study defines the role of mdROS in Ca(2+) overload-mediated cardiac arrhythmogenesis and underscores the importance of considering mitochondrial targets in designing new antiarrhythmic therapies.

Sex differences in ß-adrenergic responsiveness of action potentials and intracellular calcium handling in isolated rabbit hearts.

Cardioprotection in females, as observed in the setting of heart failure, has been attributed to sex differences in intracellular calcium handling and its modulation by ß-adrenergic signaling. However, further studies examining sex differences in ß-adrenergic responsiveness have yielded inconsistent results and have mostly been limited to studies of contractility, ion channel function, or calcium handling alone. Given the close interaction of the action potential (AP) and intracellular calcium transient (CaT) through the process of excitation-contraction coupling, the need for studies exploring the relationship between agonist-induced AP and calcium handling changes in female and male hearts is evident. Thus, the aim of this study was to use optical mapping to examine sex differences in ventricular APs and CaTs measured simultaneously from Langendorff-perfused hearts isolated from naïve adult rabbits during ß-adrenergic stimulation. The non-selective ß-agonist isoproterenol (Iso) decreased AP duration (APD90), CaT duration (CaD80), and the decay constant of the CaT (τ) in a dose-dependent manner (1-316.2 nM), with a plateau at doses ≥31.6 nM. The Iso-induced changes in APD90 and τ (but not CaD80) were significantly smaller in female than male hearts. These sex differences were more significant at faster (5.5 Hz) than resting rates (3 Hz). Treatment with Iso led to the development of spontaneous calcium release (SCR) with a dose threshold of 31.6 nM. While SCR occurrence was similar in female (49%) and male (53%) hearts, the associated ectopic beats had a lower frequency of occurrence (16% versus 40%) and higher threshold (100 nM versus 31.6 nM) in female than male hearts (p<0.05). In conclusion, female hearts had a decreased capacity to respond to ß-adrenergic stimulation, particularly under conditions of increased demand (i.e. faster pacing rates and "maximal" levels of Iso effects), however this reduced ß-adrenergic responsiveness of female hearts was associated with reduced arrhythmic activity.

Morning surge of ventricular arrhythmias in a new arrhythmogenic canine model of chronic heart failure is associated with attenuation of time-of-day dependence of heart rate and autonomic adaptation, and reduced cardiac chaos.

Patients with chronic heart failure (CHF) exhibit a morning surge in ventricular arrhythmias, but the underlying cause remains unknown. The aim of this study was to determine if heart rate dynamics, autonomic input (assessed by heart rate variability (HRV)) and nonlinear dynamics as well as their abnormal time-of-day-dependent oscillations in a newly developed arrhythmogenic canine heart failure model are associated with a morning surge in ventricular arrhythmias. CHF was induced in dogs by aortic insufficiency & aortic constriction, and assessed by echocardiography. Holter monitoring was performed to study time-of-day-dependent variation in ventricular arrhythmias (PVCs, VT), traditional HRV measures, and nonlinear dynamics (including detrended fluctuations analysis α1 and α2 (DFAα1 & DFAα2), correlation dimension (CD), and Shannon entropy (SE)) at baseline, as well as 240 days (240 d) and 720 days (720 d) following CHF induction. LV fractional shortening was decreased at both 240 d and 720 d. Both PVCs and VT increased with CHF duration and showed a morning rise (2.5-fold & 1.8-fold increase at 6 AM-noon vs midnight-6 AM) during CHF. The morning rise in HR at baseline was significantly attenuated by 52% with development of CHF (at both 240 d & 720 d). Morning rise in the ratio of low frequency to high frequency (LF/HF) HRV at baseline was markedly attenuated with CHF. DFAα1, DFAα2, CD and SE all decreased with CHF by 31, 17, 34 and 7%, respectively. Time-of-day-dependent variations in LF/HF, CD, DFA α1 and SE, observed at baseline, were lost during CHF. Thus in this new arrhythmogenic canine CHF model, attenuated morning HR rise, blunted autonomic oscillation, decreased cardiac chaos and complexity of heart rate, as well as aberrant time-of-day-dependent variations in many of these parameters were associated with a morning surge of ventricular arrhythmias.