We have developed a detailed mathematical model for Ca handling and ionic currents in the human ventricular myocyte. Our aims were to: 1) simulate basic excitation-contraction coupling phenomena; 2) use realistic repolarizing K current densities; 3) reach steady-state. The model relies on the framework of the rabbit myocyte model previously developed by our group, with subsarcolemmal and junctional compartments where ion channels sense higher [Ca] vs. bulk cytosol. Ion channels and transporters have been modeled on the basis of the most recent experimental data from human ventricular myocytes. Rapidly and slowly inactivating components of I to have been formulated to differentiate between endocardial and epicardial myocytes. Transmural gradients of Ca handling proteins and Na pump were also simulated. The model has been validated against a wide set of experimental data including action potential duration (APD) adaptation and restitution, frequencydependent increase in Ca transient peak and [Na] i . Interestingly, Na accumulation at fast heart rate is a major determinant of APD shortening, via outward shifts in Na pump and Na-Ca exchange currents. We investigated the effects of blocking K currents on APD and repolarization reserve: I Ks block does not affect the former and slightly reduces the latter; I K1 blockade modestly increases APD and more strongly reduces repolarization reserve; I Kr blockers significantly prolong APD, an effect exacerbated as pacing frequency is decreased, in good agreement with experimental results in human myocytes. We conclude that this model provides a useful framework to explore excitationcontraction coupling mechanisms and repolarization abnormalities at the single myocyte level.

Rationale Understanding atrial fibrillation (AF) requires integrated understanding of ionic currents and Ca2+ transport in remodeled human atrium, but appropriate models are limited. Objective To study AF we developed a new human atrial action potential (AP) model, derived from atrial experimental results and our human ventricular myocyte model. Methods and Results Atria vs. ventricles have lower IK1, resulting in more depolarized resting membrane potential (~7mV). We used higher Ito,fast density in atrium, removed Ito,slow, and included an atrial-specific IKur. INCX and INaK densities were reduced in atrial vs. ventricular myocytes according to experimental results. SERCA function was altered to reproduce human atrial myocyte Ca2+ transients. To simulate chronic AF, we reduced ICaL, Ito, IKur and SERCA, and increased IK1, IKs and INCX. We also investigated the link between Kv1.5 channelopathy, [Ca2+]i, and AF. The sinus rhythm model showed a typical human atrial AP morphology. Consistent with experiments, the model showed shorter APs and reduced AP duration shortening at increasing pacing frequencies in AF or when ICaL was partially blocked, suggesting a crucial role of Ca2+ and Na+ in this effect. This also explained blunted Ca2+ transient and rate-adaptation of [Ca2+]i and [Na+]i in chronic AF. Moreover, increasing [Na+]i and altered INaK and INCX causes rate-dependent atrial AP shortening. Blocking IKur to mimic Kv1.5 loss-of-function increased [Ca2+]i and caused early-afterdepolarizations under adrenergic stress, as observed experimentally. Conclusions Our study provides a novel tool and insights into ionic bases of atrio-ventricular AP differences, and shows how Na+ and Ca2+ homeostasis critically mediate abnormal repolarization in AF.

Rationale In heart failure (HF), CaMKII expression and reactive oxygen species (ROS) are increased. Both ROS and CaMKII can increase late INa leading to intracellular Na accumulation and arrhythmias. It has been shown that ROS can activate CaMKII via oxidation. Objective We tested whether CaMKIIδ is required for ROS-dependent late INa regulation and if ROS-induced Ca released from the sarcoplasmic reticulum (SR) is involved. Methods and Results 40 µmol/L H2O2 significantly increased CaMKII oxidation and autophosphorylation in permeabilized rabbit cardiomyocytes. Without free [Ca]i (5 mmol/L BAPTA/1 mmol/L Br2-BAPTA) or after SR depletion (caffeine 10 mmol/L, thapsigargin 5 µmol/L) the H2O2-dependent CaMKII oxidation and autophosphorylation was abolished. H2O2 significantly increased SR Ca spark frequency (confocal microscopy) but reduced SR Ca load. In wildtype (WT) mouse myocytes, H2O2 increased late INa (whole cell patch-clamp). This increase was abolished in CaMKIIδ−/− myocytes. H2O2-induced [Na]i and [Ca]i accumulation (SBFI and Indo-1 epifluorescence) was significantly slowed in CaMKIIδ−/− myocytes (vs. WT). CaMKIIδ−/− myocytes developed significantly less H2O2-induced arrhythmias, and were more resistant to hypercontracture. Opposite results (increased late INa, [Na]i and [Ca]i accumulation) were obtained by overexpression of CaMKIIδ in rabbit myocytes (adenoviral gene transfer) reversible with CaMKII inhibition (10 µmol/L KN93 or 0.1 µmol/L AIP). Conclusion Free [Ca]i and a functional SR are required for ROS activation of CaMKII. ROS-activated CaMKIIδ enhances late INa, which may lead to cellular Na and Ca overload. This may be of relevance in HF, where enhanced ROS production meets increased CaMKII expression.

Compartmentalized cAMP/PKA signalling is now recognized as important for physiology and pathophysiology, yet a detailed understanding of the properties, regulation and function of local cAMP/PKA signals is lacking. Here we present a fluorescence resonance energy transfer (FRET)-based sensor, CUTie, which detects compartmentalized cAMP with unprecedented accuracy. CUTie, targeted to specific multiprotein complexes at discrete plasmalemmal, sarcoplasmic reticular and myofilament sites, reveals differential kinetics and amplitudes of localized cAMP signals. This nanoscopic heterogeneity of cAMP signals is necessary to optimize cardiac contractility upon adrenergic activation. At low adrenergic levels, and those mimicking heart failure, differential local cAMP responses are exacerbated, with near abolition of cAMP signalling at certain locations. This work provides tools and fundamental mechanistic insights into subcellular adrenergic signalling in normal and pathological cardiac function.

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Hypercontractility of arterial myocytes and enhanced vascular tone during diabetes are, in part, attributed to the effects of increased glucose (hyperglycemia) on L-type CaV1.2 channels. In murine arterial myocytes, kinase-dependent mechanisms mediate the increase in CaV1.2 activity in response to increased extracellular glucose. We identified a subpopulation of the CaV1.2 channel pore-forming subunit (α1C) within nanometer proximity of protein kinase A (PKA) at the sarcolemma of murine and human arterial myocytes. This arrangement depended upon scaffolding of PKA by an A-kinase anchoring protein 150 (AKAP150) in mice. Glucose-mediated increases in CaV1.2 channel activity were associated with PKA activity, leading to α1C phosphorylation at Ser1928. Compared to arteries from low-fat diet (LFD)–fed mice and nondiabetic patients, arteries from high-fat diet (HFD)–fed mice and from diabetic patients had increased Ser1928 phosphorylation and CaV1.2 activity. Arterial myocytes and arteries from mice lacking AKAP150 or expressing mutant AKAP150 unable to bind PKA did not exhibit increased Ser1928 phosphorylation and CaV1.2 current density in response to increased glucose or to HFD. Consistent with a functional role for Ser1928 phosphorylation, arterial myocytes and arteries from knockin mice expressing a CaV1.2 with Ser1928 mutated to alanine (S1928A) lacked glucose-mediated increases in CaV1.2 activity and vasoconstriction. Furthermore, the HFD-induced increases in CaV1.2 current density and myogenic tone were prevented in S1928A knockin mice. These findings reveal an essential role for α1C phosphorylation at Ser1928 in stimulating CaV1.2 channel activity and vasoconstriction by AKAP-targeted PKA upon exposure to increased glucose and in diabetes.

Optimal cardiac function depends on proper timing of excitation and contraction in various regions of the heart, as well as on appropriate heart rate. This is accomplished via specialized electrical properties of various components of the system, including the sinoatrial node, atria, atrioventricular node, His-Purkinje system, and ventricles. Here we review the major regionally-determined electrical properties of these cardiac regions and present the available data regarding the molecular and ionic bases of regional cardiac function and dysfunction. Understanding these differences is of fundamental importance for the investigation of arrhythmia mechanisms and pharmacotherapy.

Background-Potassium currents contribute to action potential duration (APD) and arrhythmogenesis. In heart failure, Ca/calmodulin-dependent protein kinase II (CaMKII) is upregulated and can alter ion channel regulation and expression. Methods and Results-We examine the influence of overexpressing cytoplasmic CaMKII␦ C , both acutely in rabbit ventricular myocytes (24-hour adenoviral gene transfer) and chronically in CaMKII␦ C -transgenic mice, on transient outward potassium current (I to ), and inward rectifying current (I K1 ). Acute and chronic CaMKII overexpression increases I to,slow amplitude and expression of the underlying channel protein K V 1.4. Chronic but not acute CaMKII overexpression causes downregulation of I to,fast , as well as K V 4.2 and KChIP2, suggesting that K V 1.4 expression responds faster and oppositely to K V 4.2 on CaMKII activation. These amplitude changes were not reversed by CaMKII inhibition, consistent with CaMKII-dependent regulation of channel expression and/or trafficking. CaMKII (acute and chronic) greatly accelerated recovery from inactivation for both I to components, but these effects were acutely reversed by AIP (CaMKII inhibitor), suggesting that CaMKII activity directly accelerates I to recovery. Expression levels of I K1 and Kir2.1 mRNA were downregulated by CaMKII overexpression. CaMKII acutely increased I K1 , based on inhibition by AIP (in both models). CaMKII overexpression in mouse prolonged APD (consistent with reduced I to,fast and I K1 ), whereas CaMKII overexpression in rabbit shortened APD (consistent with enhanced I K1 and I to,slow and faster I to recovery). Computational models allowed discrimination of contributions of different channel effects on APD. Key Words: action potentials Ⅲ potassium Ⅲ arrhythmia Ⅲ electrophysiology Ⅲ heart failure H eart failure (HF) is accompanied by arrhythmogenic changes related to electric remodeling. This is associated with prolongation of action potential duration (APD) 1 and downregulation of transient outward K-current (I to ) and inward rectifying K-current (I K1 ). I K1 is responsible for stabilizing the diastolic membrane potential (E m ), such that decreased I K1 increases the propensity for triggered arrhythmias. 2 I to is important in early repolarization and influences the effects of other currents and transporters by affecting AP voltage-time trajectory. There are at least 2 components of I to generated by different K-channel isoforms, which can be distinguished according to their recovery and inactivation kinetics. 3,4 The fast component (I to,fast ) recovers and inactivates with time constants () of Ͻ100 ms, whereas the slow component (I to,slow ) recovers with of hundreds of milliseconds up to several seconds and inactivates with of Ϸ200 ms. 3-5 Downregulation of I to has been described in animal models of hypertrophy and human HF, 2,6,7 is associated with APD prolongation, 8 and predisposes to early afterdepolarizations. Conclusion-CaMKII Clinical Perspective on p 294In HF, expression and activity of Ca/cal...