Rabbit Left Ventricle Research Articles

Effects of neferine on Kv4.3 channels expressed in HEK293 cells and ex vivo electrophysiology of rabbit hearts.

Neferine is an isoquinoline alkaloid isolated from seed embryos of Nelumbo nucifera (Gaertn), which has a variety of biological activities. In this study we examined the effects of neferine on Kv4.3 channels, a major contributor to the transient outward current (I(to)) in rabbit heart, and on ex vivo electrophysiology of rabbit hearts. Whole-cell Kv4.3 currents were recorded in HEK293 cells expressing human cardiac Kv4.3 channels using patch-clamp technique. Arterially perfused wedges of rabbit left ventricles (LV) were prepared, and transmembrane action potentials were simultaneously recorded from epicardial (Epi) and endocardial (Endo) sites with floating microelectrodes together with transmural electrocardiography (ECG). Neferine (0.1-100 μmol/L) dose-dependently and reversibly inhibited Kv4.3 currents (the IC50 value was 8.437 μmol/L, and the maximal inhibition at 100 μmol/L was 44.12%). Neferine (10 μmol/L) caused a positive shift of the steady-state activation curve of Kv4.3 currents, and a negative shift of the steady-state inactivation curve. Furthermore, neferine (10 μmol/L) accelerated the inactivation but not the activation of Kv4.3 currents, and markedly slowed the recovery of Kv4.3 currents from inactivation. Neferine-induced blocking of Kv4.3 currents was frequency-dependent. In arterially perfused wedges of rabbit LV, neferine (1, 3, and 10 μmol/L) dose-dependently prolonged the QT intervals and action potential durations (APD) at both Epi and Endo sites, and caused dramatic increase of APD10 at Epi sites. Neferine inhibits Kv4.3 channels likely by blocking the open state and inactivating state channels, which contributes to neferine-induced dramatic increase of APD10 at Epi sites of rabbit heart.

From the Purkinje fibres to the ventricle: One dimensional computer simulation for the healthy and failing heart.

This study used one-dimensional computer simulation to investigate the influence of heart failure on action potential conduction through the left Purkinje fibres to the left ventricle. The study was based on a rabbit model of left ventricular heart failure caused by volume and pressure overload. To simulate the effect of heart failure, we began with models of the healthy rabbit Purkinje fibre action potential and healthy left ventricular (endocardial) action potential. In the absence of ionic current measurements from failing rabbit Purkinje fibres, we assumed that changes in ionic currents mirrored changes in ion channel expression (measured at the messenger RNA level): ionic conductances were adjusted based on changes in expression of the relevant ion channels. Ionic currents in the left ventricle were adjusted in the same way, but in addition, changes in ionic currents measured in the failing rabbit left ventricle by Ruijter et al. and Powizd et al. were used in simulations. The simulations predict a gradient in action potential duration from the Purkinje fibres to the ventricle and this gradient is exacerbated in heart failure. The predicted changes in the Purkinje fibre and left ventricular action potential were compared to actual changes measured using sharp microelectrodes.

Increased cell membrane capacitance is the dominant mechanism of stretch-dependent conduction slowing in the rabbit heart: a computational study.

Volume loading of the cardiac ventricles is known to slow electrical conduction in the rabbit heart, but the mechanisms remain unclear. Previous experimental and modeling studies have investigated some of these mechanisms, including stretch-activated membrane currents, reduced gap junctional conductance, and altered cell membrane capacitance. In order to quantify the relative contributions of these mechanisms, we combined a monomain model of rabbit ventricular electrophysiology with a hyperelastic model of passive ventricular mechanics. First, a simplified geometric model with prescribed homogeneous deformation was used to fit model parameters and characterize individual MEF mechanisms, and showed good qualitative agreement with experimentally measured strain-CV relations. A 3D model of the rabbit left and right ventricles was then compared with experimental measurements from optical electrical mapping studies in the isolated rabbit heart. The model was inflated to an end-diastolic pressure of 30 mmHg, resulting in epicardial strains comparable to those measured in the anterior left ventricular free wall. While the effects of stretch activated channels did alter epicardial conduction velocity, an increase in cellular capacitance was required to explain previously reported experimental results. The new results suggest that for large strains, various mechanisms can combine and produce a biphasic relationship between strain and conduction velocity. However, at the moderate strains generated by high end-diastolic pressure, a stretch-induced increase in myocyte membrane capacitance is the dominant driver of conduction slowing during ventricular volume loading.

Structural heterogeneity modulates effective refractory period: a mechanism of focal arrhythmia initiation.

Reductions in electrotonic loading around regions of structural and electrophysiological heterogeneity may facilitate capture of focal triggered activity, initiating reentrant arrhythmias. How electrotonic loading, refractoriness and capture of focal ectopics depend upon the intricate nature of physiological structural anatomy, as well as pathological tissue remodelling, however, is not well understood. In this study, we performed computational bidomain simulations with anatomically-detailed models representing the rabbit left ventricle. We used these models to quantify the relationship between local structural anatomy and spatial heterogeneity in action potential (AP) characteristics, electrotonic currents and effective refractory periods (ERPs) under pacing and restitution protocols. Regions surrounding vessel cavities, in addition to tissue surfaces, had significantly lower peak downstream electrotonic currents than well coupled myocardium ( vs A/cm2), with faster maximum AP upstroke velocities ( vs mV/ms), although noticeably very similar APDs ( vs ms) and AP restitution properties. Despite similarities in APDs, ERPs in regions of low electrotonic load in the vicinity of surfaces, intramural vessel cavities and endocardial structures were up to ms shorter compared to neighbouring well-coupled tissue, leading to regions of sharp ERP gradients. Consequently, focal extra-stimuli timed within this window of ERP heterogeneity between neighbouring regions readily induced uni-directional block, inducing reentry. Most effective induction sites were within channels of low ERPs between large vessels and epicardium. Significant differences in ERP driven by reductions in electrotonic loading due to fine-scale physiological structural heterogeneity provides an important mechanism of capture of focal activity and reentry induction. Application to pathological ventricles, particularly myocardial infarction, will have important implications in anti-arrhythmia therapy.

Analysis of unipolar electrograms in rabbit heart demonstrated the key role of ventricular apicobasal dispersion in arrhythmogenicity.

Reduced repolarization reserve and increased transmural dispersion of repolarization (TDR) are known risk factors for Torsade de Pointes development, but less is known about the role of apex-to-base (apicobasal) repolarization in arrhythmogenesis. Three needles were inserted in rabbit left ventricle to record unipolar electrograms from endocardium to epicardium and base to apex. Total repolarization interval (TRI) and peak-to-end repolarization interval (Tp) were assessed after quinidine (n=6) and D,L-sotalol (n=6) perfusion in combination with the IKs inhibitor chromanol 293B. About 30µM D,L-sotalol increased TRI and Tp more at the base (TRI +40±4%; Tp +89±11%) relative to the apex (TRI +28±3%, Tp +30±8%). Similar results were obtained with quinidine: TRI and Tp increased more at the base compared to the apex. No significant differences were recorded from the endocardium to the epicardium. Our results show that combined IKr+IKs block prolonged TRI and Tp significantly more at the ventricular base than at the apex, in the absence of transmural dispersion of refractoriness. Regional changes in TRI and Tp indicate the contribution of apicobasal dispersion to arrhythmogenicity compared to TDR in a rabbit heart model.

GW24-e1724 Verapamil reduces transmural dispersion of repolarisation and prevents Torsade de Pointes in LQT2 model of the long QT syndrome

ObjectivesTo investigate the effects of verapamil on LQT2 models and its electrophysiologic mechanism by which verapamil suppresses torsades de pointes (TdP) in a developed left ventricular wedge model of LQT2.MethodsTransmembrane...

The Integration and Functional Evaluation of Rabbit Pacing Cells Transplanted into the Left Ventricular Free Wall

To evaluate the feasibility of cell transplantation to treat bradyarrhythmia, we analyzed the in vivo integration and pacing function after transplantation of mHCN4-modified rabbit bone marrow mesenchymal stem cells (MSCs) into the rabbit left ventricle free wall epicardium. In our investigation, we injected MSCs transduced with or without mHCN4 into the rabbit left ventricle free wall epicardium. Chemical ablation of the sinoatrial node was performed and bilateral vagus nerves were sequentially stimulated to observe premature left ventricular contraction or left ventricular rhythm. We found that the mHCN4-transduced MSC group had a significantly higher ventricular rate and a shorter QRS duration than that of the control and EGFP group. Furthermore, the mHCN4-transduced MSCs, but not the control cells, gradually adapted long-spindle morphology and became indistinguishable from adjacent ventricle myocytes. The modified MSCs showed pacing function approximately 1 week after transplantation and persisted at least 4 weeks after transplantation. In conclusion, a bradyarrhythmia model can be successfully established by chemical ablation of the sinoatrial node and sequential bilateral vagus nerve stimulation. The mHCN4-modified rabbit MSCs displayed evident dynamic morphology changes after being transplanted into rabbit left ventricle free wall epicardium. Our studies may provide a promising strategy of using modified stem cell transplantation to treat bradyarrhythmia.

Adaptation of a Rabbit Myocardium Material Model for Use in a Canine Left Ventricle Simulation Study

The myocardium of the left ventricle (LV) of the heart comprises layers of muscle fibers whose orientation varies through the heart wall. Because of these fibers, accurate modeling of the myocardium stress-strain behavior requires models that are nonlinear, anisotropic, and time-varying. This article describes the development and testing of a material model of the canine LV myocardium, which will be used in ongoing simulations of the mechanics of the LV with fluid-structure interaction. The model assumes that myocardium deformation has two extreme states: one during which the muscle fibers are fully relaxed, and another during which the muscle fibers are fully contracted. During the second state, the "total" stresses are assumed to be the sum of "passive" stresses, which represent the fully relaxed muscle fibers, and "active" stresses, which are additional stresses due to the contraction of the muscle fibers. The canine LV myocardium is modeled as a transversely isotropic material for which material properties vary in the fiber and cross-fiber directions. The material behavior is considered to be hyperelastic and is modeled by a strain-energy density function in a manner that is an adaptation of an approach based on measurements of the stress-strain behavior of rabbit LV myocardia. A numerical method has been developed to calculate suitable parameter values for the passive material model using previous passive canine LV myocardium stress measurements and taking into account existing physical and numerical constraints. In the absence of published measurements of total canine LV myocardium stresses, a method has been developed to estimate these stresses from available passive and total rabbit LV myocardium stresses and then to calculate active material parameter values. Material parameter values were calculated for passive and active canine LV myocardium. Passive stresses calculated using the model compare well to previous stress measurements while active stresses calculated using the model compare well with those approximated from rabbit measurements. The adapted material model of the canine LV myocardium is deemed to be suitable for use in simulations of the operation of both idealized and realistic canine hearts. The estimated model parameter values can be easily revised to more appropriate ones if measurements of active canine LV myocardium stresses become available. The extension of this material model to a fully orthotropic one is also possible but determination of its parameters would require stress-stretch measurements in the fiber and both cross-fiber directions.

Connexin43 knockdown or overexpression modulates cell coupling in control and failing rabbit left ventricular myocytes

We have shown that failing human and rabbit left ventricle (LV) exhibits downregulation and dephosphorylation of connexin43 (Cx43) and that Cx43 dephosphorylation in heart failure (HF) contributes to reduced cell coupling. However, the role of Cx43 downregulation per se in impaired coupling in HF is unclear. First, we used adenovirus (Ad) encoding a Cx43 siRNA sequence to knock down Cx43 protein levels in cultured control rabbit LV myocytes. Cells cultured for up to 48 h with intermittent pacing maintained Cx43 protein levels and phosphorylation status. Cell coupling in Cx43 knockdown myocyte pairs (by Lucifer Yellow dye transfer) was markedly reduced after 24 h infection (associated with approximately 40% Cx43 knockdown) and after 48 h (associated with approximately 70% Cx43 knockdown). The phosphorylation status, distribution of remaining Cx43 proteins, and levels of other cardiac connexins (Cx40 and Cx45) were unchanged. Second, we overexpressed Cx43 to levels comparable to control using an adenovirus encoding wild-type Cx43 (Cx43WT) gene in isolated LV myocytes from our arrhythmogenic HF rabbit model. We found 87% more Cx43WT proteins improved dye coupling [vs. Ad-beta-galactosidase (LacZ) infected HF controls]. Overexpressed Cx43 protein was located throughout the myocyte membrane (same pattern as in controls), and the phosphorylation status of Cx43 remained comparable to that in AdLacZ infected HF controls. In addition to Cx43 dephosphorylation, downregulation of Cx43 plays an essential role in reduced cell coupling in the failing rabbit heart. Modulation of Cx43 expression could be a novel therapeutic approach to improve conduction and decrease sudden death in HF.

Detection of endogenous acetylcholine release during brief ischemia in the rabbit ventricle: A possible trigger for ischemic preconditioning

Aims To examine endogenous acetylcholine (ACh) release in the rabbit left ventricle during acute ischemia, ischemic preconditioning and electrical vagal stimulation. Main methods We measured myocardial interstitial ACh levels in the rabbit left ventricle using a cardiac microdialysis technique. In Protocol 1 ( n = 6), the left circumflex coronary artery (LCX) was occluded for 30 min and reperfused for 30 min. In Protocol 2 ( n = 5), the LCX was temporarily occluded for 5 min. Ten minutes later, the LCX was occluded for 30 min and reperfused for 30 min. In Protocol 3 ( n = 5), bilateral efferent vagal nerves were stimulated at 20 Hz and 40 Hz (10 V, 1-ms pulse duration). Key findings In Protocol 1, a 30-min coronary occlusion increased the ACh level from 0.39 ± 0.15 to 7.0 ± 2.2 nM (mean ± SE, P < 0.01). In Protocol 2, a 5-min coronary occlusion increased the ACh level from 0.33 ± 0.07 to 0.75 ± 0.11 nM ( P < 0.05). The ACh level returned to 0.48 ± 0.10 nM during the interval. After that, a 30-min coronary occlusion increased the ACh level to 2.4 ± 0.49 nM ( P < 0.01). In Protocol 3, vagal stimulation at 20 Hz and 40 Hz increased the ACh level from 0.29 ± 0.06 to 1.23 ± 0.48 ( P < 0.05) and 2.44 ± 1.13 nM ( P < 0.01), respectively. Significance Acute ischemia significantly increased the ACh levels in the rabbit left ventricle, which appeared to exceed the vagal stimulation-induced ACh release. Brief ischemia as short as 5 min can also increase the ACh level, suggesting that endogenous ACh release can be a trigger for ischemic preconditioning.

Is there a significant transmural gradient in repolarization time in the intact heart?

Is there a significant transmural gradient in repolarization time in the intact heart?

Evaluation of β1L-Adrenoceptors in Rabbit Heart by Tissue Segment Binding Assay

[(3)H]-CGP12177 biphasically bound to beta-adrenoceptors with high and low affinities in the segments and crude membranes of rabbit left ventricle. The low-affinity sites for [(3)H]-CGP12177 in the segments was double in density, compared to the density of high-affinity sites. Total abundance of the beta-adrenoceptors decreased to approximately 10% upon tissue homogenization, and the proportion of low-affinity sites was the same as that of the high-affinity sites in the membranes. The majority of the high-affinity binding sites of [(3)H]-CGP12177 in the segments and the membranes were beta(1H)-adrenoceptor, being highly sensitive to propranolol and beta(1)-antagonists (atenolol and ICI-89,406), whereas the low-affinity binding sites showed a beta(1L)-profile (less sensitive to propranolol and beta(1)-, beta(2)-, and beta(3)-antagonists). Furthermore, a part of the beta(1L)-adrenoceptors was insensitive to atenolol, ICI-89,406, and/or isoproterenol. The present binding study clearly shows that beta(1L)-adrenoceptors occur as a distinct phenotype from beta(1H)-adrenoceptors in rabbit ventricle. However, quantitative imbalance between beta(1H)- and beta(1L)-adrenoceptors and divergent ligand-beta(1L)-adrenoceptor interactions suggest a possibility that the beta(1L)-adrenoceptor may not reflect a simple conformational change or allosteric state in the beta(1)-adrenoceptor molecule.

Arrhythmogenic effects of beta2-adrenergic stimulation in the failing heart are attributable to enhanced sarcoplasmic reticulum Ca load.

Ventricular tachycardia in heart failure (HF) can initiate by nonreentrant mechanisms such as delayed afterdepolarizations. In an arrhythmogenic rabbit model of HF, we have shown that isoproterenol induces ventricular tachycardia in vivo and aftercontractions and transient inward currents in HF myocytes. To determine whether beta(2)-adrenergic receptor (beta(2)-AR) stimulation contributes, we performed in vivo drug infusion, in vitro myocyte and biochemical studies. Intravenous zinterol (2.5 microg/kg) led to ventricular arrhythmias, including ventricular tachycardia up to 13 beats long in 4 of 6 HF rabbits (versus 0 of 5 controls, P<0.01), an effect blocked by beta(2)-AR antagonist ICI-118,551 (0.2 mg/kg). In field-stimulated myocytes (0.5 to 4 Hz, 37 degrees C), beta(2)-AR stimulation (1 micromol/L zinterol+300 nmol/L beta(1)-AR antagonist CGP-29712A) induced aftercontractions and Ca aftertransients in 88% of HF versus 0% of control myocytes (P<0.01). beta(2)-AR stimulation in HF (but not control) myocytes increased Ca transient amplitude (by 29%), sarcoplasmic reticulum (SR) Ca load (by 28%), the rate of [Ca](i) decline (by 28%; n=12, all P<0.05), and phospholamban phosphorylation at Ser16, but Ca current was unchanged. All of these effects in HF myocytes were blocked by ICI-118,551 (100 nmol/L). Although total beta-AR expression was reduced by 47% in HF rabbit left ventricle, beta(2)-AR number was unchanged, indicating more potent beta(2)-AR-dependent SR Ca uptake and arrhythmogenesis in HF. Human HF myocytes showed similar beta(2)-AR-induced aftercontractions, aftertransients, and enhanced Ca transient amplitude, SR Ca load and twitch [Ca](i) decline rate. Thus, beta(2)-AR stimulation is arrhythmogenic in HF, mediated by SR Ca overload-induced spontaneous SR Ca release and aftercontractions.

Effect of α-Allocryptopine on Transient Outward Potassium Current in Rabbit Ventricular Myocytes

Objective: To investigate the mechanism of α-allocryptopine-induced inhibition of the transient outward potassium current (I_to) in rabbit left-ventricular myocytes. Methods: We used the whole-cell patch-clamp technique to record the I_to in the myocytes, which were isolated from the rabbit left ventricle by a Langendorff-perfusion device. Results: Allocryptopine decreased the amplitude and the density of the I_to in a concentration-dependent (range from 1 to 100 µM with an IC₅₀ value of 37 ± 8 µM) and frequency- or use-independent manner. At a test potential of +50 mV, the peak current density of I_to was decreased from 21.56 ± 3.24 to 13.37 ± 2.86 pA/pF by 30 µM allocryptopine. The fast time constant of I_to inactivation was reduced from 9.8 ± 1.8 to 5.7 ± 0.7 ms and the slow time constant of I_to was reduced from 50.8 ± 9.0 to 32.2 ± 12.7 ms by 30 µM allocryptopine. The inactivated curve slope was changed from –19.2 ± 7.1 to –7.5 ± 0.6 mV, while the half-activated voltage and activated curve slope and half-inactivated voltage values were not affected by allocryptopine. Transmural heterogeneity of the I_to was decreased in the presence of allocryptopine. At a test potential of +50 mV, the densities of I_to were reduced by 28.6% (epimyocardium), 50.3% (midmyocardium) and 20.1% (endocardium) after expoure to 30 µM allocryptopine.Transmural dispersion of the I_to was reduced from 11.2 ± 1.2 to 4.7 ± 0.6 pA/pF by 30 µM allocryptopine. Conclusion: Allocryptopine produced a blocking effect on the I_to in cardiac myocytes, which may be an important mechanism in its antiarrhythmic effect.

Comparison of Transcatheter Intraarterial Perfusion MR Imaging and Fluorescent Microsphere Perfusion Measurements during Transcatheter Arterial Embolization of Rabbit Liver Tumors

Transcatheter intraarterial perfusion (TRIP) magnetic resonance (MR) imaging is clinically used in the interventional MR imaging setting to verify distribution of injected embolic or chemoembolic material during liver-directed transcatheter therapies and to monitor reductions in perfusion. The accuracy of this technique remains unknown. In the present study, rabbit VX2 liver tumors were used to test the hypothesis that TRIP MR imaging accurately measures changes in tumor perfusion during transcatheter arterial embolization (TAE), with injection of fluorescent microspheres used as the gold-standard technique. Five New Zealand White rabbits were used for this study (two donor rabbits and three with VX2 liver tumors). In three rabbits with implanted VX2 liver tumors, catheters were superselectively placed under digital subtraction angiographic guidance into the left hepatic artery supplying the targeted tumor. Fluorescent microspheres were injected into each rabbit's left ventricle before and after TAE. TRIP MR images were obtained at baseline and after embolizations for all rabbits with intraarterial injections of 2.5% gadopentetate dimeglumine solution. Linear regression was used to compare relative reductions in tumor perfusion between TRIP MR imaging and fluorescent microspheres. Results were considered statistically significant at a P value less than .05. There was good correlation between TRIP MR imaging and fluorescent microsphere measurements of reduction in tumor perfusion (r = 0.722, P < .012). TRIP MR imaging provides accurate semiquantitative measurement of perfusion reduction during TAE in rabbit liver tumors.

Post‐Shock Synchronized Pacing in Isolated Rabbit Left Ventricle: Evaluation of a Novel Defibrillation Strategy

A failed near-threshold defibrillation shock is followed by an isoelectric window (IEW) and rapid repetitive responses that reinitiate ventricular fibrillation (VF). We hypothesized that properly timed (synchronized) postshock pacing stimuli (SyncP) may capture the recovered tissues during the repetitive responses and prevent postshock reinitiation of VF, resulting in improved defibrillation efficacy. We explored the effect of postshock SyncP on defibrillation efficacy in isolated rabbit hearts (n = 12). Optical recording-guided real-time detection and electrical stimulation (5 mA) of recovered tissues in anterior/posterior left ventricle (LV) were performed following IEW. The IEW duration was found to be 69 +/- 13 ms. With the same shock strength, successful and failed defibrillation episodes were associated with 50% and 15% of the myocardium, respectively, captured by the SyncP (P < 0.001). Electrical stimulation from the posterior LV resulted in 75% of episodes capturing myocardium, as compared with anterior LV stimulation (55%; P < 0.01) and higher successful defibrillation rate (14%, posterior vs. 3%, anterior LV). The overall success in terminating VF by postshock SyncP was approximately 10%. The causes for failed myocardium capture by postshock SyncP included lack of IEW after low-strength shock (42.9%), incorrect locations of reference site (25.7%) and pacing electrodes (17.9%), and others, such as wave breakthroughs (13.5%). Postshock SyncP was feasible and the larger the myocardium captured area, the more likely was the successful defibrillation. Postshock SyncP delivered to the posterior LV was more effective than anterior LV to terminate VF.

P3-12: Negative virtual electrode polarization in the rabbit left ventricle delays activation during diastolic field stimulation

P3-12: Negative virtual electrode polarization in the rabbit left ventricle delays activation during diastolic field stimulation

Connexin 43 Downregulation and Dephosphorylation in Nonischemic Heart Failure Is Associated With Enhanced Colocalized Protein Phosphatase Type 2A

In nonischemic heart failure (HF), ventricular tachycardia initiates by a nonreentrant mechanism, but there is altered conduction (that could lead to re-entry) that could arise from changes in gap junctional proteins, especially connexin43 (Cx43). We studied Cx43 expression and phosphorylation state in the left ventricle (LV) from an arrhythmogenic rabbit model of nonischemic HF and from patients with HF attributable to idiopathic dilated cardiomyopathy. We also investigated the role of protein phosphatases that dephosphorylate Cx43--PP1 and PP2A. In HF rabbit LV, Cx43 mRNA and total protein were decreased by 29% and 34%, respectively (P<0.05 and P<0.001). In controls, Cx43 was primarily in the phosphorylated state, but with HF there was a 64% increase in nonphosphorylated Cx43 (Cx43-NP, normalized to total Cx43; P<0.05). Similar results were noted in HF rabbit myocytes (P<0.05) and in human idiopathic dilated cardiomyopathy LV (P<0.05). We found that PP1 and PP2A colocalized with Cx43 in rabbit LV. With HF, the level of colocalized PP2A increased >2.5-fold (P<0.002), whereas colocalized PP1 was unchanged. We also found intercellular coupling (assessed by Lucifer Yellow dye transfer) was markedly reduced in HF. However, okadaic acid (10 nmol/L) reduced the amount of Cx43-NP and significantly improved cell coupling in HF. Thus, in nonischemic HF in rabbits and humans, there is a decrease in both Cx43 expression and phosphorylation that contributes to uncoupling. Increased levels of PP2A that colocalize with Cx43 can underlie enhanced levels of Cx43-NP in HF. Modulation of Cx43 phosphorylation may be a potential therapeutic target to improve conduction in HF.

Preclinical strategies to assess QT liability and torsadogenic potential of new drugs: The role of experimental models

The recognition of QT prolongation and torsade de pointes (TdP) in humans has resulted in the re-labeling of some drugs and the removal of others from the market in the past decade. Recent regulatory guidelines have recommended a battery of preclinical tests to assess a new drug for the QT liability in humans. The assessment includes the effect of a drug on: 1) the ionic current in stable cell lines expressing hERG channel; 2) action potential duration (APD) measured in isolated ventricular tissues; 3) the QTc interval and TdP in animals in vivo; and 4) APD, the QT interval, transmural dispersion of repolarization (TDR) and TdP potential in the isolated arterially-perfused ventricular wedge preparation. Because a noncardiac drug with an incidence of TdP even less than 0.1% can be potentially removed from the market, the experimental models used for preclinical testing have to be high sensitive and specific to the signals related to TdP. Among available experimental models, the rabbit left ventricle wedge preparation exhibits a high sensitivity and a high specificity in the identification of compounds positive and negative for QT prolongation and TdP. This is attributed to the fact that the preparation demonstrates strong signals related to QT prolongation in response to even a weaker QT prolonging agent. Signals specifically pertinent to the development of TdP, ie, early afterdepolarization (EAD) and an increase in TDR can be detected as well. The preclinical data obtained from the wedge preparation correlate well with clinical outcomes.

Ca2+-activated Cl- current reduces transmural electrical heterogeneity within the rabbit left ventricle.

Various cationic membrane channels contribute to the heterogeneity of action potential configuration between the transmural layers of the left ventricle. The role of anionic membrane channels is less intensively studied. We investigated the role of the Ca2+-activated Cl- current, ICl(Ca), in transmural electrical heterogeneity. We determined the density of ICl(Ca) and its physiological role in subepicardial and subendocardial ventricular myocytes of rabbit using the patch-clamp technique. ICl(Ca) was measured as the 4,4'diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) sensitive current. The current-voltage relationships and the densities of ICl(Ca) were similar in subepicardial and subendocardial myocytes. However, the functional role of ICl(Ca) exhibited striking differences. In subendocardial myocytes, blockade of ICl(Ca) by DIDS increased action potential duration (APD) significantly at all measured stimulus frequencies (3.33-0.2 Hz). In subepicardial myocytes, ICl(Ca) blockade increased APD only at 3.33 Hz, but not at the lower stimulus frequencies. At 1 Hz, ICl(Ca) blockade in subepicardial myocytes only caused an APD increase when the transient outward K+ current, Ito1, was blocked. The densities and gating properties of ICl(Ca) are similar in subepicardial and subendocardial myocytes. ICl(Ca) contributes to APD shortening in subendocardial, but not in subepicardial myocytes except at 3.33 Hz. These differences in functional expression of ICl(Ca) reduce the electrical heterogeneity in rabbit left ventricle.