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PubMed2005

Cellular mechanism of calcium-mediated triggered activity in the heart.

Calcium overload due to enhanced calcium entry is a mechanism for spontaneous calcium release (SCR) from the sarcoplasmic reticulum, delayed-afterdepolarizations (DAD), and triggered activity. However, the exact mechanistic relationship between elevated intracellular calcium levels and triggered activity originating from a specific location remains unclear. We hypothesize that under conditions of enhanced calcium entry, elevation of intracellular calcium will result in multiple calcium release events of which only one is more likely to initiate a triggered beat. We used optical mapping of action potentials and ratiometric calcium transients in an electromechanically-uncoupled canine wedge model of enhanced calcium entry, using I(Ks) blockade with beta-adrenergic stimulation. Under conditions of enhanced calcium entry, the rate of calcium uptake was faster compared with control conditions; however, during rapid pacing, cytoplasmic calcium elevation at the endocardium was significantly increased (15+/-4%) compared with control (10+/-3, P<0.04). Rapid pacing induced multiple simultaneous SCR events with largest amplitude and earliest onset near the endocardium compared with the epicardium. Furthermore, SCR events with largest amplitude and earliest onset served as a focus for DAD-mediated triggered activity. Interestingly, polymorphic VT occurred in some experiments when multiple SCR events occurred. In conclusion, multiple, simultaneous SCR events occur over a broad region of relatively slower calcium uptake and elevated diastolic calcium levels. However, SCR events closer to the endocardium have the largest amplitude and earliest onset and are, thereby, more likely to initiate DAD-mediated triggered activity. Finally, multiple SCR events may be a mechanism of polymorphic VT under calcium overload conditions.

Action Potentials↗
PubMed2005

Mechanisms of atrial fibrillation termination by pure sodium channel blockade in an ionically-realistic mathematical model.

The mechanisms by which Na+-channel blocking antiarrhythmic drugs terminate atrial fibrillation (AF) remain unclear. Classical "leading-circle" theory suggests that Na+-channel blockade should, if anything, promote re-entry. We used an ionically-based mathematical model of vagotonic AF to evaluate the effects of applying pure Na+-current (I(Na)) inhibition during sustained arrhythmia. Under control conditions, AF was maintained by 1 or 2 dominant spiral waves, with fibrillatory propagation at critical levels of action potential duration (APD) dispersion. I(Na) inhibition terminated AF increasingly with increasing block, terminating all AF at 65% block. During 1:1 conduction, I(Na) inhibition reduced APD (by 13% at 4 Hz and 60% block), conduction velocity (by 37%), and re-entry wavelength (by 24%). During AF, I(Na) inhibition increased the size of primary rotors and reduced re-entry rate (eg, dominant frequency decreased by 33% at 60% I(Na) inhibition) while decreasing generation of secondary wavelets by wavebreak. Three mechanisms contributed to I(Na) block-induced AF termination in the model: (1) enlargement of the center of rotation beyond the capacity of the computational substrate; (2) decreased anchoring to functional obstacles, increasing meander and extinction at boundaries; and (3) reduction in the number of secondary wavelets that could provide new primary rotors. Optical mapping in isolated sheep hearts confirmed that tetrodotoxin dose-dependently terminates AF while producing effects qualitatively like those of I(Na) inhibition in the mathematical model. We conclude that pure INa inhibition terminates AF, producing activation changes consistent with previous clinical and experimental observations. These results provide insights into previously enigmatic mechanisms of class I antiarrhythmic drug-induced AF termination. The full text of this article is available online at http://circres.ahajournals.org

Algorithms↗
PubMed2005

Cardiac-specific loss of N-cadherin leads to alteration in connexins with conduction slowing and arrhythmogenesis.

The remodeling of ventricular gap junctions, as defined by changes in size, distribution, or function, is a prominent feature of diseased myocardium. However, the regulation of assembly and maintenance of gap junctions remains poorly understood. To investigate N-cadherin function in the adult myocardium, we used a floxed N-cadherin gene in conjunction with a cardiac-specific tamoxifen-inducible Cre transgene. The mutant animals appeared active and healthy until their sudden death approximately 2 months after deleting N-cadherin from the heart. Electrophysiologic analysis revealed abnormal conduction in the ventricles of mutant animals, including diminished QRS complex amplitude consistent with loss of electrical coupling in the myocardium. A significant decrease in the gap junction proteins, connexin-43 and connexin-40, was observed in N-cadherin-depleted myocytes. Perturbation of connexin function resulted in decreased ventricular conduction velocity, as determined by optical mapping. Our data suggest that perturbation of the N-cadherin/catenin complex in heart disease may be an underlying cause, leading to the establishment of the arrythmogenic substrate by destabilizing gap junctions at the cell surface.

Animals↗
PubMed2006

Structure-related initiation of reentry by rapid pacing in monolayers of cardiac cells.

This study examines how a zigzag pattern of conduction, a form of structural heterogeneity frequently found in old or diseased hearts, affects the vulnerability to reentry during rapid pacing. A central rectangular island (8x4 mm) containing a predefined zigzag pattern was created in cultured isotropic monolayers of neonatal rat ventricular myocytes. Impulse propagation was optically mapped from 253 sites using voltage-sensitive dye and was anisotropic within the zigzag island. With increasing interval between neighboring transverse connections (a), relative to the distance between longitudinal strands (b), transverse conduction velocity (CV) decreased to 66+/-6%, 20+/-2%, and 15+/-2% of CV in the surrounding isotropic region, whereas longitudinal CV increased to 102+/-8%, 113+/-12%, and 131+/-23% for a:b ratios of 1:1, 1:5, and 1:9, respectively. During rapid pacing, propagation distal to the island was steered from the side of the island with more transverse connections ("dominant" side) toward the side with fewer connections ("weak" side). Increased asymmetry in the pattern accentuated this effect, and resulted in increased rate-dependent differences in CV on the 2 sides. Consequently, a functional obstacle formed on the weak side, followed by development of single loop reentry. The reentrant wave revolved around a line of block defined by the border of the island. Reentry chirality was determined by the weak side location, and the pacing rate needed to initiate reentry decreased with increased asymmetry in the pattern. In conclusion, reentry is readily induced by rapid pacing in confluent cardiac cell monolayers containing a central and asymmetric island of zigzag conduction.

Action Potentials↗
PubMed2006

Pacing-induced heterogeneities in intracellular Ca2+ signaling, cardiac alternans, and ventricular arrhythmias in intact rat heart.

Optical mapping studies have suggested that intracellular Ca2+ and T-wave alternans are linked through underlying alternations in Ca2+ cycling-inducing oscillations in action potential duration through Ca2+-sensitive conductances. However, these studies cannot measure single-cell behavior; therefore, the Ca2+ cycling heterogeneities within microscopic ventricular regions are unknown. The goal of this study was to measure cellular activity in intact myocardium during rapid pacing and arrhythmias. We used single-photon laser-scanning confocal microscopy to measure Ca2+ signaling in individual myocytes of intact rat myocardium during rapid pacing and during pacing-induced ventricular arrhythmias. At low rates, all myocytes demonstrate Ca2+ alternans that is synchronized but whose magnitude varies depending on recovery kinetics of Ca2+ cycling for each individual myocyte. As rate increases, some cells reverse alternans phase, giving a dyssynchronous activation pattern, even in adjoining myocytes. Increased pacing rate also induces subcellular alternans where Ca2+ alternates out of phase with different regions within the same cell. These forms of heterogeneous Ca2+ signaling also occurred during pacing-induced ventricular tachycardia. Our results demonstrate highly nonuniform Ca2+ signaling among and within individual myocytes in intact heart during rapid pacing and arrhythmias. Thus, certain pathophysiological conditions that alter Ca2+ cycling kinetics, such as heart failure, might promote ventricular arrhythmias by exaggerating these cellular heterogeneities in Ca2+ signaling.

Animals↗
PubMed2006

Relative contributions of connexins 40 and 43 to atrial impulse propagation in synthetic strands of neonatal and fetal murine cardiomyocytes.

Atrial tissue expresses both connexin 40 (Cx40) and 43 (Cx43) proteins. To assess the relative roles of Cx40 and Cx43 in atrial electrical propagation, we synthesized cultured strands of atrial myocytes derived from mice with genetic deficiency in Cx40 or Cx43 expression and measured propagation velocity (PV) by high-resolution optical mapping of voltage-sensitive dye fluorescence. The amount of Cx40 and/or Cx43 in gap junctions was measured by immunohistochemistry and total or sarcolemmal Cx43 or Cx40 protein by immunoblotting. Progressive genetic reduction in Cx43 expression decreased PV from 34+/-6 cm/sec in Cx43(+/+) to 30+/-8 cm/sec in Cx43(+/-) and 19+/-11 cm/sec in Cx43(-/-) cultures. Concomitantly, the cell area occupied by Cx40 immunosignal in gap junctions decreased from 2.0+/-1.6% in Cx43(+/+) to 1.7+/-0.5% in Cx43(+/-) and 1.0+/-0.2% in Cx43(-/-) strands. In contrast, progressive genetic reduction in Cx40 expression increased PV from 30+/-2 cm/sec in Cx40(+/+) to 40+/-7 cm/sec in Cx40(+/-) and 45+/-10 cm/sec in Cx40(-/-) cultures. Concomitantly, the cell area occupied by Cx43 immunosignal in gap junctions increased from 1.2+/-0.9% in Cx40(+/+) to 2.8+/-1.4% in Cx40(+/-) and 3.1+/-0.6% in Cx40(-/-) cultures. In accordance with the immunostaining results, immunoblots of the Triton X-100-insoluble fraction revealed an increase of Cx43 in gap junctions in extracts from Cx40-ablated atria, whereas total cellular Cx43 remained unchanged. Our results suggest that the relative abundance of Cx43 and Cx40 is an important determinant of atrial impulse propagation in neonatal hearts, whereby dominance of Cx40 decreases and dominance of Cx43 increases local propagation velocity.

Animals↗
PubMed2006

Integrin activation in the heart: a link between electrical and contractile dysfunction?

Integrins mechanically link the cytoskeleton to the extracellular matrix in cardiac myocytes and are thereby involved in mechanotransduction. Integrins appear to be necessary for cardiac myocyte hypertrophy. To determine the effect of increased integrin ligation and signaling on adult cardiac function, a heart-specific truncated alpha(5) integrin (gain of function) was conditionally expressed in mice. Four days later, we observed an 80% reduction in amplitude of the QRS complex, profound systolic dysfunction, decreased connexin43, loss of gap junctions, and abnormal intercalated discs. Surprisingly, isolated left ventricular myocytes contracted normally and exhibited normal Ca(2+) transients. This suggested that cell/cell electrical and/or mechanical coupling was disrupted. To distinguish electrical from mechanical coupling deficits, we compared the papillary muscle force generated by electrically stimulated versus rapid cooling contractions in which intracellular Ca(2+) is released without electrical depolarization. Both were decreased in the transgenic muscle. However, electrically stimulated contractions were more significantly reduced than rapid cooling contractures. This suggests a component of cell/cell electrical uncoupling. Optical mapping revealed a loss of the normal elliptical isochronal activation pattern implying a loss of preferential conduction through gap junctions. For the first time, we have shown that integrins can regulate both mechanical and electrical coupling in the adult heart, even in the absence of primary hemodynamic alterations. Furthermore, we demonstrated that unregulated integrin activation leads to both contractile dysfunction and arrhythmias.

Animals↗
PubMed1993

Spiral waves of excitation underlie reentrant activity in isolated cardiac muscle.

The mechanism of reentrant ventricular tachycardia was studied in computer simulations and in thin (approximately 20 x 20 x 0.5-mm) slices of dog and sheep ventricular epicardial muscle. A two-dimensional matrix consisting of 96 x 96 electrically coupled cells modeled by the FitzHugh-Nagumo equations was used to analyze the dynamics of self-sustaining reentrant activity in the form of elliptical spiral waves induced by premature stimulation. In homogeneous anisotropic media, spirals are stationary and may last indefinitely. However, the presence of small parameter gradients may lead to drifting and eventual termination of the spiral at the boundary of the medium. On the other hand, spirals may anchor and rotate around small discontinuities within the matrix. Similar results were obtained experimentally in 10 preparations whose electrical activity was monitored by means of a potentiometric dye and high-resolution optical mapping techniques; premature stimulation triggered reproducible episodes of sustained or nonsustained reentrant tachycardia in the form of spiral waves. As a rule, the spirals were elongated, with the major hemiaxis parallel to the longitudinal axis of the cells. The period of rotation (183 +/- 68 msec [mean +/- SD]) was longer than the refractory period (131 +/- 38 msec) and appeared to be determined by the size of the spiral's core, which was measured using a newly devised "frame-stack" plot. Drifting of spiral waves was also observed experimentally. Drift velocity was 9.8% of the velocity of wave propagation. In some cases, the core became stationary by anchoring to small arteries or other heterogeneities, and the spiral rotated rhythmically for prolonged periods of time. Yet, when drift occurred, spatiotemporal variations in the excitation period were manifested as a result of a Doppler effect, with the excitation period ahead of the core being 20 +/- 6% shorter than the excitation period behind the core. As a result of these coexisting frequencies, a pseudoelectrocardiogram of the activity in the presence of a drifting spiral wave exhibited "QRS complexes" with an undulating axis, which resembled those observed in patients with torsade de pointes. The overall results show that spiral wave activity is a property of cardiac muscle and suggest that such activity may be the common mechanism of a number of monomorphic and polymorphic tachycardias.

Animals↗
PubMed1996

Spatial changes in transmembrane potential during extracellular electrical shocks in cultured monolayers of neonatal rat ventricular myocytes.

This study investigated the role of different types of discontinuities in tissue architecture on the spatial distribution of the transmembrane potential. Specifically, we tested the occurrence of so-called "secondary sources," ie, localized hyperpolarizations and depolarizations during the application of extracellular electrical shocks (EESs). Changes in transmembrane potential relative to action potential amplitude (delta Vm/APA) were measured in patterned cultures of neonatal rat myocytes, stained with voltage-sensitive dye (RH-237), by optical mapping (96-channel photodiode array, 6- to 30-micron resolution) during the application of EES (field strength, 8 to 22 V/cm; duration, 6 ms). Across narrow cell strands (width, 218 +/- 59 [mean +/- SD] microns), EES applied during the relative refractory period produced a linear and symmetrical profile of delta Vm/APA (-65 +/- 23% maximal hyperpolarization versus +64 +/- 15% maximal depolarization). In contrast, the profile of delta Vm/APA was asymmetrical when EESs were applied during the action potential plateau (-95 +/- 32% versus +37 +/- 14%). At high magnification, no secondary sources were observed at the borders between cells. In dense isotropic cell monolayers or in monolayers and strands showing intercellular clefts, secondary sources were frequently observed. Intercellular clefts of the size of one to several myocytes were sufficient to produce secondary sources of the same magnitude as those that elicited action potentials in dense cell strands. There was a close correlation between the location of secondary sources during EES and localized conduction slowing during propagation. Thus, densely packed cultured cell strands behave as an electrical continuum with no secondary sources occurring at cell borders. Small intercellular clefts can create secondary sources of sufficient magnitude to exert a stimulatory effect.

Animals↗
PubMed1999

Virtual electrode-induced reexcitation: A mechanism of defibrillation.

Mechanisms of defibrillation remain poorly understood. Defibrillation success depends on the elimination of fibrillation without shock-induced arrhythmogenesis. We optically mapped selected epicardial regions of rabbit hearts (n=20) during shocks applied with the use of implantable defibrillator electrodes during the refractory period. Monophasic shocks resulted in virtual electrode polarization (VEP). Positive values of VEP resulted in a prolongation of the action potential duration, whereas negative polarization shortened the action potential duration, resulting in partial or complete recovery of the excitability. After a shock, new propagated wavefronts emerged at the boundary between the 2 regions and reexcited negatively polarized regions. Conduction velocity and maximum action potential upstroke rate of rise dV/dt (max) of shock-induced activation depended on the transmembrane potential at the end of the shock. Linear regression analysis showed that dV/dt(max) of postshock activation reached 50% of that of normal action potential at a V(m) value of -56.7+/-0.6 mV postshock voltage (n=9257). Less negative potentials resulted in slow conduction and blocks, whereas more negative potentials resulted in faster conduction. Although wavebreaks were produced in either condition, they degenerated into arrhythmias only when conduction was slow. Shock-induced VEP is essential in extinguishing fibrillation but can reinduce arrhythmias by producing excitable gaps. Reexcitation of these gaps through progressive increase in shock strength may provide the basis for the lower and upper limits of vulnerability. The former may correspond to the origination of slow wavefronts of reexcitation and phase singularities. The latter corresponds to fast conduction during which wavebreaks no longer produce sustained arrhythmias.

Animals↗
PubMed2000

Nonlinear changes of transmembrane potential during defibrillation shocks: role of Ca(2+) current.

Defibrillation shocks induce complex nonlinear changes of transmembrane potential (DeltaV(m)). To elucidate the ionic mechanisms of nonlinear DeltaV(m), we studied the effects of ionic channel blockers on DeltaV(m) in geometrically defined myocyte cultures. Experiments were carried out in cell strands with widths of 0.2 mm (narrow strands) and 0.8 mm (wide strands) produced using a technique of directed cell growth. Uniform-field shocks were applied across strands during the action potential (AP) plateau, and the distribution of shock-induced DeltaV(m) was measured using an optical mapping technique. Nifedipine and 4-aminopyridine were applied to inhibit the L-type calcium current (I:(Ca)) and the transient outward current (I:(to)), respectively. In control conditions, the distribution of DeltaV(m) across cell strands was highly asymmetrical with a large ratio of negative to positive DeltaV(m) (DeltaV(-)(m)/DeltaV(+)(m)) measured at the opposite strand borders. Application of nifedipine caused a large increase of DeltaV(+)(m) and a decrease of DeltaV(-)(m)/DeltaV(+)(m), indicating involvement of I:(Ca) in the asymmetrical DeltaV(m), likely as a result of the outward flow of I:(Ca) when V(m) exceeded the I:(Ca) reversal potential. DeltaV(-)(m) decreased in the narrow strands but remained unchanged in the wide strands, indicating that the changes of DeltaV(-)(m) were caused by electrotonic interaction with an area of depolarization. 4-Aminopyridine did not change DeltaV(-)(m)/DeltaV(+)(m). These results provide evidence that (1) the asymmetry of shock-induced DeltaV(m) during the AP plateau is due to outward flow of I:(Ca) in the depolarized portions of the strands, (2) I:(to) is not involved in the mechanism of DeltaV(m) asymmetry, and (3) the effects of drugs on DeltaV(m) are modulated by the tissue geometry.

4-Aminopyridine↗
PubMed2000

The role of electroporation in defibrillation.

Electric shock is the only effective therapy against ventricular fibrillation. However, shocks are also known to cause electroporation of cell membranes. We sought to determine the impact of electroporation on ventricular conduction and defibrillation. We optically mapped electrical activity in coronary-perfused rabbit hearts during electric shocks (50 to 500 V). Electroporation was evident from transient depolarization, reduction of action potential amplitude, and upstroke dV/dt. Electroporation was voltage dependent and significantly more pronounced at the endocardium versus the epicardium, with thresholds of 229+/-81 versus 318+/-84 V, respectively (P=0.01, n=10), both being above the defibrillation threshold of 181.3+/-45.8 V. Epicardial electroporation was localized to a small area near the electrode, whereas endocardial electroporation was observed at the bundles and trabeculas throughout the entire endocardium. Higher-resolution imaging revealed that papillary muscles (n=10) were most affected. Electroporation and conduction block thresholds in papillary muscles were 281+/-64 V and 380+/-79 V, respectively. We observed no arrhythmia in association with electroporation. Further, preconditioning with high-energy shocks prevented reinduction of fibrillation by 50-V shocks, which were otherwise proarrhythmic. Endocardial bundles are the most susceptible to electroporation and the resulting conduction impairment. Electroporation is not associated with proarrhythmic effects and is associated with a reduction of vulnerability.

Action Potentials↗
PubMed2001

Conduction slowing and sudden arrhythmic death in mice with cardiac-restricted inactivation of connexin43.

Cardiac arrhythmia is a common and often lethal manifestation of many forms of heart disease. Gap junction remodeling has been postulated to contribute to the increased propensity for arrhythmogenesis in diseased myocardium, although a causative role in vivo remains speculative. By generating mice with cardiac-restricted knockout of connexin43 (Cx43), we have circumvented the perinatal lethal developmental defect associated with germline inactivation of this gap junction channel gene and uncovered an essential role for Cx43 in the maintenance of electrical stability. Mice with cardiac-specific loss of Cx43 have normal heart structure and contractile function, and yet they uniformly (28 of 28 conditional Cx43 knockout mice observed) develop sudden cardiac death from spontaneous ventricular arrhythmias by 2 months of age. Optical mapping of the epicardial electrical activation pattern in Cx43 conditional knockout mice revealed that ventricular conduction velocity was significantly slowed by up to 55% in the transverse direction and 42% in the longitudinal direction, resulting in an increase in anisotropic ratio compared with control littermates (2.1+/-0.13 versus 1.66+/-0.06; P:<0.01). This novel genetic murine model of primary sudden cardiac death defines gap junctional abnormalities as a key molecular feature of the arrhythmogenic substrate.

Animals↗
PubMed2001

Fluorescent imaging of a dual-pathway atrioventricular-nodal conduction system.

A dual-pathway theory to explain atrioventricular-nodal (AVN) reentry has been proposed previously. However, the exact anatomical and functional correlates of the fast pathway (FP) and slow pathway (SP) have not yet been elucidated. We used optical mapping to reconstruct patterns of activation during retrograde conduction through the AVN and during AVN reentry in the triangles of Koch of 12 rabbits. Reentry was inducible by a premature stimulation of the bundle of His in 6 preparations (50%). A functional FP and SP appear to be anatomically correlated with posterior and posterolateral extensions of the AVN, which were recently described. Retrograde breakthrough points in 6 noninducible preparations were clustered near the apex of the triangle of Koch (FP), whereas 6 inducible preparations had either cycle length-dependent FP and SP exits (n=3) or only SP exits located near the coronary sinus orifice. The shift of breakthrough points from FP to SP during progressive shortening of the coupling interval was accompanied by a discontinuity in the conduction curve. We observed a transmural reentrant circuit involving the AVN, FP, SP, and the superficial endocardial layer of atrial and transitional cells. The presence of a functional SP during retrograde conduction was associated with inducibility of AVN reentry. The full text of this article is available at http://www.circresaha.org.

Animals↗
PubMed2005

Altered right atrial excitation and propagation in connexin40 knockout mice.

BACKGROUND: Intercellular coupling via connexin40 (Cx40) gap junction channels is an important determinant of impulse propagation in the atria. METHODS AND RESULTS: We studied the role of Cx40 in intra-atrial excitation and propagation in wild-type (Cx40(+/+)) and knockout (Cx40(-/-)) mice using high-resolution, dual-wavelength optical mapping. On ECG, the P wave was significantly prolonged in Cx40(-/-) mice (13.4+/-0.5 versus 11.4+/-0.3 ms in Cx40(+/+)). In Cx40(+/+) hearts, spontaneous right atrial (RA) activation showed a focal breakthrough at the junction of the right superior vena cava, sulcus terminalis, and RA free wall, corresponding to the location of the sinoatrial node. In contrast, Cx40(-/-) hearts displayed ectopic breakthrough sites at the base of the sulcus terminalis, RA free wall, and right superior vena cava. Progressive ablation of such sites in 4 Cx40(-/-) mice resulted in ectopic focus migration and cycle length prolongation. In all Cx40(-/-) hearts the focus ultimately shifted to the sinoatrial node at a very prolonged cycle length (initial ectopic cycle length, 182+/-20 ms; postablation sinus cycle length, 387+/-44 ms). In a second group of experiments, epicardial pacing at 10 Hz revealed slower conduction in the RA free wall of 5 Cx40(-/-) hearts than in 5 Cx40(+/+) hearts (0.61+/-0.07 versus 0.94+/-0.07 m/s; P<0.05). Dominant frequency analysis in Cx40(-/-) RA demonstrated significant reduction in the area of 1:1 conduction at 16 Hz (40+/-10% versus 69+/-5% in Cx40(+/+)) and 25 Hz (36+/-11% versus 65+/-9% in Cx40(+/+)). CONCLUSIONS: This is the first demonstration of intra-atrial block, ectopic rhythms, and altered atrial propagation in the RA of Cx40(-/-) mice.

Algorithms↗
PubMed2006

Proarrhythmic potential of mesenchymal stem cell transplantation revealed in an in vitro coculture model.

BACKGROUND: Mesenchymal stem cells (MSCs) are bone marrow stromal cells that are in phase 1 clinical studies of cellular cardiomyoplasty. However, the electrophysiological effects of MSC transplantation have not been studied. Although improvement of ventricular function would represent a positive outcome of MSC transplantation, focal application of stem cells has the potential downside of creating inhomogeneities that may predispose the heart to reentrant arrhythmias. In the present study we use an MSC and neonatal rat ventricular myocyte (NRVM) coculture system to investigate potential proarrhythmic consequences of MSC transplantation into the heart. METHODS AND RESULTS: Human MSCs were cocultured with NRVMs in ratios of 1:99, 1:9, and 1:4 and optically mapped. We found that conduction velocity was decreased in cocultures compared with controls, but action potential duration (APD80) was not affected. Reentrant arrhythmias were induced in 86% of cocultures containing 10% and 20% MSCs (n=36) but not in controls (n=7) or cocultures containing only 1% MSCs (n=4). Immunostaining, Western blot, and dye transfer revealed the presence of functional gap junctions involving MSCs. CONCLUSIONS: Our results suggest that mixtures of MSCs and NRVMs can produce an arrhythmogenic substrate. The mechanism of reentry is probably increased tissue heterogeneity resulting from electric coupling of inexcitable MSCs with myocytes.

Animals↗
PubMed2006

Matrix metalloproteinase-7 affects connexin-43 levels, electrical conduction, and survival after myocardial infarction.

BACKGROUND: Matrix metalloproteinases (MMPs) contribute to left ventricular remodeling after myocardial infarction (MI). Specific causative roles of particular MMPs, however, remain unclear. MMP-7 is abundant in cardiomyocytes and macrophages, but MMP-7 function after MI has not been defined. METHODS AND RESULTS: Wild-type (WT; n=55) and MMP-7-null (MMP-7-/-; n=32) mice underwent permanent coronary artery ligation for 7 days. MI sizes were similar, but survival was greatly improved in MMP-7-/- mice. The survival difference could not be attributed to differences in left ventricular dilation because end-diastolic volumes increased similarly. ECG analysis revealed a prolonged PR interval in WT but not in MMP-7-/- post-MI mice. Post-MI conduction velocity, determined by optically mapping electrical wavefront propagation, decreased to 78+/-6% of control for WT and was normalized in MMP-7-/- mice. In WT mice, slower conduction velocity correlated with a 53% reduction in the gap junction protein connexin-43. Direct binding of MMP-7 to connexin-43, determined by surface plasmon resonance technology, occurred in a dose-dependent manner. Connexin-43 processing by MMP-7 was confirmed by in silico and in vitro substrate analyses and MMP-7 infusion induced arrhythmias in vivo. CONCLUSIONS: MMP-7 deletion results in improved survival and myocardial conduction patterns after MI. This is the first report to implicate MMP-7 in post-MI remodeling and to demonstrate that connexin-43 is a novel MMP-7 substrate.

Amino Acid Sequence↗
PubMed2001

Heterogeneous expression of Gap junction channels in the heart leads to conduction defects and ventricular dysfunction.

BACKGROUND: - Heterogeneous remodeling of gap junctions is observed in many forms of heart disease. The consequent loss of synchronous ventricular activation has been hypothesized to result in diminished cardiac performance. To directly test this hypothesis, we designed a murine model of heterogeneous gap junction channel expression. Methods and Results-- We generated chimeric mice formed from connexin43 (Cx43)-deficient embryonic stem cells and wild-type or genetically marked ROSA26 recipient blastocysts. Chimeric mice developed normally, without histological evidence of myocardial fibrosis or hypertrophy. Heterogeneous Cx43 expression resulted in conduction defects, however, as well as markedly depressed contractile function. Optical mapping of chimeric hearts by use of voltage-sensitive dyes revealed highly irregular epicardial conduction patterns, quantified as significantly greater negative curvature of the activation wave front (-1.86+/-0.40 mm in chimeric mice versus -0.86+/-0.098 mm in controls; P<0.01; n=6 for each group). Echocardiographic studies demonstrated significantly reduced fractional shortening in chimeric mice (26.6+/-2.3% versus 36.5+/-1.6% in age-matched 129/SvxC57BL/6F1 wild-type controls; P<0.05). CONCLUSIONS: - These data suggest that heterogeneous Cx43 expression, by perturbing the normal pattern of coordinated myocardial excitation, may directly depress cardiac performance.

Animals↗