Arrhythmogenesis in heart cells involves reverse E–C coupling and reverse electrotonic conduction along T-tubules

Early afterdepolarization (EAD) is an aberrant cardiac afterpotential that underlies the development of life-threatening ventricular arrhythmias. It is believed that the development of EAD is caused by the reactivation of L-type Ca2+ current during the period of the action potential plateau; however, the cellular mechanisms that underlie the development of EAD is still controversial. One favorable alternative is the depolarizing reverse-mode operation of the Na+/Ca2+ exchanger, which is activated by aberrant Ca2+ release from the sarcoplasmic reticulum in the process of reverse E–C coupling. Since EADs develop preferentially in damaged heart cells with abnormal Ca2+-signaling, here I studied the causal link between the development of EADs and aberrant intracellular Ca2+ level ([Ca2+]i) dynamics in mouse heart cells using the whole-cell clamp technique. My results show (1) the generation of EADs was preceded by the development of depolarizing membrane potential (Vm) fluctuation, (2) the depolarizing Vm fluctuation is associated with [Ca2+]i elevation, suggesting an involvement of reverse E–C coupling via the Na+/Ca2+ exchanger, and (3) that extending the T-tubules’ length constant by decreasing the extracellular K+ level facilitated the development of the Vm fluctuation and EADs. Taken together, I conclude that EADs are caused by the depolarizing Vm fluctuation, which is induced locally in the T-tubule membrane by aberrant [Ca2+]i elevation and is conducted back electrotonically along the T-tubules.

Late I Na Blocker GS967 Supresses Polymorphic Ventricular Tachycardia in a Transgenic Rabbit Model of Long QT Type 2

Background: Long QT syndrome has been associated with sudden cardiac death likely caused by early afterdepolarizations (EADs) and polymorphic ventricular tachycardias (PVTs). Suppressing the late sodium current (I NaL ) may counterbalance the reduced repolarization reserve in long QT syndrome and prevent EADs and PVTs. Methods: We tested the effects of the selective I NaL blocker GS967 on PVT induction in a transgenic rabbit model of long QT syndrome type 2 using intact heart optical mapping, cellular electrophysiology and confocal Ca 2+ imaging, and computer modeling. Results: GS967 reduced ventricular fibrillation induction under a rapid pacing protocol (n=7/14 hearts in control versus 1/14 hearts at 100 nmol/L) without altering action potential duration or restitution and dispersion. GS967 suppressed PVT incidences by reducing Ca 2+ -mediated EADs and focal activity during isoproterenol perfusion (at 30 nmol/L, n=7/12 and 100 nmol/L n=8/12 hearts without EADs and PVTs). Confocal Ca 2+ imaging of long QT syndrome type 2 myocytes revealed that GS967 shortened Ca 2+ transient duration via accelerating Na + /Ca 2+ exchanger (I NCX )-mediated Ca 2+ efflux from cytosol, thereby reducing EADs. Computer modeling revealed that I NaL potentiates EADs in the long QT syndrome type 2 setting through (1) providing additional depolarizing currents during action potential plateau phase, (2) increasing intracellular Na + (Na i ) that decreases the depolarizing I NCX thereby suppressing the action potential plateau and delaying the activation of slowly activating delayed rectifier K + channels (I Ks ), suggesting important roles of I NaL in regulating Na i . Conclusions: Selective I NaL blockade by GS967 prevents EADs and abolishes PVT in long QT syndrome type 2 rabbits by counterbalancing the reduced repolarization reserve and normalizing Na i . Graphic Abstract: A graphic abstract is available for this article.

Dual effects of mesenteric lymph isolated from rats with burn injury on contractile function in rat ventricular myocytes

Gut-derived factors in intestinal lymph have been shown to trigger myocardial contractile dysfunction. However, the underlying cellular mechanisms remain unclear. We examined the effects of physiologically relevant concentrations of mesenteric lymph collected from rats with 40% burn injury (burn lymph) on excitation-contraction coupling in rat ventricular myocytes. Burn lymph (0.1–5%), but not control mesenteric lymph from sham-burn animals, induced dual positive and negative inotropic effects depending on the concentrations used. At lower concentrations (<0.5%), burn lymph increased the amplitude of myocyte contraction (1.6 ± 0.3-fold; n = 12). At higher concentrations (>0.5%), burn lymph initially enhanced myocyte contraction, which was followed by a block of contraction. These effects were partially reversible on washout. The initial positive inotropic effect was associated with a prolongation of action potential duration (measured at 90% repolarization, 2.5 ± 0.6-fold; n = 10), leading to significant increases in the net Ca2+ influx (1.7 ± 0.1-fold; n = 8). There were no significant changes in the resting membrane potential. The negative inotropic effect was accompanied by a decrease in the action potential plateau (overshoot decrease by 69 ± 10%; n = 4) and membrane depolarization. Voltage-clamp experiments revealed that the positive inotropic effects of burn lymph were due to an inhibition of the transient outward K+ currents that prolong action potential duration, and the inhibitory effects were due to a concentration-dependent inhibition of Ca2+ currents that lead to a reduction of action potential plateau. These burn lymph-induced changes in cardiac myocyte Ca2+ handling can contribute to burn-induced contractile dysfunction and ultimately to heart failure.

Use of translucent indium tin oxide to measure stimulatory effects of a passive conductor during field stimulation of rabbit hearts

Biomathematical models and experiments have indicated that passive extracellular conductors influence field stimulation. Because metallic conductors prevent optical mapping under the conductor, we have evaluated a passive translucent indium tin oxide (ITO) thin-film conductor to allow mapping of transmembrane potential ( Vm) and stimulatory current under the conductor. A 1-cm ITO disk was patterned photolithographically and positioned between 0.3-cm2 mesh shock electrodes on the ventricular epicardium of isolated perfused rabbit hearts stained with 4-{2-[6-(dibutylamino)-2-naphthylenal]ethenyl}-1-(3-sulfopropyl)-, hydroxide, inner salt (di-4-ANEPPS). For a 1-A, 10-ms shock during the action potential plateau, optical maps from fluorescence collected using emission ratiometry (excitation at 488 nm and emissions at 510–570 and >590 nm) indicated that the disk altered Vm by as much as the height of an action potential. Δ Vm became more positive near the edge of the disk, where the ITO conductance gradient was parallel to applied current, and more negative near the opposite edge, where the gradient was not parallel to current. For diastolic shocks, the disk expedited membrane excitation at the sites of positive Δ Vm in the heart and in a cardiac model with realistic ITO disk surface and interfacial conductances. Optical maps of ITO transmittance and the model indicated that the disk introduced anodal and cathodal stimulatory current at opposite edges of the disk. Thus ITO allows study of the stimulatory effects of a passive conductor in an electric field.

Selective attenuation by adenosine of arrhythmogenic action of isoproterenol on ventricular myocytes

We examined whether adenosine equally attenuated the stimulatory effects of isoproterenol on arrhythmic activity and twitch shortening of guinea pig isolated ventricular myocytes. Transmembrane voltages and whole cell currents were recorded with patch electrodes, and cell twitch shortening was measured using a video-motion detector. Isoproterenol increased the action potential duration at 50% repolarization (APD50), L-type Ca2+ current [ I Ca(L)], and cell twitch shortening and induced delayed afterdepolarizations (DAD), transient inward current ( I Ti), and aftercontractions. Adenosine attenuated the arrhythmogenic actions of isoproterenol more than it attenuated the effects of isoproterenol on APD50, I Ca(L), or twitch shortening. Adenosine (0.1–100 μmol/l) decreased the amplitude of DADs by 30 ± 6% to 92 ± 5% but attenuated isoproterenol-induced prolongation of the APD50 by only 14 ± 4% to 59 ± 4% and had no effect on the voltage of action potential plateau. Adenosine (30 μmol/l) inhibited I Ti by 91 ± 4% but decreased isoproterenol-stimulated I Ca(L) by only 30 ± 12%. Isoproterenol-induced aftercontractions were abolished by adenosine (10 μmol/l), whereas the amplitude of twitch shortening was not reduced. The effects of adenosine on twitch shortenings and aftercontractions were mimicked by the A1-adenosine receptor agonist CPA ( N 6-cyclopentyladenosine) and by ryanodine. In conclusion, adenosine antagonized the proarrhythmic effect of β-adrenergic stimulation on ventricular myocytes without reducing cell twitch shortening.

Virtual sources associated with linear and curved strands of cardiac cells

Transmembrane potential ( V m) responses in cardiac strands with different curvature were characterized during uniform electric-field stimulation with the use of modeling and experimental approaches. Linear and U-shaped strands (width 100–150 μm) were stained with voltage-sensitive dye. V m was measured by optical mapping across the width and at sites of beginning curvature. Field pulses were applied transverse to the strands during the action-potential plateau. For linear strands, V mcontained 1) a rapid passive component ( V m ar) nearly linear and symmetric across the width, 2) a slower hyperpolarizing component ( V m as) greater and faster on the anodal side, and 3) at high field strengths a delayed depolarizing component ( V m ad) greater on the anodal side. For U-shaped strands, V m at sites of beginning curvature also contained rapid and slow components ( V m br and V m bs, respectively) that included contributions from the linear strand response and from the fiber curvature. V m ar, V m br, and part of V m bs could be attributed to passive behavior that was modeled, and V m as, V m ad, and part of V m bs could be attributed to active membrane currents. Thus curved strands exhibit field responses separable into components with characteristic amplitude, spatial, and temporal signatures.