Congestive heart failure after myocardial infarction in the rat: cardiac force and spontaneous sarcomere activity.

The causes of reduced cardiac force development in congestive heart failure (CHF) are still uncertain. We explored the subcellular mechanisms leading to decreased force development in trabeculae from rats with a myocardial infarction. We defined CHF according to clinical and pathological criteria and compared properties of trabeculae from animals with CHF (cMI) to those of animals with a myocardial scar but without evidence of CHF (uMI), and sham-operated animals. The new findings of this study on properties of cMI trabeculae are that (1) maximal twitch force following post-extrasystolic potentiation is unchanged; (2) the sensitivity of cMI trabeculae to [Ca(2+)](o) is increased; (3) spontaneous diastolic sarcomere length (SL) fluctuations (SA) are increased in cMI at all levels of SR Ca(2+) loading; and (4) SA is accompanied by a proportional reduction of F(max). The results suggest that the probability of spontaneous diastolic opening of SR Ca(2+) channels is increased in CHF. These data provide the basis for a novel mechanism underlying systolic and diastolic dysfunction as well as arrhythmias in hearts in CHF. If SA proves to be a component of myocardial dysfunction in human CHF, our thinking about therapy of the patient with CHF may be profoundly changed.

Damage induced arrhythmias: mechanisms and implications.

Little is known about the role played by non-uniform myocardial stress and strain distributions and by non-uniform excitation contraction coupling in mechanisms underlying the premature beats that initiate an arrhythmia. We will review the evidence in support of a mechanism in which both non-uniform contraction and increased Ca2+ load of cells adjacent to acutely damaged cells are essential in the "spontaneous" generation of Ca2+ transients during the relaxation phase of the electrically driven twitch. The putative mechanism of initiation of the propagating Ca2+ waves involves feedback of rapid length (or force) changes to dissociation of Ca2+ from the contractile filaments. A novel aspect of this concept is that these mechanically elicited Ca2+ transients induce propagating Ca2+ waves that travel into the adjacent normal myocardium and cause after-depolarizations, which, in turn, may cause premature action potentials. These premature action potentials will further load the cells with Ca2+, which promotes the subsequent generation of propagating Ca2+ transients and leads to triggered arrhythmias. The damage-induced premature beats may also initiate re-entry arrhythmias in non-uniform myocardium. These observations strongly support the concept that abnormal cellular Ca2+ transport plays a crucial role in the initiation of arrhythmias in damaged and non-uniform myocardium.

Delayed rectifier K currents have reduced amplitudes and altered kinetics in myocytes from infarcted canine ventricle.

OBJECTIVE: The rapid (I(Kr)) and slow (I(Ks)) components of delayed rectifier currents play an important role in determining the cardiac action potential configuration. Abnormalities in their function may contribute to arrhythmogenesis under pathological conditions. We studied the effects of myocardial infarction on I(Kr) and I(Ks) in canine ventricular myocytes and their molecular basis. METHODS: Infarct zone myocytes (IZs) were isolated from a thin layer of surviving epicardium overlying an infarct 5 days after a total occlusion of the left anterior descending (LAD) coronary artery. Normal myocytes (NZs) were isolated from the corresponding region of control hearts for comparison. Currents were recorded under the whole-cell patch clamp conditions. RESULTS: Both I(Kr) and I(Ks) current densities were reduced in IZs versus NZs. Kinetic analysis further suggests an acceleration of I(Kr) activation and I(Ks) deactivation. RNase protection assays were used to quantify the mRNA levels of I(Kr) and I(Ks) channel subunits (dERG, dIsK and dKvLQT1) in tissue immediately adjacent to the region where myocytes were isolated. mRNA levels of all three subunits were reduced 2 days after LAD occlusion (by 48+/-9%, 68+/-5%, and 45+/-4% for dERG, dIsK and dKvLQT1, respectively, n=8 each). By day 5, the dKvLQT1 message returned to control while those of dERG and dIsK remained reduced (by 52+/-7% and 76+/-6%, respectively). CONCLUSIONS: The decrease in I(Kr) and I(Ks) amplitudes and changes in their kinetics in infarcted tissue might be due to a decrease in functional channels and/or changes in their subunit composition. Heterogeneous changes in I(Kr) and I(Ks) in infarcted hearts may impact on the effects of varying heart rate or neurohumoral modulation on repolarization.

Abnormalities in Ca(i)handling in myocytes that survive in the infarcted heart are not just due to alterations in repolarization.

Studies from our laboratory have defined alterations in Ca(i)handling in the non-dialyzed subepicardial cells that have survived in the 5 day infarcted heart (IZs). To determine whether changes in the action potential profile contributed to the observed Ca(i)changes we have used a combined voltage clamp/epifluorescent technique to determine and compare changes in fura 2 ratios in IZs compared to those of epicardial cells from the non-infarcted canine hearts (NZs). We found that Ca(i)changes in voltage clamped IZs persisted. In NZs, Ca(i)transients showed the expected voltage dependence while IZs did not. To determine whether altered NaCa exchanger activity contributed to the observed changes in Ca(i)in IZs, we measured NaCa exchanger Ca(2+)fluxes (reverse and forward mode) and ionic currents in both cell types and under different Na(i)loads (10 and 20 m m). We found that there were no significant differences in resting, peak or magnitude of fura 2 ratio changes or in outward current densities between NZs and IZs even under the different Na(i)loads. Thus, we suggest that chronic up- or downregulation of the NaCa exchanger protein does not underlie observed Ca(i)changes in IZs. Additionally, Ca(2+)released with paced voltage steps represented 79% of that released by caffeine in NZs while, in IZs, caffeine releasable Ca(2+)was equivalent to that released with step depolarization. Thus, abnormalities in Ca(i)handling in IZs appear not to arise secondarily to changes in action potential configuration nor do they appear to be due to disease-induced alteations in NaCa exchanger function.

Ca(2+) transients and Ca(2+) waves in purkinje cells : role in action potential initiation.

Purkinje cells contain sarcoplasmic reticulum (SR) directly under the surface membrane, are devoid of t-tubuli, and are packed with myofibrils surrounded by central SR. Several studies have reported that electrical excitation induces a biphasic Ca(2+) transient in Purkinje fiber bundles. We determined the nature of the biphasic Ca(2+) transient in aggregates of Purkinje cells. Aggregates (n=12) were dispersed from the subendocardial Purkinje fiber network of normal canine left ventricle, loaded with Fluo-3/AM, and studied in normal Tyrode's solution (24 degrees C). Membrane action potentials were recorded with fine-tipped microelectrodes, and spatial and temporal changes in [Ca(2+)](i) were obtained from fluorescent images with an epifluorescent microscope (x20; Nikon). Electrical stimulation elicited an action potential as well as a sudden increase in fluorescence (L(0)) compared with resting levels. This was followed by a further increase in fluorescence (L(1)) along the edges of the cells. Fluorescence then progressed toward the Purkinje cell core (velocity of propagation 180 to 313 microm/s). In 62% of the aggregates, initial fluorescent changes of L(0) were followed by focally arising Ca(2+) waves (L(2)), which propagated at 158+/-14 microm/s (n=13). Spontaneous Ca(2+) waves (L(2)*) propagated like L(2) (164+/-10 microm/s) occurred between stimuli and caused slow membrane depolarization; 28% of L(2)* elicited action potentials. Both spontaneous Ca(2+) wave propagation and resulting membrane depolarization were thapsigargin sensitive. Early afterdepolarizations were not accompanied by Ca(2+) waves. Action potentials in Purkinje aggregates induced a rapid rise of Ca(2+) through I(CaL) and release from a subsarcolemmal compartment (L(0)). Ca(2+) release during L(0) either induced further Ca(2+) release, which propagated toward the cell core (L(1)), or initiated Ca(2+) release from small regions and caused L(2) Ca(2+) waves, which propagated throughout the aggregate. Spontaneous Ca(2+) waves (L(2)*) induce action potentials.

Electrical remodeling in ischemia and infarction.

This is a review of the electrophysiologic changes occurring at different times following myocardial infarction, both in the infarcted region (substrate) and in areas remote from the infarct. Regulators of channel function which might contribute to re-modeling, including autocrine/paracrine factors involved in ion channel gene regulation, are discussed.

Steady-state and nonsteady-state action potentials in fibrillating canine atrium: abnormal rate adaptation and its possible mechanisms.

OBJECTIVE: Our goal was to study rate adaptation of atrial action potentials in non-steady and steady states to further our understanding of mechanisms determining inducibility and stability of atrial fibrillation. METHODS: We used standard microelectrode techniques to examine the characteristics of steady-state action potentials paced at regular cycle lengths (CL) and of nonsteady-state action potentials observed after an abrupt change of CL in atria from canine hearts that had been rapidly paced. RESULTS: We compared action potential characteristics among normal atria, atria in which chronic atrial fibrillation (cAF, lasting more than 3 days) had been induced and atria in which only nonsustained atrial fibrillation (nAF, lasting less than 12 h) had been induced. In steady-state, the rate adaptation of maximum diastolic potential (MDP) and action potential duration (APD) and markedly reduced in both cAF and nAF. Action potential characteristics did not differ between cAF and nAF atria, suggesting that factors other than electrophysiological properties determine the chronicity of AF. The time course of change in APD after an abrupt change of CL was altered in nAF/cAF atria; i.e., when CL was prolonged, APD also prolonged at the first beat, and then shortened during several subsequent beats (initial phase). Thereafter, APD slowly prolonged to a new steady-state (slow phase). In nAF/cAF atria, the initial phase was enhanced (greater shortening of APD) and the slow phase was reduced (less prolongation of APD). This latter phase was modified by ryanodine. CONCLUSIONS: Thus the reduced rate adaptation of steady-state APD is explained mainly by the loss of a slow phase of APD adaptation in nAF/cAF which is reversed in the presence of ryanodine. Therefore, in both nAF and cAF atria, rate adaptation of MDP as well as APD are reduced, nonsteady state as well as steady state, AP characteristics are markedly altered and these changes are partially explicable by Ca, -dependent processes.

Effects of mibefradil, a T-type calcium current antagonist, on electrophysiology of Purkinje fibers that survived in the infarcted canine heart.

INTRODUCTION: We studied the effects of mibefradil (MIB), a nondihydropyridine T-type Ca2+ channel antagonist, on T- and L-type Ca2+ (I(CaT), I(CaL)) currents in Purkinje myocytes dispersed from the subendocardium of the left ventricle of normal (NZPC) and 48-hour infarcted (IZPC) hearts. METHODS AND RESULTS: Currents were recorded with Cs+- and EGTA-rich pipettes and in Na+-K+-free external solutions to eliminate overlapping currents. In all cells, I(Ca) was reduced by MIB (0.1 to 10 microM). No change in the time course of decay of peak I(Ca) was noted. Average peak T/L ratio decreased in NZPCs but not IZPCs with 1 microM MIB. Steady-state availability of I(CaL) was altered with 1 microM MIB in both cell types (mean +/- SEM) (V0.5 = -22 +/- 4 mV for NZPC and -25 +/- 5 mV for IZPC before drug; -63 +/- 9 mV for NZPC and -67 +/- 6 mV for IZPC after drug; P < 0.05). For I(CaT), V0.5 (-50 +/- 3 mV for NZPC and -52 +/- 1 mV for IZPC before drug) shifted to -60 +/- 2 mV (NZPC) and -62 +/- 3 mV (IZPC) (P < 0.05) after drug. We also determined the effects of MIB on spontaneously beating Purkinje normal fibers and on depolarized abnormally automatic fibers from the infarcted heart using standard microelectrode techniques. When NZPC and IZPC fibers were superfused with [K+]o = 2.7 mM, MIB 3 microM and 10 microM had no effect on rate or the maximum diastolic potential, but action potential plateau shifted to more negative values, the slope of repolarization phase 3 decreased, and action potential duration increased. CONCLUSION: MIB blocks L- and T-type Ca2+ currents in Purkinje myocytes but lacks an effect on either normal or abnormal automaticity in Purkinje fibers.

Ca2+ waves during triggered propagated contractions in intact trabeculae. Determinants of the velocity of propagation.

During triggered propagated contractions, Ca2+ waves travel along cardiac trabeculae with a constant velocity (Vprop) ranging from 0. 34 to 5.47 mm/s. To explore the determinants of Vprop, we studied (1) the relationship between [Ca2+]i and Vprop and (2) the effect of low concentrations of caffeine on Vprop. Trabeculae were dissected from the right ventricle of rat hearts. [Ca2+]i was measured using electrophoretically injected fura-2 and an image-intensified CCD camera. Force was measured using a silicon strain gauge, and sarcomere length was measured using laser diffraction techniques. After induction of reproducible Ca2+ waves by trains of electrical stimuli (2.5 Hz) at 21.9+/-0.2 degrees C, the number of stimuli or [Ca2+]o was varied in 9 trabeculae. In 5 trabeculae, the effects of caffeine (0.1 to 1.0 mmol/L) at [Ca2+]o of 2.2+/-0.3 mmol/L were determined. All images were recorded under stable conditions of wave propagation. The increment in [Ca2+]i during the last electrically stimulated transient (DeltaCaT) and [Ca2+]i just before onset of the Ca2+ waves (CaD) were used to estimate the Ca2+ loading of the sarcoplasmic reticulum (SR) and the myoplasm, respectively. The ratio (DeltaCaW/DeltaCaT) of the [Ca2+]i increment during the waves (DeltaCaW) to DeltaCaT was used to estimate the probability of opening of the SR-Ca2+ release channel during wave propagation. As a result of an increase of the number of stimuli or [Ca2+]o, Vprop increased in proportion to (1) DeltaCaT (r=0.82); (2) CaD (r=0.88); (3) DeltaCaW (r=0.85); and (4) DeltaCaW/DeltaCaT (r=0.74). The addition of caffeine (

Effects of Bay Y5959 on Ca2+ currents and intracellular Ca2+ in cells that have survived in the epicardial border of the infarcted canine heart.

We determined and compared the effects of the dihydropyridine agonist, Bay Y5959, on the amplitude of L-type Ca2+ currents and intracellular Ca2+ transients in epicardial cells from noninfarcted hearts (NZs) and surviving cells from the epicardial border zone of 5-day infarcted canine hearts (IZs). We determined the effects of Bay Y5959 on the L-type Ca2+ current by using single cells and a whole-cell voltage-clamp approach. To elucidate the effects of Bay Y5959 on the amplitude and time course of the spatially averaged intracellular Ca2+ transient (Ca(i)T), myocytes from the two cell groups were loaded and studied by using the Ca2+-sensitive indicator fura-2/AM. Bay Y5959 increased the amplitude of the L-type Ca2+ current in both cell groups, but peak amplitude in NZs was always greater than that in IZs. Bay Y5959 also increased Ca(i)T amplitude in both NZs and IZs and significantly accelerated the Ca(i)T time course in IZs, particularly at the faster pacing-cycle length. We suggest that the Bay Y5959 effect to restore L-type Ca2+ currents in IZs contributes to its observed antiarrhythmic effects during the reentrant ventricular tachycardias that are known to occur in the epicardial border zone of the infarcted heart.

Lidocaine action on Na+ currents in ventricular myocytes from the epicardial border zone of the infarcted heart.

Myocytes overlying a zone of infarction form the primary substrate for serious reentrant ventricular arrhythmias. In vitro and in vivo studies suggest that antiarrhythmic agents affect Na+ channels of cells from the epicardial border zone (EBZ) of the 5-day infarcted heart differently than they affect those of normal muscle. However, the mechanisms responsible for this difference remain unclear. Previous studies have revealed differences in Na+ current (INa) density and inactivation gating kinetics in myocytes dispersed from the EBZ (IZs). Since changes in inactivation gating could influence lidocaine action, we examined the effects of lidocaine on INa of IZs (n=38) and epicardial myocytes from the noninfarcted heart (NZs) (n=50) using the whole-cell variation of the patch-clamp technique. In drug-free conditions, the voltage dependence of steady-state inactivation of IZs was shifted negative to that of NZs, causing greater inactivation of IZ channels at depolarized (> or = -100-mV) holding potentials. Consistent with a high affinity for the inactivated channel conformation, lidocaine produced more tonic block in IZs than NZs at depolarized holding potentials. Additionally, in drug-free conditions, IZ INa exhibited an enhanced rate of inactivation from closed states, a delay in recovery from inactivation, and increased use-dependent reduction in amplitude during rapid (1- to 3-Hz) pulse trains. In both IZs and NZs, lidocaine (20 to 120 micromol/L) accelerated the rate of time-dependent loss of availability and markedly delayed recovery from availability, inducing significant use-dependent reduction of INa. However, at drug concentrations > or =60 micromol/L, the difference in use-dependent current reduction between IZs and NZs was minimized. The action of lidocaine to render Na+ channel inactivation in NZs more similar to that of IZs may be central to its (pro)antiarrhythmic effects.

Reduced inward rectifying and increased E-4031-sensitive K+ current density in arrhythmogenic subendocardial purkinje myocytes from the infarcted heart.

INTRODUCTION: Subendocardial Purkinje myocytes from the 48-hour infarcted heart (IZPCs) have reduced resting potentials, possibly due to altered inwardly rectifying K+ currents IK1. Abnormal depolarization-activated outward K+ currents could contribute to long triangularly shaped action potentials of IZPCs. METHODS AND RESULTS: We used whole cell patch recordings to compare cesium-sensitive IK1 and 4-aminopyridine (4-AP)-resistant, noninactivating sustained IK between normal Purkinje myocytes (NZPCs) and IZPCs. IZPCs showed decreased net membrane currents. Two IZPC groups were distinguished, based on 4-AP-resistant outward K+ currents. IZPC-I had isochronal IK1 current-voltage relations similar to NZPCs whereas IZPC-II showed significantly reduced IK1 and increased outward plateau currents. To study the sustained IK in the presence of the Class III antiarrhythmic agent E-4031, a two-pulse protocol was used to inactivate transient outward currents, followed by step depolarizations. E-4031-sensitive currents were significantly greater in IZPCs at depolarized potentials (> 0 mV). Similar to NZPCs, IZPC E-4031 currents showed time dependence during depolarization, lack of rectification at positive steps, and voltage-dependent recovery from block. CONCLUSION: Decreased IK1 may account for reduced resting potentials in IZPCs. E-4031-sensitive currents in NZPCs, unlike those in canine ventricular myocytes, are sensitive to 4-AP and are larger in IZPCs.

Ca2+ waves during triggered propagated contractions in intact trabeculae.

Triggered propagated contractions (TPCs) starting from damaged regions travel along multicellular cardiac muscle preparations. We have reported that octanol (100 microM) inhibits TPCs. The inhibitory effect of octanol on propagation of TPCs could be due to an effect of octanol on Ca(2+)-induced Ca2+ release (CICR) mediated by Ca2+ diffusion inside the single cell or on the diffusion of Ca2+ from cell to cell via gap junctions (GJs). Therefore, we studied the regional changes in intracellular Ca2+ concentration ([Ca2+]i) during TPCs and the effect of octanol on the permeability of gap junctions (PGJ) in rat cardiac trabeculae. [Ca2+]i was measured using electrophoretically injected fura 2 and an image-intensified charge-coupled device camera. PGJ was calculated from the diffusion coefficient for fura 2 in trabeculae (Dtrab) and in the myoplasm (Dmyop). After 1- and 3-h superfusion with 100 microM 1-octanol, Dmyop showed no significant changes, whereas Dtrab was reduced significantly. Therefore, calculated PGJ was reduced from 4.15 x 10(-5) to 2.10 x 10(-5) and 0.86 x 10(-5) cm/s, respectively. The propagation velocity of the regional increases in [Ca2+]i during TPCs was constant, averaging 1.69 +/- 1.48 mm/s (range 0.34-5.47 mm/s, n = 10). These observations support the hypothesis that TPCs are initiated near the damaged ends of trabeculae and are propagated by CICR from the sarcoplasmic reticulum mediated by diffusion of Ca2+ through cells and from cell to cell through GJs.

Ca(2+)-dependent outward currents in myocytes from epicardial border zone of 5-day infarcted canine heart.

Myocytes from the epicardial border zone (EBZ) of the 5-day infarcted canine heart (IZ) have abnormal transmembrane action potentials, reduced L-type Ca2+ currents (ICa,L) and altered intracellular Ca2+ (Cai) transients compared with those of normal epicardial myocytes (NZ). We hypothesized that altered Cai cycling might be reflected in differences in Cai-dependent outward currents (Ito2). We recorded Ito2 in NZ and IZ using whole cell patch-clamp techniques. Ito2 was defined as the amplitude of the 4-aminopyridine-resistant transient outward current that was blocked by 200 microM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) or DIDS+ ryanodine (2 microM). Ito2 were present in both NZ and IZ, but peak density was significantly reduced in IZ, particularly at positive plateau voltages. Time course of decay of Ito2 was biexponential and similar in NZ and IZ. A given peak ICa,L was usually associated with a smaller peak Ito2 in IZ. These differences were exaggerated when Ito2 and Cai transients were determined in rapidly paced cells. In summary, myocytes surviving in the EBZ of the infarcted heart have Ito2, yet they are reduced in density and can vary, particularly at fast pacing rates.