András Varró

Asymmetrical distribution of ion channels in canine and human left-ventricular wall: Epicardium versus midmyocardium

The aim of the present study was to compare the distribution of ion currents and the major underlying ion channel proteins in canine and human subepicardial (EPI) and midmyocardial (MID) left-ventricular muscle. Ion currents and action potentials were recorded from canine cardiomyocytes derived from the very superficial EPI and central MID regions of the left ventricle. Amplitude, duration and the maximum velocity of depolarization of the action potential were significantly greater in MID than EPI myocytes, whereas phase-1 repolarization was more pronounced in the EPI cells. Amplitudes of the transient outwards K+ current (29.5±1.5 vs. 19.0±2.3xa0pA/pF at +50xa0mV) and the slow component of the delayed rectifier K+ current (10.3±2.3 vs. 6.5±1.0xa0pA/pF at +50xa0mV) were significantly larger in EPI than in MID myocytes under whole-cell voltage-clamp conditions. The densities of the inwards rectifier K+ current, rapid delayed rectifier K+ current and L-type Ca2+ current were similar in both cell types. Expression of channel proteins in both canine and human ventricular myocardium was determined by Western blotting. In the canine heart, the expression of Kv4.3, Kv1.4, KChIP2 and KvLQT1 was significantly higher, and that of Nav1.5 and MinK much lower, in EPI than in MID. No significant EPI-MID differences were observed in the expression of the other channel proteins studied (Kir2.1, α1C, HERG and MiRP1). Similar results were obtained in human hearts, although the HERG was more abundant in the EPI than in the MID layer. In the canine heart, the EPI-MID differences in ion current densities were proportional to differences in channel protein expression. Except for the density of HERG, the pattern of EPI-MID distribution of ion-channel proteins was identical in canine and human ventricles.

Rate and concentration-dependent effects of UK-68,798, a potent new class III antiarrhythmic, on canine Purkinje fibre action potential duration and Vmax.

1 The frequency‐dependent electrophysiological effects of UK‐68,798 in concentrations of 1, 3, 10 and 30 nm were examined in isolated cardiac Purkinje fibres of the dog at both a number of constant rates of stimulation and following abrupt changes in pacing cycle length. 2 In all concentrations evaluated, UK‐68,798 lengthened action potential duration in a concentration‐and rate‐dependent manner (e.g., at a cycle length = 500 ms, control APD90 = 234.0 ± 3.3 ms, while after 10 nm UK‐68,798, APD90 = 315.0 ± 5.9 ms). 3 The duration of action potentials evoked following abrupt changes in pacing rate were also increased in a concentration‐dependent manner at all diastolic intervals tested. 4 The fast and slow time constants for restitution of APD were not altered by UK‐68,798. However, the amplitude terms for this relation were increased. 5 In addition, the maximum upstroke velocity (V̇max) was not significantly affected by exposure to UK‐68,798 at any concentration or diastolic interval. The kinetics for recovery of V̇max were thus unaffected. 6 These findings are similar to those previously reported for recognized class III antiarrhythmic agents (e.g., bretylium, clofilium, and sotalol); however, UK‐68,798 was 1,000 to 10,000 times more potent. 7 The combined potency and selectivity of this agent seem to make it an ideal tool for the investigation of cardiac potassium channels believed responsible for controlling the duration of the action potential. 8 This potent and highly selective compound may prove extremely useful in the control of cardiac arrhythmias.

Rate-dependent electrophysiological effects of OPC-8212: comparison to sotalol.

The rate-dependent effects of OPC-8212 (6 microM), a new positive inotropic drug which lengthens cardiac action potential duration were studied in dog cardiac Purkinje fibers and compared to the effects of sotalol (30 microM), a class III antiarrhythmic using conventional microelectrode technique. Both OPC-8212 and sotalol lengthened action potential duration in a frequency-dependent manner. Both had a greater effect at slower pacing frequencies than at fast. The maximum rising velocity of the action potential upstroke was not affected by either compound at any of the pacing cycle lengths applied (300-2000 ms). The effects of OPC-8212 and sotalol on action potential duration and maximum rising velocity were also compared following abrupt changes in pacing cycle length (i.e. coupling intervals ranging between approximately 250-3000 ms). The effects of both OPC-8212 and sotalol following abrupt changes in pacing cycle length were similar to one another. Neither OPC-8212 nor sotalol altered the time constants for restitution of action potential duration or the kinetics for recovery of the maximum rising velocity of the action potential upstroke. These results indicate that the rate-dependent electrophysiological effects of OPC-8212 closely mimic those of sotalol and suggest that OPC-8212 may be an effective class III antiarrhythmic in addition to being a positive inotropic compound.

In vitro cardiac models of dog Purkinje fibre triggered and spontaneous electrical activity: effects of nicorandil.

1 The effects of nicorandil (30 μm and 100 μm) on two models of triggered activity [early afterdepolarizations (EADs) and delayed afterdepolarizations (DADs)] and on spontaneous automaticity occurring from both normal and depolarized levels of membrane potential were examined in isolated cardiac Purkinje fibres of the dog. Standard intracellular microelectrode techniques were used. 2 Nicorandil (30 μm) abolished EADs provoked by superfusion with Tyrode solution containing 2.7 mm K+ and 3 mm Cs. 3 DADs were induced by 0.2 μm acetylstrophanthidin in Tyrode solution containing 5.4 mm K+. Nicorandil (30 μm) significantly reduced the amplitude of these DADs from 12.5 ± 2.5 mV to 5.5 ± 0.2 mV (P > 0.02, n = 6), while DADs were fully abolished by 100 μm nicorandil. 4 In unstimulated Purkinje strands, superfused with 2.7 mm K+ containing Tyrode solution having a pH of either 7.4 or 6.8, spontaneous depolarizations developed with a mean maximum diastolic potential (MDP) of −84.6 ± 1.6 mV (n = 9) or −54.0 ± 1.2 mV (n = 9), respectively. Nicorandil significantly reduced the frequency of this automatic activity and caused its cessation, at either level of MDP. Nicorandil, however, produced significant hyperpolarization only when automaticity occurred from the depolarized level of potential. 5 These results suggest that nicorandil may exert significant antiarrhythmic actions in vivo by abolishing both spontaneous and triggered electrical activity.

Ionic basis for OPC-8212-induced increase in action potential duration in isolated rabbit, guinea pig and human ventricular myocytes.

Changes in transmembrane ionic currents induced by OPC-8212 (3,4-dihydro-6-[4-(3,4-dimethoxybenzoyl)-1-piperazinyl]- 2(1H)-quinoline), a recently introduced positive inotropic agent which lengthens cardiac action potential duration, were examined using whole-cell voltage-clamp techniques in single rabbit, guinea pig and human ventricular myocytes. In rabbit, OPC-8212 (12 μmol/l) significantly increased membrane action potential duration measured at 90% of repolarization by an average of 88 ms (from 462 ± 25 to 550 ± 35 ms, n = 4; P < 0.05). In rabbit this increase in duration was not associated with significant changes in either the inward rectifier or transient outward K+ currents. The magnitude of the secondary inward current evoked from a holding potential of −50 mV was significantly increased by 97 ± 8% (n = 6; P < 0.01) while a demonstrable delayed rectifier outward current could not be identified in the rabbit myocytes examined at room temperature. In guinea pig ventricular myocytes, where the delayed rectifier was large, 12 μmol/l OPC-8212 significantly depressed the current by 58 ± 10% (n = 6; P < 0.01). The effects of OPC-8212 in human ventricular myocytes obtained from the explanted heart of a single patient having an idiopathic cardiomyopathy most closely resembled those observed in isolated rabbit ventricular myocytes. Thus, in rabbit and a few human ventricular myocytes examined at room temperature, OPC-8212 appeared to lengthen cardiac membrane action potential duration primarily by increasing the amplitude of the secondary inward current believed to primarily represent current through L-type Ca2+ channels. In guinea pig preparations, OPC-8212 also decreased the delayed rectifier outward K+ current which also would account for an increase in action potential duration. OPC-8212 could not be demonstrated to affect Na+ current inactivation in a manner similar to that produced by 1 mg/l veratrine, a recognized Na + channel agonist, which dramatically slowed this process.

The combined electrophysiological effects of lignocaine and sotalol in canine isolated cardiac Purkinje fibres are rate‐dependent

1 The frequency‐dependent electrophysiological effects of lignocaine, sotalol, and their combination were studied in dog isolated cardiac Purkinje fibres, both at various constant rates of stimulation and following abrupt changes in pacing cycle length. 2 The combined effect of 18 μm lignocaine and 30 μm sotalol selectively lengthened duration of premature action potentials evoked at a diastolic interval of 40 ms (from 172.2 ± 5.4 to 201.7 ± 4.9 ms, n = 6, P > 0.01) without significantly changing the durations of action potentials evoked at the basic cycle length of 500 ms (259.1 ± 7.7 vs 251.9 ± 3.9 ms, n = 11). 3 The combination of lignocaine with sotalol, like lignocaine alone, displayed a use‐dependent depression of and revealed a slow component for a recovery of (τ = 173.5 ± 16.0 ms, n = 5). 4 The kinetics for restitution of action potential duration were also slowed by the combination of the two dwo drugs (τf = 173.6 ± 16.7, before, vs 268.5 ± 8.5 ms, after; n = 5, P > 0.01), while the maximum action potential duration attained in this relation was not increased as it was by sotalol alone. 5 Lignocaine, therefore, appeared to inhibit the sotalol‐induced lengthening of action potential duration at slow pacing rates and at long diastolic intervals. The combination of lignocaine with sotalol also completely abolished the occurrence of sotalol‐induced early afterdepolarizations. 6 Finally, sotalol alone moderately increased the range of premature action potential durations, while the combination of the two drugs significantly decreased this parameter. 7 These findings indicate that the combination of lignocaine with sotalol may provide important, and unique, beneficial electrophysiological alterations that might be expected to provide enhanced antiarrhythmic efficacy in patients.

Electrophysiologic effects of FPL 13210 on canine Purkinje fiber action potential duration and Vmax comparison to disopyramide

SummaryThe frequency-dependent effects of FPL 13210, a new disopyramide derivative, were examined in isolated canine cardiac Purkinje fibers paced at a frequency of 2 Hz and following abrupt changes in pacing cycle length. At 2 Hz, FPL 13210 depressed Vmax while shortening action potential duration measured at 50% of repolarization (APD50) and not affecting duration measured at 90% of repolarization (APD90). These effects were concentration dependent over the range of 1–30 μM. The depression of Vmax produced by 5μM FPL 13210 was not significantly different than that produced by 18 μM disopyramide while the preparations were paced constantly at 2 Hz. At these concentrations, recovery of Vmax was slowed by both FPL 13210 and disopyramide. The slow time constant estimated for this relation after exposure to FPL 13210 was approximately 6.5 times longer than that estimated following administration of disopyramide. In addition, APD90s evoked by early premature stimuli in the presence of 5 μM FPL 13210 were longer than those produced in the absence of drug when the diastolic interval was less than 60 ms. Later extra stimuli evoked at diastolic intervals longer than 100 ms produced shorter APD90s after FPL 13210 administration. Therefore, when FPL 13210 is compared to disopyramide using concentrations selected to produce equivalent degrees of Vmax depression, FPL 13210 produced effects on APD90 that were opposite to those produced by disopyramide when the diastolic interval was longer than normal. These effects of FPL 13210 would suggest that this compound should be classified as a class Ic antiarrhythmic agent, unlike disopyramide, a class Ia antiarrhythmic agent.

Comparison of the Electromechanical Effects of Vesnarinone and Amrinone in Isolated Dog Purkinje Strands and Ventricular Trabeculae.

Background: Conventional microelectrode techniques were used to compare the concentration-dependent effects of vesnarinone (0.1-100 μM) and amrinone (1 μM-1 mm) on action potential duration (APD) and developed force in both isolated dog ventricular trabeculae and Purkinje strands. Methods and Results: Both drugs increased contractility of trabecular muscle preparations, while, in Purkinje strands, vesnarinone failed to increase developed force during continuous pacing at 2 Hz. Vesnarinone lengthened APD in both preparations; although this effect was more marked in Purkinje strands. Ventricular muscle APD was not affected by amrinone (1 μM to 1 mM), while, in Purkinje strands, amrinone produced a biphasic effect on APD. Low concentrations (1-100 μM) of amrinone shortened Purkinje fiber APD, while only the highest concentration (1 mM) used lengthened APD. In addition, in Purkinje strand preparations the effects of vesnarinone (10 μM) on APD and developed force were proportional to pacing cycle length at frequencies slower than 2 Hz; however, at frequencies faster than 2 Hz vesnarinone decreased developed force while APD was lengthened. In ventricular trabecular muscle preparations, the effects of vesnarinone were not affected by frequency. Conclusions: These results indicate clear differences between the effects of vesnarinone and amrinone in isolated cardiac preparations. These differences in experimental effects in isolated cardiac preparations may help provide an explanation for the disappointing clinical response of patients in heart failure to amrinone, while vesnarinone has appeared to be beneficial.

Use-dependent action of antiarrhythmic drugs in frog skeletal muscle and canine cardiac Purkinje fiber

1. Conventional microelectrode techniques were used to study the effect of quinidine (10 microM), lidocaine (20 microM), and verapamil (3-10 microM) on action potential upstroke (V+ max) in frog skeletal muscle and dog Purkinje fiber. 2. The frequency-dependent nature of V+ max depression induced by these drugs was similar in both preparations, however, quinidine was more potent in skeletal muscle while lidocaine was in Purkinje fibers. 3. In skeletal muscle tetrodotoxin (3 and 15 nM) and low concentrations of antiarrhythmic drugs proportionally reduced the maximum velocity of depolarization and repolarization (V+ max and V- max, respectively), whereas V- max was more depressed than V+ max by high concentrations (50-200 microM) of antiarrhythmics. Decreases in the overshoot potential were proportional to the V+ max block in the case of each drug. 4. These results indicate that therapeutically relevant concentrations of quinidine and lidocaine inhibit skeletal muscle Na+ channels in a use-dependent manner similar to heart, while at higher concentrations the K+ channels may also be blocked. Therapeutic implications of the results are discussed.

Biphasic effect of tetraethylammonium on canine Purkinje fibre action potential configuration

1. Using conventional microelectrode techniques a biphasic effect of tetraethylammonium (5 mmol/l) on the configuration of action potentials recorded from isolated canine Purkinje fibres: action potentials were first shortened (early effect) and then lengthened (late effect) by tetraethylammonium. 2. The early effect of tetraethylammonium also included lengthening of phase 1 duration and elevation of the plateau amplitude. These early effects reached steady-state within the first 3 min of superfusion and were readily reversed within 3 min of initiating washout of the drug. 3. The late effect (gradual lengthening of repolarisation during phase 3) failed to reach steady-state within the initial 60 min of superfusion and was not reversible. 4. The early effects of tetraethylammonium were more marked at slow driving rates and were not affected by blockade of alpha- and beta-adrenoceptors using 1 mumol/l phentolamine and 1 mumol/l propranolol. 5. The early effects of tetraethylammonium were mimicked by 4-aminopyridine (0.5 mmol/l), and in the presence of 4-aminopyridine tetraethylammonium failed to induce further changes in action potential morphology. 6. The early effects of tetraethylammonium may be due to inhibition of the transient outward current. 7. The rapid onset and reversibility of these early effects suggest that tetraethylammonium may act from outside the cell membrane.