Canine Purkinje Fibers Research Articles

Pacemaker current i(f) in adult canine cardiac ventricular myocytes.

1. Single cells enzymatically isolated from canine ventricle and canine Purkinje fibres were studied with the whole-cell patch clamp technique, and the properties of the pacemaker current i(f) compared. 2. Steady-state i(f) activation occurred in canine ventricular myocytes at more negative potentials (-120 to -170 mV) than in canine Purkinje cells (-80 to -130 mV). 3. Reversal potentials were obtained in various extracellular Na+ (140, 79 or 37 mM) and K+ concentrations (25, 9 or 5.4 mM) to determine the ionic selectivity of i(f) in the ventricle. The results suggest that this current was carried by both sodium and potassium ions. 4. The plots of the time constants of i(f) activation against voltage were 'bell shaped' in both canine ventricular and Purkinje myocytes. The curve for the ventricular myocytes was shifted about 30 mV in the negative direction. In both ventricular and Purkinje myocytes, the fully activated I-V relationship exhibited outward rectification in 5.4 mM extracellular K+. 5. Calyculin A (0.5 microM) increased i(f) by shifting its activation to more positive potentials in ventricular myocytes. Protein kinase inhibition by H-7 (200 microM) or H-8 (100 microM) reversed the positive voltage shift of i(f) activation. This effect of calyculin A also occurred when the permeabilized patch was used for whole-cell recording. 6. These results indicate i(f) is present in ventricular myocytes. If shifted to more positive potentials i(f) could play a role in ischaemia-induced ventricular arrhythmias. The negative shift of i(f) in the ventricle might play a role in differentiating non-pacing regions of the heart from those regions that pace.

A Subtype of α1Adrenoceptor Mediates Depression of Conduction in Purkinje Fibers Exposed to Halothane

An action of epinephrine at alpha adrenoceptors has been reported to slow conduction in Purkinje fibers exposed to halothane. In Purkinje fibers one pharmacologically distinguishable alpha 1-adrenoceptor subtype (alpha 1B) sensitive to the noncompetitive antagonist chloroethylcholinidine mediates decreases in automaticity. Another alpha 1 subtype (alpha 1A), sensitive to the competitive antagonist WB4101, increases spontaneous rate and action potential duration by a mechanism thought to involve hydrolysis of membrane phosphoinositides by phospholipase C. This study examined the dose-response relation and receptor-effector mechanisms underlying depression of conduction in canine Purkinje fibers by epinephrine with halothane. Conduction velocity was determined in vitro by measuring the conduction time between action potentials recorded from two Purkinje fibers located about 6 mm apart along the length of free running portions of the ventricular conduction system, the false tendons. Velocity was evaluated at 1-min intervals during trials of rapid exposure to different agonists in groups of 6-12 preparations. Epinephrine (0.2-5.0 microM) transiently decreased Purkinje conduction velocity in a dose-related manner by as much as 33% (at 5 microM epinephrine with 0.86 mM (2.8%) halothane). Velocity decreased by 5% (P < or = 0.01) at an epinephrine concentration similar to "just-threshold" dysrhythmogenic plasma epinephrine concentrations (0.2 microM epinephrine with 0.46 mM halothane) reported in halothane-anesthetized dogs. The decreases of conduction velocity were blocked by prazosin but not by metoprolol, were produced by phenylephrine but not by clonidine, and were antagonized by equimolar (0.5 microM) concentrations of WB4101 more so (P < or = 0.01) than by chloroethylclonidine. WB4101 (0.1 microM) produced 87% inhibition of the response to 0.2 microM epinephrine after chloroethylclonidine pretreatment, indicating mediation by the alpha 1A subtype. Other agonists linked to cardiac phospholipase C activation, including endothelin 1 (40 nM) and the muscarinic agonist carbamylcholine (1 mM), also decreased conduction velocity in fibers exposed to halothane. Clinically relevant concentrations of epinephrine transiently depress conduction in Purkinje fibers exposed to halothane by activating cardiac alpha 1 adrenoceptors, largely but not exclusively the WB4101-sensitive alpha 1A subtype, reportedly coupled to stimulation of phospholipase C and generation of the second messengers diacylglycerol and inositol trisphosphate. Anesthetic potentiation of cardiac alpha 1-adrenoceptor effects may contribute to the generation of halothane-epinephrine dysrhythmias by abnormally slowing conduction and facilitating reentry.

In Vivo and In Vitro Electrophysiologic Effects of Terodiline on Dog Myocardium

Terodiline hydrochloride, widely prescribed for urinary incontinence, has been reported to cause bradycardia and torsades de pointes. In this study, we characterized the electrophysiologic effects of terodiline in dog cardiac tissues in vivo and in isolated canine cardiac Purkinje fibers. Terodiline (1 to 10 microM) resulted in dose-dependent reduction of action potential amplitude and maximal upstroke velocity (Vmax). The threshold for these effects was approximately 2 microM (0.6 mg/L), and the changes were cycle-length dependent. Terodiline (> or = 2 microM) also depressed the action potential plateau but did not significantly alter action potential duration at concentrations < or = 10 microM. In vivo studies demonstrated that high doses of terodiline (3 mg/kg) lengthened AH and HV intervals, slowed spontaneous sinus rate, prolonged ventricular refractoriness, and inhibited vagally induced slowing of the sinus node. Sympathetic effects on spontaneous sinus rate were unchanged. In both isolated canine Purkinje fibers and anesthetized dogs, terodiline did not evoke afterdepolarizations, repetitive firing, or ventricular tachyarrhythmias under normal or hypokalemic conditions. Our findings suggest that terodiline (> or = 1 to 2 microM) leads to blockade of sodium and calcium channels as well as muscarinic receptors in canine cardiac tissues.

Electrophysiological effects of alpha 2-adrenergic stimulation in canine cardiac Purkinje fibers

The effects of alpha 2-adrenergic stimulation on action potentials were measured in isolated canine Purkinje fibers. Action potential durations at 50 and 90% of repolarization (APD50 and APD90) were significantly prolonged by 0.25 microM l-norepinephrine + 0.5 microM dl-propranolol (NE+P) from baseline values of 166 +/- 7 and 249 +/- 9 (SE) ms (n = 7) to 174 +/- 7 and 265 +/- 9 ms, respectively (P < 0.05 for both). Selective alpha 2-blockade with 0.01 microM yohimbine (YO) reduced this prolongation by NE+P in APD50 and APD90 to 169 +/- 7 and 256 +/- 8 ms, respectively (P < 0.05 compared with NE+P). Additional selective alpha 1-blockade with 0.01 microM prazosin (PZ) completely blocked the effects of NE+P, returning APD50 and APD90 to 163 +/- 7 and 250 +/- 9 ms (not different from baseline). After incubation of isolated Purkinje fibers with pertussis toxin (1 microgram/ml), which reduced the availability of a 41-kDa membrane protein for ADP ribosylation by 70 +/- 7% (n = 4, P < 0.05), YO failed to reverse the prolongation in action potential durations brought on by NE+P, but the effects of PZ were intact. The effects of alpha 2-stimulation on beta-adrenergic-induced delayed afterdepolarizations (DADs) were studied by burst pacing of Purkinje fibers in Tyrode solution containing 7.5 mM Ca2+. The DADs induced in the presence of NE+PZ (beta- + alpha 2-stimulation) were significantly smaller in amplitude and required a shorter pacing cycle length to reach threshold than those induced in the presence of NE+PZ+YO (unopposed beta-adrenergic stimulation). Furthermore sustained triggered activity, seen in five of eight preparations under beta-stimulation, could no longer be elicited in the presence of beta- + alpha 2-stimulation. These results suggest that the postjunctional alpha 2-adrenergic receptors in canine Purkinje fibers are coupled to a pertussis toxin-sensitive G protein and that stimulation of these receptors leads to action potential prolongation and suppression of DADs induced by beta-adrenergic stimulation.

Effects of Hypothermia, Potassium, and Verapamil on the Action Potential Characteristics of Canine Cardiac Purkinje Fibers

Hypothermia may induce hypokalemia and increase intracellular Ca2+ by affecting serum K+ and Ca2+ fluxes across the cell membrane. These ionic alterations may significantly change the electrophysiologic characteristics of the cardiac action potential and may induce cardiac arrhythmias. The current study was undertaken to determine whether electrophysiologic changes in Purkinje fibers induced by hypothermia could be reversed by manipulating the extracellular K+ and transmembrane Ca2+ fluxes by Ca2+ channel blockade with verapamil. A conventional microelectrode method was used to determine the effects of hypothermia (32 +/- 0.5 degrees C and 28 +/- 0.5 degrees C) and various external K+ concentrations ([K+]o) (2.3, 3.8, and 6.8 mM) on maximum diastolic potential, maximum rate of phase 0 depolarization (Vmax), and action potential duration (APD) at 50% (APD50) and at 95% (APD95) repolarization in isolated canine cardiac Purkinje fibers. To evaluate the contribution of the slow inward Ca2+ current to action potential changes in hypothermia, the experiments were repeated in the presence of the Ca(2+)-channel antagonist verapamil (1 microM). Variations of [K+]o induced the expected shifts in maximum diastolic potential, and hypothermia (28 degrees C) induced moderate depolarization, but only when [K+]o was > or = 3.9 mM (P < 0.05). Hypothermia decreased Vmax at all [K+]o studied (P < 0.05). Regardless of the temperature, Vmax was not affected by verapamil when [K+]o was < or = 3.9 mM, but at 6.8 mM [K+]o in hypothermia Vmax was significantly lower in the presence of verapamil. Hypothermia increased both the APD50 and the APD95. The effects of verapamil on APD were temperature and [K+]o dependent; between 37 degrees C and 28 degrees C with 2.3 mM [K+]o in the superfusate, verapamil did not affect APD. At 28 degrees C in the presence of verapamil, the APD50 and APD95 decreased only if the [K+]o was > or = 3.9 mM. Verapamil and K+ supplementation in hypothermia may exert an antiarrhythmic effect, primarily by reducing the dispersion fo prolonged APD.

771-2 Isoproterenol Amplification of Flecainide-induced Conduction Slowing in Depolarized Canine Purkinje Fibers: Elucidation of Flecainide Proarrhythmia in CAST

To elucidate the potential mechanism for the flecainide (FLEC) proarrhythmia observed in CAST, the voltage-dependence of β -adrenergic modulation of impulse propagation in 10 FLEC-superfused canine Purkinje fibers was examined using a dual microelectrode technique. At physiologic membrane potentials (V m ) ([K + ]&z.omicr; = 5.4 mM), 1 μ M FLEC decreased V max from 677 ± 72 to 576 ± 97 V/sec (p = 0.04) and squared conduction velocity (θ 2 ) from 1.28 ± 0.5 to 1.04 ± 0.4 (M/s) 2 . With K+ depolarization to V m = -70 mV, FLEC further reduced V max from 324 ± 100 to 241 ± 64 V/s and θ 2 from 0.90 ± 0.3 to 0.83 ± 0.5 (M/s) 2 and produced a 1.9 mV hyperpolarizing shift of apparent Na+ channel availability curves derived from θ 2 . The addition of 1 μM isoproterenol (ISO) to FLEC-superfused fibers at physiologic V m increased θ 2 by 10% to 1.14 ± 0.5 (M/s) 2 (p = 0.009) without altering V max . At -70 mV, the addition of ISO magnified the FLEC-induced reduction of V max an additional 26% to 177 ± 49 V/s (p = 0.001) and θ 2 by 20% to 0.67 ± 0.4 (M/s) 2 (p = 0.008), producing an additional 1.8 mV (p = 0.001) and 1.9 mV (p = 0.001) hyperpolarizing shift in the apparent Na +channel inactivation curves generated from V max and θ 2 , respectively. At physiologic V m , the action potential duration (APD95) was reduced from 308 ± 32 to 266 ± 25 (p &lt; 0.001) by FLEC and subsequently to 217 ± 4 (p &lt; 0.001) with ISO addition. With 12 mM K + , APD95 decreased from 193 ± 24 to 179 ± 20 ms (p = 0.001) with FLEC and to 162 ± 14 ms (p = 0.001) with ISO. Thus. at depolarized V m , ISO amplified the FLEC-induced reduction of V max and θ 2 , suggesting a further adrenergic-mediated reduction of Na+ current. Consequently, the synergy between catecholamines and FLEC at depolarized Vm and the shortened APD 95 could facilitate double-wave reentrant arrhythmias in the presence of FLEC and superimposed ischemia.

771-3 Direct Characterization of F1ecalnide Binding Rates from Use-Dependent Conduction Delay In Canine Purkinje Fibers

To quantitatively characterize flecainide-induced channel blockade from usedependent conduction delay (CD), 12 canine purkinje fibers were studied using a dual microelectrode technique. During 60 sec of pacing at interstimulus intervals (ISI) of 1.25–0.4 sec with 2 μ M flecainide (FLEC), incremental CD followed a monoexponential time course, the rates of which were linearly related to the interpulse recovery interval (t r = ISI — action potential duration). Steady state block was an exponential function of the recovery rates. Use-dependent block derived from incremental CD and decremental squared conduction velocity ( θ 2 ) was characterized by the forward (k) and reverse (I) rate constants for the activated (a) and resting (r) states: k a (× 10 6 ) (mol -1 s -1 ) l a (s -1 ) k r (× 10 2 ) (mol -1 s -1 ) I r (s -1 ) CD 7.0 ± 2.6 12.0 ± 4.4 0.6 ± 1.7 4.01 ± 1.63 θ 2 10.0 ± 3.4 14.7 ± 2.5 2.8 ± 5.7 3.66 ± 1.40 V max(prox) 6.8 ± 2.3 15.9 ± 5.0 5.1 ± 10.3 4.22 ± 1.11 These rates reflect marked open state Na+ channel block and closed channel trapping at resting membrane potentials with FLEC. The addition of 1 μ M isoproterenol (ISO) to FLEC-superfused fibers reversed the FLEC-induced reduction of θ 2 from 1.79 ± 0.7 to 1.89 ± 0.89 (m/s) 2 (p = 0.017) without changing V ˙ max . The rate constants for FLEC binding and unbinding were not altered by ISO. Thus FLEC's apparent binding rates can be quantified from its use-{jependent effects on conduction. Both ISO's selective reversal of FLEC effect on θ 2 but not V ˙ max and the absence of changes in the rate constants suggest that the modulation of FLEC effect is due to an alteration in passive membrane properties. These characterizations will facilitate subsequent comparisons of FLEC interactions in pathologic and hyperadrenergic states in vivo .

Modulation of the Electrical Restitution of Canine Purkinje Fibers by Local Anesthetic Drugs: A Study with Flecainide and Propafenone

Some antiarrhythmic drugs that depress conduction also influence action potential duration (APD). This could modify the time course of changes in the APD of premature stimuli (electrical restitution) and affect dispersion of repolarization and homogeneity of refractoriness. The effects of two potent local anesthetic drugs, i.e., flecainide and propafenone, on electrical restitution were studied in canine Purkinje fibers, superfused with Tyrode's solution and impaled with glass microelectrodes. APD was measured at 90% repolarization (APD90): fibers were stimulated for 3 minutes at cycle lengths (CLs) between 350 msec and 1.5 seconds, and kept quiescent for 5 minutes in between. For each run we calculated the percent ratio of the second APD (APD test = APDt) to the first APD after quiescence (APDo) (APDt/APDo x 100). The ratio was correlated with the CL of the run (i.e., the coupling interval between APDo and APDt = CI) by the monoexponential function APDt/APDo x 100 = 100-exp(-tau/CI), whose time constant tau indicates the speed of electrical restitution. At 1 microM, flecainide decreased tau by 26% +/- 4% (to 310 +/- 41 msec from 245 +/- 30 msec, n = 6, P < 0.05), while propafenone did not change it. Thus, unlike propafenone, flecainide slows the process of electrical restitution in Purkinje fibers. This may derive from the drug's action on currents other than Na current (i.e., IK), relevant to the duration of action potential. In a reentrant circuit, despite similar effects on conduction, APD and refractoriness of the first beat would be differently modulated by flecainide and propafenone.

Electrophysiologic effects of alprafenone on canine cardiac tissue.

We studied the actions of the propafenone derivative alprafenone on transmembrane action potentials (APs) of canine Purkinje fibers (PF) and ventricular and atrial muscle. In PF with normal maximum diastolic potentials (MDPs), alprafenone (5 x 10(-8)-1 x 10(-6) M) produced concentration-dependent decreases in AP amplitude (APA), Vmax, AP duration (APD), and conduction velocity. The effects on Vmax were use dependent. The decrease in PF APD was the result of the drug's action on the slope of phase 2. In contrast to its effect on PF, alprafenone induced a significant increase in APD in ventricular epicardial and endocardial muscle and had no effect on atrial APD. In PF, alprafenone prolonged the effective refractory period (ERP) with respect to APD, and at high [K+]0, prolonged refractoriness beyond repolarization. Alprafenone decreased Vmax and slow-response APA. It had no effect on normal automaticity or isoproterenol (ISO)-induced automaticity, but significantly suppressed BaCl2-induced abnormal automaticity at low levels of membrane potential. Alprafenone inhibited both early and delayed afterdepolarizations and eliminated triggered activity. We conclude that alprafenone's range of actions is consistent with that of other effective antiarrhythmic drugs and is very similar to that of propafenone.

Hypothermia modifies anesthetic effect on contractile force and Ca2+ transients in cardiac Purkinje fibers.

Mild hypothermia enhances cardiac contractility, and volatile anesthetics depress contractility. Contractile force (tension) and Ca2+ transients were measured in canine Purkinje fibers at 35 and 25 degrees C with and without halothane and isoflurane to examine how anesthetics attenuate the positive inotropic effect of mild hypothermia. Isometric tension and light emitted from the photoprotein aequorin were used to assess contractility and intracellular Ca2+ transients in fibers stimulated at 40-60 pulses/min. At 35 degrees C, each anesthetic depressed peak tension and peak Ca2+ transients and decreased contractile force duration but, for halothane, increased Ca2+ transient duration. Decreases in tension by both anesthetics at 35 degrees C were converted to marked increases in tension at 25 degrees C, whereas Ca2+ transients were little changed. Removal of anesthetics at 25 degrees C greatly increased tension with a small increase in Ca2+ transients that was much lower than that at 35 degrees C. The curve relating peak contractile force as a function of Ca2+ transients at 25 degrees C during stepwise increases in extracellular CaCl2 was shifted steeper and leftward of the curve at 35 degrees C. These studies suggest that the positive inotropic effect of mild hypothermia is due primarily to increased myofibrillar Ca2+ sensitivity and that anesthetics decrease tension during hypothermia by decreasing myofibrillar Ca2+ sensitivity. Reduced influx of transsarcolemmal or sarcoplasmic reticular Ca2+ may also play a role during mild hypothermia.

Role of junctional zone cells between Purkinje fibres and ventricular muscle in arrhythmogenesis.

The aim was to examine under abnormal conditions the flow of currents across junctional (J-) cells found between Purkinje fibre cells and ventricular muscle cells, and determine whether these currents play a role in increasing the excitation rate of ventricular muscle. Canine Purkinje fibre and papillary muscle preparations were mapped to locate the Purkinje fibre-ventricular muscle cell junction (PVJ). Using standard techniques, action potentials from Purkinje fibres, J-cells, and ventricular muscle cells and extracellular electrograms were simultaneously recorded at the PVJ. The tissue was then superfused with Tyrode solution plus 4.5-5.0 mmol of ethylenediamine tetra-acetate (EDTA). EDTA prolonged the action potential duration mainly in Purkinje fibres. Secondary plateaus were recorded at membrane potentials of -63.5(SD 7.6) mV (n = 16) in J-cells, and at membrane potentials of -74.9(4.3) mV (n = 9) in ventricular muscle cells. Triggered activations appeared on both secondary plateaus with the earliest site of activation at J-cells (n = 12), at ventricular muscle cells (n = 4), or in both (n = 6). Tetrodotoxin (3-9 x 10(-7) M) and verapamil (1 x 10(-6)-10(-5) M) suppressed triggered activations. The PVJ zone appears to be an important site for the generation of triggered activations. Interventions suggest that triggered activations originating in the J-cell depend on delayed repolarisations which trigger the activation of sodium and/or calcium channels.

Effect of halothane on in vivo and in vitro cardiotoxicity of an aminocardenolide.

Halothane opposes cardiotoxicity of neutral-sugar digitalis compounds in intact animals, presumably by depressing a sympathetic component of arrhythmogenesis. However, halothane also produces a dose-related reduction in arrhythmogenicity of ouabain in isolated canine Purkinje fibers, suggesting that the anesthetic may oppose direct mechanisms of cardiotoxicity as well. The present study examined in vivo and in vitro the effect of halothane on the arrhythmogenicity of ASI-222 (3-beta-O[4-amino-4-6-dideoxy-beta-D-galactopyranosyl] digitoxigen in HCl), a highly polar aminocardenolide with no sympathetic component to cardiotoxicity. For in vivo studies, ASI-222 was infused at a rate of 1 microgram/kg/min until appearance of third-degree atrioventricular (AV) block or sustained ventricular arrhythmias in 5 conscious (control) and 6 halothane-anesthetized (1.4% end-tidal) dogs. For in vitro studies, standard microelectrode techniques were used to measure action potentials (AP) in seven excised canine Purkinje fibers superfused with oxygenated Krebs-Henseleit buffer. AP were recorded during control superfusion, after induction of toxicity with 10(-7) M ASI-222, and during exposure to 0.5, 1.0, and 2.0% halothane. Purkinje fibers were paced at 500-ms cycle lengths (CL) for 20 beats, and the amplitude of delayed afterdepolarizations (DAD) were recorded. Pacing at 250 ms CL was used to trigger ectopy. In vivo studies showed no difference in the cardiotoxic dose of ASI-222 between control dogs and those anesthetized with 1.4% halothane. However, in 4 of 6 anesthetized dogs, acutely increasing the inspired halothane concentration suppressed arrhythmias once end-tidal concentration were >2.2%.(ABSTRACT TRUNCATED AT 250 WORDS)

Beta-adrenergic regulation of action potentials and automaticity in young and adult canine purkinje fibers

To investigate developmental changes in the cellular electrophysiological effects of beta 1- and beta 2-adrenoceptor stimulation on canine Purkinje fibers (PF), we studied the effects of isoproterenol, a nonselective beta-agonist, and salbutamol, a preponderantly beta 2-agonist, alone or in presence of the selective beta 1-antagonist CGP-20712A or the beta 2-antagonist ICI-118551. Standard microelectrode techniques were used to study adult and neonatal (< 11 days) PF paced at a cycle length of 1 s or allowed to beat spontaneously. In paced adult PF, isoproterenol significantly increased the maximum diastolic potential and significantly decreased action potential duration at 60 and 90% of full repolarization (APD60 and APD90) in a concentration-dependent fashion. These effects were not observed in neonatal PF, which instead manifested a significant increase in phase 2 amplitude and APD30 that was not observed in adult PF. All these effects occurred as well with salbutamol but were less pronounced and required higher agonist concentrations. Isoproterenol decreased the automatic cycle length of adult fibers from 4,079 +/- 796 ms during control to 2,190 +/- 229 ms at 10(-5) M (P < 0.05) and from 1,535 +/- 105 to 1,249 +/- 90 ms in neonatal PF (P < 0.05). In both adults and neonates, > 90% of this effect was reached at a concentration of 10(-8) M. Salbutamol had the same effect but required higher concentrations. In both adults and neonates, the beta 2-antagonist ICI-118551 did not modify any of the effects of isoproterenol and salbutamol, whereas the beta 1-antagonist CGP-20712A significantly antagonized them. In conclusion, 1) the effects of beta-adrenergic stimulation on transmembrane potentials of canine PF change during development, both qualitatively (in paced PF) and quantitatively (in automatic PF) and 2) the responses seen are attributable to the activation of beta 1- and not beta 2-adrenoceptors.

Quinidine but Not Disopyramide Prolongs Cardiac Purkinje Fiber Action Potentials After a Pause

Prolongation of the cardiac action potential (AP), leading eventually to early afterdepolarizations (EADs), is believed to underlie drug-induced long QT syndromes and torsade de pointes. Episodes of torsade de pointes frequently occur after a prolonged pause. We studied the effects of quinidine and disopyramide on AP duration (APD) in canine cardiac Purkinje fibers after pauses of 2,000-10,000 ms. Standard intracellular microelectrode techniques were used to record APs from canine Purkinje fibers at an interstimulus interval (ISI) of 1,000 ms. Pauses of 2,000-10,000 ms were introduced into the basic drive cycle in the presence and absence of subtherapeutic and therapeutic concentrations of quinidine and disopyramide. We observed a biphasic response in APD to quinidine and disopyramide at ISI = 1,000 ms. Quinidine but not disopyramide produced a marked dose- and time-dependent additional prolongation of APD immediately after the pauses. This effect was highly statistically significant. We conclude that disopyramide and quinidine have qualitatively different effects on APD after a pause and that this observation may cast some light on the apparently greater frequency of torsade de pointes occurring with quinidine than with disopyramide. Possible mechanisms include differential drug effects on outward potassium or inward sodium channels.

Direct quantification of apparent binding indices from quinidine-induced in vivo conduction delay in canine myocardium.

To characterize quantitatively the quinidine (QUIN)-induced conduction delay (CD) in vivo, canine ventricular activation times were examined with an epicardial mapping technique. A high-resolution index of normalized (N) QUIN CD, derived from all 56 recording sites, was used to quantify QUIN effect. Repetitive stimulation elicited monoexponential increases in CD(N), the rates of which were a linear function of interpulse recovery interval, tr. Steady-state CD(N) was also linearly related to an exponential function of tr and drug uptake rates. The frequency-dependent properties of QUIN in 14 dogs were characterized by apparent binding and unbinding rates of ka = 7.1 +/- 3.5 x 10(6) M-1 s-1, la = 81 +/- 51 s-1 for activated, and kr = 12.6 +/- 11.3 x 10(3) M-1 s-1, lr = 0.51 +/- 0.26 s-1 for resting states. ka and la were similar to values previously derived in canine Purkinje fibers. Drug unbinding at resting potentials was faster in vivo than previously observed in vitro. The time constant of recovery from QUIN block extracted from the interpulse recovery rate was also identical to that determined from post-mature stimulus diastolic scanning. As predicted by the two-state model, similar binding rates were also derived from declining CD(N) elicited by step decreases in heart rate. These findings represent a complete quantitative description of use-dependent QUIN CD in vivo and provide a firm foundation for characterizing antiarrhythmic drug action under physiologic and pathologic conditions.

Alpha1‐Adrenergic Receptor Modulation of Repolarization in Canine Purkinje Fibers

Alpha 1-adrenergic receptor stimulation increases contractility and prolongs repolarization. These effects are modulated by alpha 1-adrenergic receptor-mediated inhibition of transsarcolemmal potassium currents. We used standard microelectrode techniques to study the actions of 4-aminopyridine (4-AP), which blocks the transient outward current, I(to), and WAY-123,398, which blocks the delayed rectifier, Ik, on canine Purkinje fiber action potential prolongation induced by phenylephrine. At a basic cycle length of 1 second, phenylephrine (0.1 to 10 microM) dose-dependently prolonged action potential duration at 90% repolarization (APD90) from 331 +/- 10 msec to 400 +/- 12 msec (P < 0.05) at phenylephrine, 10 microM. Phenylephrine did not change phase 1 or plateau height. 4-AP (0.1 mM) decreased phase 1 magnitude, shifted plateau height to more positive potentials (from 0.1 +/- 1.8 mV to 14.3 +/- 1.1 mV [P < 0.05]), and shortened APD90 from 318 +/- 9 msec to 294 +/- 8 msec (P < 0.05). 4-AP did not block phenylephrine effects on APD90, which increased, at 10 microM phenylephrine, from 294 +/- 8 msec to 342 +/- 6 msec (P < 0.05). In contrast, WAY-123,398 (0.1 microM) prolonged APD90 from 360 +/- 6 msec to 452 +/- 6 msec (P < 0.05), and had no effect on plateau height. In the presence of WAY-123,398, phenylephrine no longer increased APD90. (1) Agents that block I(to) shorten APD in Purkinje fibers; and (2) the alpha-agonist mediated increase of APD in canine Purkinje fibers can be explained by inhibition of Ik.

Developmental electrophysiology of encainide and its major metabolites on the Purkinje fiber action potential.

With clinical data suggesting that neonates may be more prone to developing electrophysiologic side effects from encainide, this study investigates the in vitro developmental electrophysiologic effects of encainide and its major metabolites on the action potential parameters of the canine cardiac Purkinje fiber. With standard microelectrode techniques, the in vitro tonic and rate-related effects of encainide, and its major metabolites (3-methoxy-4-hydroxy encainide, MODE, and O-dimethyl encainide, ODE) were investigated using mature and immature canine cardiac Purkinje fibers. The significant developmental differences in the effects of these compounds on the canine Purkinje fiber illustrated in this study are: (1) 1 x 10(-6) M encainide depresses Vmax in neonatal Purkinje fibers, yet not in the adult. (2) 1 x 10(-6) M MODE lengthens APD90 in the neonate, yet it has no substantial effect in the adult. (3) 1 x 10(-6) M ODE shortens APD90 in the adult, yet it has no appreciable effect on the neonate. (4) Rate-related effects of encainide and ODE are more pronounced in adult Purkinje fibers. In contrast to other in vitro studies on class I antiarrhythmic agents, neonatal canine Purkinje fibers seem to be more sensitive than the adult to the tonic depolarization depressant effect of encainide. This in vitro sensitivity parallels clinical experience with the drug in neonatal patients. Although encainide is no longer available for clinical use, these findings highlight the fact that the immature conduction system may show markedly different sensitivities to different class I agents despite the fact that these agents share similar qualitative pharmacologic properties.

Actions of vasoactive intestinal peptide and neuropeptide Y on the pacemaker current in canine Purkinje fibers.

We have investigated the actions of vasoactive intestinal peptide (VIP) and neuropeptide Y (NPY) on the pacemaker current (I(f)) in canine Purkinje fibers. On voltage pulses to the middle of the I(f) activation range, VIP reversibly increases I(f), whereas NPY reversibly decreases I(f). A three-pulse voltage protocol suggests that VIP shifts I(f) activation in the positive direction and that NPY shifts I(f) activation in the negative direction on the voltage axis without changing maximal I(f) conductance. These effects of VIP and NPY on I(f) are exerted through their specific peptide receptors, since the effects are blocked by VIP and NPY receptor antagonists. VIP and NPY are colocalized in cardiac parasympathetic and sympathetic nerve endings, respectively, and can be released preferentially on high and long-lasting nerve stimulation. Given this colocalization and frequency-dependent release, these results suggest a role for these neuropeptides in controlling cardiac I(f) and consequently heart rate.

Interaction of Anesthetics and Catecholamines on Conduction in the Canine His--Purkinje System

The findings in papillary muscles that epinephrine facilitates conduction at Purkinje fiber-muscle junctions and in the endocardium are consistent with older observations that activation of myocardial beta-adrenergic receptors speeds conduction and activation in the heart and thereby increases the synergy of contraction (46,47). The cellular mechanism underlying this action is probably increased cell-to-cell coupling between muscle fibers secondary to elevation of cyclic AMP (19,48). However, the findings that epinephrine alone or with halothane transiently slows conduction in the Purkinje layer while simultaneously improving conduction across Purkinje-muscle junctions and in the endocardium may represent proarrhythmic actions. These actions could facilitate arrhythmogenesis by transiently increasing regional differences of activation and repolarization times in the conduction system and myocardium and thereby increasing vulnerability to induction of reentry by premature impulses. Such a proarrhythmic effect could explain an older observation that low-dose norepinephrine infusions decrease the threshold for induction of fibrillation by two premature beats in pentobarbital-anesthetized animals (49). The cellular basis underlying the different responses of Purkinje fibers and the endocardial muscle layer to catecholamines, in which velocity decreased and increased, respectively, is not known. Our working hypothesis to explain this action in canine Purkinje fibers is a mechanism involving activation of WB4101-sensitive alpha 1-adrenoceptor, G-protein coupling to phospholipase C and the generation of DAG and IP3 leading to modulation of cell-to-cell coupling, which is potentiated in the presence of partial uncoupling by halothane. The different responses of Purkinje and myocardial fibers are speculated to result from differences in the relative density of this subtype of alpha 1-adrenoceptor, differences in the subcellular effector coupling mechanisms, or differences in the specific connexin proteins forming gap junctions between Purkinje and myocardial fibers (50).

Early after-depolarizations and torsade de pointes: implications for the control of cardiac arrhythmias by prolonging repolarization.

Common clinical features in drug-induced torsade de pointes include hypokalemia and cycle-length prolongation just prior to initiation of the arrhythmia. In canine Purkinje fibres, drugs known to be associated with torsade de pointes, such as quinidine, sotalol or N-acetylprocainamide, consistently produce early after-depolarizations (EADs) and triggered activity at slow drive rates; for quinidine, these abnormalities are exaggerated by low extracellular potassium. Triggered activity can be abolished in vitro in two ways. First, action-potential shortening with abolition of EADs can be accomplished by increasing stimulation rates, beta-stimulation and action-potential shortening antiarrhythmics such as lidocaine. Second, triggered activity can be suppressed, with less prominent effects on EADs, by magnesium, alpha- and/or beta-adrenergic blockade and calcium-channel blockers. The parallels between these in vitro findings and clinical torsade de pointes suggest that EADs and triggered activity play a role in the genesis of the clinical arrhythmia. Further research directed at determining the mechanisms underlying the cellular abnormalities and their propagation to the whole heart should yield information that will increase the safety of antiarrhythmic therapy.