Beat dependent alteration of Ca2+-activated Cl- current during rapid stimulation in rabbit ventricular myocytes.

The transient outward currents (Ito) play an important role in action potential repolarization in cardiac myocytes. Two components of Ito have been identified as 4-AP-sensitive but Ca2+-insensitive Ito carried by K, and Ca2+-sensitive but 4-AP insensitive Ito carried by Cl- (I(Cl(Ca))). It is known that the amplitudes of Ito change depending on the stimulation frequency. In this study we investigated the beat dependent alteration of I(Cl(Ca)) during rapid stimulation using the whole cell patch clamp technique in rabbit ventricular myocytes. The cells were internally perfused with a solution containing 0.1 microM free Ca2+ to develop I(Cl(Ca)) and all internal K+ was replaced with Cs+ to block 4-AP-sensitive Ito and other K+ currents. By applying depolarizing pulses at a high frequency of 2.5 Hz, the amplitudes of I(Cl(Ca)) gradually increased as the number of pulses increased following a transient decrease in the 2nd pulse and reached a plateau level at the 20th pulse. The shape of the current-voltage curve of I(Cl(Ca)) was not overly different for different numbers of preceding pulses. The recovery from inactivation of I(Cl(Ca)) could be fitted to a single exponential curve and full recovery was achieved after > 1 sec with a time constant of 368 ms. The ramp clamp experiments showed that the conductance of the background I(Cl(Ca)) increased with the preceding pulse numbers, indicating that the resting level of [Ca2]i increased with the pulses applied. From these results, we conclude that beat dependent alteration of I(Cl(Ca)) is determined by not only its apparent kinetic property, but also the resting level of [Ca2+]i during rapid stimulation.

A hyperpolarization- and acid-activated nonselective cation current in Xenopus oocytes.

Heterologous expression of a variety of membrane proteins in Xenopus oocytes sometimes results in the appearance of a hyperpolarization-activated inward current. The nature of this current remains incompletely understood. Some investigators have suggested that this current is a Cl current, whereas others have identified it as a nonselective cation current. The purpose of this investigation was to characterize this current in more detail. The hyperpolarization-activated inward current (I(IN)) present in native oocytes was composed of a current carried at least partly by Ca and Mg under physiological ionic conditions plus a Ca-activated Cl current. The Ca-activated Cl current was blocked by chelation of cytosolic Ca with 1,2-bis(2-aminophenoxy)ethane-N,N,N', N'-tetraacetic acid. When Cl currents were blocked, the cation current could be carried by Ca, Mg, or Co, but not appreciably by Ba, Mn, or Cd. I(IN) was stimulated by intracellular acidification. The properties of I(IN) were quite different from those of the store-operated Ca current. Heterologous expression of transient receptor potential-like gene product (TRPL), one of the members of the transient receptor potential family of putative store-operated Ca channels, apparently resulted in alteration of the voltage sensitivity of the endogenous I(IN).

Pharmacokinetics of landiolol hydrochloride, a new ultra-short-acting beta-blocker, in patients with cardiac arrhythmias.

OBJECTIVES: To elucidate pharmacokinetics and pharmacodynamics of landiolol hydrochloride, newer developed ultra-short-acting beta-blocker, in patients with various cardiac tachyarrhythmias. BACKGROUND: The short duration of action and titratability of landiolol hydrochloride make it ideal for use in patients with a clinical need for beta-blockers. METHODS: In a total of 31 examinations we infused the drug in 19 patients (mean age, 55 +/- 14 years). After the persistence of the tachyarrhythmias was confirmed, continuous infusion was started at rates of 0.005, 0.01, 0.02, 0.04, and 0.08 mg/kg/min for 5 minutes (for paroxysmal atrial fibrillation, paroxysmal supraventricular tachycardia, and ventricular tachycardia) or 15 minutes (for ventricular premature complex). We analyzed the pharmacokinetics of 16 examinations. A one-compartment model provided a close fit for each blood concentration-time curve. RESULTS: The maximum blood concentrations obtained clearly showed the dose dependency and revealed very short half-lives (range, 2.3 to 4.0 minutes). Area under the blood concentration-time curves also increased, showing dose dependency. In paroxysmal atrial fibrillation, landiolol hydrochloride reduced the heart rate from 111 +/- 20 to 90 +/- 10/min. Sinus rhythm was restored, without any adverse effects, in three of five patients with paroxysmal supraventricular tachycardia and one patient with ventricular tachycardia. There was no significant change in peripheral blood pressure. CONCLUSIONS: Landiolol hydrochloride has a shorter elimination half-life than any other beta-blocker, and it can be administered safely to patients with various tachyarrhythmias.

Bimodal control of a Ca(2+)-activated Cl(-) channel by different Ca(2+) signals.

Ca(2+)-activated Cl(-) channels play important roles in a variety of physiological processes, including epithelial secretion, maintenance of smooth muscle tone, and repolarization of the cardiac action potential. It remains unclear, however, exactly how these channels are controlled by Ca(2+) and voltage. Excised inside-out patches containing many Ca(2+)-activated Cl(-) channels from Xenopus oocytes were used to study channel regulation. The currents were mediated by a single type of Cl(-) channel that exhibited an anionic selectivity of I(-) > Br(-) > Cl(-) (3.6:1.9:1.0), irrespective of the direction of the current flow or [Ca(2+)]. However, depending on the amplitude of the Ca(2+) signal, this channel exhibited qualitatively different behaviors. At [Ca(2+)] < 1 microM, the currents activated slowly upon depolarization and deactivated upon hyperpolarization and the steady state current-voltage relationship was strongly outwardly rectifying. At higher [Ca(2+)], the currents did not rectify and were time independent. This difference in behavior at different [Ca(2+)] was explained by an apparent voltage-dependent Ca(2+) sensitivity of the channel. At +120 mV, the EC(50) for channel activation by Ca(2+) was approximately fourfold less than at -120 mV (0.9 vs. 4 microM). Thus, at [Ca(2+)] < 1 microM, inward current was smaller than outward current and the currents were time dependent as a consequence of voltage-dependent changes in Ca(2+) binding. The voltage-dependent Ca(2+) sensitivity was explained by a kinetic gating scheme in which channel activation was Ca(2+) dependent and channel closing was voltage sensitive. This scheme was supported by the observation that deactivation time constants of currents produced by rapid Ca(2+) concentration jumps were voltage sensitive, but that the activation time constants were Ca(2+) sensitive. The deactivation time constants increased linearly with the log of membrane potential. The qualitatively different behaviors of this channel in response to different Ca(2+) concentrations adds a new dimension to Ca(2+) signaling: the same channel can mediate either excitatory or inhibitory responses, depending on the amplitude of the cellular Ca(2+) signal.

Anion permeability and conduction of adenine nucleotides through a chloride channel in cardiac sarcoplasmic reticulum.

Cardiac sarcoplasmic reticulum (SR) membrane contains several chloride (Cl-) channels. We have characterized a 116-pS Cl- channel (500 mM cis, 50 mM trans Cl-) in cardiac SR that is activated by protein kinase A-dependent phosphorylation. To understand its function further, we examined the permeation of various anions and adenine nucleotides using the planar lipid bilayer-vesicle fusion technique. This Cl- channel showed a high selectivity to anions and its permeability sequence was Br- > Cl- > I- > NO3- > F-. When all anions were replaced with ATP in the cis solution, channel activity persisted. The conductance was 78 pS with 200 mM ATP and 68 pS with 100 mM ATP. The reversal potentials were +63 mV and +41 mV in 200 mM ATP and in 100 mM ATP, respectively. With 100 mM ADP or AMP in the cis solution, channel activities were also observed. The conductances were 87 pS with 100 mM ADP and 115 pS with 100 mM AMP. The apparent adenine selectivity of this channel was ATP > ADP > AMP, assuming that they exist as divalent anions. These results suggest that the SR Cl- channel in cardiac cells may serve as a transporter for the movement of adenine nucleotides between cytosol and SR lumen.

Dynamics of calcium regulation of chloride currents in Xenopus oocytes.

Ca-activated Cl currents are widely expressed in many cell types and play diverse and important physiological roles. The Xenopus oocyte is a good model system for studying the regulation of these currents. We previously showed that inositol 1,4,5-trisphosphate (IP3) injection into Xenopus oocytes rapidly elicits a noninactivating outward Cl current (ICl1-S) followed several minutes later by the development of slow inward (ICl2) and transient outward (ICl1-T) Cl currents. In this paper, we investigate whether these three currents are mediated by the same or different Cl channels. Outward Cl currents were more sensitive to Ca than inward Cl currents, as shown by injection of different amounts of Ca or by Ca influx through a heterologously expressed ligand-gated Ca channel, the ionotropic glutamate receptor iGluR3. These data could be explained by two channels with different Ca affinities or one channel with a higher Ca affinity at depolarized potentials. To distinguish between these possibilities, we determined the anion selectivity of the three currents. The anion selectivity sequences for the three currents were the same (I > Br > Cl), but ICl1-S had an I-to-Cl permeability ratio more than twofold smaller than the other two currents. The different anion selectivities and instantaneous current-voltage relationships were consistent with at least two different channels mediating these currents. However, after consideration of possible errors, the hypothesis that a single type of Cl channel underlies the complex waveforms of the three different macroscopic Ca-activated Cl currents in Xenopus oocytes remains a viable alternative.

Activation of Ca2+-sensitive Cl- current by reverse mode Na+/Ca2+ exchange in rabbit ventricular myocytes.

We investigated how Ca2+-sensitive transient outward current, Ito(Ca), is activated in rabbit ventricular myocytes in the presence of intracellular Na+ (Na+i) using the whole-cell patch-clamp technique at 36 degreesC. In cells dialysed with Na+-free solutions, the application of nicardipine (5 microM) to block L-type Ca2+ current (ICa) completely inhibited Ito(Ca). In cells dialysed with a [Na+]i>/=5 mM, however, Ito(Ca) could be observed after blockade of ICa, indicating the activity of an ICa-independent component. The amplitude of ICa-independent Ito(Ca) increased with voltage in a [Na+]i-dependent manner. The block of Ca2+ release from the sarcoplasmic reticulum by caffeine, ryanodine or thapsigargin blocked ICa-independent Ito(Ca). In Ca2+-free bath solution Ito(Ca) was completely abolished. The application of 2 mM Ni2+ or the newly synthesized compound KBR7943, a selective blocker of the reverse mode of Na+/Ca2+ exchange, or perfusion with pipette solution containing XIP (10 microM), a selective blocker of the exchanger, blocked ICa-independent Ito(Ca). From these results we conclude that, in the presence of Na+i, Ito(Ca) can be activated via Ca2+-induced Ca2+ release triggered by Na+/Ca2+ exchange operating in the reverse mode after blockade of ICa.

Dual regulation of the skeletal muscle ryanodine receptor by triadin and calsequestrin.

Triadin, a calsequestrin-anchoring transmembrane protein of the sarcoplasmic reticulum (SR), was successfully purified from the heavy fraction of SR (HSR) of rabbit skeletal muscle with an anti-triadin immunoaffinity column. Since depletion of triadin from solubilized HSR with the column increased the [3H]ryanodine binding activity, we tested a possibility of triadin for a negative regulator of the ryanodine receptor/Ca2+ release channel (RyR). Purified triadin not only inhibited [3H]ryanodine binding to the solubilized HSR but also reduced openings of purified RyR incorporated into the planar lipid bilayers. On the other hand, calsequestrin, an endogenous activator of RyR [Kawasaki and Kasai (1994) Biochem. Biophys. Res. Commun. 199, 1120-1127; Ohkura et al. (1995) Can. J. Physiol. Pharmacol. 73, 1181-1185] potentiated [3H]ryanodine binding to the solubilized HSR. Ca2+ dependency of [3H]ryanodine binding to the solubilized HSR was reduced by triadin, whereas that was enhanced by calsequestrin. Interestingly, [3H]ryanodine binding to the solubilized HSR potentiated by calsequestrin was reduced by triadin. Immunostaining with anti-triadin antibody proved that calsequestrin inhibited the formation of oligomeric structure of triadin. These results suggest that triadin inhibits the RyR activity and that RyR is regulated by both triadin and calsequestrin, probably through an interaction between them. In this paper, triadin has been first demonstrated to have an inhibitory role in the regulatory mechanism of the RyR.

Electrophysiologic and hemodynamic effects of a single oral dose of pilsicainide hydrochloride, a new class 1c antiarrhythmic agent.

To establish the clinical efficacy of pilsicainide, we evaluated its electrophysiologic and hemodynamic effects after a single oral administration to 18 patients with documented supraventricular tachycardia (SVT). To determine the minimal effective blood level, changes in efficacy with time were evaluated by serial reinduction studies with venous blood sampling for measurement of the plasma pilsicainide level. Sixty minutes after administration of a single oral dose of pilsicainide, the sinoatrial conduction time, AH and HV intervals, and the effective refractory period of the right ventricle were prolonged. Ventriculoatrial conduction was blocked in 11 patients [nine of 12 via accessory pathway and two of six via the atrioventricular (AV) node], resulting in the suppression of SVT induction in nine of 13 patients. Pilsicainide increased the heart rate and mean pulmonary arterial pressure and decreased the stroke volume index at 60 min. PQ interval, QRS width, and QTc were significantly prolonged after pilsicainide, and the percentage prolongations of the PQ interval were well correlated with the plasma pilsicainide levels. The plasma level effective for suppression of SVT was considered to be >0.5 microg/ml. We concluded that a single oral administration of pilsicainide is well tolerated and effective in suppressing SVT.

Clinical and electrophysiologic effects of dofetilide in patients with supraventricular tachyarrhythmias.

The clinical and electrophysiologic effect of intravenous dofetilide was evaluated in patients with paroxysmal atrial fibrillation (AF) of recent onset (< 7 days) and paroxysmal supraventricular tachycardia (PSVT). From 2.5 to 5.0 micrograms/kg of dofetilide was administered intravenously for the termination of arrhythmias. For the electrophysiologic study (EPS), 3.0 micrograms for loading and subsequently 2 micrograms/kg was injected for 45 min as a maintenance dose. The EPSs were performed before the loading and during the maintenance dose. AF was successfully converted to sinus rhythm in seven (54%) of 13 patients. The duration of AF from its onset was significantly shorter in responders than that of nonresponders (p < 0.05). Dofetilide also terminated PSVT in four of six patients. In the EPS, dofetilide proportionately lengthened the effective refractory period of the atrium, ventricle, and the accessory pathways without slowing of the intracardiac conduction. Dofetilide completely suppressed the induction of PSVT in seven of 13 patients, restricted the induction zone in five, and inhibited perpetuation of the arrhythmia in the remaining one. The cycle length of PSVT remained unchanged after dofetilide. These results imply that the suppression of the development and maintenance of reentrant arrhythmias may result from the lengthening effect of dofetilide on the refractoriness and the consequent elimination of the excitable gap at the critical part of the reentrant loop.

Purification and Some Properties of an α-Amylase from an Anaerobic Bacterium Isolated from Coastal Sediment.

A marine, obligate anaerobic bacterium, SS71, isolated from a coastal sediment, was a Gram-positive, asporogenous, rod-shaped organism with a G + C mol% of 37.3 ±0.1. This bacterium produced an α-amylase with a molecular mass of 91 kDa and an isoelectric point of 4.3. The α-amylase had an optimal pH of 7.0 and an optimal temperature of 35°C. The enzyme activity was promoted by 0.5-2.0% NaCl.

Curative percutaneous catheter ablation for various supraventricular and ventricular tachyarrhythmias. Results in 187 consecutive patients during the first five years.

Closed-chest transcatheter electrical ablation (catheter ablation) has been applied to various supraventricular and ventricular tachyarrhythmias as a radical therapeutic technique since its introduction in 1982. Currently, it has become a first line therapy for supraventricular tachyarrhythmias except atrial fibrillation and uncommon types of atrial flutter. We first carried out the ablation procedure in 1991 for the treatment of ventricular tachycardia. Up to February 1997, a total of 187 patients underwent catheter ablation in our institution. The aims of this study are to demonstrate our results of catheter ablation in the early 5 years and to show the usefulness of this new curative method. Successful results were obtained in 168 of 187 patients (overall final success rate: 89.8%). The success rates of each category of tachyarrhythmias were 100/105 patients (95%) with WPW syndrome, 41/46 (89%) with atrioventricular nodal reentrant tachycardia, 7/10 (70%) with atrial flutter, 4/4 (100%) with atrial tachycardia, 2/2 (100%) with medically refractory atrial fibrillation, 13/15 (85%) with idiopathic ventricular tachycardia and 3/7 (43%) with sustained ventricular tachycardia associated with structural heart disease, respectively. Complications that required invasive treatments were observed in 3 patients (2 hemopericardium and 1 complete atrioventricular block). Our results indicate that catheter ablation is highly effective in most categories of tachyarrhythmias and can be applied safely without lethal complications.

Clinical effects and pharmacokinetics of a single oral dose of pirmenol hydrochloride.

To establish the clinical efficacy of a single oral dose of pirmenol, we evaluated electrophysiologic and hemodynamic effects simultaneously after drug administration, performing electrophysiologic testing in 20 patients with ECG-documented paroxysmal supraventricular tachycardia (PSVT) before and after a single oral 200-mg dose of pirmenol. Hemodynamic measurements were made with a Swan-Ganz catheter in the first 10 consecutive patients. In a different series of patients, we administered a single 200-mg oral dose of pirmenol to evaluate its acute termination effect in 7 patients with PSVT and 9 with paroxysmal atrial fibrillation. Pirmenol prolonged the refractory period of the retrograde conduction system in patients with or without an accessory pathway, and supraventricular tachycardia was no longer inducible at 60 min in 11 patients [8 of 11 with atrioventricular (AV) reentrant tachycardia and 3 of 5 with AV nodal reentrant tachycardia]. Pirmenol increased the heart rate (p < 0.01) and total systemic resistance (p < 0.05), and reduced the stroke volume index (p < 0.01), all significantly. The plasma concentration of pirmenol at 1 h after administration was 0.75 +/- 0.48 microgram/ml. A single oral dose of pirmenol during tachyarrhythmia successfully restored sinus rhythm in 4 of 7 (57%) patients with PSVT and 4 of 9 (44%) patients with paroxysmal atrial fibrillation. A single oral dose of pirmenol was well tolerated as episodic treatment in patients with supraventricular tachyarrhythmias.

Use of intravenous dofetilide in atrial flutter with hemodynamic instability.

Paroxysmal atrial flutter is a drug-resistant supraventricular tachyarrhythmia that is often accompanied by hemodynamic instability due to an excessive ventricular response. We report here a case of atrial flutter with 1:1 atrioventricular conduction, in which we were able to rescue the patient from a state of cardiogenic shock by using intravenous dofetilide, a new class III antiarrhythmic agent. Dofetilide was suitable for the treatment because it immediately increased atrioventricular nodal refractoriness without any negative inotropic action.

A case of cardiac foreign bodies associated with four types of tachycardias.

A case of cardiac foreign bodies leading to development of four varieties of automatic tachycardia is reported. A 51-year-old male with aortic regurgitation was admitted to our hospital because of palpitations. An electrocardiogram revealed junctional tachycardia with or without left bundle branch block, and two types of fascicular tachycardias. Computed tomography showed metallic foreign bodies from a fractured guidewire in the membranous portion of the interventricular septum, which was inadvertently retained when he underwent diagnostic cardiac catheterization at the age of 27.