Rate-dependent blockade of a potassium current in human atrium by the antihistamine loratadine

Abstract

1. The antihistamine loratadine is widely prescribed for the treatment of symptoms associated with allergies. Although generally believed to be free of adverse cardiac effects, there are a number of recent reports suggesting that loratadine use may be associated with arrhythmias, in particular atrial arrhythmias.

2. Nothing is known regarding the potassium channel blocking properties of loratadine in human cardiac cells. Using the whole-cell patch clamp technique, the effects of loratadine on the transient outward K current (I_to), sustained current (I_sus), and current measured at −100 mV (I_K1 and I_ns), the major inward and outward potassium currents present in human atrial myocytes, were examined in order to provide a possible molecular mechanism for the observed atrial arrhythmias reported with loratadine use.

3. Loratadine rate-dependently inhibited I_to at therapeutic concentrations with 10 nM loratadine reducing I_to amplitude at a pacing rate of 2 Hz by 34.9±6.0%. In contrast, loratadine had no effect on either I_sus or current measured at −100 mV.

4. These results may provide a possible mechanism for the incidences of supraventricular arrhythmias reported with the use of loratadine.

Introduction

Antagonists to H₁-histamine receptors are commonly prescribed for the treatment of allergies, urticarial diseases, and symptomatic relief of upper respiratory infections. Recently, members of this family of compounds such as terfenadine and astemizole have been associated with rare but potentially life-threatening adverse cardiac events (Craft, 1985; Davies et al., 1989; Bishop & Gaudry, 1989; Monahan et al., 1990; Lindquist & Edwards, 1997). For instance, terfenadine use has been associated with serious ventricular arrhythmias such as torsade de pointe and sudden death (Davies et al., 1989; Monahan et al., 1990). Terfenadine and astemizole have subsequently been shown to block one or more cardiac potassium channels and it is believed this action may underlie the observed arrhythmias (Woosley et al., 1993; Salata et al., 1995; Rampe et al., 1993; Crumb et al., 1995c).

In contrast to terfenadine and astemizole, the antihistamine loratadine is generally believed to be free of adverse cardiac effects. However, there are a number of recent reports suggesting that loratadine use may be associatd with arrhythmias, in particular atrial arrhythmias (Lindquist & Edwards, 1997; Aust Adv Drug React Bull, 1996; Haria et al., 1994). In human atrium, the transient outward K current (I_to), sustained current (I_sus), and current measured at −100 mV (which consists of the inwardly rectifying K current, I_K1, and the non-selective cation current, I_ns), represent the major inward and outward K currents and thus play an important role in the resting potential, shape and duration of the human atrial action potential. Blockade of any of these potassium currents may be potentially arrhythmogenic. To date, nothing is known regarding the potassium channel blocking properties of loratadine in human cardiac cells. Therefore, the present study was undertaken to characterize the effects of loratadine on I_to, I_sus, and current measured at −100 mV in isolated human atrial myocytes in order to provide a possible molecular mechanism for the observed arrhythmias reported with loratadine use.

Methods

Human tissue was obtained, in accordance with Tulane University School of Medicine Institutional guidelines. Myocytes were isolated from specimens of human right atrial appendage obtained during surgery from hearts of five patients (ages 40–67 years) undergoing cardiopulmonary bypass. All atrial specimens were described as grossly normal at the time of excision and all patients had normal P waves on electrocardiography. Some patients had received cardioactive drugs including calcium channel blockers and digitalis. The cell isolation procedure has been described in detail in Crumb et al. (1995a).

Isolated human atrial myocytes were superfused with an ‘external' solution that consisted of (in mmol l⁻¹): 137 NaCl, 4 KCl, 1 MgCl₂, 1.8 CaCl₂, 11 Glucose, 10 HEPES; adjusted to a pH of 7.4 with NaOH. Glass pipettes were filled with an ‘internal' solution that consisted of (in mmol l⁻¹): 120 K-aspartate, 20 KCl, 4 Na-ATP, 5 EGTA, 5 HEPES; adjusted to a pH of 7.2 with KOH. Loratadine was kindly provided by Almirall Prodesfarma (Barcelona, Spain) and dissolved in 100% DMSO to make concentrated stock solutions (1 mM). Loratadine was added to the bath solution from this concentrated stock (final DMSO concentration less than 0.01%) or from another more diluted stock solution (100 μM) made in distilled, deionized water.

Experiments were performed in the presence of 200 μM Cd²⁺ to block the L-type Ca current and at a holding potential of −60 mV to inactivate the sodium current. All experiments were performed at room temperature (22–23°C) so that peak I_to could be accurately distinguished from the capacitive current. The transient outward current was measured by subtracting the amplitude of the current measured at the end of the depolarizing voltage pulse (sustained current) from peak current amplitude. The sustained current was measured at the end of a depolarizing pulse to +60 mV.

Acceptable atrial myocytes were rod-shaped and lacked any visible blebs on the surface. Currents were measured using the whole-cell variant of the patch clamp method (Hamill et al., 1981). Currents were digitized at 3–5 kHz and filtered at 1 kHz. Pipette tip resistance were approximately 1.0–2.0 MΩ when the pipettes were filled with the internal solution. Analogue capacity compensation and 40–60% series resistance (R_s) compensation was used in all experiments to yield voltage drops across uncompensated R_s of less than 3 mV. Paired and unpaired Student's t-test was used for statistical analysis. Data are presented as mean±s.e.mean.

For exponential fits of data (e.g., fits of time course of current decay and I_to recovery), a single exponential fit was accepted as the fit of choice whenever the following criteria were met: (a) the amplitude parameters obtained from the least squares fit were all of the same sign and (b) when a negative value for the AIC (asymptotic information criteria) statistic was obtained when comparing a one versus a two exponential fit (Akaike, 1974; Horn, 1987).

Results

Figure 1 shows the effects of loratadine on K⁺ currents recorded from an isolated human atrial myocyte. In the absence of drug, application of voltage pulses from −100 mV to +60 mV elicited a family of currents characteristic of human atrial myocytes. At potentials negative to −70 mV, an inward current, which reflects the combination of the inwardly rectifying potassium current (I_K1) and the inward component of a non-selective cation current (Crumb et al., 1995b), was elicited. At more depolarized potentials, a current was elicited which decays rapidly to a steady-state (see Figure 1, control). The decaying current represents the transient outward potassium current (I_to) while the current remaining after the decay of I_to, commonly referred to as the sustained current (I_sus) is distinct and consists of two currents, a potassium current designed Kv 1.5 (Wang et al., 1993) and the non-selective cation current (Crumb et al., 1995b). A reduction in the amplitude of any of these currents can alter the shape and duration of the cardiac action potential and thus may be potentially arrhythmogenic.

Figure 1.

Effects of 100 nM loratadine on potassium currents recorded from an isolated human atrial myocyte. A family of currents were elicited by the protocol shown before and after addition of 100 nM loratadine to the bath solution. Currents were elicited at a pacing rate of 0.1 Hz. The drug-sensitive currents represent control-loratadine currents. Note the lack of effect on current amplitude.

While pacing slowly (0.1 Hz), addition of 100 nM loratadine to the solution bathing the cell, a concentration associated with arrhythmias in humans (Hilbert et al., 1987), produced no noticeable change in the amplitude of either inward or outward currents. This is clearly illustrated by the lack of a definable loratadine-sensitive current (Figure 1). Similar results were observed in four additional cells.

Since I_to is sensitive to pacing rate and the actions of many ion channel blocking drugs are rate-dependent, it is possible that the effects of loratadine on I_to may be frequency-dependent. To test this hypothesis, the effects of loratadine on the timecourse of recovery of I_to were examined using a 2-pulse protocol consisting of a 500 ms conditioning pulse to +60 mV followed by a test pulse to +60 mV after a variable recovery period at the holding potential (−60 mV) (Figure 2). As illustrated in Figure 2, the timecourse of I_to recovery was significantly slower in the presence of loratadine (100 nM) compared with control and was best fit with a single exponential relationship. The time constant of recovery (τ) obtained in the presence of drug was 107.7±9.3 ms, significantly slower than in the absence of drug (63.5±5.0 ms) (n=6, P<0.05). These results suggest that at higher pacing rates a reduction in I_to amplitude might be observed in the presence of loratadine.

Figure 2.

Recovery kinetics of I_to before and after 100 nM loratadine. A plot of the I_to recovery kinetics obtained from a human atrial myocyte before and after exposure to 100 nM loratadine. Currents were elicited by the indicated protocol. Data points were fit with a single exponential function of the form −0.85 exp (−×/104.1 ms)+0.998 after loratadine and −0.92 exp (−×/59.0 ms)+1.00 before loratadine.

Figure 3 illustrates the effects of pacing rate on loratadine inhibition of I_to. To more accurately characterize the effects of loratadine on I_to, the current measured at the end of the voltage pulse was subtracted from the peak current (Crumb et al., 1995b). After a 20 s interval at the holding potential in the presence of 100 nM loratadine, application of a depolarizing voltage pulse elicited a current which was virtually identical in amplitude to that observed in the absence of drug indicating a lack of tonic block (Figure 3B). In 100 nM loratadine, tonic reduction of I_to was 4.1±1.3%, I_sus was 6.3±2.1, and the current measured at −100 mV was 6.6±4.2% (n=5). At pacing rates which simulate either a resting heart rate (1 Hz) or a heart rate achieved during moderate exercise (2 Hz), 100 nM loratadine caused a pronounced reduction in the amplitude of I_to (Figure 3B and C). Interestingly, the rate-dependent effects of loratadine were also observed at therapeutic concentrations (i.e. 10 nM) (Figure 3C). For instance, at a frequency of 1 Hz, 10 nM loratadine produced a 15.3±2.4% (n=5) reduction in I_to amplitude, not significantly different from control (8.8±2.3%, n=16) (Figure 3C). At higher pacing rates, (2 Hz), 10 nM loratadine markedly reduced I_to amplitude by 34.9±6.0% (n=5), significantly greater than that observed in the absence of loratadine (19.9±1.5%, n=16) (P<0.05) (Figure 3C). In contrast, loratadine produced only a negligibly greater blockade of I_sus when compared to control even at the highest pacing rate tested (2 Hz) (1.1±0.7% greater than control with 10 nM, 3.6±2.6% with 100 nM, 6.9±2.9% with 1 μM, n=5–6). Even at a concentration of 10 μM, loratadine produced only a small reduction in the amplitude of the current measured at −100 mV when paced at 2 Hz (8.3±3.7%, n=5).

Figure 3.

Effects of pacing rate in the presence of 100 nM loratadine on I_to recorded from an isolated human atrial myocyte. (A) Pulse protocol. (B) Reduction in amplitude of I_to in a cell paced at 1 and 2 Hz. Illustrated are the first and steady-state currents of a 30 pulse rate train. Currents were elicited by the pulse protocol in panel A. (C) Rate- and concentration-dependent reduction of I_to by loratadine. % reduction of I_to at a given pacing rate was calculated as the reduction in current amplitude of the last pulse in the rate train (30 pulses) relative to the first pulse in the rate train. Symbols are mean±s.e. (n=5–16). Asterisks indicate value is significantly different from control.

As suggested in Figure 3, the kinetics of I_to decay appeared to be faster in the presence of loratadine than in its absence. The kinetics of I_to decay were measured once a steady-state level of current reduction was achieved at either 1 or 2 Hz in the presence of loratadine and compared to that measured in the absence of loratadine (control) at the same pacing rate. The decay of I_to could be well fit with a single exponential function with values ranging between 50 and 80 ms in the absence of drug. Fits obtained from cells before and after exposure to loratadine are shown in Figure 4. As indicated, in the presence of loratadine I_to current decay tended to be faster, although this tendency did not reach significance at any concentration or pacing rate (P=0.06).

Figure 4.

Effect of loratadine on I_to decay kinetics. Examples of current traces recorded in the absence and presence of 100 nM loratadine are shown at top of figure. Currents illustrated were recorded after current amplitude had reached a steady-state and were elicited by a series of 30 pulses (320 ms) to +60 mV from a holding potential of −60 mV (pacing rate=2 Hz). For clarity, end of pulse current has been subtracted from peak current and currents have been superimposed. Plots of decay time constants measured from cells before and after exposure to either 10 or 100 nM loratadine are also shown. Currents were fit with a single exponential function.

The combination of a reduction in I_to amplitude and a tendency to ‘speed-up' I_to decay kinetics suggests that in the presence of loratadine the amount of current exiting the cell through transient outward channels during depolarizations at pacing rates of 1 Hz or greater will be reduced. This hypothesis was tested by calculating the I_to current integral in the presence and absence of loratadine at pacing rates of 1 and 2 Hz. As illustrated in Figure 5, in the presence of 100 nM loratadine the total amount of current leaving the cell during a depolarizing pulse was dramatically and significantly reduced when compard to control (Figures 5A and B) (P<0.05). Interestingly, the ability of 10 nM loratadine to reduce total current exiting the cell was not significantly different from that observed for 100 nM (Figure 5D).

Figure 5.

Effect of loratadine on I_to total charge movement in human atrial myocytes. (A) Example of the integrated current in the absence of drug (26 pA ms⁻¹). Steady-state current has been subtracted from peak current. Line indicates zero current. Currents illustrated were recorded after current amplitude had reached a steady-state and were elicited by a series of 30 pulses (320 ms) to +60 mV from a holding potential of −60 mV (pacing rate=2 Hz). (B) Current recorded from same cell after addition of 100 nM loratadine (15.5 pA ms⁻¹). (C) Change in total charge exiting through I_to channels induced by 100 nM loratadine. Shaded area indicates the loratadine-sensitive integrated current. (D) Plot of the % reduction in total charge movement measured in cells before and after exposure to loratadine at the indicated pacing rates. All values were significantly different from control, indicated by asterisks. Data are mean±s.e.mean (n=5).

Discussion

The major finding of this study is that loratadine blocks I_to recorded from human atrial myocytes and significantly reduces the amount of I_to current exiting the myocyte in a rate-dependent manner at concentrations achieved therapeutically and during an overdose (Hilbert et al., 1978). At therapeutic concentrations (10 nM), loratadine significantly blocked I_to at pacing rates which mimic elevated heart rates (2 Hz) while at concentrations which may be achieved during an overdose (100 nM) (Hilbert et al., 1987), I_to was blocked at pacing rates which simulate both normal (1 Hz) and elevated heart rates (Figure 3). These results provide a possible molecular mechanism for the reported supraventricular arrhythmias associated with the use of loratadine. The reduction caused by loratadine in the total amount of I_to current exiting the myocyte (Figure 5) may be arrhythmogenic since I_to is an important repolarizing current in human atrium and has been shown to modulate atrial action potential shape and duration (Shibata et al., 1989).

The block of I_to by loratadine (10–100 nM) described in the present study is in marked contrast to a previous report in which loratadine at concentrations up to 1 μM exhibited no blocking actions on I_to recorded from rat ventricle (Ducic et al., 1997). These differences may reflect species differences in the molecular nature of I_to or in the fact that the previous report did not examine the effects of rate on the blocking action of loratadine. In fact, in the present study when cells were paced slowly (0.1 Hz), loratadine even at concentrations up to 1 μM produced only a very modest reduction in the amplitude of I_to similar to previous reports (Figure 3).

Use of non-sedating antihistamines has been associated with cardiac arrhythmias (Craft, 1985; Davies et al., 1989; Bishop & Gaudry, 1989; Monahan et al., 1990; Lindquist & Edwards, 1997). Although the occurrence of such cardiotoxic side effects is rare, it is nonetheless important to recognize since these arrhythmias can be life threatening. The adverse cardiac effects associated with antihistamines are typified by terfenadine which has been associated with prolongation of the QT interval and torsade de pointes (Davies et al., 1989; Monahan et al., 1990; Lindquist & Edwards, 1997). Providing a possible mechanism for these arrhythmias, terfenadine has been shown to block several cardiac potassium channels including I_to, I_sus and the delayed rectifier (I_Kr) in human heart as well as in other species and expression systems (Woosley et al., 1993; Salata et al., 1995; Rampe et al., 1993; Crumb et al., 1995c). Block of I_Kr and/or I_sus by terfenadine is believed to be in part responsible for the reported incidences of QT prolongation and torsade de pointes.

There are several reports of loratadine associated adverse cardiac events in patients in particular atrial arrhythmias. Reports of palpitations and arrhythmias have been reported to the Australian ADR Advisory Committee (1996) and a number of reports of supraventricular arrhythmias have been reported to the United States Food and Drug Administration (Haria et al., 1994). Recently, an examination of adverse drug reaction (ADR) reports obtained from the World Health Organization ADR database indicates several hundred incidences of loratadine associated ‘rate and rhythm disorders' and more than a dozen incidences of sudden cardiac death (Lindquist & Edwards, 1997). In addition to atrial arrhythmias, loratadine use has also been associated with ventricular arrhythmias. One case report associates loratadine use with QT prolongation and ventricular arrhythmias in a patient with a history of both atrial and ventricular arrhythmias (Good et al., 1994).

In summary, the present study describes the block of the potassium current I_to in human atrial myocytes by loratadine. This potassium channel blockade may provide a mechanism for the rare incidences of supraventricular arrhythmias reported with the use of this antihistamine. Although untested, it is intriguing to speculate that block of I_to in human ventricle may be similarly arrhythmogenic. The observation that a marked reduction in repolarizing current is observed with therapeutic concentrations of loratadine at rates mimicking elevated heart rates suggests that care should be taken in administering this antihistamine in patients with an existing tachyarrhythmia or under conditions where heart rate may be elevated (i.e. exercise or stress).

Abbreviations

I_K1

inwardly rectifying potassium current

I_ns

non-selective cation current

I_sus

sustained current

I_to

transient outward potassium current