Effects of rosiglitazone on the configuration of action potentials and ion currents in canine ventricular cells

Abstract

Background and purpose: In spite of its widespread clinical application, there is little information on the cellular cardiac effects of the antidiabetic drug rosiglitazone in larger experimental animals. In the present study therefore concentration-dependent effects of rosiglitazone on action potential morphology and the underlying ion currents were studied in dog hearts.

Experimental approach: Standard microelectrode techniques, conventional whole cell patch clamp and action potential voltage clamp techniques were applied in enzymatically dispersed ventricular cells from dog hearts.

Key results: At concentrations ≥10 µM rosiglitazone decreased the amplitude of phase-1 repolarization, reduced the maximum velocity of depolarization and caused depression of the plateau potential. These effects developed rapidly and were readily reversible upon washout. Rosiglitazone suppressed several transmembrane ion currents, concentration-dependently, under conventional voltage clamp conditions and altered their kinetic properties. The EC(50) value for this inhibition was 25.2 ± 2.7 µM for the transient outward K(+) current (I(to)), 72.3 ± 9.3 µM for the rapid delayed rectifier K(+) current (I(Kr)) and 82.5 ± 9.4 µM for the L-type Ca(2+) current (I(Ca) ) with Hill coefficients close to unity. The inward rectifier K(+) current (I(K1)) was not affected by rosiglitazone up to concentrations of 100 µM. Suppression of I(to), I(Kr), and I(Ca) was confirmed also under action potential voltage clamp conditions.

Conclusions and implications: Alterations in the densities and kinetic properties of ion currents may carry serious pro-arrhythmic risk in case of overdose with rosiglitazone, especially in patients having multiple cardiovascular risk factors, like elderly diabetic patients.

Figure 1

(A) Representative set of superimposed action potentials showing the cumulative concentration-dependent effects of rosiglitazone on action potential configuration. Early events of the action potential are enlarged in the inset, the first time derivatives of action potential upstrokes are depicted on the right. The action potentials were superimposed so as to match their upstrokes. (B–E) Cumulative concentration-dependent effects of rosiglitazone on the maximum rate of depolarization (V_max), amplitude of phase-1 repolarization, action potential duration measured at 50% (APD₅₀) and 90% (APD₉₀) level of repolarization, and action potential amplitude respectively. Phase-1 amplitude was determined as a difference of overshoot potential and the deepest point of the incisura. Each concentration of rosiglitazone was superfused for 3 min, the washout (Wo) lasted for 10 min. Symbols and bars represent mean ± SEM values of eight myocytes, obtained from four dogs. *P < 0.05, significant changes from pre-drug control values, which are also indicated by dotted lines.

Figure 2

Effect of rosiglitazone on I_to. (A), (B): Cumulative concentration-dependent effects of rosiglitazone on I_to measured under conventional voltage clamp conditions. Representative superimposed I_to current traces (A) recorded before and after superfusion with increasing concentrations of rosiglitazone, and the dose–response curve (B) obtained for I_to blockade in five cells, each from a different animal, including the results of the Hill plot. (C): Time course of development and reversibility of the effect of rosiglitazone on I_to measured in a representative cell. (D)–(F): Effect of 30 µM rosiglitazone on kinetic properties of I_to studied in five myocytes, each from a different dog. (D): Time course of inactivation of I_to. The current decay was fitted as a sum of two (fast and slow) exponential components. (E): Current–voltage relationship obtained for I_to. Peak values of I_to were plotted against the respective test potential shown on abscissa. (F): Effect of rosiglitazone on the voltage-dependence of steady-state inactivation of I_to. Test depolarizations to +50 mV were preceded by a set of prepulses clamped to various voltages between −70 and +10 mV. Peak currents measured after these prepulses were normalized to the peak current measured after the −70 mV prepulse and plotted against the respective prepulse potential. Solid lines were obtained by fitting data to the two-state Boltzmann function. Symbols, columns and bars are means ± SEM. *P < 0.05, significant changes from control.

Figure 3

Effect of rosiglitazone on I_Kr. (A), (B): Concentration-dependent effects of rosiglitazone on I_Kr measured under conventional voltage clamp conditions. Representative superimposed I_Kr tail current traces (A) recorded before and after superfusion with increasing concentrations of rosiglitazone, and the dose–response curve (B) obtained for I_Kr blockade in five cells, each from a different animal, including the results of the Hill plot. (C): Time course of development and reversibility of the effect of rosiglitazone on I_Kr recorded from a representative cell. (D)–(F): Effect of 100 µM rosiglitazone on kinetic properties of I_Kr studied in five myocytes, each obtained from a different dog. (D): Time course of deactivation of I_Kr. Decay of tail currents was fitted as a sum of two (fast and slow) exponential components. (E): Voltage-dependence of activation of I_Kr was determined by varying the potential for I_Kr activation as indicated on the abscissa. The results were fitted to the two-state Boltzmann function denoted by solid lines. (F): Time constant of activation was determined by monoexponential fitting of data obtained using the tail envelope test (applying depolarizations to +50 mV and determining tail current amplitudes at −40 mV). Durations of these depolarizing pulses are displayed on the abscissa. Symbols, columns and bars are mean values ± SEM. *P < 0.05, significant changes from control.

Figure 4

Effect of rosiglitazone on I_Ca. (A), (B): Cumulative concentration-dependent effects of rosiglitazone on I_Ca measured under conventional voltage clamp conditions. Representative superimposed I_Ca current traces (A) recorded before and after superfusion with increasing concentrations of rosiglitazone, and the dose–response curve (B) obtained for I_Ca blockade in six cells from five dogs, including the results of the Hill plot. (C): Time course of development and reversibility of the effect of rosiglitazone on I_Ca measured in a representative cell. (D)–(F): Effect of 100 µM rosiglitazone on kinetic properties of I_Ca studied in five myocytes, derived from five different animals. (D): Time course of inactivation of I_Ca. The current decay was fitted as a sum of two (fast and slow) exponential components. (E): Current–voltage relationship obtained for I_Ca. Amplitudes of I_Ca were plotted against the respective test potential shown on abscissa. (F): Effect of rosiglitazone on the voltage-dependence of steady-state inactivation of I_Ca. Test depolarizations to +5 mV were preceded by a set of prepulses clamped to various voltages between −50 and +5 mV. Peak currents measured after these prepulses were normalized to the peak current measured after the −50 mV prepulse and plotted against the respective prepulse potential. Solid lines were obtained by fitting data to the two-state Boltzmann function. Symbols, columns and bars are means ± SEM. *P < 0.05, significant changes from control.

Figure 5

Cumulative concentration-dependent effects of rosiglitazone on I_K1 measured under conventional voltage clamp conditions. Representative superimposed I_K1 current traces (A) recorded before and after superfusion with increasing concentrations of rosiglitazone, and the dose–response curve (B) obtained for I_K1 blockade in four cells, each from a different dog. Symbols and bars are means ± SEM. *P < 0.05, significant changes from control.

Figure 6

Effects of 1, 10 and 100 µM rosiglitazone on ion currents under action potential voltage clamp conditions. Representative records of a command signal (A), and the underlying current traces obtained before (B), in the presence of (C)–(E), and after washout of rosiglitazone (F). Because ion currents were obtained from the same cell that provided the command action potential, the pre-drug current record was a horizontal line at the zero level. The post-drug difference currents have been inverted in order to present the currents with their conventional polarity. Dotted lines indicate zero current level for each trace except for (A), where it represents zero voltage.

Figure 7

Effects of 1, 10 and 100 µM rosiglitazone on the early outward current peak, indicator of I_to (A), the inward current peak, indicator of I_Ca (B), and the late outward current peak, indicator predominantly of I_Kr (C), obtained under action potential voltage clamp conditions in five myocytes, each from a different animal. Columns and bars are means ± SEM. *P < 0.05, significant changes from control.