TASK-1 channels may modulate action potential duration of human atrial cardiomyocytes

Abstract

Background/aims: Atrial fibrillation is the most common arrhythmia in the elderly, and potassium channels with atrium-specific expression have been discussed as targets to treat atrial fibrillation. Our aim was to characterize TASK-1 channels in human heart and to functionally describe the role of the atrial whole cell current I(TASK-1).

Methods and results: Using quantitative PCR, we show that TASK-1 is predominantly expressed in the atria, auricles and atrio-ventricular node of the human heart. Single channel recordings show the functional expression of TASK-1 in right human auricles. In addition, we describe for the first time the whole cell current carried by TASK-1 channels (I(TASK-1)) in human atrial tissue. We show that I(TASK-1) contributes to the sustained outward current I(Ksus) and that I(TASK-1) is a major component of the background conductance in human atrial cardiomyocytes. Using patch clamp recordings and mathematical modeling of action potentials, we demonstrate that modulation of I(TASK-1) can alter human atrial action potential duration.

Conclusion: Due to the lack of ventricular expression and the ability to alter human atrial action potential duration, TASK-1 might be a drug target for the treatment of atrial fibrillation.

Fig. 1

Expression analysis of TASK-1 in the human heart. (A), Quantitative mRNA expression analysis of human TASK-1 in heart tissue pooled from 12 different donors (n = 8 qPCR runs). Relative expression was quantified as 1/2 ^‡CT, where ‡CT is CT (GAPDH) - CT (K⁺ channel). (B), Sample amplification blots of quantitative PCR analysis of human TASK-1 in different cardiac tissues. (C), Quantitative PCR data analyzing the TASK-1 expression in various cardiac regions, normalized to left atrium (n = 5 qPCR runs). For patient information see Table 1 or Material and Methods section.

Fig. 2

Electrophysiological characterization of human cardiac I_TASK-1. (A), Cell attached recordings from a TASK-1 like channel in human atrial cardiomyocytes. The lower panel shows a magnification of the events designated in the top panel by dotted squares. (B), Representative current traces elicited by the voltage protocol illustrated in panel C. After a pre-step of 70 ms to −50 mV, the voltage was stepped for 300 ms from −60 mV to +50 mV in 10 mV increments. The holding potential was −80 mV and the sweep time interval was 10 s. Control traces are shown in black and traces after administration of 200 nM A293 in grey. The chemical structure of A293 is illustrated below. (C), Mean current voltage relationships in the absence and presence of A293 derived at the end of the 300 ms test pulse (as indicated by the arrow in panel B). Significance was analyzed using a paired Student's t-test. (D), Isolation of the A293-sensitive TASK-1 current, using 200 nM A293. Currents were recorded using the shown voltage ramps from +40 mV to −80 mV (duration 9.5 s) from a holding potential of 0 mV. Control traces are shown in black. The bar graph in the middle panel analyzes the currents measured at +30 mV (as illustrated by the arrow) and the difference current analyzed after application of the TASK blocker A293 (which corresponds to I_TASK-1). (E), Depicted is the average difference current with S.E.M., isolated with 200 nM A293 (n = 7). The solid line indicates a fit to the GHK equation. (F), Recordings of a background conductance after application of a blocker mixture (light grey), using the same protocols and analysis as described above. 2 µM A293 (in the presence of the blocker mixture) was used to isolate the A293-sensitive TASK-1 currents and the relative contribution to the background current.

Fig. 3

Patch clamp recordings of right human auricle cardiomyocytes. (A), Right auricle cardiomyocytes were injected with a small negative current of about −20 pA in order to hyperpolarize the cells to −80 m V. Action potentials were elicited by injection of a 2 − 5 ms current pulse of 2 nA amplitude. Action potentials were evoked with a frequency of 1 Hz. After action potential duration reached a steady state (black), 2 µM A293 was administered, until steady-state was reached (grey). (B), Relative increase in APD₅₀ and APD₉₀ by 2 µM A293. (C), Dynamic patch clamp experiments of single myocytes were performed as previously described [37]. Right human auricle cardiomyocytes were held in current clamp mode and resting membrane potential was adjusted to −80 mV via the injection of a negative offset current, as described above. Action potentials were elicited at a frequency of 1 Hz via injection of a positive current pulse (2 − 3 nA) of 2 ms duration. Dynamic patch clamp experiments with subtraction of I_TASK-1 led to prolonged action potential (dark grey line), injection of an additional I_TASK-1 shortened APD (light grey line). (D), Bar graph showing prolongation of APD₅₀ and APD₉₀, respectively after subtraction of I_TASK-1. (E), Shortening of APD₅₀ and APD₉₀ after injecting additional I_TASK-1.

Fig. 4

Development of a TASK-1 Markov model. TEVC measurements of TASK-1 channels in Xenopus oocytes served to create a Markov model based on a closed-closed-open assumption. The figure illustrates the comparison of biophysical parameters of TASK-1 injected oocytes measured by TEVC (squares) and the Markov model (circles). (A), Representative TASK-1 current voltage relationship recordings elicited by the voltage protocol illustrated. The recordings in (A) served as input for the comparison of TASK-1 model data with TEVC recordings (B-E). (B), Current voltage relationship. (C), Time constants of activation. (D), Ratio of the amplitudes of the time constants of activation. (E), Percentage of instantaneous current. (F), Representative recordings of TASK-1 deactivation, using the illustrated voltage protocol. The recordings in (F) served as input for the comparison to the TASK-1 model data (G-H). (G), Fraction of current showing deactivation. (H), Time constant of deactivation. (I), Scheme of the Markov model with simulated currents for activation (left) and deactivation (right). Currents were normalized.

Fig. 5

Action potential simulations using the TASK-1 Markov model show the influence of I_TASK-1 on human atrial action potential. (A), Computational simulation of a normal action potential (black). Subtraction of I_TASK-1 led to prolongation of APD and an increase in plateau voltage (blue). A 2fold increased I_TASK-1 current shortened APD and decreased plateau voltage (red). (B), Net membrane currents with (black) and without (blue) I_TASK-1. (C), L-type calcium current (I_CaL) with (black) and without (blue) I_TASK-1. (D-F), The effects of I_TASK-1 subtraction on other currents. (D), Effects on I_Kur (black/blue). (E), Effects on I_Kr (black/grey) and I_Ks (green/blue). (F), Effects on the inward rectifier current (I_K1) (black/blue).