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. 2019 Feb;597(3):723-740.
doi: 10.1113/JP277188. Epub 2018 Dec 5.

Action potential shortening rescues atrial calcium alternans

Giedrius Kanaporis  1 , Zane M Kalik  1 , Lothar A Blatter  1
Affiliations

Action potential shortening rescues atrial calcium alternans

Giedrius Kanaporis et al. J Physiol. 2019 Feb.

Abstract

Key points: Cardiac alternans refers to a beat-to-beat alternation in contraction, action potential (AP) morphology and Ca2+ transient (CaT) amplitude, and represents a risk factor for cardiac arrhythmia, including atrial fibrillation. We developed strategies to pharmacologically manipulate the AP waveform with the goal to reduce or eliminate the occurrence of CaT and contraction alternans in atrial tissue. With combined patch-clamp and intracellular Ca2+ measurements we investigated the effect of specific ion channel inhibitors and activators on alternans. In single rabbit atrial myocytes, suppression of Ca2+ -activated Cl- channels eliminated AP duration alternans, but prolonged the AP and failed to eliminate CaT alternans. In contrast, activation of K+ currents (IKs and IKr ) shortened the AP and eliminated both AP duration and CaT alternans. As demonstrated also at the whole heart level, activation of K+ conductances represents a promising strategy to suppress alternans, and thus reducing a risk factor for atrial fibrillation.

Abstract: At the cellular level alternans is observed as beat-to-beat alternations in contraction, action potential (AP) morphology and magnitude of the Ca2+ transient (CaT). Alternans is a well-established risk factor for cardiac arrhythmia, including atrial fibrillation. This study investigates whether pharmacological manipulation of AP morphology is a viable strategy to reduce the risk of arrhythmogenic CaT alternans. Pacing-induced AP and CaT alternans were studied in rabbit atrial myocytes using combined Ca2+ imaging and electrophysiological measurements. Increased AP duration (APD) and beat-to-beat alternations in AP morphology lowered the pacing frequency threshold and increased the degree of CaT alternans. Inhibition of Ca2+ -activated Cl- channels reduced beat-to-beat AP alternations, but prolonged APD and failed to suppress CaT alternans. In contrast, AP shortening induced by activators of two K+ channels (ML277 for Kv7.1 and NS1643 for Kv11.1) abolished both APD and CaT alternans in field-stimulated and current-clamped myocytes. K+ channel activators had no effect on the degree of Ca2+ alternans in AP voltage-clamped cells, confirming that suppression of Ca2+ alternans was caused by the changes in AP morphology. Finally, activation of Kv11.1 channel significantly attenuated or even abolished atrial T-wave alternans in isolated Langendorff perfused hearts. In summary, AP shortening suppressed or completely eliminated both CaT and APD alternans in single atrial myocytes and atrial T-wave alternans at the whole heart level. Therefore, we suggest that AP shortening is a potential intervention to avert development of alternans with important ramifications for arrhythmia prevention and therapy.

Keywords: K+ current; action potential duration; alternans; arrhythmia; atria; calcium; excitation-contraction coupling.

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Figures

Figure 1
Figure 1. Action potential waveforms used as voltage commands for voltage clamp experiments
A, simultaneous recordings of APD and CaT alternans from current‐clamped single atrial myocyte. B, voltage commands used to pace cells during AP voltage clamp experiments representing AP waveforms observed during large (APCaT_Large) and small (APCaT_Small) alternans CaTs. These AP waveforms were used in all AP voltage clamp experiments in this study.
Figure 2
Figure 2. Severity of CaT alternans is determined by AP morphology
A, CaTs elicited with the same‐shape APCaT_Large, the same‐shape APCaT_Small and the alternans AP clamp protocols recorded from the same atrial myocyte paced at cycle lengths (CL) of 740 ms (left) and 540 ms (right). B, CaT alternans ratios recorded from 15 individual cells (10 rabbits) stimulated with the same‐shape APCaT_Large, the same‐shape APCaT_Small and the alternans AP clamp protocols at CL = 540 ms. Individual symbols represent individual cells. C, pacing with the same‐shape APCaT_Small waveforms (grey bars) enhances the degree of CaT alternans compared to the APCaT_Large protocol (white bars). CaT alternans ratio (AR) is further increased during the alternans AP voltage clamp protocol (black bars). n for individual data sets varies from 7 to 34.
Figure 3
Figure 3. Suppression of Ca2+‐activated chloride channels does not prevent CaT alternans
Aa, CaT alternans recorded in control and in the presence of Cl− channel blocker DIDS (500 μmol l−1) in a field stimulated atrial myocyte; A b, CaT alternans ratios (ARs) in control and after the application of DIDS in five individual field‐stimulated cells; Ac, mean CaT ARs in control and in the presence of DIDS (paired t test, P = 0.89, n = 5, 3 rabbits). B, CaCC inhibition with DIDS leads to APD prolongation. Ba, raw traces of APs recorded in control and after application of DIDS. Bb, overlay of APs recorded in control (black) and after the application of DIDS (grey).
Figure 4
Figure 4. K+ channel agonists increase K+ current in atrial myocytes
Aa, K+ current traces elicited by voltage steps (−70 to +50 mV) from V H = −60 mV before and after application of Kv11.1 agonist NS1643 (20 μmol l−1). Arrows indicate where the amplitudes of K+ and tail currents were measured. I–V curves of K+ currents (Ab) and tail currents (Ac) recorded in control and in the presence of NS1643 (n = 5, 4 rabbits). B, representative current traces (Ba) elicited by the same protocol as in A before and after application of Kv7.1 agonist ML277 (0.5 μmol l−1). I–V curves of K+ currents (Bb) and tail currents (Bc) recorded in control and after application of ML277 (n = 8, 3 rabbits).
Figure 5
Figure 5. K+ channel agonists shorten AP duration (APD)
Aa, traces of APs recorded in control and after the application of NS1643 (20 μmol l−1); Ab, overlay of APs recorded in control (black) and after the application of NS1643 (grey); Ac, mean APD at 30, 50 and 90% repolarization level before and after application of NS1643 (n = 9, 7 rabbits). Ba, traces of APs recorded in control and after the application of ML2773 (0.5 μmol l−1); Bb, overlay of APs recorded in control and after the application of ML277; Bc, mean APD at 30, 50 and 90% repolarization level before and after the application of ML277 (n = 10, 6 rabbits).
Figure 6
Figure 6. AP shortening reduces degree of CaT alternans in field‐stimulated atrial myocytes
Aa, CaTs recorded before and after application of Kv11.1 channel agonist NS1643 in field‐stimulated atrial myocytes. Ab, changes of CaT alternans ratio (AR) in control and in the presence of NS1643 from 11 individual cells. Ac, mean CaT AR in control and after application of NS1643 (n = 11, 4 rabbits). Ba, CaTs recorded before and after application of Kv7.1 channel agonist ML277 in field stimulated atrial myocytes. Bb, changes in CaT AR from 10 individual cells. Bc, mean CaT AR in control and after application of ML277 (n = 10, 6 rabbits).
Figure 7
Figure 7. Degree of CaT alternans remains stable during prolonged pacing
A, mean CaT alternans ratio (AR) over time in field‐stimulated atrial myocytes (n = 11, 4 rabbits) continuously paced for 10 min. B, application of ML277 (0.5 μmol l−1), following 10 min recording of stable CaT alternans, leads to fast reduction in CaT AR in field‐stimulated atrial myocytes. Recovery of CaT AR is observed during the washout of ML277. Top panel: representative CaT traces observed at the times indicated by the corresponding letter on the main graph.
Figure 8
Figure 8. AP shortening suppresses CaT and APD alternans in current‐clamped atrial myocytes
A and B, effect of K+ channel agonists NS1643 (A) and ML277 (B) on CaT alternans in current‐clamped atrial myocytes. a, action potential and [Ca2+]i traces observed before and after application of NS1643 (n = 4, 3 rabbits) or ML277 (n = 5, 5 rabbits). b, CaT alternans ratio in individual cells in control and after the application of K+ channel agonists. c, mean CaT alternans ratio in control and in the presence of K+ channel agonists. d, in addition to CaT alternans, APD alternans also was abolished in the presence of NS1643 and ML277. Ratios of APDs of odd and even beats at 30, 50 and 90% of AP repolarization are shown. In the presence of K+ channel agonists the ratio approaches 1 (marked by dashed line), indicating elimination of APD alternans.
Figure 9
Figure 9. K+ channel agonists have no effect on CaT alternans under voltage clamp conditions
A and B, application of NS1643 (n = 8, 7 rabbits) (A) and ML277 (n = 5, 5 rabbits) (B) has no effect on the degree of CaT alternans in AP voltage‐clamped atrial myocytes, demonstrating that the effect of these compounds is caused by shortening of APD. a, CaT alternans ratio (AR) in individual atrial myocytes before and after application of agonists of K+ channels observed during the alternans AP stimulation protocol. b, mean CaT ARs in the absence and presence of the K+ agonists during the same‐shape APCaT_Small, the same‐shape APCaT_Large and the alternans AP protocol. In all groups mean CaT ARs in control and in the presence of agonists are not statistically different.
Figure 10
Figure 10. NS1643 abolishes atrial T‐wave (Ta) alternans in Langendorff‐perfused hearts
Atrial electrograms recorded from the same Langendorf‐perfused isolated rabbit heart in control and after the application of NS1643 (20 μmol l−1). In the presence of NS1643 beat‐to‐beat alternation of the Ta‐wave is eliminated.
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