Atrial tachyarrhythmia in Rgs5-null mice

Abstract

Aims: The aim of this study was to elucidate the effects of regulator of G-protein signaling 5 (Rgs5), a negative regulator of G protein-mediated signaling, on atrial repolarization and tachyarrhythmia (ATA) in mice.

Methods and results: In present study, the incidence of ATA were increased in Rgs5(-/-) Langendorff-perfused mouse hearts during program electrical stimulation (PES) (46.7%, 7 of 15) and burst pacing (26.7%, 4 of 15) compared with wild-type (WT) mice (PES: 7.1%,1 of 14; burst:7.1%,1 of 14) (P<0.05). And the duration of ATA also shown longer in Rgs5(-/-) heart than that in WT, 2 out of 15 hearts exhibited sustained ATA (>30 s) but none of them observed in WT mice. Atrial prolonged repolarization was observed in Rgs5(-/-) hearts including widened P wave in surface ECG recording, increased action potential duration (APD) and atrial effective refractory periods (AERP), all of them showed significant difference with WT mice (P<0.05). At the cellular level, whole-cell patch clamp recorded markedly decreased densities of repolarizing K(+) currents including I(Kur) (at +60 mV: 14.0±2.2 pF/pA) and I(to) (at +60 mV: 16.7±1.3 pA/pF) in Rgs5(-/-) atrial cardiomyocytes, compared to those of WT mice (at +60 mV I(to): 20.4±2.0 pA/pF; I(kur): 17.9±2.0 pF/pA) (P<0.05).

Conclusion: These results suggest that Rgs5 is an important regulator of arrhythmogenesis in the mouse atrium and that the enhanced susceptibility to atrial tachyarrhythmias in Rgs5(-/-) mice may contribute to abnormalities of atrial repolarization.

Figure 1. ECG and MAP recordings in Rgs5-/- and WT mice.

Two representative electrocardiograms recorded from telemetry ECG showed the measurements of Pdur and PR intervals (A). Compared with wild-type mice, Rgs5^−/− showed widened P wave, and the action potential duration (APD) prolonged in the atria of Rgs5^−/− from 50% (APD₅₀) to 90% (APD₉₀) (B,C). The effective refractory periods (ERP) also significantly increased during S₁ cycle length of PES at 150 ms, 125 ms and 100 ms.

Figure 2. Transmembrane action potentials (TAP) from WT and Rgs5^−/− atrial samples.

The TAP was recorded at paced frequency from 1 Hz to 6.7 Hz (A–D). Mean±SEM APD values at 90% repolarization (APD₉₀) show APD prolongation at different pacing frequency (E). Early after-depolarizations (EADs) occurred in Rgs5^−/− group at 1 Hz, 2 Hz and 3.3 Hz. EADs occurrence was quantified as percentage of preparations (F).

Figure 3. Sinus node recovery time (SNRT) in WT (A) and Rgs5^−/− (B) heart.

The SNRT was measured by a 2 s pacing train at a cycle length of 100 ms, and defined as the interval between end of the stimuli and recovered sinus rhythm. The bipolar recording electrode was placed on the atrial appendage.

Figure 4. Atrial tachyarrhythmia (ATA) inducibility in Rgs5^−/− and WT hearts.

PES (A) and Burst stimuli (50 Hz, 2 s) (B) were applied to induce atrial tachyarrhythmia (ATA). The sustained rapid atrial activities (duration>30 s) were elicited in Rgs5^−/− hearts, but Wild-type hearts showed non-sustained rapid atrial activities during both procedures. Overall incidence of ATA induced by PES and burst pacing in WT and Rgs5^−/− hearts (C), and the incidence was sorted by the ATA duration (<10 s, 10–30 s and >30 s) during PES (D) and burst pacing (E).

Figure 5. Alterations in K⁺ current were recorded in atrial myocytes of Rgs5^−/− and WT mice.

Representative whole-cell total outward K⁺ currents (I_peak) (A,B), transient outward currents (Ito) (C, D) and ultra-rapid delayed rectifier currents (I_kur) (E, F) evoked during 500 ms test potential steps from −40 mV to +60 mV from a holding potential (HP) of −80 mV. The inwardly rectified K⁺ currents (I_K1) (G, H), evoked during 350 ms voltage steps to potentials between −120 and −40 mV (HP = −80 mV). The current-voltage relationship (I–V) of I_peak and I_K1 were plotted (B, D). Membrane capacity (C_m) (I) and maximal densities of currents (J) were compared between wild-type and Rgs5^−/− mice. *P<0.05 WT vs Rgs5^−/−.

Figure 6. Comparison of magnitude of I_peak, I_to, I_Kur and I_K1 in atrial myocytes of WT and Rgs5^−/− mice recorded at 22°C and 37°C (A).

Bar graph represents mean (±S.E.M) maximal current density of each of the four outward K⁺ current recorded at 22°C and 37°C in WT (B) and Rgs5^−/− (C) group.

Figure 7. Channel kinetics of I_to and I_Kur were analyzed in Rgs5^−/− and WT cardiomyocytes, respectively.

The steady-state inactivation (A) and reactivation (B) of curves in I_to plotted similar tendency between Rgs5^−/− and wild-type atrial myocytes. However, the property of I_Kur in Rgs5^−/− cardiomyocytes showed markedly modification with significant difference of half-inactivated voltage (V_1/2) (C) and reactivated time constant (τ) (D). Histogram summarized the statistical significance of V_1/2 and τ in WT and Rgs5^−/− groups (E, F).

Figure 8. Fibrosis assay in Rgs5^−/− and WT atrium.

mRNA expression levels of mediators of fibrosis including Tgfβ1, procollagen, type Iα1 (Col1α1) and procollagen, type IIIα1 (Col3α1) were analysis in WT and Rgs5^−/− mouse atrium (A, B). Relative abundance was calculated with the value of wild-type as a reference of 100%. PSR staining on histological sections of the atrial appendage and ventricle was performed in WT and Rgs5^−/− mice (C).