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Transient Outward K+ Current (Ito) Underlies the Right Ventricular Initiation of Polymorphic Ventricular Tachycardia in a Transgenic Rabbit Model of Long-QT Syndrome Type 1.

ABSTRACT: BACKGROUND:Sudden death in long-QT syndrome type 1 (LQT1), an inherited disease caused by loss-of-function mutations in KCNQ1, is triggered by early afterdepolarizations (EADs) that initiate polymorphic ventricular tachycardia (pVT). We investigated ionic mechanisms that underlie pVT in LQT1 using a transgenic rabbit model of LQT1. METHODS:Optical mapping, cellular patch clamping, and computer modeling were used to elucidate the mechanisms of EADs in transgenic LQT1 rabbits. RESULTS:The results showed that shorter action potential duration in the right ventricle (RV) was associated with focal activity during pVT initiation. RV cardiomyocytes demonstrated higher incidence of EADs under 50 nmol/L isoproterenol. Voltage-clamp studies revealed that the transient outward potassium current (Ito) magnitude was 28% greater in RV associated with KChiP2 but with no differences in terms of calcium-cycling kinetics and other sarcolemmal currents. Perfusing with the Ito blocker 4-aminopyridine changed the initial focal sites of pVT from the RV to the left ventricle, corroborating the role of Ito in pVT initiation. Computer modeling showed that EADs occur preferentially in the RV because of the larger conductance of the slow-inactivating component of Ito, which repolarizes the membrane potential sufficiently rapidly to allow reactivation of ICa,L before IKr has had sufficient time to activate. CONCLUSIONS:Ito heterogeneity creates both triggers and an arrhythmogenic substrate in LQT1. In the absence of IKs, Ito interactions with ICa,L and IKr promote EADs in the RV while prolonging action potential duration in the left ventricle. This heterogeneity of action potential enhances dispersion of refractoriness and facilitates conduction blocks that initiate pVTs.

SUBMITTER: Choi BR

PROVIDER: S-EPMC6081959 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature

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Publications

Transient Outward K<sup>+</sup> Current (I<sub>to</sub>) Underlies the Right Ventricular Initiation of Polymorphic Ventricular Tachycardia in a Transgenic Rabbit Model of Long-QT Syndrome Type 1.

Choi Bum-Rak BR Li Weiyan W Terentyev Dmitry D Kabakov Anatoli Y AY Zhong Mingwang M Rees Colin M CM Terentyeva Radmila R Kim Tae Yun TY Qu Zhilin Z Peng Xuwen X Karma Alain A Koren Gideon G

Circulation. Arrhythmia and electrophysiology 20180601 6

<h4>Background</h4>Sudden death in long-QT syndrome type 1 (LQT1), an inherited disease caused by loss-of-function mutations in KCNQ1, is triggered by early afterdepolarizations (EADs) that initiate polymorphic ventricular tachycardia (pVT). We investigated ionic mechanisms that underlie pVT in LQT1 using a transgenic rabbit model of LQT1.<h4>Methods</h4>Optical mapping, cellular patch clamping, and computer modeling were used to elucidate the mechanisms of EADs in transgenic LQT1 rabbits.<h4>Re ...[more]

PMID: 29769222

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Project description:RationaleAutosomal-dominant mutations in ryanodine receptor type 2 ( RYR2) are responsible for ≈60% of all catecholaminergic polymorphic ventricular tachycardia. Dysfunctional RyR2 subunits trigger inappropriate calcium leak from the tetrameric channel resulting in potentially lethal ventricular tachycardia. In vivo CRISPR/Cas9-mediated gene editing is a promising strategy that could be used to eliminate the disease-causing Ryr2 allele and hence rescue catecholaminergic polymorphic ventricular tachycardia.ObjectiveTo determine if somatic in vivo genome editing using the CRISPR/Cas9 system delivered by adeno-associated viral (AAV) vectors could correct catecholaminergic polymorphic ventricular tachycardia arrhythmias in mice heterozygous for RyR2 mutation R176Q (R176Q/+).Methods and resultsGuide RNAs were designed to specifically disrupt the R176Q allele in the R176Q/+ mice using the SaCas9 ( Staphylococcus aureus Cas9) genome editing system. AAV serotype 9 was used to deliver Cas9 and guide RNA to neonatal mice by single subcutaneous injection at postnatal day 10. Strikingly, none of the R176Q/+ mice treated with AAV-CRISPR developed arrhythmias, compared with 71% of R176Q/+ mice receiving control AAV serotype 9. Total Ryr2 mRNA and protein levels were significantly reduced in R176Q/+ mice, but not in wild-type littermates. Targeted deep sequencing confirmed successful and highly specific editing of the disease-causing R176Q allele. No detectable off-target mutagenesis was observed in the wild-type Ryr2 allele or the predicted putative off-target site, confirming high specificity for SaCas9 in vivo. In addition, confocal imaging revealed that gene editing normalized the enhanced Ca2+ spark frequency observed in untreated R176Q/+ mice without affecting systolic Ca2+ transients.ConclusionsAAV serotype 9-based delivery of the SaCas9 system can efficiently disrupt a disease-causing allele in cardiomyocytes in vivo. This work highlights the potential of somatic genome editing approaches for the treatment of lethal autosomal-dominant inherited cardiac disorders, such as catecholaminergic polymorphic ventricular tachycardia.

Dataset Information

Transient Outward K+ Current (Ito) Underlies the Right Ventricular Initiation of Polymorphic Ventricular Tachycardia in a Transgenic Rabbit Model of Long-QT Syndrome Type 1.

Publications

Transient Outward K<sup>+</sup> Current (I<sub>to</sub>) Underlies the Right Ventricular Initiation of Polymorphic Ventricular Tachycardia in a Transgenic Rabbit Model of Long-QT Syndrome Type 1.

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