Background Genotype-positive patients who suffer from the cardiac channelopathy Long QT Syndrome (LQTS) may display a spectrum of clinical phenotypes, with often unknown causes. Therefore, there is a need to identify factors influencing disease severity to move towards an individualized clinical management of LQTS. One possible factor influencing the disease phenotype is the endocannabinoid system, which has emerged as a modulator of cardio-vascular function. In this study, we aim to elucidate whether endocannabinoids target the cardiac voltage-gated potassium channel KV7.1/KCNE1, which is the most frequently mutated ion channel in LQTS.Methods We used two-electrode voltage clamp, molecular dynamics simulations and the E4031 drug-induced LQT2 model of ex-vivo guinea pig hearts.Findings We found a set of endocannabinoids that facilitate channel activation, seen as a shifted voltage-dependence of channel opening and increased overall current amplitude and conductance. We propose that negatively charged endocannabinoids interact with known lipid binding sites at positively charged amino acids on the channel, providing structural insights into why only specific endocannabinoids modulate KV7.1/KCNE1. Using the endocannabinoid ARA-S as a prototype, we show that the effect is not dependent on the KCNE1 subunit or the phosphorylation state of the channel. In guinea pig hearts, ARA-S was found to reverse the E4031-prolonged action potential duration and QT interval. Interpretation We consider the endocannabinoids as an interesting class of hKV7.1/KCNE1 channel modulators with putative protective effects in LQTS contexts.Copyright (c) 2023 The Author(s). Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

Background and PurposeCongenital long QT syndrome (LQTS) involves genetic mutations affecting ion channels, leading to a prolonged QT interval and increased risk of potentially lethal ventricular arrhythmias. Mutations in the genes encoding KV7.1/KCNE1 are the most frequent, with channel loss-of-function contributing to LQTS. The endocannabinoid N-arachidonoyl-L-serine (ARA-S) has been shown to facilitate activation of wild type KV7.1/KCNE1 channels and to counteract a prolonged QT interval in isolated guinea pig hearts. In this study, we examine the ability of ARA-S to facilitate activation of LQTS-associated mutations, in various regions of the channel, and hence to counteract loss-of-function.Experimental ApproachThe two-electrode voltage clamp technique on Xenopus oocytes expressing human KV7.1/KCNE1 channels was used to investigate the effects of ARA-S in 20 LQTS type 1-associated mutations distributed across the channel. Thereafter, different electrophysiology was used to assess ARA-S effects in mammalian cells.Key ResultsARA-S enhanced the function of all mutated channels by shifting V50 and increasing current amplitude. However, the magnitude of effect varied, related to whether mutations were in one of the two putative ARA-S binding sites on the channel as suggested by molecular dynamics simulations. ARA-S displayed translational potential by facilitating channel opening in mammalian cells and shortening the action potential duration in cardiomyocytes.Conclusions and ImplicationsThis study demonstrates the rescuing capability of ARA-S on a diverse set of LQTS mutants. These insights may aid in developing drug compounds using ARA-S sites and mechanisms and guide interpretation of which LQTS mutants respond well to such compounds.

The endogenous endocannabinoid-like compound N-arachidonoyl-L-serine (ARA-S) facilitates activation of the human Kv7.1/KCNE1 channel and shortens a prolonged action potential duration and QT interval in guinea pig hearts. Hence, ARA-S is interesting to study further in cardiac models to explore the functional impact of such Kv7.1/KCNE1-mediated effects. To guide which animal models would be suitable for assessing ARA-S effects, and to aid interpretation of findings in different experimental models, it is useful to know whether Kv7.1/KCNE1 channels from relevant species respond similarly to ARA-S. To this end, we used the two-electrode voltage clamp technique to compare the effects of ARA-S on Kv7.1/KCNE1 channels from guinea pig, rabbit, and human Kv7.1/KCNE1, when expressed in Xenopus laevis oocytes. We found that the activation of Kv7.1/KCNE1 channels from all tested species was facilitated by ARA-S, seen as a concentration-dependent shift in the voltage-dependence of channel opening and increase in current amplitude and conductance over a broad voltage range. The rabbit channel displayed quantitatively similar effects as the human channel, whereas the guinea pig channel responded with more prominent increase in current amplitude and maximal conductance. This study suggests that rabbit and guinea pig models are both suitable for studying ARA-S effects mediated via Kv7.1/KCNE1.