Hysteretic hERG channel gating current recorded at physiological temperature

Abstract

Cardiac hERG channels comprise at least two subunits, hERG 1a and hERG 1b, and drive cardiac action potential repolarization. hERG 1a subunits contain a cytoplasmic PAS domain that is absent in hERG 1b. The hERG 1a PAS domain regulates voltage sensor domain (VSD) movement, but hERG VSD behavior and its regulation by the hERG 1a PAS domain have not been studied at physiological temperatures. We recorded gating charge from homomeric hERG 1a and heteromeric hERG 1a/1b channels at near physiological temperatures (36 ± 1 °C) using pulse durations comparable in length to the human ventricular action potential. The voltage dependence of deactivation was hyperpolarized relative to activation, reflecting VSD relaxation at positive potentials. These data suggest that relaxation (hysteresis) works to delay pore closure during repolarization. Interestingly, hERG 1a VSD deactivation displayed a double Boltzmann distribution, but hERG 1a/1b deactivation displayed a single Boltzmann. Disabling the hERG 1a PAS domain using a PAS-targeting antibody similarly transformed hERG 1a deactivation from a double to a single Boltzmann, highlighting the contribution of the PAS in regulating VSD movement. These data represent, to our knowledge, the first recordings of hERG gating charge at physiological temperature and demonstrate that VSD relaxation (hysteresis) is present in hERG channels at physiological temperature.

Development of automated patch clamp assays to overcome the burden of variants of uncertain significance in inheritable arrhythmia syndromes

“…At an experimental level, temperature control can also be an issue, which is important to study ion channel kinetics of temperature-sensitive variants ( Vandenberg et al, 2006 ; Abdelsayed et al, 2015 ; Lei et al, 2019 ; Jones, 2022 ; Ren et al, 2022 ; Kriegeskorte et al, 2023 ). Whilst temperature can be preset on most APC systems (e.g., Biolin Scientific, Fluxion and Nanion Technologies, Table 1 ), most APC assays for cardiac ion channels have been conducted at room temperature ( Vanoye et al, 2018 ; Glazer et al, 2020b ; Ng et al, 2021 ) due to an inherent instability of cell membranes at physiological temperatures leading to lower success rates.…”

The Novel Long QT Syndrome Type 2-associated F129I Mutation in the KCNH2 Gene Significantly Affects I Kr Through the hERG1 Homomeric and Heteromeric Potassium Channels

“…[11,12] Previous studies showed homomeric herg1a channel and heteromeric hERG1a/hERG1b channel has distinct effect to drugs and temperature. [13,14] Although the formation of the homomeric and heteromeric hERG1 channels has been shown by various studies, the details of the in vivo oligomerization and stoichiometry are not confirmed and require further investigation. Currently, studies have focused on the relationship between the functionality of the hERG1a channels and the LQTS type 2-associated mutations.…”