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Membranes (Basel)
2023 May 19;135:. doi: 10.3390/membranes13050529.
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Recording Sodium Self-Inhibition of Epithelial Sodium Channels Using Automated Electrophysiology in Xenopus Oocytes.
Lawong RY
,
May F
,
Etang EC
,
Vorrat P
,
George J
,
Weder J
,
Kockler D
,
Preller M
,
Althaus M
.
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The epithelial sodium channel (ENaC) is a key regulator of sodium homeostasis that contributes to blood pressure control. ENaC open probability is adjusted by extracellular sodium ions, a mechanism referred to as sodium self-inhibition (SSI). With a growing number of identified ENaC gene variants associated with hypertension, there is an increasing demand for medium- to high-throughput assays allowing the detection of alterations in ENaC activity and SSI. We evaluated a commercially available automated two-electrode voltage-clamp (TEVC) system that records transmembrane currents of ENaC-expressing Xenopus oocytes in 96-well microtiter plates. We employed guinea pig, human and Xenopus laevis ENaC orthologs that display specific magnitudes of SSI. While demonstrating some limitations over traditional TEVC systems with customized perfusion chambers, the automated TEVC system was able to detect the established SSI characteristics of the employed ENaC orthologs. We were able to confirm a reduced SSI in a gene variant, leading to C479R substitution in the human α-ENaC subunit that has been reported in Liddle syndrome. In conclusion, automated TEVC in Xenopus oocytes can detect SSI of ENaC orthologs and variants associated with hypertension. For precise mechanistic and kinetic analyses of SSI, optimization for faster solution exchange rates is recommended.
FKZ 005-2101-0144 Ministry of Culture and Science of the State of North Rhine-Westphalia, Startförderung der H-BRS 2022 Bonn-Rhein-Sieg University of Applied Sciences
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