Electrophysiological measurement of ion channels on plasma/organelle membranes using an on-chip lipid bilayer system

Koki Kamiya,Masatoshi Hayakawa,Nobuo Misawa,Ryuji Kawano,Toshihisa Osaki,Satoshi Fujii,Shoji Takeuchi,Kenji Nakao

doi:10.1038/s41598-018-35316-4

Koki Kamiya, Masatoshi Hayakawa + Show 6 more

Open Access

https://doi.org/10.1038/s41598-018-35316-4

Copy DOI

Abstract

Ion channels are located in plasma membranes as well as on mitochondrial, lysosomal, and endoplasmic reticulum membranes. They play a critical role in physiology and drug targeting. It is particularly challenging to measure the current mediated by ion channels in the lysosomal and the endoplasmic reticulum membranes using the conventional patch clamp method. In this study, we show that our proposed device is applicable for an electrophysiological measurement of various types of ion channel in plasma and organelle membranes. We designed an on-chip device that can form multiple electrical contacts with a measurement system when placed on a mount system. Using crude cell membranes containing ion channels extracted from cultured cells without detergents, we detected open/close signals of the hERG, TRPV1, and NMDA channels on plasma membranes, those of the TRPML1 channels on lysosomal membranes, and open/close signals of the RyR channels on SR membranes. This method will provide a highly versatile drug screening system for ion channels expressed by various cell membranes, including plasma, SR, mitochondrial, Golgi, and lysosomal membranes.

Highlights

Ion channels in cell membranes form pores that facilitate the transport of ions across membranes[1]
We modify our previously developed device to improve its reliability and make it easier to handle for electrical measurements, and apply it to the electrophysiological measurement of various types of ion channels on plasma and organelle membranes (the human Ether-a-go-go-Related Gene[12,13], Transient Receptor Potential Vanilloid 1 (TRPV1)[14], N-methyl-D-aspartate (NMDA)[15], Ryanodine Receptor (RyR)[16], and Transient
Combined with the crude cell membrane fractions containing the target ion channels, as a proof of concept, the capability of the proposed on-chip bilayer lipid membrane (BLM) system was verified in terms of signal detection for various types of ion channels obtained from the plasma and organellar membranes

Summary

Introduction

Ion channels in cell membranes form pores that facilitate the transport of ions (sodium, potassium, calcium, and chloride) across membranes[1]. The functions of a single ion channel have been investigated based on current using an electrophysiological technique known as the patch clamp method[5,6,7] This technique involves manual operation to patch a glass microcapillary to the surface of the cell membrane to form a high-resistance seal in the giga-ohm range. Artificial planar bilayer lipid membrane (BLM) systems have recently been developed to study the ion channel current in a more engineering-oriented approach[8,9] In these systems, the target ion channels are purified from living cells, reconstituted in the BLM, and ion current is measured by a patch clamp amplifier. We recorded the ion current signals of TRPML1 and RyR, which are normally expressed on lysosome membranes and SR membranes, respectively

Methods

Results

Conclusion