Monitoring voltage fluctuations of intracellular membranes

Masoud Sepehri Rad,Bradley J Baker,Oliver Braubach,Lawrence B Cohen

doi:10.1038/s41598-018-25083-7

Abstract

In eukaryotic cells, the endoplasmic reticulum (ER) is the largest continuous membrane-enclosed network which surrounds a single lumen. Using a new genetically encoded voltage indicator (GEVI), we applied the patch clamp technique to cultured HEK293 cells and neurons and found that there is a very fast electrical interaction between the plasma membrane and internal membrane(s). This discovery suggests a novel mechanism for interaction between the external membrane and internal membranes as well as mechanisms for interactions between the various internal membranes. The ER may transfer electrical signals between the plasma membrane and other internal organelles. The internal membrane optical signal is reversed in polarity but has a time course similar to that of the plasma membrane signal. The optical signal of the GEVI in the plasma membrane is consistent from trial to trial. However, the internal signal decreases in size with repeated trials suggesting that the electrical coupling is degrading and/or the resistance of the internal membrane is decaying.

Highlights

In eukaryotic cells, the endoplasmic reticulum (ER) is the largest continuous membrane-enclosed network which surrounds a single lumen
In this paper we introduce mutations to the loop regions of the Voltage Sensing Domain (VSD) of an ArcLight-type GEVI14 resulting in a distinct intracellular optical signal that responds to changes in the plasma membrane potential
The plasma membrane fluorescence gets dimmer during depolarization steps while the internal signal gets brighter when expressed in HEK cells

Summary

Introduction

The endoplasmic reticulum (ER) is the largest continuous membrane-enclosed network which surrounds a single lumen. GEVIs are fluorescent proteins which yield optical signals in response to changes in membrane potential. In this paper we introduce mutations to the loop regions of the Voltage Sensing Domain (VSD) of an ArcLight-type GEVI14 resulting in a distinct intracellular optical signal that responds to changes in the plasma membrane potential. The possibility that the other 80% of the cells do not experience a change in internal membrane potentials is real but relative low given that high intracellular fluorescence is an excellent predictor for observation of an internal signal.

Methods

Results

Conclusion