An automated planar patch-clamp approach to measure the membrane potential and resting membrane currents in a human cerebrovascular endothelial cell line

Abstract

Background

The conventional “whole-cell patch-clamp” recording technique is widely used to measure the resting membrane potential (V_M) and to dissect the underlying membrane ionic conductances in isolated vascular endothelial cells.

New method

Herein, we assessed whether the automated patch-clamp (APC) technology, which replaces the traditional patch-pipette with a planar substrate to permit researchers lacking formal training in electrophysiology to generate large amounts of data in a relatively short time, can be used to characterize the bioelectrical activity of vascular endothelial cells. We assessed whether the Port-a-Patch planar patch-clamp system, which is regarded as the smallest electrophysiological rig available on the market, can be used to measure the V_M and resting membrane currents in the human cerebrovascular endothelial cell line, hCMEC/D3.

Comparison with existing methods

We demonstrated that the Port-a-Patch planar patch-clamp system provides the same values of the resting V_M as those provided by the conventional patch-clamp technique. Furthermore, the APC technology provides preliminary data demonstrating that the resting V_M of hCMEC/D3 cells is primarily contributed by Cl^- and Na⁺, as demonstrated with the patch-clamp technique for many other endothelial cell types.

Conclusions

The Port-a-Patch planar patch-clamp system can be successfully used to measure the resting V_M and the underlying membrane ionic conductances in hCMEC/D3 cells. We envisage that this easy-to-use APC system could also be extremely useful for the investigation of the membrane currents that can be activated by chemical, thermal, optical, and mechanical stimuli in this cell line as well as in other types of isolated vascular endothelial cells.