The Touch and Zap method for in vivo whole-cell patch recording of intrinsic and visual responses of cortical neurons and glial cells.

Adrien E Schramm,Lyle J Graham,Daniele Marinazzo,Thomas Gener

doi:10.1371/journal.pone.0097310

Abstract

Whole-cell patch recording is an essential tool for quantitatively establishing the biophysics of brain function, particularly in vivo. This method is of particular interest for studying the functional roles of cortical glial cells in the intact brain, which cannot be assessed with extracellular recordings. Nevertheless, a reasonable success rate remains a challenge because of stability, recording duration and electrical quality constraints, particularly for voltage clamp, dynamic clamp or conductance measurements. To address this, we describe “Touch and Zap”, an alternative method for whole-cell patch clamp recordings, with the goal of being simpler, quicker and more gentle to brain tissue than previous approaches. Under current clamp mode with a continuous train of hyperpolarizing current pulses, seal formation is initiated immediately upon cell contact, thus the “Touch”. By maintaining the current injection, whole-cell access is spontaneously achieved within seconds from the cell-attached configuration by a self-limited membrane electroporation, or “Zap”, as seal resistance increases. We present examples of intrinsic and visual responses of neurons and putative glial cells obtained with the revised method from cat and rat cortices in vivo. Recording parameters and biophysical properties obtained with the Touch and Zap method compare favourably with those obtained with the traditional blind patch approach, demonstrating that the revised approach does not compromise the recorded cell. We find that the method is particularly well-suited for whole-cell patch recordings of cortical glial cells in vivo, targeting a wider population of this cell type than the standard method, with better access resistance. Overall, the gentler Touch and Zap method is promising for studying quantitative functional properties in the intact brain with minimal perturbation of the cell's intrinsic properties and local network. Because the Touch and Zap method is performed semi-automatically, this approach is more reproducible and less dependent on experimenter technique.

Highlights

The patch-clamp electrophysiological technique was developed to record currents from single membrane channels [1] and later applied to record macroscopic currents and voltages in the socalled whole-cell configuration [2]
There were no significant differences in measured properties as function of species; unless mentioned neuron and glial cell recordings are pooled across species
We have described Touch and Zap, a revised method for obtaining the whole-cell patch clamp configuration for neurons and glial cells, here applied to in vivo blind patch recordings in rat and cat cortex

Summary

Introduction

The patch-clamp electrophysiological technique was developed to record currents from single membrane channels [1] and later applied to record macroscopic currents and voltages in the socalled whole-cell configuration [2]. The introduction of in vivo two-photon microscopy has allowed visual monitoring of whole-cell recordings in vivo [11,12,13,14,15], but since this approach is limited to upper layers of the cortex, the blind patch method remains an important technique in vivo. The refinement of this technique has recently reached a new level, with the automated in vivo protocol described by Kodandaramaiah et al [16]. For reviews on the methodological aspect of the technique see [17,18,19]

Methods

Results

Conclusion