A combined protocol for isolation, culture, and patch-clamp recording of dorsal root ganglion neurons

Abstract

The dorsal root ganglion (DRG) neurons are crucial in transmitting sensory information from the peripheral nervous system to the central nervous system, including touch, pain, temperature, and proprioception. Understanding the functions and mechanisms of DRG neurons is essential for studying sensory processing and developing efficient treatments for sensory disorders. In addition, electrophysiological patch-clamp recording is a powerful and classical tool to study the functions and mechanisms of the nervous system. Building upon the strategies outlined in published works and our group's abundant research experience in DRG neurons' functions by patch-clamp, we have summarized and put forward a comprehensive step-by-step protocol combining juvenile rat DRG neuron isolation and culture, and patch-clamp recording. This protocol would be a powerful guidance document for neuroscience researchers to study sensory DRG neurons' physiological and pathological functions using electrophysiological tools.

Keywords: Culture; Isolation; Patch-clamp recording; Sensory dorsal root ganglion neurons.

Figure 1

Images show each step of isolating DRG neurons from postnatal 5-day Sprague-Dawley rats. See more details in Step 2

Figure 2

Images show acutely isolated DRG neurons (after ~2 h culture). A Acutely isolated DRG neurons. B An enlarged image from Panel A

Figure 3

Images show the PatchMaster software windows of each step of the whole-cell patch-clamp recording mode. The insert in Panel A is a schematic diagram of the patch-clamp setup. The inserts in Panels B–D are photographs showing the relative position of a patch-clamp electrode and the DRG neuron. See more details in Step 5

Figure 4

The action potentials, currents, and membrane capacitance signals from acutely isolated DRG neurons. A Membrane potential (Vm) signals (Upper), evoked by 500 pA current injection (Lower protocol, stim). B Membrane current (Im) signals (Upper), evoked by ramp depolarization (Lower protocol, stim). C Membrane capacitance (Cm, Upper) signals, evoked by 200 ms depolarization from –70 mV to 0 mV (Lower protocol, stim), from Ca²⁺-free solution (Left) and 2 mmol/L Ca²⁺-containing solution (Right), respectively. The Gm and Gs reflect the quality of capacitance recording, and Im is the current signal (Middle)