Electrophysiological Methods to Assess Peripheral Pain Block in an Anesthetized Rat

Summary

Here, we present a method for assessing the activation and block of sensory axons in peripheral nerves in acute, anesthetized, in vivo rat experiments using two types of spinal electrophysiological measurements that can discriminate the activity of nerve fiber subtypes: Local Field Potentials and Wide Dynamic Range single neuron recordings.

Abstract

In vivo electrophysiological recordings from the lumbar region of the rat spine can be used to study the activation of sensory neural signals in the hindlimbs, as well as the efficacy of therapeutic interventions that interrupt ascending pain signals.

Here, we describe the surgical procedures (laminectomy and sciatic nerve exposure), data collection, and analysis of two complementary electrophysiological signals: local field potential recordings and wide dynamic range single neuron recordings. Both differentiate activation of nerve fiber subclasses based on conduction velocity relative to an electrical stimulation pulse.

Local field potentials (LFP) can differentiate between A α/β and C nerve fibers but cannot detect Aδ activity. They provide information about the global activation of the whole sciatic. Following electrical direct stimulation of the sciatic nerve using a J-cuff style bipolar electrode, area under the curve measurements of the two troughs corresponding to each component are calculated.

Wide dynamic range (WDR) single neuron recordings assess the activity of a subset of sciatic activation, and can differentiate Aα/β, Aδ, and C fiber types. Electrical stimulation can be applied directly to the sciatic as with LFP, or via needle electrodes placed in the plantar surface of the hindpaw. This higher-frequency signal yields high signal-to-noise action potential spikes. Spike counts within predefined latency windows reflect the level of activity of the various fiber types.

To date, these techniques have been used to assess the temporal properties of three different pain block modalities: Direct current electrical nerve block using a custom-made Carbon Separated Nerve Interface Electrode (CSINE), Kilohertz Frequency Alternating Current, and photobiomodulation using an 830 nm laser. This surgical preparation has been used for nerve block interventions for up to 5 h.

Introduction

Directly blocking pain signals in peripheral nerves prior to reaching the spinal cord has potential for several pain therapy applications, such as knee osteoarthritis and post-amputation phantom limb syndrome^1,2,3,4. In general, such pain therapy interventions would reduce/eliminate the systemic side effects noted in current pharmaceutical anesthetics and analgesics^1,2.

Two types of complementary electrophysiological measurements can be recorded from the l....

Protocol

This protocol involving animal experiments was approved by the Institutional Animal Care and Use Committee (IACUC) of Case Western Reserve University (CWRU). Case Western Reserve University is an AAALAC-accredited institution and conforms to appropriate federal, state, and local laws and regulations, as well as institutional policies. Our research group was trained and certified by CWRU staff. When working on IACUC-approved protocols, we adhere to Public Health Service policy and the Guide for the Care and Use of Laboratory Animals (8th edition)³². This procudure is terminal and euthanasia would be triggered (through IACUC approved protocol) up....

Results

The data generated using this protocol allow one to assess the relative amount of block for subclasses of sensory nerve fibers. It also allows the temporal properties of the block to be assessed; both the rate at which block in induced, and the rate of recovery following the cessation of the blocking stimulus³⁵. For the LFP measurements the activation of fibers is determined by calculating the area under the curve for the specific time period corresponding to the fiber type of interest. In

Discussion

This method describes the collection of two complementary electrophysiological signals that can be used to measure the efficacy of potentially therapeutic interventions to block sensory signals in peripheral nerves.

Local field potentials (LFP) can be recorded from the spinal dorsal columns using high impedance (1 MΩ) microelectrodes. They are used to measure the activation of the whole sciatic nerve; the LFP represents the responses of many cells, including excitatory postsynaptic poten.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
1 x 8 multielectrode array, 500 µm spacing between shafts	Microprobes	Custom product	Custom microelectrode array
14 G Angio catheter	Bencton Dickinson	SKU: 381164, GTIN: 382903811649
8-channel bioamplifier	World Precision Instruments	ISO-DAM8A	Head stage
AmScope SM-6T series trinocular zoom stereo microscope 3.5-45x Magnification with articulating arm with ring light	Amscope	SM-8TW2-144S	Surgical microscope
Data acquisition device	Cambridge Electronic Design	Power1401-3A
Dumont #5 forceps	Fine Science Tools	11251-20
Friedman-Pearson Rongeurs (1mm cup size)	Fine Science Tools	16021-14
Harvard apparatus ventilator	Harvard Apparatus	Model 683
Isoflurane, USP	Piramel Critical Care	66794-017-25
Laryngoscope	Sarnova HC	301-B802
Miller 00 laryngoscope blade	Sarnova HC	301-B5100-20
Paired subdermal needles	RhythmLink	RLSND121-1.0	13 mm stainless steel needle electrodes
Rocuronium bromium, USP	Solco	43547-530-10
Spike2	Cambridge Electronic Design
Square wave isolated pulse stimulator	AM-Systems	Model 2100	Biphasic rectangular isolated pulse stimulator
Stereotaxic platform  with adjustable base mounts with post and clamp	David Kopf Instruments	Model 982
SureSite IV 22G x 1" safety catheter	Medline	DYNA22100	22 G intravenous catheter
Syringe driver	Cole Parmer	Model 74900
Ultra precise micro manipulator 10 micron resolution	David Kopf Instruments	Model 961
Universal holder	David Kopf Instruments	Model 1772	Alternate 3D printed connector
U-shaped stereotaxic frame	David Kopf Instruments	Model 1430
Vannas spring scissors - 2 mm cutting edge	Fine Science Tools	15000-03
Vertebrae clamps	David Kopf Instruments	Model 986C