Polygraphic Recording Procedure for Measuring Sleep in Mice

Summary

The recording of electroencephalogram (EEG) and electromyogram (EMG) in freely behaving mice is a critical step to correlate behavior and physiology with sleep and wakefulness. The experimental protocol described herein provides a cable-based system for acquiring EEG and EMG recordings in mice.

Abstract

Recording of the epidural electroencephalogram (EEG) and electromyogram (EMG) in small animals, like mice and rats, has been pivotal to study the homeodynamics and circuitry of sleep-wake regulation. In many laboratories, a cable-based sleep recording system is used to monitor the EEG and EMG in freely behaving mice in combination with computer software for automatic scoring of the vigilance states on the basis of power spectrum analysis of EEG data. A description of this system is detailed herein. Steel screws are implanted over the frontal cortical area and the parietal area of 1 hemisphere for monitoring EEG signals. In addition, EMG activity is monitored by the bilateral placement of wires in both neck muscles. Non-rapid eye movement (Non-REM; NREM) sleep is characterized by large, slow brain waves with delta activity below 4 Hz in the EEG, whereas a shift from low-frequency delta activity to a rapid low-voltage EEG in the theta range between 6 and 10 Hz can be observed at the transition from NREM to REM sleep. By contrast, wakefulness is identified by low- to moderate-voltage brain waves in the EEG trace and significant EMG activity.

Introduction

Technical advances have often precipitated quantum leaps in the understanding of neurobiological processes. For example, Hans Berger's discovery in 1929 that electrical potentials recorded from the human scalp took the form of sinusoidal waves, the frequency of which was directly related to the level of wakefulness of the subject, led to rapid advances in the understanding of sleep-wake regulation, in both animals and humans alike.¹ To this day the electroencephlogram (EEG), in conjunction with the electromyogram (EMG), i.e., electrical activity produced by skeletal muscles, represents the data "backbone" of nearly every experimental an....

Protocol

Ethics Statement: Procedures involving animal subjects have been approved by the Institutional Animal Experiment Committee at the University of Tsukuba.

1. Preparation of Electrodes and Cables for EEG/EMG Recordings

1. Prepare EEG/EMG recording electrode according to the following procedure.
   Note: The electrode is disposable and can be used only for 1 animal. Plan carefully the wiring configuration for all connectors. Place marks on the connectors for the correct orientation.
   1. Solder each pin of a 4-pin header to a 2-cm stainless steel wire. In brief, hold one end of the wire to the pin, place a hot soldering iron....

Results

Figure 1B illustrates examples of the mouse EEG in the different vigilance states. As shown in Table 1, epochs are classified as NREM sleep if the EEG shows large, slow brain waves with a delta rhythm below 4 Hz and the EMG has only a weak or no signal. Epochs are classified as REM sleep if the EEG shows rapid low-voltage brain waves in the theta range between 6 and 10 Hz and the EMG shows low amplitude. Other epochs should be classified as wakefulness (<.......

Discussion

This protocol describes a set-up for EEG/EMG recordings that allows the assessment of sleep and wakefulness under low-noise, cost-effective, and high-throughput conditions. Due to the small size of the EEG/EMG electrode head assembly, this system can be combined with other implants for intra-brain experiments, including optogenetics (optical fiber implantation) or, in conjunction with simultaneous cannula implantation, microinfusion of drugs into the mouse brain.³¹ Moreover, the design of the electrode head as.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
4-pin header	Hirose	A3B-4PA-2DSA(71)
Ampicillin	Meiji Seika
Analog-to-digital converter	Contec	AD16-16U(PCIEV)
Caffeine	Sigma	C0750
Carbide cutter	Minitor	B1055
Crimp housing	Hirose	DF11-4DS-2C
Crimp socket	Hirose	DF11-30SC
Dental cement (Toughron Rebase)	Miki Chemical Product
Epoxy adhesive	Konishi	#16351
FFC/FPC connector	Honda Tsushin Kogyo	FFC-10BMEP1(B)
Flat cable	Hitachi Cable	20528-ST LF
Instant glue (Aron Alpha A)	Toagosei	N/A
Meloxicam	Boehringer Ingelheim	N/A
Pentobarbital	Kyoritsu Seiyaku	N/A
Signal amplifier	Biotex	N/A
Sleep recording chamber	APL	N/A
SleepSign software	Kissei Comtec	N/A	for EEG/EMG recording/analysis
Slip ring	Biotex	N/A
Stainless steel screw	Yamazaki	N/A	φ1.0 × 2.0
Stainless steel wire	Cooner Wire	AS633