Continuous Video Electroencephalogram during Hypoxia-Ischemia in Neonatal Mice

Summary

This manuscript describes a method for continuous video EEG recordings using multiple depth electrodes in neonatal mice undergoing hypoxia-ischemia.

Abstract

Hypoxia ischemia is the most common cause of neonatal seizures. Animal models are crucial for understanding the mechanisms and physiology underlying neonatal seizures and hypoxia ischemia. This manuscript describes a method for continuous video electroencephalogram (EEG) monitoring in neonatal mice to detect seizures and analyze EEG background during hypoxia ischemia. Use of video and EEG in conjunction allows description of seizure semiology and confirmation of seizures. This method also allows analysis of power spectrograms and EEG background pattern trends over the experimental time period. In this hypoxia ischemia model, the method allows EEG recording prior to injury to obtain a normative baseline and during injury and recovery. Total monitoring time is limited by the inability to separate pups from the mother for longer than four hours. Although, we have used a model of hypoxic-ischemic seizures in this manuscript, this method for neonatal video EEG monitoring could be applied to diverse disease and seizure models in rodents.

Introduction

Hypoxic ischemic encephalopathy (HIE) is a condition that affects 1.5 in 1000 newborns annually and is the most common cause of neonatal seizures^1,2. Infants who survive are at risk for various neurological disabilities such as cerebral palsy, intellectual disability, and epilepsy^3,4,5.

Animal models play a critical role in understanding and investigating the pathophysiology of hypoxia ischemia and neonatal seizures^6,7. A modified V....

Protocol

All animal studies were approved by the Institutional Animal Care and Use Committee (IACUC) of the University of Virginia.

1. Electrode building/cable building

1. Use a unipolar insulated stainless-steel wire (0.005” bare diameter, 0.008” coated) to make an electrode that is connected with a female socket connector (female receptacle connector 0.079).
2. Use a special custom-made cable to connect animals to the amplifier.
   1. Attach a male 4-pin connector (Male connector 0.079”) to 4 channel unity gain impedance matching operational amplifier (op-amp). Attach a 10K resistor to the wires that connect....

Results

Seizure semiology

Neonatal hypoxia-ischemia exposure results in both generalized and focal seizures in mice (Figure 1A-C). Video EEG recordings allow electrographic findings to be correlated to behavior on video. These behaviors were scored using a previously published neonatal rodent behavioral seizure score (BSS)¹⁶. In addition to BSS, we categorized events based on whether the behavior was focal/unilateral, bila.......

Discussion

We have presented a model for continuous video-EEG monitoring in neonatal mice during hypoxic-ischemic seizures. Video analysis in conjunction with EEG allows characterization of seizure semiology. Analysis of EEG allows for extraction of power spectrograms and background amplitude analysis.

Correct and careful placement of electrodes is crucial in this protocol, as injury during electrode placement or inaccurate placement can significantly affect results. Assessment of normal baseline EEG act.......

Materials

Name	Company	Catalog Number	Comments
SURGERY
Ball Point Applicator	Metrex Research	8300-F	i-bond applicator
Cranioplast (Powder/Resin)	Coltene	H00383	Perm Reline/Power
I-Bond	Kulzer GmbH, Germany
LOOK Silk Suture	Surgical Specialities Corporation	SP115	LOOK SP115 Black Braided Silk Non absorbable surgical suture
RS-5168 Botvin Forceps	Roboz Surgical Instrument	RS5168	Forcep for surgery/ligation
RS-5138 Graefe Forceps	Roboz Surgical Instrument	RS5138	Forcep for surgery/ligation
UV light for I-Bond	Blast Lite By First Media	BL778	UV ligth for I-bond
Vannas Microdissecting Scissor	Roboz Surgical Instrument	RS5618	Scissor for ligation
Vet Bond	3M Vetbond	1469SB	Vet Glue
HYPOXIA
Hypoxidial	Starr Life Science
Oxygen sensor	Medical Products		MiniOxI- oxygen analyzer/sensor for hypoxia rig
EEG RECORDING
Female receptacle connector 0.079"	Mill-Max Manufacturing Corp	832-10-024-10-001000	Ordered from Digikey
Grass Amplifier	Natus Neurology Incorporated		Grass Product
LabChart Pro	ADI Instruments		Software to run the system
Male Socket Connector 0.079"	Mill-Max Manufacturing Corp	833-43-024-20-001000	Ordered from Digikey
Operational Amplifier	Texas Instruments, Dallas, TX, USA	TLC2274CD	TLC2274 Quad Low‐Noise Rail‐to Rail Operational Amplifier
Operational Amplifier	Texas Instruments, Dallas, TX, USA	TLC2272ACDR	TLC2274 Quad Low‐Noise Rail‐to Rail Operational Amplifier
Stainless Steel wire	A-M Systems	791400	0.005" Bare/0.008" Coated 100 ft
Ultra-Flexible Wire	McMaster-Carr	9564T1	36 Gauze wire of various color