The Pilocarpine Model of Temporal Lobe Epilepsy and EEG Monitoring Using Radiotelemetry System in Mice

Summary

This manuscript describes a method of inducing status epilepticus by systemic pilocarpine injection and of monitoring spontaneous recurrent seizures in live animals using a wireless telemetry video and electroencephalogram system. This protocol can be utilized for studying the pathophysiologic mechanisms of chronic epilepsy, epileptogenesis, and acute seizures.

Abstract

Temporal lobe epilepsy (TLE) is a common neurological disorder in adulthood. For translational studies of chronic epilepsy, pilocarpine-induced status epilepticus (SE) is frequently selected to recapitulate spontaneous recurrent seizures (SRS). Here we present a protocol of SE induction by intraperitoneal (i.p.) injection of pilocarpine and monitoring of chronic recurring seizures in live animals using a wireless telemetry video and electroencephalogram (EEG) system. We demonstrated notable behavioral changes that need attention after pilocarpine injection and their correlation with hippocampal neuronal loss at 7 days and 6 weeks post-pilocarpine. We also describe the experimental procedures of electrode implantation for video and EEG recording, and analysis of the frequency and duration of chronic recurrent seizures. Finally, we discuss the possible reasons why the expected results are not achieved in each case. This provides a basic overview of modeling chronic epilepsy in mice and guidelines for troubleshooting. We believe this protocol can serve as a baseline for suitable models of chronic epilepsy and epileptogenesis.

Introduction

TLE is one of the most common acquired epilepsies¹. People with epilepsy experience recurrent seizures as a result of abnormal neuronal activities in the brain^2,3. Given that TLE is often intractable, it is crucial to understand the basic mechanisms underlying the development of epilepsy.

Animal models that can recapitulate the key characteristics of human TLE can offer better appreciation of TLE pathophysiology, allowing us to readily monitor and manipulate critical factors in epileptogenesis. Among them, chemoconvulsants-induced SE has been widely used

Protocol

All experimental procedures were approved by the Ethics Committee of the Catholic University of Korea and were carried out in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals (NIH Publications No. 80-23).

1. SE Induction

1. Purchase 8-week-old male C57BL/6NHsd mice and weigh each mouse. Then, use a marker pen to mark the tails of all mice for their easy identification during SE induction.
2. Calculate the amount of scopolamine m.......

Discussion

This work describes the experimental procedures for the SE induction and the evaluation of chronic seizures.

Several factors can affect successful SE induction. Accurate behavioral monitoring according to the Racine scale is critical for the development of SRS. Head nodding, forelimb clonus, rearing, and falling are the behavioral hallmarks of acute seizures developing into SE phase^4,16. Once the first motor seizure is detected, the le.......

Materials

Name	Company	Catalog Number	Comments
C57BL/6	Envigo	C57BL/6NHsd
Scopolamine methyl nitrate	Sigma	S2250	Make 10X stock
Terbutaline hemisulfate salt	Sigma	T2528	Make 10X stock
Pilocarpine hydrochloride	Sigma	P6503
Intensive care unit	Daejong instrument industry Co., Ltd.		28~30℃
Ketamine hydrochloride	Yuhan corporation
Xylazine hydrochloride	Bayer Korea
Diazepam	SAMJIN
Castor oil (Kolliphor EL)	Sigma	C5135	Polyoxyl 35 hydrogenated castor oil
Saline	Daihan pharm. Co.
5% Dextrose	Daihan pharm. Co.
Iodine solution (Povidin)	Firson
vet ointment (Terramycin)	Pfizer
Blue Nylon	AILEE	NB617
Mupirocin (Bearoban)	Daewoong Pharmaceutical Co., Ltd
Ketoprofen	Samchundang Pharm. Co., Ltd		5 mg/kg
Gentamicin	Huons, Ltd.		5 mg/kg
1 mL syringe	Sung shim medial Co., Ltd.
26 guage needle	Sung shim medial Co., Ltd.		26 G * 13 mm (1/2")
30 guage needle	Sung shim medial Co., Ltd.		30 G * 13 mm (1/2")
Razor blade	Dorco
Drill	Saeshin precision Co., Ltd.		207A, 35K (speed)
Telemetry video/EEG system	Data sciences International. Inc.	Version 5.20-SP6
Implantable transmitter	Data sciences International. Inc.	ETA-F10
Screw	Sungho Steel		M1.4, 2 mm length stainless steel
Vertex dental material	Dentimex
Acetone	Duksan pure chemicals Co., Ltd.		CAS 67-64-1
Paraformaldehyde (PFA)	millipore	1.04005.1000	4 %
Sucrose	Sigma	S9378	30 % solution in 0.01 M PBS
Cresyl violet acetate	Sigma	C5042
Ethanol	EMD Millipore Co.	UN1170
xylene	Duksan pure chemicals Co., Ltd.	UN1307
Acetic acid glacial	Junsei chemical	31010-0350
FSC33 Clear	Leica biosystems		OCT compound for tissue freezing
DPX Mounting for histology	Sigma	6522
Forceps	Fine science tools	11002-12
Scissors	Solco biomedical	02-2445
Stereotaxic frame	David Kopf Instruments	E51070012