Modeling Stroke in Mice: Transient Middle Cerebral Artery Occlusion via the External Carotid Artery

Summary

Different models of middle cerebral artery occlusion (MCAo) are used in experimental stroke research. Here, an experimental stroke model of transient MCAo via the external carotid artery (ECA) is described, which aims to mimic human stroke, in which the cerebrovascular thrombus is removed due to spontaneous clot lysis or therapy.

Abstract

Stroke is the third most common cause of mortality and the leading cause of acquired adult disability in developed countries. To date, therapeutic options are limited to a small proportion of stroke patients within the first hours after stroke. Novel therapeutic strategies are being extensively investigated, especially to prolong the therapeutic time window. These current investigations include the study of important pathophysiological pathways after stroke, such as post-stroke inflammation, angiogenesis, neuronal plasticity, and regeneration. Over the last decade, there has been increasing concern about the poor reproducibility of experimental results and scientific findings among independent research groups. To overcome the so-called “replication crisis”, detailed standardized models for all procedures are urgently needed. As an effort within the “ImmunoStroke” research consortium (https://immunostroke.de/), a standardized mouse model of transient middle cerebral artery occlusion (MCAo) is proposed. This model allows the complete restoration of the blood flow upon removal of the filament, simulating the therapeutic or spontaneous clot lysis that occurs in a large proportion of human strokes. The surgical procedure of this “filament” stroke model and tools for its functional analysis are demonstrated in the accompanying video.

Introduction

Stroke is one of the most common causes of death and disability worldwide. Although there are mainly two distinct forms of stroke, ischemic and hemorrhagic, 80–85% of all stroke cases are ischemic¹. Currently, only two treatments are available for patients with ischemic stroke: pharmacological treatment with recombinant tissue plasminogen activator (rtPA) or mechanical thrombectomy. However, due to the narrow therapeutic time window and multiple exclusion criteria, only a select number of patients can benefit from these specific treatment options. Over the last two decades, preclinical and translational stroke research has focused on the ....

Protocol

The experiments reported in this video were conducted following the national guidelines for the use of experimental animals, and the protocols were approved by the German governmental committees (Regierung von Oberbayern, Munich, Germany). Ten-week-old male C57Bl/6J mice were used and housed under controlled temperature (22 ± 2 °C), with a 12 h light-dark cycle period and access to pelleted food and water ad libitum.

1. Preparation of the material and instruments

Discussion

The present protocol describes an experimental stroke model based on the consensus agreement of a German multicenter research consortium (“ImmunoStroke”) to establish a standardized transient MCAo model. The transient MCAo model established by introducing a silicon-coated filament through the ECA to the origin of the MCA is one of the most widely used stroke models to achieve arterial reperfusion after a delimitated occlusion period. Therefore, this procedure can be considered a translationally relevant strok.......

Materials

Name	Company	Catalog Number	Comments
45° ramp	H&S Kunststofftechnik		height: 18 cm
5/0 threat	Pearsalls	10C103000
5 mL Syringe	Braun
Acetic Acid	Sigma Life Science	695092
Anesthesia system for isoflurane	Drager
Bepanthen pomade	Bayer
C57Bl/6J mice	Charles River	000664
Clamp	FST	12500-12
Clip	FST	18055-04
Clip holder	FST	18057-14
Cotons	NOBA Verbondmitel Danz	974116
Cresyl violet	Sigma Life Science	C5042-10G
Cryostat	Thermo Scientific CryoStarNX70
Ethanol 70%	CLN Chemikalien Laborbedorf	521005
Ethanol 96%	CLN Chemikalien Laborbedorf	522078
Ethanol 99%	CLN Chemikalien Laborbedorf	ETO-5000-99-1
Filaments	Doccol	602112PK5Re
Fine 45 angled forceps	FST	11251-35
Fine forceps	FST	11252-23
Fine Scissors	FST	14094-11
Glue	Orechseln	BSI-112
Hardener Glue	Drechseln & Mehr	BSI-151
Heating blanket	FHC DC Temperature Controller
Isoflurane	Abbot	B506
Isopentane	Fluka	59070
Ketamine	Inresa Arzneimittel GmbH
Laser Doppler	Perimed	PF 5010 LDPM, Periflux System 5000
Laser Doppler probe	Perimed	91-00123
Phosphate Buffered Saline pH: 7.4	Apotheke Innestadt Uni Munchen	P32799
Recovery chamber	Mediheat
Roti-Histokit mounting medium	Roth	6638.1
Saline solution	Braun	131321
Scalpel	Feather	02.001.30.011
Silicon-coated filaments	Doccol	602112PK5Re
Stereomicropscope	Leica	M80
Superfrost Plus Slides	Thermo Scientific	J1800AMNZ
Vannas Spring Scissors	FST	15000-00
Xylacine	Albrecht