A Murine Model of Subarachnoid Hemorrhage

Summary

A standardized mouse model of subarachnoid hemorrhage by intraluminal Circle of Willis perforation is described. Vessel perforation and subarachnoid bleeding are monitored by intracranial pressure monitoring. In addition various vital parameters are recorded and controlled to maintain physiologic conditions.

Abstract

In this video publication a standardized mouse model of subarachnoid hemorrhage (SAH) is presented. Bleeding is induced by endovascular Circle of Willis perforation (CWp) and proven by intracranial pressure (ICP) monitoring. Thereby a homogenous blood distribution in subarachnoid spaces surrounding the arterial circulation and cerebellar fissures is achieved. Animal physiology is maintained by intubation, mechanical ventilation, and continuous on-line monitoring of various physiological and cardiovascular parameters: body temperature, systemic blood pressure, heart rate, and hemoglobin saturation. Thereby the cerebral perfusion pressure can be tightly monitored resulting in a less variable volume of extravasated blood. This allows a better standardization of endovascular filament perforation in mice and makes the whole model highly reproducible. Thus it is readily available for pharmacological and pathophysiological studies in wild type and genetically altered mice.

Introduction

SAH is the stroke subtype with the least beneficial outcome for patients: 40% of the patients die within a month after the bleeding¹ and survivors rarely have a clinically favorable outcome.

The large majority of spontaneous SAHs (80%) are caused by rupture of intracranial aneurysms which are mostly located along the anterior and posterior communicating artery, the basilar artery, and middle cerebral artery (MCA)².

Such aneurysms are difficult to model in animals and therefore animal models of SAH are either performed by injection of blood into the subarachnoid space/cerebral ventricles or b....

Protocol

All surgical procedures were subjected to ethical review and approved by the government of Upper Bavaria (reference number: 55.2-1-54-2532.3-13-13 and -2532-136-11). Animals are male C57BL/6 mice with a body weight of approximately 25 g.

1. Animal Preparation

1. Induce anesthesia by putting the mouse into a chamber. Flush the chamber with 5% isoflurane until the animal loses consciousness.
2. Inject premixed anesthetics intraperitoneally: fentanyl (0.05 mg/kg), midazolam (5 mg/kg) and medetomidine (0.5 mg/kg). Check reflexes before and regularly during the procedure. Reinject one third of the initial amount hourly to maintai....

Results

Mortality

Once the surgery technique is mastered the procedure does not elicit any intraoperative mortality. Also bleeding can be achieved in virtually all animals. Postoperative mortality is 30-40% with most animals dying on day 1 after surgery (Figure 5).

ICP values after SAH

The ICP before bleeding is around 4 mmHg. Bleeding results in a sharp increase of the ICP up to 120 mmHg. ICP values then stabilize within 5 min at appr.......

Discussion

Treatment options after SAH are scarce and mostly inefficacious. Therefore the pathophysiology of post-hemorrhagic brain damage needs to be further understood in order to identify new therapeutic targets and develop novel therapeutic approaches. Standardized and well reproducible animal models in genetically modified animals, i.e. mice, are crucial for such investigations. The CWp model has become a widely used model for SAH as it resembles the pathophysiology in humans closely; however, its use in mice is hampe.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Equipment
operation microscope	Leica	KL2500
isoflurane vaporizer	Harvard Instruments	Continuous Flow Vaporizer
respirator	Hugo Sachs	Minivent 845
microcapnograph	Hugo Sachs	Type 340
temperature controller	FHC	DC Temperature Controller
dental drill	Paggen	Labset- N
ICP monitor	Codman	ICP monitor
blood pressure monitor	AD Instruments	Bridge Amp FE221
syringe pump	World Precision Instruments	SP101IZ
pulsoximeter	Kent Scientific	MouseSTAT
LDF	Perimed	Periflux 5000
analog data monitor	AD Instruments	Power Lab 16/35
Material
cement for ICP probe fixation	Speiko	Carboxylate cement
glue for LDF probe fixation	Bob Smith Industries	Cyanoacrylate glue (Maxi Cure and Insta Set)
venous catheter	Johnson Johnson	Jelco winged i.v. catheter; REF 4076	modified intubation tube
tubing for femoral catheter	Smiths Medical	Fine Bore Polythene Tubing; ID 0.28 mm OD 0.61 mm; REF 800/100/100	cut to 30 cm length
filament for vessel perforation	Ethicon	Prolene 5-0	cut to 12 mm length
surgical equipment	Fine Scientific Instruments	forceps medical #5, vessel scissors 8 cm, microclip 4 mm jaw