Laser-Induced Brain Injury in the Motor Cortex of Rats

Summary

The protocol presented here shows a technique to create a rodent model of brain injury. The method described here uses laser irradiation and targets motor cortex.

Abstract

A common technique for inducing stroke in experimental rodent models involves the transient (often denoted as MCAO-t) or permanent (designated as MCAO-p) occlusion of the middle cerebral artery (MCA) using a catheter. This generally accepted technique, however, has some limitations, thereby limiting its extensive use. Stroke induction by this method is often characterized by high variability in the localization and size of the ischemic area, periodical occurrences of hemorrhage, and high death rates. Also, the successful completion of any of the transient or permanent procedures requires expertise and often lasts for about 30 minutes. In this protocol, a laser irradiation technique is presented that can serve as an alternative method for inducing and studying brain injury in rodent models.

When compared to rats in the control and MCAO groups, the brain injury by laser induction showed reduced variability in body temperature, infarct volume, brain edema, intracranial hemorrhage, and mortality. Furthermore, the use of a laser-induced injury caused damage to the brain tissues only in the motor cortex unlike in the MCAO experiments where destruction of both the motor cortex and striatal tissues is observed.

Findings from this investigation suggest that laser irradiation could serve as an alternative and effective technique for inducing brain injury in the motor cortex. The method also shortens the time for completing the procedure and does not require expert handlers.

Introduction

Globally, stroke is the second leading cause of death and the third leading cause of disability¹. Stroke also leads to severe disability, often requiring extra care from medical staff and relatives. There is, therefore, a need to understand the complications associated with the disorder and improve the potential for more positive outcomes. 

The use of animal models is the initial step to understanding diseases. To ensure the best research outcomes, a typical model would include a simple technique, affordability, high reproducibility, and minimal variability. The determinants in ischemic stroke models include bra....

Protocol

The following procedure was conducted according to the Guidelines of the Use of Experimental Animals of the European Community. The experiments were also approved by the Animal Care Committee at the Ben-Gurion University of the Negev.

1. Animal selection and preparation

1. Select 65 male Sprague-Dawley rats weighing 300 to 350 g with no overt pathology for this procedure. The smaller size poses technical difficulties for the MCAO procedure.
2. Assign 3 rats per cage and let the.......

Discussion

It is fair to assume that the laser technique is minimally invasive, given that no deaths or SAH occurred in the laser group. The primary cause of death and SAH is the damage to blood vessels that leads to an elevation of intracranial pressure (ICP), as shown in the original MCAO techniques¹⁰. The absence of death and SAH in the laser group is likely due to the specific effects of lasers: they do not have direct impact on blood vessels and can induce coagulation in case of leakage. Low infarct vol.......

Materials

Name	Company	Catalog Number	Comments
2,3,5-Triphenyltetrazolium chloride	SIGMA - ALDRICH	298-96-4
50% trichloroacetic acid	SIGMA - ALDRICH	76-03-9
Brain & Tissue Matrices	SIGMA - ALDRICH	15013
Cannula Venflon 22 G	KD-FIX	1.83604E+11
Centrifuge Sigma 2-16P	SIGMA - ALDRICH	Sigma 2-16P
Compact Analytical Balances	SIGMA - ALDRICH	HR-AZ/HR-A
Digital Weighing Scale	SIGMA - ALDRICH	Rs 4,000
Dissecting scissors	SIGMA - ALDRICH	Z265969
Eppendorf pipette	SIGMA - ALDRICH	Z683884
Eppendorf Tube	SIGMA - ALDRICH	EP0030119460
Ethanol 96 %	ROMICAL		Flammable Liquid
Evans Blue 2%	SIGMA - ALDRICH	314-13-6
Fluorescence detector	Tecan, Männedorf Switzerland	model Infinite 200 PRO multimode reader
Heater with thermometer	Heatingpad-1	Model: HEATINGPAD-1/2
Infusion Cuff	ABN	IC-500
Isofluran, USP 100%	Piramamal Critical Care, Inc	NDC 66794-017
Multiset	TEVA MEDICAL	998702
Olympus BX 40 microscope	Olympus
Optical scanner	Canon	Cano Scan 4200F
Petri dishes	SIGMA - ALDRICH	P5606
Scalpel blades 11	SIGMA - ALDRICH	S2771
Sharplan 3000 Nd:YAG (neodymium-doped yttrium aluminum garnet) laser machine	Laser Industries Ltd
Stereotaxic head holder	KOPF	900LS
Sterile Syringe 2 ml	Braun	4606027V
Syringe-needle 27 G	Braun	305620