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In This Article

  • Summary
  • Abstract
  • Introduction
  • Protocol
  • Representative Results
  • Discussion
  • Acknowledgements
  • Materials
  • References
  • Reprints and Permissions

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.......

Representative Results

No deaths or SAH were registered in either the control or experimental groups (Table 1). The MCAO group had a 20% rate of both mortality and SAH.

The relative body temperature changes in the rats of both groups were also similar, despite a difference in the variability of both groups (Table 1).

There was a significantly worse NSS in both the laser (16 ± 1.1) and MCAO (20 ± 1.5) models, compared to the sham-operated control.......

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.......

Acknowledgements

We would like to thank the Department of Anesthesiology of Soroka University Medical Center and the laboratory staff of Ben-Gurion University of the Negev for their help in the performance of this experiment.

....

Materials

NameCompanyCatalog NumberComments
2,3,5-Triphenyltetrazolium chlorideSIGMA - ALDRICH298-96-4
50% trichloroacetic acidSIGMA - ALDRICH76-03-9
Brain & Tissue MatricesSIGMA - ALDRICH15013
Cannula Venflon 22 GKD-FIX1.83604E+11
Centrifuge Sigma 2-16PSIGMA - ALDRICHSigma 2-16P
Compact Analytical BalancesSIGMA - ALDRICHHR-AZ/HR-A
Digital Weighing ScaleSIGMA - ALDRICHRs 4,000
Dissecting scissorsSIGMA - ALDRICHZ265969
Eppendorf pipetteSIGMA - ALDRICHZ683884
Eppendorf TubeSIGMA - ALDRICHEP0030119460
Ethanol 96 %ROMICALFlammable Liquid
Evans Blue 2%SIGMA - ALDRICH314-13-6
Fluorescence detectorTecan, Männedorf Switzerlandmodel Infinite 200 PRO multimode reader
Heater with thermometerHeatingpad-1Model: HEATINGPAD-1/2
Infusion CuffABNIC-500
Isofluran, USP 100%Piramamal Critical Care, IncNDC 66794-017
MultisetTEVA MEDICAL998702
Olympus BX 40 microscopeOlympus
Optical scannerCanonCano Scan 4200F
Petri dishesSIGMA - ALDRICHP5606
Scalpel blades 11SIGMA - ALDRICHS2771
Sharplan 3000 Nd:YAG (neodymium-doped yttrium aluminum garnet) laser machineLaser Industries Ltd
Stereotaxic head holderKOPF900LS
Sterile Syringe 2 mlBraun4606027V
Syringe-needle 27 GBraun305620

References

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Laser induced Brain InjuryMotor CortexSprague Dawley RatsNd YAG LaserNeurological Severity ScoreTTC StainingEvans Blue DyeTraumatic Brain InjuryBrain Infarct Volume

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