An <em>In Vivo</em> Blood-brain Barrier Permeability Assay in Mice Using Fluorescently Labeled Tracers

Kavi Devraj; Sylvaine Gu&#233;rit; Jakranka Macas; Yvonne Reiss

doi:10.3791/57038

In Vivo barriera emato - encefalica permeabilità saggio nei topi utilizzando fluorescent...

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JoVE Journal Neuroscience

An In Vivo Blood-brain Barrier Permeability Assay in Mice Using Fluorescently Labeled Tracers

In Vivo barriera emato - encefalica permeabilità saggio nei topi utilizzando fluorescente etichettati traccianti

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09:35 min

February 26, 2018

DOI: 10.3791/57038-v

Kavi Devraj^1,2, Sylvaine Guérit¹, Jakranka Macas¹, Yvonne Reiss¹

¹Institute of Neurology (Edinger Institute),Goethe University Hospital, ²Pharmazentrum Frankfurt, Institute for General Pharmacology and Toxicology,Goethe University Hospital

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Overview

This study presents a method for assessing mouse brain vascular permeability using intraperitoneally injected fluorescent tracers. It aims to investigate blood-brain barrier dysfunction, relevant in conditions such as stroke and Alzheimer's disease, by providing quantitative and visual data on neurovascular permeability.

Key Study Components

Area of Science

Neuroscience
Vascular Biology
Biomedical Research

Background

Assessing blood-brain barrier dysfunction is critical for understanding various neurological disorders.
The use of fluorescent tracers allows for both quantitative and qualitative analysis of permeability.
This method can provide insights on recovery from vascular dysfunction after therapy.
Implications extend towards the diagnosis and treatment of brain edema occurring in multiple diseases.

Purpose of Study

To develop a reliable assay for measuring vascular permeability in mouse brain models.
To evaluate therapeutic interventions aimed at restoring blood-brain barrier functionality.
To facilitate deeper investigations into the underlying mechanisms of neurovascular permeability changes.

Methods Used

This study employs an intraperitoneal injection method using fluorescent tracers.
A mouse model is utilized to examine the integrity of the blood-brain barrier.
Concurrent quantitative assessment of permeability is performed via fluorometry, alongside fluorescence microscopy for regional visualization.
The complete procedure, from injection to analysis, takes approximately 5–6 hours for 10 mice.
Detailed steps include anesthesia, cardiac perfusion, tissue harvesting, and subsequent fluorescence measurements.

Main Results

The method allows for effective quantification of blood-brain barrier permeability.
Fluorescent microscopy provides visual confirmation of tracer localization and permeability assessments.
The procedure has shown potential applicability in studying recovery processes following various neurological injuries.

Conclusions

This study demonstrates a specific, efficient assay for examining vascular permeability in mouse models.
It enables researchers to further explore therapeutic impacts on blood-brain barrier integrity.
The method contributes to understanding the mechanisms of vascular dysfunction in neurological disorders.

Frequently Asked Questions

What are the advantages of using this permeability assay?

This assay provides a quantitative approach to measure vascular permeability and allows for both fluorometric and microscopic methods for comprehensive analysis.

How is the mouse model prepared for the assay?

Mice are injected with a fluorescent tracer, deeply anesthetized, and undergo cardiac perfusion before tissue harvesting for analysis.

What types of data can be obtained from this method?

The method yields quantitative permeability data from fluorometric readings and detailed visualizations from fluorescence microscopy.

How can this technique be applied in research?

It can be used to study the blood-brain barrier's response to treatments and its role in various neurological diseases, like strokes or brain tumors.

Are there any limitations to this method?

The assay's effectiveness relies on proper perfusion technique and timely processing, which requires careful attention to procedural details.

What are the key safety considerations for handling mice during the procedure?

Ensure adherence to institutional animal care protocols, including proper anesthesia and humane treatment throughout the procedure.

Qui presentiamo un'analisi di permeabilità vascolare del cervello del mouse usando iniezione intraperitoneale di traccianti fluorescenti seguita da aspersione che è applicabile ai modelli animali di disfunzione della barriera emato - encefalica. Un hemi-cervello è usato per valutare quantitativamente permeabilità e l'altro per la visualizzazione dell'elemento tracciante/immunostaining. La procedura richiede 5-6 h per 10 topi.

L'obiettivo generale di questa procedura è valutare la permeabilità della vascolarizzazione cerebrale nei topi utilizzando traccianti marcati in fluorescenza per valutare la disfunzione della barriera emato-encefalica in modelli animali. Questo metodo può aiutare a rispondere a domande chiave nel campo della barriera emato-encefalica relative alla permeabilità neuro-vascolare nelle malattie e al suo recupero dopo la terapia. Il vantaggio principale di questo metodo è che è semplice e quantitativo.

Offre la possibilità di valutare contemporaneamente la permeabilità mediante fluorimetria, che è quantitativa, e anche la microscopia a fluorescenza per le difese regionali. Le implicazioni di questa tecnica si estendono alla diagnosi e alla terapia di diversi disturbi neurologici, tra cui i tumori cerebrali, l'ictus e il morbo di Alzheimer, perché queste malattie sono associate a un edema cerebrale pericoloso per la vita e al miglioramento della funzione BBD come obiettivo terapeutico chiave. A dimostrare parte della procedura sarà la signorina Jadranka Macas, responsabile della tecnologia scientifica presso l'istituto.

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