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Implantation of a Cranial Window for Repeated In Vivo Imaging in Awake Mice
Published on: June 22, 2021
Studying protein degradation pathways in vivo using a cranial window-based approach
Vivek K Unni1, Darius Ebrahimi-Fakhari, Charles R Vanderburg
1Alzheimer's Research Unit, MassGeneral Institute for Neurodegenerative Disease, MGH, Harvard Medical School, CNY114, 16th St., Charlestown, MA 02129, USA.
Methods (San Diego, Calif.)
|December 29, 2010
Summary
Researchers developed a new method to study protein degradation in the brain using a cranial window to deliver a ubiquitin-proteasome system (UPS) inhibitor. This technique allows for in vivo testing of UPS roles in neurodegenerative diseases.
Area of Science:
- Neuroscience
- Molecular Biology
- Biochemistry
Background:
- The ubiquitin-proteasome system (UPS) is crucial for protein degradation in neurons.
- UPS dysfunction is linked to neurodegenerative diseases like Alzheimer's and Parkinson's.
- Systemic UPS inhibitors often fail to cross the blood-brain barrier (BBB), hindering in vivo research.
Purpose of the Study:
- To develop and validate a novel in vivo method for studying UPS function in the mammalian brain.
- To overcome the limitations of systemic drug delivery across the BBB for UPS inhibition.
Main Methods:
- A "cranial window" technique was employed to deliver the UPS inhibitor clasto-lactacystin beta-lactone (CLBL) directly to the mouse cortex.
- Two tissue recovery methods were developed: vibratome/laser capture microscopy (LCM) and vibratome-only.
- Immunoblot analysis was used to confirm UPS inhibition.
Main Results:
- The cranial window approach successfully delivered CLBL into the mouse cortex.
- UPS inhibition was documented up to 600 micrometers deep into the cortex.
- The developed methods allow for targeted in vivo investigation of protein degradation.
Conclusions:
- This novel cranial window strategy provides a valuable tool for in vivo investigation of the UPS in the brain.
- The technique complements existing methods and expands the research toolkit for neurodegenerative disease studies.
- It enables direct testing of hypotheses regarding UPS-mediated protein degradation in living animal models.
