Isolation and comparison of neural stem cells from the adult rat brain and spinal cord canonical neurogenic niches

STAR Protocols, 2022 · DOI: 10.1016/j.xpro.2022.101426 · Published: June 17, 2022

Regenerative Medicine Neurology Research Methodology & Design

Simple Explanation

This protocol outlines the process of extracting, culturing, and characterizing neural stem cells (NSCs) from the adult rat brain and spinal cord. It focuses on three specific areas known as neurogenic niches. The method involves carefully dissecting and breaking down tissue, followed by cell culture techniques and EdU labeling to identify and study NSCs. This protocol is designed to yield a large number of viable cells, allowing for the creation of substantial cell banks, which can reduce costs, labor, and the number of animals needed for research.

Study Duration

Not specified

Participants

10-week-old female Sprague Dawley rats

Evidence Level

Not specified

Key Findings

1
The protocol provides a unified method for extracting NSCs from three neurogenic niches: the subventricular zone (SVZ), subgranular zone (SGZ), and central canal (CC).
2
The surface topography of the culture vessel significantly impacts NSC survival and proliferation, with SVZ-derived NSCs requiring completely smooth surfaces and SGZ-derived NSCs thriving on rougher surfaces.
3
CHIR 99021, a chemical compound, is crucial for the survival of SGZ-derived NSCs and prevents spontaneous differentiation in SVZ-derived NSCs, but it induces differentiation in CC-derived NSCs.

Research Summary

The protocol details a method for isolating and comparing neural stem cells (NSCs) from the adult rat brain and spinal cord canonical neurogenic niches. It involves tissue dissection, dissociation, cell culture, EdU labeling, and NSC characterization. The protocol aims to provide a cost- and labor-efficient method that reduces the number of animals used while yielding a high number of viable cells for long-term cell banks.

Practical Implications

CNS Injury and Disease Modeling

The protocol enables researchers to extract, maintain, and differentiate NSCs, which can be used in studies of neuroplasticity and regeneration, including CNS injury and disease modeling.

Optimized Culture Conditions

Understanding the specific surface requirements for different NSCs (smooth for SVZ, rough for SGZ) can lead to more effective in vitro studies.

Targeted Differentiation Strategies

The differential effects of CHIR 99021 on NSCs from different niches provide insights for targeted differentiation strategies.

Study Limitations

1
The protocol is optimized for 10-week-old female Sprague Dawley rats, and results may vary with different strains and ages.
2
NSCs are highly sensitive to the culture vessel's material and surface treatment, requiring specific vessels for optimal results.
3
NSC characterization lacks universally recognized markers, requiring a combination of proliferation, marker expression, and neural lineage multipotency assessments.

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