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Methods for culturing primary sympathetic neurons and for determining neuronal viability
Jonathan Whitfield1, Stephen J Neame, Jonathan Ham
1Eisai London Research Laboratories, University College London, UK.
Methods in Molecular Biology (Clifton, N.J.)
|April 24, 2004
Summary
This study details a method for inducing and measuring neuronal apoptosis in developing sympathetic neurons. It provides a reliable in vitro model for testing inhibitors of cell death pathways.
Area of Science:
- Neuroscience
- Cell Biology
- Developmental Biology
Background:
- Developing nerve growth factor (NGF)-dependent sympathetic neurons serve as a key model for studying neuronal apoptosis.
- Understanding neuronal cell death pathways is crucial for neurodegenerative disease research.
Purpose of the Study:
- To describe the preparation of purified primary sympathetic neuron cultures.
- To outline a method for inducing apoptosis via NGF deprivation.
- To present a simple assay for measuring neuronal viability.
Main Methods:
- Purification of primary sympathetic neurons.
- Induction of apoptosis through NGF withdrawal.
- Live/Dead assay for assessing neuronal viability.
Main Results:
- Established a reproducible in vitro model for neuronal apoptosis.
- Demonstrated a method for quantifying cell death in response to NGF deprivation.
- Validated the use of the Live/Dead assay for assessing inhibitor efficacy.
Conclusions:
- The described protocol provides a robust system for studying NGF withdrawal-induced neuronal apoptosis.
- This model is suitable for screening small molecule inhibitors targeting cell death pathways.
- The Live/Dead assay offers a straightforward approach to evaluate the effects of inhibitors on neuronal survival.