Genetic Study of Axon Regeneration with Cultured Adult Dorsal Root Ganglion Neurons

Journal of Visualized Experiments, 2012 · DOI: 10.3791/4141 · Published: August 17, 2012

Simple Explanation

Mature neurons in the central nervous system (CNS) do not regenerate axons after injury due to reduced growth ability and a hostile environment. Adult dorsal root ganglion (DRG) neurons are used as a model to study regenerative axon growth, because they regenerate robustly after peripheral nerve injuries. The described method allows genetic manipulation of adult DRG neurons using electroporation to study axon regeneration in vitro.

Study Duration

1-4 days

Participants

6- to 10-weeks old adult mouse

Evidence Level

Not specified

Key Findings

1
Re-plated neurons extend axons quickly with reduced branching, mimicking the effects of a conditioning lesion.
2
Knockdown of c-Jun significantly reduces axon growth in re-plated DRG neurons, confirming its role in axon regeneration.
3
Cultured adult DRG neurons provide a useful model system to study axon growth from adult neurons.

Research Summary

Adult DRG neurons provide a useful system to study axon regeneration due to their ability to regenerate axons robustly after peripheral nerve injury. The in vitro procedure presented enables rapid and effective genetic study of regenerative axon growth in adult DRG neurons. The re-suspension and re-plating procedure is particularly useful for loss-of-function studies and mimics an in vivo conditioning lesion effect.

Practical Implications

Model system for regeneration studies

Adult DRG neurons provide a valuable in vitro tool to investigate the roles of regeneration-associated genes in regulation of axon regeneration.

Loss-of-function studies

The re-suspension and re-plating procedure is particularly useful for loss-of-function studies

Mimicking conditioning lesion

Re-plating cultured DRG neurons mimics an in vivo conditioning lesion effect, thus providing a more physiological relevant approach to study axon regeneration in vitro.

Study Limitations

1
Transfection efficiency with electroporation is 20-50% for DNA plasmids.
2
The effect of siRNAs reduces after longer period due to the degradation of the siRNA oligos.
3
Plasmid size can affect co-transfection efficiency.

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