Obtaining a Mixed Oligodendrocyte and Astrocyte Culture From Adult Mouse Neural Stem Cells
Transcription
Take a coronal mouse brain slice.
Identify the lateral ventricles, the C-shaped cavities in the brain, and isolate the ventricle walls.
Place the tissue in a buffer containing trypsin to degrade the extracellular matrix, releasing the cells.
Centrifuge and remove the supernatant. Resuspend the cells in a medium.
Plate the cells and incubate them vertically to maintain them in suspension.
Add growth factors that induce neural stem cell proliferation and aggregation to form three-dimensional neurospheres.
Harvest the neurospheres, centrifuge, and discard the supernatant.
Add fresh medium and mechanically dissociate the neurospheres.
Plate the cells and add a medium containing growth factors that induce neural stem cell differentiation into oligodendrocyte precursors.
The precursors proliferate and form oligospheres.
Mechanically dissociate the oligospheres to release the precursors.
Seed the precursors in extracellular matrix protein-coated wells to facilitate cellular attachment.
Add a medium containing specific hormones and neurotrophic factors to induce precursor maturation into oligodendrocytes and astrocytes.
