Culturing and Maintaining Dopaminergic Neurons from Mouse Embryonic Brain Tissue
Transcript
Treat the embryonic midbrain tissue, rich in developing dopaminergic neurons, with a trypsin-EDTA solution to detach them from the tissue matrix.
Add a deactivation medium to inhibit enzyme activity, and then wash to remove residual enzymes.
Pipette the tissue repeatedly. The embryonic neurons possess fewer projections, reducing the stress during this disturbance and generating a single-cell suspension.
Centrifuge and remove the supernatant, then resuspend the neurons in a complete medium. Transfer these neurons onto a polymer-coated coverslip for cell attachment.
Place the coverslip in a multi-well plate with a complete medium, providing nutrients and growth factors essential for neuron growth.
The antibiotics in the medium prevent bacterial growth, supporting neuronal viability.
As neurons grow and mature, they develop cellular projections, such as axons and dendrites, forming synaptic connections.
Periodically, remove half of the used media to eliminate toxic by-products.
Add a fresh medium for the long-term maintenance of the dopaminergic neurons.
