Preparation of primary rat cortical neuron and astrocyte co-culture

1-3 days postpartum Sprague Dawley rats (University College London breeding colony) are used.

Experimental procedures are performed according to the United Kingdom Animal (Scientific Procedures) Act of 1986.

Rat cortices are placed in an ice-cold Dissecting buffer (described in Materials).

Wash five times with Washing buffer (described in Materials).

Tissues are digested with a Disgesting buffer (described in Materials) for 0h 15m 0s

Digested tissues are neutralized with a dissecting buffer.

Washing twice with a Washing buffer,

Dissociate with a Washing buffer supplemented with DNAse.

Dissociated pellets are collected in Neurobasal completed medium.

Approximately 600,000 cells are plated on25 mm Poly-D-Lysin (PDL) coated coverslips and 200,000 cells for 8-well ibidi chambers (PDL coated).

The cultures are maintained at 37°C (5% volume CO2), and the media are changed every 4-5 days.

Cells can be used at 12-16 days.