Dissociation of the spinal cord followed the procedure described in Häring et al., 2018. In short, following the isolation of gray matter (from cervical to sacral levels), the tissue was transferred into a 3cm plastic dish with 2.5ml of pre heated (37°C) digestion solution (300μl TrypLE Express (Life Technologies), 2000μl Papain (Worthington; 25U/ml in aCSF), 100μl DNase I (Worthington; 1mM in aCSF) and 100μl aCSF. Meninges were removed and the gray matter cut into pieces 1-2mm2. After 30 min incubation at 37°C, pieces were triturated with the first Pasteur pipette (see above). This procedure was repeated every 20min using Pasteur pipettes with decreasing diameter appropriate to the dissociation state. Depending on the progress of spinal cord dissociation, 100μl of TrypLE solution was added.
