At the experimental endpoint, cells were fixed with 4% paraformaldehyde and stained using standard protocols and the following antibodies: β‐tubulin III (mouse, Sigma Aldrich T8660, 1:1000), TH (rabbit, PelFreez Biologicals P40101, 1:1000), GFAP (rabbit, Dako Z0334, 1:1000; mouse, BiolegendSM121, 1:500), synapsin (rabbit, Invitrogen 72732524, 1:2000). For fluorescent stainings, Alexa 546 (goat anti‐mouse, Thermofisher Scientific A‐11030, 1:500), Alexa 647 (goat anti‐rabbit, ThermoFisher Scientific A‐21245, 1:500), Cy5 (donkey anti‐rabbit, Jackson Immunoresearch, 711605152, 1:500). Nuclei were counterstained using DAPI (4′,6‐diamidino‐2‐phenylindole) (1:1000) or 2 µg mL−1 Hoechst 33342.
Live and dead staining was done by incubating cells for 30 min at 37 °C in 2 × 10−6m Calcein AM (Sigma Aldrich) and 2 × 10−6m Ethidium homodimer‐1 (EthD‐1, ThermoFisher).
The middle image in Figure2d was acquired via a 5 × 5 cm tile scan with online image stitching on the above mentioned Ti2 microscope using an objective with 4× magnification. The rest of the images were acquired on an LSM 710 (Carl Zeiss) inverted confocal microscope. Objectives with 10×, 20×, and 60× magnification were used. Images were processed and analyzed in ImageJ software.
Live and dead staining was done by incubating cells for 30 min at 37 °C in 2 × 10−6
The middle image in Figure