Figure 1.

OTC dissection and culture

(A) Illustration of the dissection of the brain from the mouse head (step 7). From left to right: cut the skin longitudinally with microdissection scissors and pull it off with curved forceps. Proceed similarly with the skull. Cut the cerebellum with a scalpel.

(B) Horizontal section of a mouse brain. Examine sections under a binocular microscope (step 9) and cut out the region of interest for OTCs according to the red square (step 10). Scale bar: 2 mm.

(C) The entorhinal-hippocampal formation after dissection. Scale bar: 2 mm.

(D) Place the OTCs with appropriate morphology on a Millipore insert in a 6-well plate for subsequent culture (step 12).

(E) Images of different stages during culture of OTCs – DIV 0, 1, 7, 14, 21. OTCs flatten in the first days after their dissection. Up to 6 OTCs can be placed on an insert, safely spaced and with sufficient nutrients from the incubation medium (note that OTCs will have moved slightly after the first image; this may occur during transfer from the hood to the incubator before they are attached to the insert). DIV: day in vitro (where the day of the dissection is DIV0). Scale bar: 2 mm.