original stains

landmark basedco-registration: MRI to Perls’

co-registered images

mask T2*w CAL (B ch) TH iron NM

T2*w 3D

CAL TH Perls’

Perls’ T2*w overlapped on Perls’

T2*w CAL TH Perls’

masked,in gray scale

binarized , thresholdedmanually corrected (white removed)

CAL (110) TH (125) Iron (185) NM (250)

Figure e-2. Image processing steps (on sample from HC2). The stages were: co-registration based on landmarks, 'cleanup' (masking the whole stem, removing small veins, artifacts etc.), converting to gray scale, thresholding and thus binarizing the images, and hence making the masks for iron, NM, TH, calbindin. The final masks were also corrected to remove remaining artifacts.

Image processing steps: stains and MRI