Fig. 1. Sample preparation.

A Biopsy preparation of the laminar site of the biopsy from BA46 is depicted in this illustration, with an average height of 2.5 mm (i). The six layers of BA46 can be seen in a cross-section of a biopsy after it has been stained with toluidine blue. Scale bar = 500 µm. Layer III is located within 2 mm of the pial surface (ii). A razor knife with a 2 mm gap between each blade was flipped on top of the biopsy. The red area shows the removed tissue (iii). A razor knife with a 1 mm gap between each blade was flipped on top of the biopsy. The red area shows the removed tissue (iv). The dimension of the tissue can be seen after the excess tissue has been removed. Before inserting the tissue within the embedding mold, it was flipped 90^◦ to make sure the layers were parallel to the cutting direction (v). The illustration shows the tissue in the embedding mold. The resin block was photographed after it was trimmed with a glass knife, so it only contained layers I–IV (vi). B Tissue block preparation of DLPFC containing BA46 was rotated uniformly around a vertical axis (VA), perpendicular to the block’s central pial surface (i). The tissue block was embedded in a box filled with 7% agarose (ii). Next, the tissue block was cut systematically into uniformly random into 2.5 mm slabs orthogonal to the anterior-posterior direction (iii). The tissue block was then reassembled and every second the slab was systematically sampled (iv). After that, each slab was documented and various gyri positions were specified, e.g., medial frontal gyrus (MFG), inferior frontal gyrus (IFG) (v). Finally, the slabs were embedded in glycolmethacrylate (vi). C The section were stained with toluidine blue and VA was positioned based on documentation. Scale bar = 6 mm (i). The stereological analysis was then performed in layer III of BA46 based on the cytoarchitecture. Scale bar = 500 µm (ii).