For histology, brain samples were embedded in 4% agarose and sectioned into 100 μm slices using a vibratome (Mikrom HM 650 V, Thermo Scientific). Nerve and whisker samples were prepared for cryo-sectioning by embedding in 10% sucrose and 4% agarose and then left in 30% Sucrose solution until they sunk to the bottom for cryo-protection. Samples were then sectioned into 40–60 μm slices at −15°C using a Leica Frigomobil 0247/08.1996 cryo-sectioning device.
Sections were immediately mounted on gelatine-coated slides with 0.1 M PB and imaged using a Leica DM5500B fluorescence microscope (Wetzlar, Germany). For neural tracing with Carbocyanine dyes, immediate imaging after sectioning is preferred as the dye continues to diffuse through the tissue. Leaving sections for too long can result in leaking of the dye out of the cut cells leading to diffuse and non-specific labeling. After imaging, the sections can be kept in 0.1 M PB for further histological staining.
Sections were immediately mounted on gelatine-coated slides with 0.1 M PB and imaged using a Leica DM5500B fluorescence microscope (Wetzlar, Germany). For neural tracing with Carbocyanine dyes, immediate imaging after sectioning is preferred as the dye continues to diffuse through the tissue. Leaving sections for too long can result in leaking of the dye out of the cut cells leading to diffuse and non-specific labeling. After imaging, the sections can be kept in 0.1 M PB for further histological staining.