Distribution Patterns of Three Molecularly Defined Classes of GABAergic Neurons Across Columnar Compartments in Mouse Barrel Cortex

This study was carried out in accordance with the principles of the Basel Declaration and German laws on animal research (TierSchG and TierSchVersV, 2013). The protocol was approved by the LAVES (Niedersächsisches Landesamt für Verbraucherschutz und Lebensmittelsicherheit). Tissue Preparation and Immunocytochemistry The animals were anesthetized with an overdose of ketamin (Essex Tierarznei) and transcardially perfused with 0.9% NaCl solution to remove blood from vessels, followed by 4% paraformaldehyde dissolved in 0.1 M phosphate buffer (PB, pH 7.4). The brains were removed and the two hemispheres separated. The left hemispheres were flattened (Welker and Woolsey, 1974) whereas the right ones were kept unmodified, and all tissue was postfixed for 2 h in the same fixative. The right hemispheres were cut in the coronal plane, the left hemispheres tangentially on a vibratome (VT 1200S, Leica), both with a nominal section thickness of 50 μm. The sections of PV-GIN animals were incubated with rabbit anti-GFP (Invitrogen) 1:2500 in TRIS-buffered saline with 0.3% Triton X-100 (TBST) for 2 days (in a cold room), followed by an anti-rabbit IgG coupled with Alexa-488 (Invitrogen) 1:500 in TBST for 2 h. The PV+ cells contained a sufficient amount of tdTomato, therefore they did not need further signal amplification. The sections of SSTcre/tdTomato and VIPcre/tdTomato animals were stained with guinea pig anti-vesicular glutamate transporter 2 (vGlut2; Millipore)...
Source: Frontiers in Neuroanatomy - Category: Neurology Source Type: research