Mouse anti parvalbumin pv

Coronal sections of the brain were cut at a thickness of 30 µm using a vibrating microtome (vibratome 1000, the Vibratome Company, St. Louis, MO, USA). Primary antibodies, including mouse anti-tyrosine hydroxylase (TH; 1:2000; Sigma-Aldrich) or mouse anti-Parvalbumin (PV) (1:2000; Sigma-Aldrich), were used. Biotinylated goat anti-mouse IgG (1:500 the Jackson ImmunoResearch Laboratories, West Grove, PA, USA) was used as the secondary antibody. Immunoreactivity was amplified using an avidin-biotin complex system (1:100, Vector Labs, Burlingame, CA, USA). TH is the key enzyme for dopamine synthesis, and TH immuno-histochemistry was used to label dopaminergic neurons. PV immunostaining was used to label PV-positive GABAergic interneurons. The densities of immunopositive cells were estimated using StereoInvestigator system (MicroBrightField Bioscience, Williston, VT, USA) [35 (link)].

In some experiments, two sections each from the anterior, middle, and posterior hippocampus were processed for fluorescence labeling. Sections were blocked in 3% w/v BSA in TBS solution and incubated for 70 h at 4°C with rabbit anti-c-Fos primary antibody (1:2500, Cell Signaling, United States) together with one of the following antibodies – (i) mouse anti-parvalbumin (PV; 1:2000; Sigma, United States), (ii) mouse anti-somatostatin (SOM; 1:25; GeneTex Inc., United States), or (iii) mouse anti-neuronal nitric oxide synthase (nNOS; 1:50, Santa Cruz Biotechnology, United States). PV, SOM, and nNOS are markers of hippocampal interneurons (Freund and Buzsáki, 1996 (link)). After washing with TBS with 0.1% v/v Triton (TBS-T; 5 min × 3), sections were incubated with Alexa Fluor 488 tagged donkey anti-rabbit secondary antibody and Alexa Fluor 647 tagged donkey anti-mouse secondary antibody (1:400; Molecular Probes, United States) for 1 h at room temperature in the dark. Sections were then washed with TBS-T (5 min × 3), mounted on gelatin-coated slides, and cover slipped with Gelmount (Sigma, United States) aqueous mounting medium. Sections were kept in the dark at 4°C before imaging.