Las af 3 analysis software

Histological experiments for single-labeled (c-Fos) cells, double-labeled (c-Fos + ChAT or PARV) cells, and AChE background staining were visualized using a Nikon E600 microscope fitted with a CoolSNAP digital camera (Roper Scientific, Trenton, NJ, USA). Fluorescence images were visualized using a Nikon E600 microscope or a Leica SP8 multiphoton confocal microscope (Leica Microsystems; Wetzlar, Germany) equipped with LAS AF 3 analysis software (Leica Microsystems; Wetzlar, Germany). Immunoperoxidase photomicrographs were captured using IP Lab Software (Scanalytics; Trenton, NJ, USA). Images were imported into Adobe Photoshop 6.0 (Adobe Systems; San Jose, CA, USA) to adjust the image size and to make minor alterations to contrast and brightness. To indicate the approximate brain region where photomicrographs were obtained, diagrams and schematics of brain regions in the results section were adapted from the third edition of The Rat Brain by Paxinos and Watson (Paxinos and Watson 1998 ).