*4.7. Data Analysis*

Four sections per antibody between 2.0 mm and 2.7 mm caudal to the bregma [27] were examined using an image analysis system and ImageJ software v. 1.5 (National Institutes of Health, Bethesda, MD, USA). Digital images of the whole dentate gyrus and midpoint of the CA1 region were captured with a BX51 light microscope (Olympus, Tokyo, Japan) equipped with a digital camera (DP72, Olympus). The number of NeuN- and Ki67-positive nuclei were counted in the hippocampal CA1 region and dentate gyrus, respectively, using an image analysis system (Optimas 6.5, CyberMetrics, Scottsdale, AZ, USA). Intensities for GFAP, Iba-1, and DCX were evaluated by ROD obtained after transforming the mean gray level using this formula: ROD = log(256/mean gray level). The ROD of background staining was determined in unlabeled portions of the sections using Photoshop CC software (Adobe Systems Inc., San Jose, CA, USA), and this value was subtracted to correct for nonspecific staining using ImageJ v. 1.50 software (National Institutes of Health, Bethesda, MD, USA). The data are expressed as the percentage of the vehicle-treated group values (set to 100%).
