Mice were placed in the center of an open-field arena (40 × 40 × 30 cm3) for 60 min and then habituated by daily placing in the same chamber for 10 min per day for three consecutive days. After the 3-day habituation, the general locomotion was assessed for a 30 min period. Specifically, we measured, 1) distance moved in the arena, 2) rearing, as the number of times the mice lifted both forepaws from

the ground, and 3) grooming, as the time that the mice groomed with its mouth and/or paws. Additional adult male mice at 90 ± 2 days old of age were used in the hidden platform versions of the Morris water maze tests using Stoelting poo-60135 as described before (Tu et al., 2010, see also Supplemental Experimental Procedures). Kinase Inhibitor Library supplier The forebrain was isolated from mice and placed

in RNAlater solution (QIAGEN) and tissues were homogenized using a TissueRuptor homogenizer (50/60 Hz) according Selleck Autophagy inhibitor to the manufacturer’s protocol. Total RNA was prepared from harvested hippocampal tissue with the miRNeasy Mini Kit (QIAGEN) and treated with an RNase-free DNase (QIAGEN) according to the manufacturer’s instructions. The RNA concentration was measured by spectrophotometer (DU 640; BECKMAN). RNA integrity was verified by electrophoresis in a 1% agarose gel. For microRNA expression profiling analysis, Mouse OneArray v2 (RmiOA.2) chips with 785 unique miRNA probes and 105 experimental control probe sets (Phalanx Biotech Group) were used. For gene expression profiling analysis, mouse whole genome OneArray microarray Resveratrol v2 (MOA-002) chips with 26,423 mouse genome probes and 872 experimental control probes were used. Datasets for single and dual channel experiments were analyzed with GeneChip Robust Multichip

Average (GC-RMA) and normalized using OneArray Software (Phalanx Biotech Group). All hybridized chips met standard quality control criteria were calculated as the fold-changes. The statistical relevance was expressed as the p values. The miscript cDNA synthesis kit (QIAGEN) was used for the reverse transcription reaction according to the manufacturer’s instructions. We used 1 μg total RNA, with 4 μl 5 × miscript reaction mix and 1 μl miscript reverse transcriptase. The total volume was 20 μl. Samples were incubated for 5 min at 25°C. All samples were then heated to 42°C for 30 min, and reactions were stopped by heating to 85°C for 5 min. For quantitative PCR (qPCR) all specific primers were selected using Beacon Designer Software (BioRad) and synthesized by IDT (Coralville, IA), as listed in Table S1. The PCR amplification of each product was further assessed using 10-fold dilutions of mouse brain cDNA library as a template and found to be linear over five orders of magnitude and at greater than 95% efficiency. All the PCR products were verified by sequencing.