C57bl 6n male mice

Male C57BL/6N mice (6 weeks old, Koatech) were used to assess spatiotemporal LIN28A expression patterns. For the functional analysis of LIN28A in epilepsy, we crossed Nestin-Cre mice (47 (link)) with Lin28a^loxP/+ mice (a gift from Hao Zhu at UT Southwestern Medical Center, Dallas, Texas, USA), followed by the breeding of Nestin-Cre:Lin28a^loxP/+ mice with Lin28a^loxP/+ mice to generate male and female LIN28A-cKO and LIN28A WT mice. Mice were genotyped by polymerase chain reaction (PCR) using genomic DNA and primers for Lin28a (forward: 5′-TCC AAC CAG CAG TTT GCA G-3′, reverse: 5′-GCA GCT GGT AAG AAC AAA CCT-3′), and Nestin-Cre (forward: 5′-GGT CGA TGC AAC GAG TGA TGA GG-3′, reverse: 5′-GCT AAG TGC CTT CTC TAC ACC TGC G-3′). LIN28A WT and LIN28A-cKO mice were backcrossed to C57BL/6N mice for at least 5 generations prior to beginning the studies. All mice were bred and housed in an animal facility with a 12-hour light, 12-hour dark cycle and had access to food and water ad libitum.

Sixty specific pathogen-free C57BL/6N male mice (12 weeks old; 24–27 g) were purchased from Koatech (Gyeonggido, Korea). They received food (Cargill Agri Furina, Gyeonggido, Korea) and water ad libitum, and were housed in a room maintained at 23 ± 2°C with a 12-h light-dark cycle. After acclimatization for one week, the mice were randomly divided into six groups (n = 10 per group): naïve, scopolamine, PNE pretreatment group (25, 50 or 100 mg/kg) and tacrine group (10 mg/kg, as a positive control). All groups were orally administered with distilled water (naïve and control), PNE, or tacrine for seven days prior to scopolamine injection. Except naïve group, scopolamine was dissolved in 0.9% physiological saline (2 mg/kg), and injected once intraperitoneally, 30 min before the Morris water maze task on first day.
The dose of PNE was determined based on prescreening results. The protocol was approved by the Institutional Animal Care and Use Committee of Daejeon University (DJUARB 2013-031), and was conducted in accordance with the Guide for the Care and Use of Laboratory Animals published by the United States National Institutes of Health (NIH).