β d arabinofuranoside hydrochloride

Primary cortical neuron cultures were prepared from the cerebral cortex of Wistar rat embryo of 17 days as described previously (Kim et al., 1998 (link)). Briefly, pregnant Wistar rats were anesthetized with sodium pentobarbital (30 mg/kg, i.p., Sigma–Aldrich) and sacrificed by cervical dislocation. The cerebral cortex of fetal rats was rapidly removed bilaterally and collected. Tissues were then gently minced using a sterile razor blade and digested in PBS (0.1 M, pH 7.4, Sigma–Aldrich) for 15 min. A Pasteur pipette was used for dissociation of cells (approximately 5–10 times). After centrifugation (200 × g for 3 min), cells were re-suspended in DMEM (Sigma–Aldrich) supplemented with FBS (15%, Carlsbad), L-glutamine (2 mM, Sigma–Aldrich), sodium bicarbonate (4.2 mM, Sigma–Aldrich), BSA (0.3 g/l, Sigma–Aldrich), β-mercaptoethanol (0.1 mM, Sigma–Aldrich), penicillin (1%, Sanofi Aventis), streptomycin (50 μg/ml, Sanofi Aventis) and grown on 0.1% poly-L-Lysine (Sigma–Aldrich) coated plates. Cultures were incubated at 37°C in a humidified 5% CO₂ atmosphere. To prevent proliferation of non-neuronal cells, cytosine β-D-arabinofuranoside hydrochloride (10 μM, Sigma–Aldrich) was added 3 days after plating. In all experiments, 11 days mature cells were used.

β-D-arabinofuranoside hydrochloride, Hoechst 33342, propidium iodide, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), oxamate, dimethyl sulfoxide (DMSO), sodium bicarbonate, phosphate buffer saline (PBS), nicotinamide dinucleotide adenine (NAD), diaphorase, L-glutamic acid monosodium salt hydrate, poly-L-lysine, Dulbecco’s modified Eagle’s medium (DMEM), β-mercaptoethanol, lactate, L-glutamine were purchased from Sigma–Aldrich (St. Louis, MO, United States). Fetal bovine serum (FBS) and bovine serum albumin (BSA) were purchased from Gibo/Invitrogen (Carlsbad, CA, United States). Penicillin and streptomycin were purchased from Sanofi-Aventis (Guildford, United Kingdom).