Rat hippocampal neurons were isolated from E18 Sprague-Dawley rat embryos and cultured at a density of 2 × 105 cells per coverslip on poly-l-lysine–coated 18-mm glass coverslips as previously described with minor modifications (25). Briefly, neurons were cultured in Neurobasal medium (Invitrogen) supplemented with B27 that was conditioned by separate cultures of primary rat astrocytes and glia. Neurons were treated at DIV 4 with 5 μM cytosine β-d-arabinofuranoside hydrochloride (Sigma-Aldrich) to eliminate the presence of native astrocytes and glia on the glass coverslips. Medium was changed every 3 to 4 days with new glia-conditioned Neurobasal medium for proper culture maintenance. At DIV 12, neurons were cotransfected with plasmids using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s instructions. Neuro-2A mouse neuroblastoma cells were maintained in Dulbecco’s minimum essential medium with 10% fetal bovine serum and 1% penicillin/streptomycin.

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