Primary cultures of hippocampal neurons were obtained from rats at embryonic age E 18. The entire hippocampus was isolated and dissociated with trypsin. Cells were plated in a 24-well plate at a density of 50.000 cells/well on poly-D-lysine coated coverslips. Cells were grown in glia-conditioned Neurobasal medium supplemented with 2% B27, 1% Na-pyruvate, 1% glutamax, 1% penicillin/streptomycin, 1% Fungizone (all Invitrogen) at 37°C and 5% CO2. At days 12–14 in vitro (DIV 12–14), cells were transduced with CNIH-2 by high-titer lentiviral preparations or at DIV 15–20 by semliki forest viral particles.
