Proteins were denatured by boiling in ( Laemmli, 1970) sample buffer containing 100 mM DTT ( De Souza et al., 2003). After this,

0.2 mg of inhibitors protein extracts obtained from each tissue were separated by SDS–PAGE, transferred to nitrocellulose membranes learn more and blotted with anti-AKT, anti-Bcl-2 and anti-GSK-3β. Antibodies were from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Chemiluminescent detection was performed with horseradish peroxidase-conjugate secondary antibodies. Visualization of the protein bands was performed by exposure of the membranes to RX-films. The original membrane was stripped and reblotted with actin loading protein (bands not showing). After transfer, the membrane was stained with Ponceau and bands were visualized, photographed and quantified before the primary

antibody, to control the transfer. Band intensities were quantitated by optical densitometry (Scion Image software, ScionCorp, Frederick, MD) of the developed autoradiographs. All data are presented as mean ± SEM. Differences among experimental groups in the forced swimming and open field tests and in the assessment of the biochemical analysis were determined by one-way ANOVA, followed by Tukey post-hoc test when ANOVA was significant; P values <0.05 were considered to be statistically significant. The effects of the acute and chronic administration of lamotrigine on the Lonafarnib manufacturer immobility times are illustrated in Fig. 1A. In the acute (F(3–21) = 6.148; p = 0.04 Fig. 1A) and chronic (F(3–66) = 6.222; p = 0.01 Fig. 1A) treatments we observed a decrease in the immobility time with imipramine at the dose of 30 mg/kg and lamotrigine at the doses of 10 and 20 mg/kg, compared with saline. Interestingly, in the open-field test both acute aminophylline and chronic treatments with imipramine or lamotrigine did not modify the number of crossings (acute; F(3–55) = 0.595; p = 0.62; Fig. 1B; chronic; F(3–53) = 3.411; p = 0.24 Fig. 1B) and rearings (acute; F(3–55) = 0.393; p = 0.75;

chronic; F(3–53) = 0.844; p = 0.47 Fig. 1B), compared with saline. With regards to the acute treatment, there was an increase the BDNF levels in the prefrontal cortex with lamotrigine at the dose of 20 mg/kg (F(3–16) = 5.501; p = 0,009 Fig. 2A), compared with saline, but BDNF protein levels did not alter in the prefrontal cortex with imipramine at the dose of 30 mg/kg (F(3–16) = 5.501; p = 0.22 Fig. 2A) and with lamotrigine at the dose of 10 mg/kg (F(3–16) = 5.501; p = 0.91 Fig. 2A), compared with saline. The amygdala (F(3–16) = 1.292; p = 0,31 Fig. 2A) and the hippocampus (F(3–16) = 2.844; p = 0.71 Fig. 2A) did not have any alterations in their BDNF levels after acute treatment. In the chronic treatment data, we found an increase occurred in the BDNF levels in the prefrontal cortex with lamotrigine at the dose of 10 and 20 mg/kg (F(3–16) = 8.478; p = 0.01 Fig.