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. 2013 Jan 11;8(1):e53793. doi: 10.1371/journal.pone.0053793

Figure 3. TrkB receptor inhibition blocks BDNF upregulation of VGLUT1 and VGLUT2.

Figure 3

(A–C) Cultured hippocampal neurons at DIV7 (A, B) and DIV14 (C) were stimulated with BDNF (100 ng/ml), for the indicated periods of time, in the presence or absence of a selective inhibitor of tyrosine kinase activity, K252a (200 nM), and VGLUT1 (A, C) and VGLUT2 (B) protein levels were determined by western blot. Quantification of the indicated number of experiments, performed in independent preparations, is presented as mean percentage ± SEM compared to the control (unstimulated neurons). (D–F) DIV7 hippocampal neurons were stimulated with IGF-1 or bFGF, for 4 or 20 h, and VGLUT1 (D) and VGLUT2 (E) protein levels were determined by western blot. Quantification of 4 different experiments, performed in independent preparations, is presented as mean percentage ± SEM compared to the control. Statistical significance was determined by One Way ANOVA followed by Bonferronís multiple comparison test with a confidence interval of 99% (*p<0.05, **p<0.01, ***p<0.001). (F) DIV7 hippocampal neurons were stimulated with IGF-1 or bFGF, for 15 or 30 min, and the levels of ERK1/2 phosphorylation were determined by western blot. The antibody used specifically recognizes the phosphorylated isoforms 1 and 2 of ERK, but not the nonphosphorylated (presumably inactive) proteins.