Synaptic activity increases hnRNP A2/B1 in dendrites and induces the delivery of hnRNP A2/B1 to the synapse by a BDNF-dependent mechanism.

<p>Cultured hippocampal neurons were stimulated or not with bicuculline (50 µM), 4-AP (2.5 mM) and glycine (10 µM), for 30 min. Where indicated, neurons were treated with TrkB-Fc (1 µg/ml) for 30 min and then stimulated or not with bicuculline in the presence of the BDNF scavenger. The cells were immunostained for hnRNP A2/B1 (red), MAP2 (blue), and PSD95 (green) (A). White arrows indicate PSD95-positive clusters that also contain hnRNP A2/B1 (A). The integrated fluorescence intensity, area and number of hnRNP A2/B1 puncta in dendrites (B, C, and D) and at the synapse (as defined by the signal of hnRNP A2/B1 that overlaps with PSD95) (E, F, and G) was analysed using ImageJ software and represented per dendritic area. The percentage of dendritic hnRNP A2/B1 signal that colocalizes with PSD95 was also analysed (H). Results are normalized to control and are averaged of 3–7 different experiments performed in independent preparations. The following number of cells was used in the analysis of the dendritic-localized hnRNP A2/B1 puncta: Ctr (n = 87 cells), Bic (n = 64 cells), TrkB-Fc (n = 47 cells), TrkB-Fc+Bic (n = 48 cells). For the analysis of synaptic-localized hnRNP A2/B1 puncta the following number of cells was analysed: Ctr (n = 76 cells), Bic (n = 54 cells), TrkB-Fc (n = 35 cells), TrkB-Fc+Bic (n = 35 cells). Error bars, mean ± SEM. Statistical analysis was performed by one-way ANOVA, followed by Bonferroni's test. n.s. Not significant, * P<0.05; ** P<0.01; *** P<0.001. Scale bar = 4 µm.