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HuD Interacts with Bdnf mRNA and Is Essential for Activity-Induced BDNF Synthesis in Dendrites

Figure 5

Changes in HuD expression and short-term PMA treatment do not affect levels of dendritic mRNAs with Bdnf 5' and 3'-UTRs.

(A) Dual Bdnf mRNA FISH and HuD immunocytochemistry on hippocampal cultures transfected with recombinant GFP-mHuD at 14 DIV and fixed, stained and imaged at 16 DIV, using antisense probe against the Bdnf coding sequence for in situ and anti-GFP for immunocytochemistry (n = 30; scale bar, 20 μm). Co-localization data was obtained using the JACoP plugin on ImageJ. (B) Colorized dendritic in situ signal using a probe against the Bdnf mRNA coding sequence in hippocampal cultures transfected with the indicated plasmids at 14 DIV and then fixed at 16 DIV (sense, sense in situ probe; scale bar, 20 μm). (C) Mean ± SEM in 20-μm bins from images represented in (B), normalized to the mean dendritic signal from 20–40 μm away from the soma of untransfected neurons on the same coverslip (n = 30; *, p<0.05; **, p<0.01; student's t-test). (D) Colorized dendritic in situ signal using a probe against the GFP coding sequence in hippocampal cultures transfected with the reporter construct ExIV A*B at 14 DIV and then treated for 1 or 3 hours with PMA or vehicle, then fixed at 16 DIV (sense, sense in situ probe; scale bar, 20 μm). (E) Mean ± SEM in 20-μm bins from images represented in (D) (n = 30; *, p<0.01; **, p<0.001; student's t-test).

Figure 5

doi: https://doi.org/10.1371/journal.pone.0117264.g005