Abstract
BDNF is a pro-survival protein involved in neuronal development and synaptic plasticity. BDNF strengthens excitatory synapses and contributes to LTP, presynaptically, through enhancement of glutamate release, and postsynaptically, via phosphorylation of neurotransmitter receptors, modulation of receptor traffic and activation of the translation machinery. We examined whether BDNF upregulated vesicular glutamate receptor (VGLUT) 1 and 2 expression, which would partly account for the increased glutamate release in LTP. Cultured rat hippocampal neurons were incubated with 100 ng/ml BDNF, for different periods of time, and VGLUT gene and protein expression were assessed by real-time PCR and immunoblotting, respectively. At DIV7, exogenous application of BDNF rapidly increased VGLUT2 mRNA and protein levels, in a dose-dependent manner. VGLUT1 expression also increased but only transiently. However, at DIV14, BDNF stably increased VGLUT1 expression, whilst VGLUT2 levels remained low. Transcription inhibition with actinomycin-D or α-amanitine, and translation inhibition with emetine or anisomycin, fully blocked BDNF-induced VGLUT upregulation. Fluorescence microscopy imaging showed that BDNF stimulation upregulates the number, integrated density and intensity of VGLUT1 and VGLUT2 puncta in neurites of cultured hippocampal neurons (DIV7), indicating that the neurotrophin also affects the subcellular distribution of the transporter in developing neurons. Increased VGLUT1 somatic signals were also found 3 h after stimulation with BDNF, further suggesting an increased de novo transcription and translation. BDNF regulation of VGLUT expression was specifically mediated by BDNF, as no effect was found upon application of IGF-1 or bFGF, which activate other receptor tyrosine kinases. Moreover, inhibition of TrkB receptors with K252a and PLCγ signaling with U-73122 precluded BDNF-induced VGLUT upregulation. Hippocampal neurons express both isoforms during embryonic and neonatal development in contrast to adult tissue expressing only VGLUT1. These results suggest that BDNF regulates VGLUT expression during development and its effect on VGLUT1 may contribute to enhance glutamate release in LTP.
Highlights
BDNF is a pro-survival protein that promotes neuronal differentiation and synaptic plasticity [1,2], in addition to neuroprotection [3,4]
The sustained increase in VGLUT1 and VGLUT2 protein levels observed in hippocampal neurons (DIV14 and DIV7, respectively) subjected to a chronic stimulation with BDNF was not observed when the incubation was limited to 4 h, and followed by 14 h incubation in culture conditioned medium (p.0.05) (Fig. S1)
We have shown that BDNF regulates VGLUT1 and VGLUT2 gene and protein expression, during development of cultured hippocampal neurons, through activation of TrkB receptors and the phospholipase C-c (PLCc) signaling pathway
Summary
BDNF (brain-derived neurotrophic factor) is a pro-survival protein that promotes neuronal differentiation and synaptic plasticity [1,2], in addition to neuroprotection [3,4]. Activation of TrkB receptors by BDNF leads to receptor dimerization and trans-autophosphorylation of several tyrosine residues in the intracellular domain, including Y490 and Y816, which allow recruiting proteins containing PTB and SH2 (Src homology-type 2) domains, activating in parallel the Ras-ERK (extracellular signal-regulated kinase), PI3-K (phosphatidylinositol 3-kinase)/Akt and phospholipase C-c (PLCc) signaling pathways [20]. Trans-autophosphorylation of Y490 enables recruitment of Shc (Src homology 2-containing protein), IRS1 (insulin receptor substrate 1) and IRS2 linker proteins, thereby activating the Ras-ERK and PI3K/Akt cascades [26]. The Ras-ERK signaling pathway is crucial for neurogenesis [28], inhibition of proapoptotic proteins [29], stimulation of pro-survival gene expression [30] and protein synthesis-dependent plasticity [31]. The PI3K/Akt pathway has a pivotal role in cell survival [32], neuroprotection [3], trafficking of synaptic proteins [33] and can directly control protein synthesis through mTOR (mammalian target of rapamycin) activation and 4EBP phosphorylation [34]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
