Rab3A Is Required for Brain-Derived Neurotrophic Factor-Induced Synaptic Plasticity: Transcriptional Analysis at the Population and Single-Cell Levels

Abstract

Brain-derived neurotrophic factor (BDNF) modulates synaptic strength in hippocampal neurons, in addition to promoting survival and differentiation. To identify genes involved in trophic regulation of synaptic plasticity, we have used a multidisciplinary approach of differential display and family-specific slot blots in combination with whole-cell patch-clamp recordings of dissociated hippocampal neurons. Three hour exposure to BDNF elicited a 2.6-fold increase in synaptic charge and a concomitant induction of 11 genes as revealed by differential display, including the small GTP-binding vesicular trafficking protein Rab3A and the enzyme guanylate cyclase (GC). Slot blot analysis on a population of neurons confirmed an average of 3.1-fold induction of these clones. In contrast, individual pyramidal-like neurons that were first characterized electrophysiologically in the presence of BDNF and subjected to transcriptional analysis displayed more robust increases (4.8-fold), emphasizing the neuronal heterogeneity. Transcriptional changes of Rab3A and GC were accompanied by translational regulation, shown by Western blot analysis. Furthermore, a number of GC-associated and Rab3A effector molecules were induced by BDNF at either the gene or protein levels. The functional role of Rab3A in BDNF-induced synaptic plasticity was assessed using cells derived from Rab3A knock-out mice. These neurons failed to show an increase in synaptic charge in response to BDNF at 10 min; however a late response to BDNF was detected at 20 min. This late response was similar in time course to that induced by postsynaptic activation of glutamate receptors. Our results demonstrate a requirement for Rab3A and may reveal a temporal distinction between presynaptic and postsynaptic mechanisms of BDNF-induced synaptic plasticity associated with learning and memory.

Site-Specific Synapsin I Phosphorylation Participates in the Expression of Post-Tetanic Potentiation and Its Enhancement by BDNF

“…However, in the former study neurotransmitter release was evoked by a longlasting depolarization with 4-aminopyridine, which is known to mobilize not only the RRP, but also a large fraction of the recycling pool of SVs, while here, instead, we have used either singleaction potentials or short stimulation trains, which mobilize only a fraction of RRP or, at most, the "whole" RRP. Probably, the rapid effect on RRP involves other presynaptic molecular actors, as was suggested by recent results showing that the BDNF-induced increase in neurotransmitter release is hindered in Rab3A (Thakker-Varia et al, 2001;Alder et al, 2005) and Rim1alfa (Simsek-Duran and Lonart, 2008) KO mice. Moreover, other presynaptic mechanisms, such as the increase in intraterminal Ca 2ϩ following activation of phospholipase C␥ and/or transient receptor potential cation channels (TRPC) or fastdepolarizing Na ϩ currents could underlie the rapid increase of Pr and RRP evoked by BDNF (Li et al, 1999;Blum et al, 2002;Amaral and Pozzo-Miller, 2007).…”

Early Presynaptic and Late Postsynaptic Components Contribute Independently to Brain-Derived Neurotrophic Factor-Induced Synaptic Plasticity

“…In addition, the knockout displays increased synaptic depression after short trains of repetitive stimuli, suggesting that Rab3A plays a role in the recruitment of synaptic vesicles to the active zone, possibly at a very late step in the secretion process (Geppert et al, 1997). Using Rab3A knock-out mice, we demonstrated a requirement for this synaptic vesicle protein in the early increase in synaptic charge induced by BDNF (Thakker-Varia et al, 2001). In contrast, a late response of Rab3A mutant cells to BDNF was observed and is temporally similar to that produced by pairing glutamate iontophoresis and BDNF exposure (Crozier et al, 1999).…”

BDNF-induced local protein synthesis and synaptic plasticity

“…Rab3a is associated with synaptic vesicles and the BDNF-induced upregulation of neurotransmitter release was shown to be impaired in cultured hippocampal neurons form Rab3a knockout mice (Alder et al, 2005;Thakker-Varia et al, 2001). A role for Rab3a in the presynaptic effects of BDNF is further suggested by the results showing an impairment of the BDNF-induced potentiation of glutamate release in synaptosomes isolated from the CA1 region of Rim1a (Rab3a interacting molecule 1a) knockout mice (Simsek-Duran and Lonart, 2008).…”

“…Synaptic vesicle proteins and proteins related to their traffic were also shown to be upregulated by BDNF (Melo et al, 2013;Tartaglia et al, 2001;Thakker-Varia et al, 2001) (see Section 4.6), but additional studies are required to determine whether some of these alterations are extended to the synapse. Taken together, the BDNF-induced changes in the neuronal proteome described above suggest several putative mechanisms that may contribute to synaptic potentiation by BDNF.…”