Activation of the CREB/c-Fos Pathway during Long-Term Synaptic Plasticity in the Cerebellum Granular Layer.

Daniela Gandolfi,Albertino Bigiani,Marie-Therese Fuzzati-Armentero,Egidio D’Angelo,Lisa Mapelli,Silvia Cerri,Jonathan Mapelli,Simona Tritto,Fabio Blandini,Mariarosa Polimeni

doi:10.3389/fncel.2017.00184

Abstract

The induction of long-term potentiation and depression (LTP and LTD) is thought to trigger gene expression and protein synthesis, leading to consolidation of synaptic and neuronal changes. However, while LTP and LTD have been proposed to play important roles for sensori-motor learning in the cerebellum granular layer, their association with these mechanisms remained unclear. Here, we have investigated phosphorylation of the cAMP-responsive element binding protein (CREB) and activation of the immediate early gene c-Fos pathway following the induction of synaptic plasticity by theta-burst stimulation (TBS) in acute cerebellar slices. LTP and LTD were localized using voltage-sensitive dye imaging (VSDi). At two time points following TBS (15 min and 120 min), corresponding to the early and late phases of plasticity, slices were fixed and processed to evaluate CREB phosphorylation (P-CREB) and c-FOS protein levels, as well as Creb and c-Fos mRNA expression. High levels of P-CREB and Creb/c-Fos were detected before those of c-FOS, as expected if CREB phosphorylation triggered gene expression followed by protein synthesis. No differences between control slices and slices stimulated with TBS were observed in the presence of an N-methyl-D-aspartate receptor (NMDAR) antagonist. Interestingly, activation of the CREB/c-Fos system showed a relevant degree of colocalization with long-term synaptic plasticity. These results show that NMDAR-dependent plasticity at the cerebellum input stage bears about transcriptional and post-transcriptional processes potentially contributing to cerebellar learning and memory consolidation.

Highlights

Long-term plasticity consists of changes in synaptic transmission and neuronal excitability (Hebb, 1949; Bliss and Lomo, 1973; Bienenstock et al, 1982; Bear and Abraham, 1996) that are thought to provide the basis for learning and memory in the brain (Bliss and Collingridge, 1993; Bliss et al, 2003; Bliss and Collingridge, 2013; Sweatt, 2016)
We addressed the issue as to whether, in the cerebellar granular layer, long-term synaptic plasticity is accompanied by activation of the cAMP-responsive element binding protein (CREB)/c-Fos system
By combining in situ hybridization and immunohistochemistry with voltage-sensitive dye imaging (VSDi; Gandolfi et al, 2015), our results reveal a relevant degree of colocalization of long-term synaptic plasticity with activation of the CREB/c-Fos system through an N-methyl-Daspartate receptor (NMDAR)-dependent mechanism

Summary

Introduction

Long-term plasticity consists of changes in synaptic transmission and neuronal excitability (Hebb, 1949; Bliss and Lomo, 1973; Bienenstock et al, 1982; Bear and Abraham, 1996) that are thought to provide the basis for learning and memory in the brain (Bliss and Collingridge, 1993; Bliss et al, 2003; Bliss and Collingridge, 2013; Sweatt, 2016). Consolidation often begins through an N-methyl-D-aspartate receptor (NMDAR)-mediated calcium increase promoting phosphorylation of the cAMP-responsive element binding protein (CREB), which is involved both in synaptic plasticity and in plasticity of intrinsic excitability (Flavell and Greenberg, 2008; Benito and Barco, 2010). CREB is a constitutive transcription factor (TF) that regulates the transcription of genes with a CRE site in their promoter leading to the expression of inducible TFs including the immediately early gene (IEG) c-Fos (Brindle and Montminy, 1992; West et al, 2002; Alberini, 2009). Whether the CREB/c-Fos system is activated in response to activity patterns inducing long-term plasticity of synaptic transmission and intrinsic excitability at the mossy fiber—granule cell (MF-GrC) relay (D’Angelo et al, 1999; Armano et al, 2000; Gall et al, 2005; Seja et al, 2012) is unknown. Enough, GrCs show the highest NMDAR and CREB expression levels amongst all cerebellar neurons (Monaghan and Anderson, 1991; Brodie et al, 2004), the role of CREB in cerebellar synaptic plasticity has been so far investigated only for parallel fiber long-term depression (LTD) in Purkinje cells (PC; Ahn et al, 1999), which do not express postsynaptic NMDARs at these synapses (Piochon et al, 2010)

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Discussion

Conclusion