Elsevier

Brain Research

Volume 1435, 30 January 2012, Pages 1-7
Brain Research

Research Report
Developmental AMPA receptor subunit specificity during experience-driven synaptic plasticity in the rat barrel cortex

https://doi.org/10.1016/j.brainres.2011.11.033Get rights and content

Abstract

During early postnatal brain development, experience-driven delivery of AMPA receptors to synapses participates in the initial organization of cortical function. By combining virus-mediated in vivo gene delivery with in vitro whole cell recordings, we identified a subunit-specific developmental program of experience-driven AMPA receptor delivery to synapses in rat barrel cortex. We expressed green fluorescent protein (GFP)-tagged AMPA receptors (GFP-GluR1, or GFP-GluR4) into layer 2/3 pyramidal neurons at two distinct developmental periods, postnatal day (P)8–P10 and P12–P14. Two days after viral infection, acute brain slices were prepared, and synaptic transmission from layer 4 to layer 2/3 was analyzed by whole cell recordings. We found that whisker experience drives GluR4 but not GluR1 into these synapses early in postnatal development (P8–P10). However, at P12–14, GluR1 but not GluR4 is delivered into synapses by whisker experience. This precise developmental plan suggests unique plasticity properties endowed in different AMPA receptor subunits which shape the initial experience-driven organization of cortical function.

Highlights

► We identified a developmental program of experience-driven AMPAR delivery. ► We found whisker experience drives GluR4 but not GluR1 early in development (P8–P10). ► We found experience drives GluR1 but not GluR4 later in development (P12–P14).

Introduction

AMPA-type glutamate receptors are mainly responsible for the fast neurotransmission of glutamatergic synapses (Derkach et al., 2007, Malinow and Malenka, 2002). AMPA receptors in the central nervous system (CNS) form tetramers comprised of four subunits (GluR1,2,3 and 4) (Barry and Ziff, 2002, Hollmann and Heinemann, 1994). These four receptors can be classified into two groups. While GluR1 and 4 possess long cytoplasmic carboxyl terminal (C-tail), GluR2 and GluR3 have short cytoplasmic C-tail (Esaki et al., 2005, Sheng and Lee, 2001). In addition, GluR2 has a splicing variant which contains long C-tail, named GluR2long (Kohler et al., 1994).

Previous studies revealed that synaptic activity in vitro (Barry and Ziff, 2002, Boehm et al., 2006, Bredt and Nicoll, 2003, Hayashi et al., 2000, Kakegawa et al., 2004, Malinow and Malenka, 2002, Scannevin and Huganir, 2000, Shi et al., 2001, Zhu et al., 2000) and experience in vivo (Clem and Barth, 2006, Clem et al., 2010, Jitsuki et al., 2011, Kessels and Malinow, 2009, Mitsushima et al., 2011, Rumpel et al., 2005, Takahashi et al., 2003) drive AMPA receptors with long C-tail into synapses. In in vitro hippocampal slice cultures, spontaneous activity is sufficient to drive GluR4 and GluR2long into synapses (Kolleker et al., 2003, Zhu et al., 2000), while synaptic delivery of GluR1 requires robust synaptic activity such as LTP inducing stimuli. In vivo, GluR1 is driven by whisker experience into synapses from layer 4 to layer 2/3 pyramidal neurons in rat barrel cortex (Takahashi et al., 2003). However, the developmental constraints for activity-driven synaptic delivery of GluR1, or other long-C-tail receptor subunits, have not been established.

Here we characterized the experience dependent delivery of long-tailed AMPA receptors into synapses formed from layer 4 to 2/3 in the developing rat barrel cortex in vivo. We identify two periods characterized by activity-driven synaptic incorporation of different long-tailed AMPA receptor subunits. Between P8 and P10, whisker experience drives GluR4 but not GluR1 into synapses. This corresponds to a time when the initial synaptic connection of these layers in the rat barrel cortex is being established (Raymond et al., 2001, Stern et al., 2001). Later, between P12 and P14, GluR1 but not GluR4 is delivered into synapses by whisker experience at a time when the fine mapping in the barrel field is formed. Thus, our results suggest that the initial connections can be made functional rather permissively (given the low level of activity required for Glur4 synaptic incorporation). However, their fine tuning is conducted with a greater amount of quality control, since GluR1 has higher requirements for synaptic incorporation.

Section snippets

Expression of AMPA receptors in the developing rat barrel cortex

In order to elucidate roles of synaptic delivery of long-tailed AMPA receptors on the establishment of cortical circuit in the developing barrel cortex, we first investigated mRNA expression of GluR1 and GluR4 in the developing rat barrel cortex by RT-PCR. mRNA of both long-tailed AMPA receptors was observed at P9 and P14 (Fig. 1A).

Next, we examined protein expression of these receptors. Tissues from layers 1 to 3 area of barrel region at either P10 or P14 were obtained, and P2 fraction was

Discussion

In this study, we have examined the developmental profile of experience-driven AMPA receptor trafficking at the synapse between cortical layers 4 and 2/3. At a time when synapses are initially being constructed (P8 to P10) (Raymond et al., 2001), GluR4-containing receptors can be driven into synapses by experience; GluR1-containing receptors cannot. A few days later (P12 to P14), when connections at these synapses are being fine tuned, experience can drive GluR1-containing receptors into

RT-PCR

Nine to fourteen-day-old Wistar rats (Charles River) were decapitated under halothane anesthesia and barrel cortex including from layes 1 to 4 were microdissected. Total RNAs were extracted with TriZol (Invitrogen) and reverse transcribed to cDNA with random primers (SuperScript VILO cDNA Syntesis kit, Invitrogen) according to the manufacturer's instructions. The cDNAs were amplified by PCR (GoTaq Green Master Mix, Promega) with sequence specific primers according to Dijk et al.(Bureau et al.,

Acknowledgment

We thank Yoshiko Kanno for excellent technical assistance. This project was supported by Grant-in-Aid for Young Scientists (start-up) (18800037), Grant-in-Aid for Scientific Research (B) (20300131) (T.T.),JST, CREST (T.T.), Special Coordination Funds for Promoting Science and Technology (T.T.), the Sumitomo Foundation (T.T.), KANAE Foundation (T.T.), Takeda Science Foundation (T.T.), NARSAD (T.T.), the grant for 2007 Strategic Research Project (K19025) of Yokohama City University (T.T.),

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