Elsevier

Neuropharmacology

Volume 45, Issue 6, November 2003, Pages 703-713
Neuropharmacology

Regulation of AMPA receptor dephosphorylation by glutamate receptor agonists

https://doi.org/10.1016/S0028-3908(03)00319-8Get rights and content

Abstract

Phosphorylation of the α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor subunit GluR1 at Ser845 enhances AMPA channel activity. This study demonstrates that Ser845 is rapidly dephosphorylated upon AMPA receptor activation in nucleus accumbens slices. AMPA-induced dephosphorylation at Ser845 was blocked by CNQX, an AMPA receptor antagonist, by nifedipine, an L-type Ca2+ channel antagonist, or by cyclosporin A, a calcineurin inhibitor. N-methyl-d-aspartate (NMDA) treatment also decreased phosphorylation of Ser845, an effect that was blocked by MK-801, an NMDA receptor antagonist, but not by nifedipine. Accumbens neurons are enriched for dopamine- and cyclic AMP (cAMP)-regulated phosphoprotein, Mr 32,000 (DARPP-32), a potent inhibitor of protein phosphatase 1 (PP1) when phosphorylated by PKA (at Thr34). We tested the hypothesis that the AMPA/KA or NMDA-stimulated dephosphorylation of DARPP-32 via calcineurin, leading to increased PP1 activity and dephosphorylation of GluR1. AMPA or NMDA treatment decreased phospho-Thr34-DARPP-32 levels, effects that were blocked by receptor antagonists, or cyclosporin A. However, dephosphorylation of Ser845 mediated by AMPA or NMDA receptors was unaffected in DARPP-32/inhibitor-1 knockout mice. These data suggest that AMPA- or NMDA-induced dephosphorylation of GluR1 at Ser845 occurs by a mechanism that is independent of DARPP-32 and PP1, but involves activation of calcineurin. Thus, Ca2+-dependent dephosphorylation of GluR1 may serve as a negative feedback mechanism for the regulation of AMPA receptor activity in neurons.

Introduction

The neostriatum, including the dorsal striatum and the nucleus accumbens, plays a critical role in regulating functions as diverse as the control of motor activity (Albin et al., 1989) and the development of drug addiction (Nestler, 2001). Ascending dopamine (DA)-containing neurons from the midbrain innervate the predominant cell type of the striatum and nucleus accumbens, the medium spiny-type neuron (Kemp and Powell, 1971). DA, released from these neurons, modulates the excitability of medium spiny neurons in response to glutamate, released from afferents from the cortex and other brain regions.

Three subtypes of ligand-gated ion channels, N-methyl-d-aspartate (NMDA), (±)-α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA), and kainate subclasses, mediate actions of glutamate in the brain (Hollamnn and Heinemann, 1994). Each of these receptor subtypes is expressed in medium spiny neurons of the striatum and nucleus accumbens (Petralia and Wenthold, 1992, Martin et al., 1993, Tallaksen-Greene and Albin, 1994, Ghasemzadeh et al., 1996, Bernard et al., 1997).

AMPA receptors, in particular, are believed to mediate a large proportion of glutamatergic excitation of medium spiny neurons (Cepeda et al., 1993). Phosphorylation of the GluR1 subunit of the AMPA receptor at Ser845 by cAMP-dependent protein kinase (PKA) (Roche et al., 1996) and at Ser831 by protein kinase C (PKC) or Ca2+/calmodulin-dependent protein kinase II (CaMKII) (Barria et al., 1997) enhances the activity of AMPA channels. Moreover, the increased phosphorylation of GluR1 at Ser845 (Snyder et al., 2000) in response to D1-type DA receptor activation has been implicated in the enhancement of AMPA receptor currents in medium spiny neurons (Yan et al., 1999). The ability of DA to regulate both GluR1 phosphorylation and AMPA channel activity requires the inhibition of protein phosphatase 1 (PP1), a major protein serine/threonine phosphatase in the brain. Inhibition of PP1 activity is mediated via a biochemical cascade involving the phosphorylation by PKA of dopamine- and cyclic AMP (c-AMP)-regulated phosphoprotein, Mr 32,000 (DARPP-32), a protein that is highly enriched in medium-sized spiny neurons (Ouimet et al., 1984). DARPP-32, when phosphorylated by PKA on a single threonine residue (Thr34), is converted into a potent inhibitor of PP1 (Hemmings et al., 1984). As a result, DARPP-32 plays a critical role in regulating the excitability of medium spiny neostriatal neurons through its functional regulation of a number of cellular effectors. In addition to AMPA receptors, these include voltage-dependent Ca2+ and Na+ channels, the electrogenic ion pump Na+, K+-ATPase, and NMDA and GABAA receptors (Fienberg et al., 1998, Greengard et al., 1999, Snyder et al., 2000, Flores-Hernandez et al., 2000).

It has been shown that activation of NMDA receptors in a model of hippocampal long-term depression (LTD) results in a marked dephosphorylation of GluR1 at Ser845 (Lee et al., 1998) suggesting that glutamate-induced dephosphorylation of GluR1 may be a critical mechanism for regulating the functional activity of AMPA receptors. Activation of NMDA receptors on striatal neurons has been shown to increase Ca2+ influx into neurons (MacDermott et al., 1986), leading to the dephosphorylation of DARPP-32 by calcineurin, the Ca2+/calmodulin-dependent protein phosphatase (Halpain et al., 1990). The aim of the present study was to determine whether glutamate, via activation of NMDA and/or non-NMDA subclasses of glutamate receptors, might also regulate the dephosphorylation of GluR1 in neostriatal neurons and whether this might involve DARPP-32 and PP1.

Section snippets

Materials

(±)-α-Amino-3-hydroxy-5-methylisoxazole-4-propionic acid hydrobromide (AMPA hydrobromide), 6-cyano-7-nitroquinoxaline-2,3-dione, disodium salt (CNQX), N-methyl-D-aspartate, d(−)-2-amino-5-phosphonopentanoic acid (AP-5), and MK-801 hydrogen maleate were obtained from Research Biochemicals, Inc. (Natick, MA) and Sigma Chemical Co. (St. Louis, MO). Nifedipine and cyclosporin A were purchased from Alexis Biochemicals (San Diego, CA). Goat anti-mouse IgG was purchased from Pierce, nitrocellulose

AMPA-induced dephosphorylation of GluR1 at Ser845: calcineurin-dependence

The regulation of phosphorylation of Ser845, a site that controls channel open time probability (Roche et al., 1996), was examined in nucleus accumbens slices. The GluR1 subunit of the AMPA receptor was detected as a band of Mr~106 kDa in homogenates of rat nucleus accumbens (Fig. 1A). A basal level of phosphorylation was detected in untreated slices. Treatment of slices with AMPA, an agonist at AMPA and kainate subclasses of glutamate receptors, promoted a rapid dephosphorylation of Ser845

Discussion

The present study demonstrates that the activation of AMPA/kainate subclass of glutamate receptor, an expected result of D1-mediated phosphorylation of GluR1, induces a rapid dephosphorylation of GluR1 at Ser845 and of DARPP-32 at Thr34. However, dephosphorylation of GluR1 in response to AMPA did not appear to require regulation of PP1 activity. This conclusion is based on the inability of calyculin A, at concentrations that are known to block most PP1 activity in slices, to prevent

Acknowledgements

These studies were supported by U.S.P.H.S. Grant 40899 (to P.G., G.L.S., and A.C.N.). The authors wish to thank Dr. Akinori Nishi for helpful comments and Carmina Valle and Peter Ingrassia for excellent technical assistance.

References (40)

  • A. Barria et al.

    Regulatory phosphorylation of AMPA-type glutamate receptors by CaM-KI1 during long-term potentiation

    Science

    (1997)
  • V. Bernard et al.

    Cellular, subcellular, and subsynaptic distribution of AMPA-type glutamate receptor subunits in the neostriatum of the rat

    J. Neurosci.

    (1997)
  • J.A. Bibb et al.

    Phosphorylation of DARPP-32 by Cdk5 modulates dopamine signaling in neurons

    Nature

    (1999)
  • A.J. Borgdorff et al.

    Regulation of AMPA receptor lateral movements

    Nature

    (2002)
  • C. Cepeda et al.

    Neuromodulatory actions of dopamine in the neostriatum are dependent upon the excitatory amino acid subtypes activated

    Proc. Natl. Acad Sci., USA

    (1993)
  • A.A. Fienberg et al.

    DARPP-32: regulator of the efficacy of dopaminergic neurotransmission

    Science

    (1998)
  • J. Flores-Hernandez et al.

    D1 dopamine receptor activation reduced GABAA receptor currents in neostriatal neurons through a PKA/DARPP-32/PP1 signaling cascade

    J. Neurophysiol.

    (2000)
  • M.B. Ghasemzadeh et al.

    Multiplicity of glutamate receptor subunits in single striatal neurons: an RNA amplification study

    Molec. Pharmacol.

    (1996)
  • S. Halpain et al.

    Activation of NMDA receptors induced dephosphorylation of DARPP-32 in rat striatal slices

    Nature

    (1990)
  • H.C. Hemmings et al.

    DARPP-32, a dopamine- and adenosine 3′:5′-monophosphate-regulated phosphoprotein: regional, tissue and phylogenetic distribution

    J. Neurosci.

    (1986)
  • Cited by (0)

    View full text