Subunit-Specific Regulation of NMDA Receptor Endocytosis

At excitatory synapses, both NMDA and AMPA receptors are localized to the postsynaptic density (PSD). However, unlike AMPA receptors, synaptic NMDA receptors are stable components of the PSD. Even so, surface-expressed NMDA receptors undergo endocytosis, which is more robust early in development and declines during synaptic development. We investigated the subunit-specific contributions to NMDA receptor endocytosis, specifically defining the endocytic motifs and endocytic pathways preferred by the NR2A and NR2B subunits. We find that NR2A and NR2B have distinct endocytic motifs encoded in their distal C termini and that these interact with clathrin adaptor complexes with differing affinities. We also find that NR2A and NR2B sort into different intracellular pathways after endocytosis, with NR2B preferentially trafficking through recycling endosomes. In mature cultures, we find that NR2B undergoes more robust endocytosis than NR2A, consistent with previous studies showing that NR2A is more highly expressed at stable synaptic sites. Our findings demonstrate fundamental differences between NR2A and NR2B that help clarify developmental changes in NMDA receptor trafficking and surface expression.

[1]  Y. Jan,et al.  Clustering of Shaker-type K+ channels by interaction with a family of membrane-associated guanylate kinases , 1995, Nature.

[2]  Ann Marie Craig,et al.  Activity Regulates the Synaptic Localization of the NMDA Receptor in Hippocampal Neurons , 1997, Neuron.

[3]  K. Roche,et al.  PDZ Domain Suppression of an ER Retention Signal in NMDA Receptor NR1 Splice Variants , 2000, Neuron.

[4]  P. Seeburg,et al.  Domain interaction between NMDA receptor subunits and the postsynaptic density protein PSD-95. , 1995, Science.

[5]  R. Wenthold,et al.  Turnover Analysis of Glutamate Receptors Identifies a Rapidly Degraded Pool of the N-Methyl-d-aspartate Receptor Subunit, NR1, in Cultured Cerebellar Granule Cells* , 1999, The Journal of Biological Chemistry.

[6]  S. Coultrap,et al.  LTP leads to rapid surface expression of NMDA but not AMPA receptors in adult rat CA1 , 2002, Nature Neuroscience.

[7]  J. Bonifacino,et al.  Signals for sorting of transmembrane proteins to endosomes and lysosomes. , 2003, Annual review of biochemistry.

[8]  S. Vicini,et al.  Functional and pharmacological differences between recombinant N-methyl-D-aspartate receptors. , 1998, Journal of neurophysiology.

[9]  G. Westbrook,et al.  A use-dependent tyrosine dephosphorylation of NMDA receptors is independent of ion flux , 2001, Nature Neuroscience.

[10]  J. Bonifacino,et al.  Signal-binding Specificity of the μ4 Subunit of the Adaptor Protein Complex AP-4* , 2001, The Journal of Biological Chemistry.

[11]  R. Petralia,et al.  Trafficking of NMDA receptors. , 2003, Annual review of pharmacology and toxicology.

[12]  S. Vicini,et al.  Developmental changes in localization of NMDA receptor subunits in primary cultures of cortical neurons , 1998, The European journal of neuroscience.

[13]  K. Roche,et al.  Molecular determinants of NMDA receptor internalization , 2001, Nature Neuroscience.

[14]  L. Raymond,et al.  Differential regulation of synaptic and extra-synaptic NMDA receptors , 2002, Nature Neuroscience.

[15]  W. Ju,et al.  Glycine binding primes NMDA receptor internalization , 2003, Nature.

[16]  S. Vicini,et al.  Increased contribution of NR2A subunit to synaptic NMDA receptors in developing rat cortical neurons , 1998, The Journal of physiology.

[17]  G. Westbrook,et al.  Mobile NMDA Receptors at Hippocampal Synapses , 2002, Neuron.

[18]  Mark von Zastrow,et al.  Role of AMPA Receptor Cycling in Synaptic Transmission and Plasticity , 1999, Neuron.

[19]  G. Rumbaugh,et al.  Distinct Synaptic and Extrasynaptic NMDA Receptors in Developing Cerebellar Granule Neurons , 1999, The Journal of Neuroscience.

[20]  R. Huganir,et al.  Redistribution and Stabilization of Cell Surface Glutamate Receptors during Synapse Formation , 1997, The Journal of Neuroscience.

[21]  M. Ehlers,et al.  An NMDA Receptor ER Retention Signal Regulated by Phosphorylation and Alternative Splicing , 2001, The Journal of Neuroscience.

[22]  G. Westbrook,et al.  The Incorporation of NMDA Receptors with a Distinct Subunit Composition at Nascent Hippocampal Synapses In Vitro , 1999, The Journal of Neuroscience.

[23]  M. Salter,et al.  Ca(2+)-independent reduction of N-methyl-D-aspartate channel activity by protein tyrosine phosphatase. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[24]  J. Bonifacino,et al.  Interaction of tyrosine-based sorting signals with clathrin-associated proteins. , 1995, Science.

[25]  R. Malenka,et al.  An ER retention signal explains differences in surface expression of NMDA and AMPA receptor subunits , 2001, Neuropharmacology.

[26]  R. Wenthold,et al.  Export from the Endoplasmic Reticulum of Assembled N-Methyl-D-aspartic Acid Receptors Is Controlled by a Motif in the C Terminus of the NR2 Subunit* , 2004, Journal of Biological Chemistry.

[27]  J. Bonifacino,et al.  Structural Determinants of Interaction of Tyrosine-based Sorting Signals with the Adaptor Medium Chains* , 1996, The Journal of Biological Chemistry.

[28]  R. Malenka,et al.  AMPA receptor trafficking and synaptic plasticity. , 2002, Annual review of neuroscience.

[29]  M. Sheng,et al.  Molecular organization of the postsynaptic specialization , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[30]  Mark F. Bear,et al.  Internalization of ionotropic glutamate receptors in response to mGluR activation , 2001, Nature Neuroscience.

[31]  K. Roche,et al.  Synaptic expression of the high-affinity kainate receptor subunit KA2 in hippocampal cultures , 1995, Neuroscience.

[32]  M. Sheng,et al.  PDZ Domains: Structural Modules for Protein Complex Assembly* , 2002, The Journal of Biological Chemistry.

[33]  A. McAllister,et al.  Rapid recruitment of NMDA receptor transport packets to nascent synapses , 2002, Nature Neuroscience.

[34]  E. Molnár,et al.  Assembly intracellular targeting and cell surface expression of the human N-methyl-d-aspartate receptor subunits NR1a and NR2A in transfected cells , 1998, Neuropharmacology.

[35]  Z. Fu,et al.  Relationship between Availability of NMDA Receptor Subunits and Their Expression at the Synapse , 2002, The Journal of Neuroscience.

[36]  T. Soderling,et al.  Quantitation of AMPA receptor surface expression in cultured hippocampal neurons , 1997, Neuroscience.

[37]  K. Roche,et al.  Differential binding of the AP-2 adaptor complex and PSD-95 to the C-terminus of the NMDA receptor subunit NR2B regulates surface expression , 2003, Neuropharmacology.

[38]  J. Kemp,et al.  Developmental Changes in NMDA Receptor Glycine Affinity and Ifenprodil Sensitivity Reveal Three Distinct Populations of NMDA Receptors in Individual Rat Cortical Neurons , 1998, The Journal of Neuroscience.