Modulation of N-methyl-D-aspartate receptors in the brain by protein phosphorylation may play a central role in the regulation of synaptic plasticity. To examine the phosphorylation of the NR1 subunit of N-methyl-Daspartate receptors in situ, we have generated several polyclonal antibodies that recognize the NR1 subunit only when specific serine residues are phosphorylated. Using these antibodies, we demonstrate that protein kinase C (PKC) phosphorylates serine residues 890 and 896 and cAMP-dependent protein kinase (PKA) phosphorylates serine residue 897 of the NR1 subunit. Activation of PKC and PKA together lead to the simultaneous phosphorylation of neighboring serine residues 896 and 897. Phosphorylation of serine 890 by PKC results in the dispersion of surface-associated clusters of the NR1 subunit expressed in fibroblasts, while phosphorylation of serine 896 and 897 has no effect on the subcellular distribution of NR1. The PKC-induced redistribution of the NR1 subunit in cells occurs within minutes of serine 890 phosphorylation and reverses upon dephosphorylation. These results demonstrate that PKA and PKC phosphorylate distinct residues within a small region of the NR1 subunit and differentially affect the subcellular distribution of the NR1 subunit.Ionotropic glutamate receptors mediate most rapid excitatory transmission in the central nervous system and play important roles in synaptic plasticity, neuronal development, and neurological disorders (1-5). Glutamate receptors have been divided into NMDA 1 (N-methyl-D-aspartate) and non-NMDA (kainate or AMPA) receptors based on their pharmacological and physiological properties (1, 2). Non-NMDA glutamate receptors activate and desensitize rapidly and mediate excitatory synaptic transmission. NMDA receptors are more slowly activated and desensitized and have a high Ca 2ϩ permeability and a voltage-dependent Mg 2ϩ block, two properties thought to underlie use-dependent synaptic plasticity in the brain (1-3). Molecular cloning studies have recently identified the genes encoding subunits for the NMDA and non-NMDA receptors (1, 2). NMDA receptors consist of two families of homologous subunits, the NR1 and NR2A-D subunits (6 -9), and are thought to be pentameric or tetrameric complexes of the NR1 subunit with one or more of the NR2 subunits (10, 11). The differential expression of NR2 subunits in the various regions of the brain may account for the diversity of NMDA receptor subtypes (1). In addition, the NR1 subunit is highly alternatively spliced giving rise to at least seven forms of NR1 (NR1A-G) increasing the potential diversity of NMDA receptors in the brain (12-14).Protein phosphorylation has been recognized as a major mechanism for the regulation of glutamate receptor function (15). NMDA receptors appear to be regulated by a number of protein kinases and phosphatases. Activation of protein kinase C (PKC) by phorbol esters have been demonstrated to activate (16,17) or depress (18, 19) neuronal NMDA receptors. In addition, intracellular perfusion of purified ...

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