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The clustering of neurotransmitter receptors at postsynaptic sites is considered to play an important role in modulating synaptic efficacy. To investigate the mechanisms underlying neurotransmitter receptor clustering, we expressed the NMDA ( N -methyl- d -aspartate) receptor NR1A subunit in human embryonic kidney (HEK) 293 cells. As previously shown, the cells exhibited subcellular clusters of the receptor protein with a mean diameter of approximately 0.7 μm. To examine the involvement of cytoskeletal structures on this clustering, we disrupted actin filaments or microtubules by treating the cells with alkaloids. In the actin filament-disrupted cells, the receptor protein shifted from the cellular membrane to the cytoplasm where it formed macroclusters (approximate diameter 3 μm). In the microtubule-disrupted cells, the subcellular clusters of NR1A could not be detected and the protein was diffusely distributed throughout the cytoplasm. Similar results were obtained by coexpression of the receptor protein with fusion proteins harboring various C-terminal exon cassettes. These results suggest that subcellular clustering of the NR1 subunit of the NMDA receptors is regulated by the interaction of its C-terminus with cytoskeletal components, where differentially spliced cassettes interact separately with actin filaments or microtubules. Modulation of the interaction between the neurotransmitter receptors and the cytoskeleton leads to the rearrangement of the receptor clusters and may contribute to certain types of synaptic plasticity.

Cerebellar long-term depression (LTD) induced at parallel fiber-Purkinje neuron synapses is proposed to underlie certain types of motor learning. α-Amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptors, which mediate chemical transmission in these synapses, are clustered on the postsynaptic membrane. By increasing local density of the receptors, clustering is believed to increase synaptic efficacy. This article focuses on molecular mechanisms regulating the synaptic AMPA receptor clustering in Purkinje cells, which could underlie the expression of cerebellar LTD. Synaptic AMPA receptor clusters in dendritic spines of Purkinje cells are disrupted upon protein kinase C (PKC)-mediated phosphorylation of serine 880 in the C-terminal domain of GluR2. Phosphorylation of this residue causes significant reduction in the affinity of GluR2 C-terminal tail for glutamate receptor interacting protein (GRIP), a molecule known to be crucial for AMPA receptor clustering. Consequently, AMPA receptors on the synaptic membrane are destabilized and internalized by endocytosis. Based on these findings, a model for the expression of cerebellar LTD is proposed, in which a decrease in the number of postsynaptic AMPA receptors, initiated by phosphorylation of GluR2 serine 880, is the major mechanism underlying cerebellar LTD.

The interaction of neurotransmitter receptors with the underlying cytoskeleton via subsynaptic proteins is an important mechanism for the targeting of the receptors to synapses in the central nervous system. We show that delta glutamate receptors (delta receptors), expressed predominantly in the dendritic spines of cerebellar Purkinje cells, directly interact with spectrin, a member of the actin-binding family of proteins. Moreover, the interaction between spectrin and C-terminal domain of the delta receptor is 50% inhibited by 1 microM of Ca2+ in vitro, compared with that in the absence of Ca2+. These results suggest that delta receptors on the postsynaptic membrane of the dendritic spines of cerebellar Purkinje cells are anchored to the actin cytoskeleton via spectrin, and that Ca2+ elevation in the dendritic spines causes delta receptor declustering by dissociation of the receptors from spectrin. This mechanism for receptor anchoring at postsynaptic sites may regulate synaptogenesis and/or synaptic plasticity.