Molecular mechanisms of AMPA receptor synaptic recruitment

The formation of synapses involve the recruitment of both pre- and postsynaptic components to the nascent synapse. Neurons in the central nervous system (CNS) form both excitatory and inhibitory synapses, and the appropriate neurotransmitter receptors must be recruited to postsynaptic specializations opposing presynaptic terminals releasing the correct neurotransmitter for functional synaptic transmission. Inhibitory neurotransmission in the CNS is mediated mainly by the glycine and GABAA receptors, while excitatory neurotransmission is mediated mainly by the NMDA and AMPA subtypes of ionotropic glutamate receptors. Despite our knowledge of the structure and function of the AMPA receptors, the molecular mechanisms required for synaptic recruitment of these receptors remains poorly characterized. In this study, we ask the following questions: (1) what domains on the AMPA receptor is required for synaptic recruitment, and (2)what axonal factors are required for synaptic recruitment of AMPA receptors. To answer these questions, we have established a hippocampal neuron-glia coculture system where neurons express factors which recruit GluR4 overexpressed in glia to sites of contact between excitatory axons and glia. Using this system, we have mapped the domain on GluR4 which is necessary and sufficient for synaptic recruitment. We found that GluR4 lacking the N-terminal domain (NTD) fail to get recruited by excitatory axons when overexpressed in glia, and fail to get recruited to synaptic sites when expressed in neurons, whereas the NTD alone expressed on the surface of glia or neurons gets recruited to excitatory axons and synaptic sites respectively. We have also investigated the involvement of the neuronal pentraxins in the synaptic recruitment of GluR4. Neuronal pentraxin 1 (NP1) and neuronal pentraxin receptor (NPR) are colocalized with glial clusters of GluR4 induced by excitatory axons, and are colocalized with endogenous GluR4 in hippocampal dissociated cultures. NP1 also causes coclustering of GluR4 and the NTD when coexpressed in heterologous cells. Knockdown of NP1 and NPR by RNAi decreases the ability of axons to recruit GluR4 to synapses. These results suggest that the neuronal pentraxins may recruit GluR4 to synapses through the NTD.