Protein Kinase C Induced Glutamate Receptor Trafficking And Insertion To Postsynaptic Membrane

Glutamate receptors are the predominant excitatory neurotransmitter in hippocampus. Ionotropic glutamate receptors (iGluRs) are a major class of heteromeric ligand-gated ion channels and mediate the majority of the excitatory neurotransmission in the vertebrate central nervous system (CNS). iGluRs can be classified as N-methyl-D-aspartate (NMDA) receptor or the non-NMDA receptor, which can be further subdivided intoα-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptor, or kainate (KA) receptor. These receptors are important for neural development, learning and memory and cognition. One of the major functions of glutamate receptors appears to be the modulation of synaptic transmission, resulting in synaptic plasticity. An important cellular models of synaptic plasticity in the mammalian brain are the long-term depression (LTD) and long-term potentiation (LTP) that are generated at excitatory synapses on hippocampal CA1 pyramidal cells.An increase or decrease in the number of ionotropic glutamate receptors on a post-synaptic cell may lead to long-term potentiation or long-term depression of that cell, respectively. NMDA receptor, as a"molecular switch", triggers synaptic plasticity and contributes to the induction of LTP. While AMPA receptor is important for memory storage and participates in the expression and maintainance of LTP. During LTP, excitatory postsynaptic current (EPSC) is enhanced by the increased numbers and efficiency of AMPA receptor. Therefore, the trafficking of NMDA or AMPA receptor is critical for the regulation of syanptic plasticity.NMDA and AMPA receptor can be phosphorylated by a series of protein kinase (such as PKC, PKA, CaMKII et al.), which directly influence their functional property, including channel localization, conductance, and open probability. Recent studies have shown that PKC promotes delivery of NMDA or AMPA receptor to surface membrane. However, the detailed molecular mechanisms remain unclear. Thus, The primary aim of the study was to investigate the possible molecular mechanism of NMDA or AMPA receptor trafficking regulated by PKC.1. Protein kinase C promotes NMDA receptor trafficking by indirectly triggering CaMKII autophosphorylationThe NMDA receptor (NMDAR) plays a central role in the function of excitatory synapses. Many forms of synaptic plasticity depend on NMDA receptor activation and subsequent increase of intracellular Ca2+. Therefore, it is very important for synaptic plasticity to regulate functional NMDAR. Using the methods of western blotting, immunoprecipitation, immunofluorescene and electrophysiology, we demonstrate the possible mechanism of NMDAR trafficking induced by protein kinase C (PKC). Our results showed that PKC promotes NMDA receptor trafficking to the cell surface or indirectly activates Src and upregulates NMDA receptor function. PKC promotes NMDA receptor trafficking to the cell surface via interaction with NMDAR-associated proteins (NAPs). Here we show that CaMKII is such a NAP that mediates potentiation of NMDAR trafficking by PKC. PKC activation elicits concurrent potentiation in both CaMKII autophosphorylation and association between CaMKII and NMDARs, accompanying by functional NMDAR insertion, at postsynaptic site. This PKC-induced potentiation was abolished by CaMKII antagonist AIP or by selectively disturbing the interaction between CaMKII and NR2A or NR2B with peptide Tat-NR2A and Tat-NR2B. The PKC-tyrosine kinase Src signaling pathway contributed to the potentiation of NMDA channel activity but was not required for PKC-induced NMDAR trafficking. Moreover, Tat-NR2A and Tat-NR2B completely abolished PKC-induced long-term potentiation (LTP) respectively. Further mutual occluding experiments demonstrate that PKC and CaMKII share common signaling pathway in potentiation of NMDAR trafficking and LTP production. Taken together, our present results support the interpretation that PKC promotes NMDA receptor trafficking by indirectly triggering CaMKII autophosphorylation and subsequent increased association with NMDARs, , which appears to be critical for synaptic palsticity..2. Activation of aPKCλis required for AMPA receptors insertion during PI3K-induced LTP.AMPA receptors mediate fast synaptic transmission at a majority of excitatory synapses, and participate in the expression and maintainance of LTP. The number and functional property of AMPA receptor (AMPAR) in postsynaptic sites is widely regarded as a major determinant of synaptic plasticity. Therefore, it is very important for us to understand an underlying mechanism of AMPA receptor trafficking. Using the methods of western blotting, immunoprecipitation, immunofluorescene and electrophysiology, we illustrate the role of PKC in modulating the trafficking of AMPA receptor in synapse. Our results demonstrated that aPKCλ, a specifically atypical PKC, is necessary and sufficient for PI3K-induced LTP. aPKCλmay be activated indirectly by PI3K and promotes GluR1 insertion to postsynaptic membrane. Further studies have shown that P62, as adaptor protein, mediateds the formation of GluR1-P62-aPKCλcomplex, and recruits GluR1 closer to aPKCλ. Then aPKCλphosphorylates GluR1 at ser818 and regulates AMPA receptor trafficking to postsynaptic membrane.