TNF alpha Signaling: Effects on AMPA Receptor Location

描述（由申请人提供）：本提案将研究肿瘤坏死因子α (tnf - α)信号级联及其对突触活性的快速影响。tnf - α已被证明可增加未患病神经元的突触活性，并可在疾病或创伤期间诱导神经元的兴奋性毒性。在兴奋性神经元中表达的nmda型和ampa型谷氨酸受体（NMDARs和AMPARs）在调节突触活动中起主要作用。因此，这些受体是tnf - α信号调节的可能靶点。特别是，ampar已被发现是突触后密度的高度调控和移动组成部分，并被认为通过快速定位到含有nmda的突触来增加突触增强。最近我们发现，快速（15分钟）向培养的大鼠海马神经元提供tnf - α增加了突触活性和AMPAR对突触的定位，但没有改变NMDARs的定位。这背后的信号机制尚不清楚。

DESCRIPTION (provided by applicant): This proposal will study the tumor necrosis factor alpha (TNF-alpha) signaling cascade and its rapid effects on synaptic activity. TNF-alpha has been shown to increase synaptic activity in non-diseased neurons, and can induce excitotoxicity in neurons during disease or trauma. NMDA-type and AMPA-type glutamate receptors (NMDARs and AMPARs) expressed in excitatory neurons are known to have major roles in modulating synaptic activity. These receptors therefore are possible targets of TNF-alpha signaling regulation. In particular, AMPARs have been found to be a highly regulated and mobile component of the postsynaptic density and are thought to increase synaptic potentiation via rapid localization to NMDAR-containing synapses. Recently we have shown that TNF-alpha supplied to rat hippocampal neurons in culture rapidly (15 minutes) increased synaptic activity and AMPAR localization to synapses, but did not alter the localization of NMDARs. The signaling mechanisms underlying this are unknown. This proposal will examine TNF-alpha signaling pathways potentially responsible for the TNF-alpha-induced increase in AMPAR localization and synaptic activity. Subcellular AMPAR localization will be examined after various treatments in cultured neurons from two CNS regions, the hippocampus and spinal cord. The analysis will utilize cell biological, immunohistochemical, biochemical and electrophysiological techniques. The first three aims in this proposal will examine TNF-alpha signaling in cultured rat and mouse hippocampal neurons. Specific aim 1 will determine if signaling from one of the two TNF-alpha receptors- TNFR1, TNFR2, or both contribute to increased AMPAR surface localization. TNFR1 and TNFR2 are both expressed in hippocampal neurons and elicit overlapping yet distinct signaling pathways. Specific aim 2 will test the hypothesis that CamKII and a calcium mediated pathway are responsible for this activity. Aim 3 will investigate the intracellular origin of the AMPARs that appear on the neuron surface after TNF-alpha signaling. Finally, Aim 4 will ask if TNF-alpha is able to cause increases in surface AMPAR localization on neurons of different regions of the CNS, the cortex and spinal cord. If so, the details of TNF-alpha's signaling on AMPAR localization in motorneurons and cortical neurons will be assessed as hippocampal neurons were assessed in aims 1 through 3. These aims outline the logical next step extending our preliminary data showing that TNF-alpha increases synaptic AMPAR localization. The results will significantly increase our understanding of synaptic function in the CNS in both health and disease. CNS neurons that undergo excitotoxic cell death are oversupplied with TNF-alpha after stroke or spinal cord injury as well as during diseases such as Alzheimer's, Parkinson's, and ALS. Recent data linking TNF-alpha signaling to increased synaptic activity suggests that an oversupply TNF-alpha may kill neurons by moving excessive numbers of AMPARs to synapses. Thus, specific knowledge of how TNF-alpha signaling increases AMPAR surface localization in particular classes of CNS neurons may allow us to more effectively prevent excitotoxicity in a range of diseases and trauma.