Functional Characterization of synaptic calcium/calmodulin kinase II (CaMKII) acc

描述（由申请人提供）：钙/钙调素依赖性激酶II （CaMKII）广泛表达，CaMKII的激活在细胞分裂、碳水化合物代谢、突触可塑性和学习记忆等多种细胞过程中起着重要作用（见Lisman et al.， 2002和Colomer and Means， 2007）。本研究的长期目标是探讨钙/钙调蛋白激酶II （CaMKII）的局部诱导增殖突触（L-IPS）易位的新现象的作用。这种异突触过程可以被描述为一种缓慢的、波状的突触CaMKII积累传播，源于谷氨酸和甘氨酸局部应用于一小部分树突，激活一部分相邻突触。在刺激后的10分钟内，CaMKII不仅在刺激部位的突触中积累，而且在整个树突乔木的突触中积累。这一过程依赖于NMDA受体的激活，l型Ca2+通道，并导致ampa型谷氨酸受体亚基GluR1在突触的表达增加（比基线高出18%）。具体目的包括：确定L-IPS易位后突触积累诱导的异突触可塑性是否阻断了长期增强（LTP，一种已被充分表征的突触可塑性机制）。此外，我们还将对CaMKII L-IPS易位后其他突触蛋白（如GluR2和NMDA受体亚基）的表达水平进行研究。目前的研究旨在证实L-IPS易位导致的突触CaMKII积累的稳定性和可逆性。最后，脊柱形态的变化也将在CaMKII L-IPS易位期间和之后的一段时间内进行检查。分离海马细胞培养中表达的报告结构的活细胞成像将是研究这些问题的主要方法，这些问题涉及CaMKII的L-IPS易位的神经元功能。局部刺激小树突区域导致CaMKII在整个树突乔木的非刺激突触的突触积累，这一发现暗示了可塑性和细胞内信号传导的新特性；因此，需要对这一过程进行功能表征。包括阿尔茨海默病、癫痫、帕金森氏病和因缺血导致的细胞损失在内的几种疾病都与CaMKII有关。揭示这种异突触现象的作用可以进一步阐明细胞范围内CaMKII激活的动力学，并为发现这些疾病的一些机制提供可能的途径。

DESCRIPTION (provided by applicant): Calcium/calmodulin dependent kinase II (CaMKII) is widely expressed and activation of CaMKII plays a fundamental role in several cellular processes ranging from proper cell division and carbohydrate metabolism to synaptic plasticity and learning and memory (see Lisman et al., 2002 and Colomer and Means, 2007). The long-term goal of this research is to investigate the role of novel phenomenon called locally-induced propagating synaptic (L-IPS) translocation of calcium/calmodulin kinase II (CaMKII). This heterosynaptic process can be described as a slow, wave-like propagation of synaptic CaMKII accumulation that stems from localized application of glutamate and glycine to a small portion of dendrite, activating a subset of contiguous synapses. Within 10 minutes after this stimulation, CaMKII accumulates at synapses not only at the stimulation site but at synapses throughout the dendritic arbor. This process relies on NMDA receptor activation, L-type Ca2+ channels and results in an increase of expression of the AMPA-type glutamate receptor subunit GluR1 at synapses (~18% above baseline). Specific aims include: determining if heterosynaptic plasticity induced by synaptic accumulation following L-IPS translocation occludes long-term potentiation (LTP; a well-characterized proposed mechanism of synaptic plasticity). As well, investigation into expression levels of other synaptic proteins (e.g., GluR2, and NMDA receptor subunits) following L-IPS translocation of CaMKII will be conducted. The current research proposes studies to confirm stability and reversibility of synaptic CaMKII accumulation resulting from L-IPS translocation. Finally, changes in spine morphology will also be examined both during and at periods after L-IPS translocation of CaMKII. Live-cell imaging of reporter constructs expressed in dissociated hippocampal cell culture will be the predominate method employed to investigate these questions addressing the neuronal function of L-IPS translocation of CaMKII. The finding that local stimulation of a small dendrite area results in synaptic accumulation of CaMKII at non-stimulated synapses throughout the dendritic arbor alludes to a novel property of plasticity and intracellular signaling; thus, functional characterization of this process needs to be conducted. Several diseases implicate a role for CaMKII including Alzheimer's disease, epilepsy, Parkinson's disease and cell loss due to ischemia. Uncovering the role of this heterosynaptic phenomenon could elucidate further the dynamics of cell-wide CaMKII activation and provide possible avenues of discovery into the mechanisms of some of these disorders. PUBLIC HEALTH RELEVANCE: Calcium/calmodulin dependent kinase II (CaMKII) migrates to synapses throughout a neuron following stimulation of a small segment of dendrite. This locally-induced propagating synaptic (L-IPS) translocation of CaMKII requires further functional characterization to determine its role in neuron function. As CaMKII activation is involved in several physiological processes, ranging from myocardial contraction to synaptic plasticity, as well as neurological disorders like Alzheimer's, epilepsy and cell death due to stroke/ischemia, it is of interest to uncover why CaMKII signals activity from one local site to all the synapses of a neuron.