CALCIUM AND CALMODULIN DEPENDENT PROTEIN KINASE

钙和钙调蛋白依赖性蛋白激酶

• 批准号: 6393323
• 负责人: MARY B KENNEDY
• 金额: $ 35.08万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 1981
• 资助国家: 美国
• 起止时间: 1981-07-01 至 2003-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6393323
• 关键词: NMDA receptors; calmodulin dependent protein kinase; enzyme activity; enzyme induction /repression; genetically modified animals; guanine nucleotide binding protein; guanosinetriphosphatase activating protein; hippocampus; immunocytochemistry; laboratory mouse; long term potentiation; neural transmission; phosphoproteins; synapses; voltage /patch clamp

DESCRIPTION (Adapted from the applicant's abstract): Derangements in synaptic transmission are part of the pathology of several neurological and mental diseases including epilepsy, schizophrenia, depression and Alzheimer's disease. The investigator is studying the molecular mechanisms underlying regulation of synaptic transmission. The investigator proposes to study the spatial and temporal regulation at glutamatergic synapses of Ca2+/calmodulin-dependent protein kinase II (CaMKII) and one of its principal postsynaptic substrates, SynGAP. CaMKII is concentrated in glutamatergic neurons in the forebrain and is present in the postsynaptic density where it can be activated by Ca2+ influx through NMDA receptors. The message for alpha-CaMKII is abundant in dendrites in the hippocampus. The investigator has recently found that activation of NMDA receptors produces a long-lasting increase in activated autophosphorylated CaMKII and a rapid rise in CaMKII protein in dendrites. In the first aim, the investigator proposes to use quantitative immunocytochemistry and whole cell patch clamp recordings to study spatial and temporal control of synthesis and autophosphorylation of CaMKII in single neurons in hippocampal slices. The investigator will produce long-term potentiation or long-term depression in single neurons and determine how these and other stimulation paradigms affect the regulation CaMKII throughout the neuron. In the second aim, the investigator proposes to measure the degradation rate of CaMKII in hippocampal slices and determine whether neuronal stimulation alters degradation as well as synthesis of CaMKII in dendrites. SynGAP is a Ras GTPase-activating protein that is concentrated in the postsynaptic density along with CaMKII. The investigator recently found that active CaMKII phosphorylates and inactivates SynGAP's GTPase-activating activity. This inactivation could lead to an increase in active Ras, and thus to an increase in activation of the MAP kinase pathway. The investigator will study the spatial and temporal control of SynGAP by phosphorylation by CaMKII and its link to activation of the MAP kinase pathway by NMDA receptors. In the third aim the amino acid residues on SynGAP that inhibit its GAP activity upon their phosphorylation by CaMKII will be identified. The investigator will then raise phosphosite specific antibodies that recognize the phosphorylated, inhibited form of SynGAP. In the fourth aim, the investigator proposes to use the antibodies to study spatial and temporal regulation of phosphorylation of SynGAP and MAP kinase in hippocampal slices. In the fifth aim, the investigator proposes to create mutant mice lacking SynGAP. The investigator will test the effect of the SynGAP knockout on regulation of the MAP kinase pathway by NMDA receptors.

描述（改编自申请人的摘要）：突触中的紊乱 传播是一些神经和精神疾病病理学的一部分， 癫痫、精神分裂症、抑郁症和阿尔茨海默病等疾病。 研究人员正在研究调节的分子机制， 突触传递研究人员建议研究空间和 Ca ~（2+）/钙调素依赖性突触时间调节 蛋白激酶II（CaMKII）及其主要的突触后底物之一， SynGAP。CaMKII集中在前脑的多巴胺能神经元中， 存在于突触后致密物中，在那里它可以被Ca 2+内流激活 通过NMDA受体。α-CaMKII的信息在树突中丰富 在海马区。研究人员最近发现， 受体产生一个长期持续的增加激活自磷酸化 CaMKII和CaMKII蛋白在树突中的快速上升。在第一个目标中， 研究者建议使用定量免疫细胞化学和全细胞 膜片钳记录研究合成的空间和时间控制， 海马切片中单个神经元中CaMK II的自身磷酸化。的 研究者将产生长时程增强或长时程抑制 单个神经元，并确定这些和其他刺激模式如何影响 调节整个神经元的CaMKII。在第二个目标中， 研究者建议测量海马中CaMKII的降解速率， 切片，并确定神经元刺激是否改变降解以及 CaMKII在树突中的合成。 SynGAP是一种Ras GTP酶激活蛋白， 突触后密度沿着与CaMK Ⅱ。调查人员最近发现， 活性CaMKII磷酸化并灭活SynGAP的GTP酶激活 活动这种失活可能导致活性Ras的增加，因此 MAP激酶通路的激活增加。研究者将 研究通过CaMK II磷酸化对SynGAP的空间和时间控制 以及它与NMDA受体激活MAP激酶途径的联系。在 第三个目标是SynGAP上抑制其GAP活性的氨基酸残基， 将鉴定它们被CaMKII磷酸化。之后研究者要 产生识别磷酸化的磷酸化位点特异性抗体， SynGAP的抑制形式。在第四个目标中，研究人员建议使用 抗体研究磷酸化的空间和时间调节， 海马切片中的SynGAP和MAP激酶。在第五个目标中，调查员 提出创造缺乏SynGAP的突变小鼠。研究人员将测试 SynGAP基因敲除对NMDA调节MAP激酶通路的影响 受体。