DEVELOPMENTAL EXPRESSION OF CAM KINASE II ISOFORMS

谷氨酸能突触和突触神经传递的修饰 依赖于钙离子的第二信使系统的激活有助于 学习和记忆的过程以及观察到的神经元功能障碍 中风和脑缺血、局灶性癫痫和阿尔茨海默病。 此外，这些系统发挥着重要的作用，但人们对此知之甚少。 在神经元发育、再生和有丝分裂中。多功能机型 钙离子和钙调蛋白依赖的蛋白激酶(CaM KII)及其底物 对于这个激酶，微管相关蛋白MAP-2已经被 与这些过程有牵连。在目前的提案中，初级神经元 培养和转化的细胞将被用于潜在地检查 调控基因表达和靶向的重要生理因子 神经元发育过程中的CaM Kii亚型和MAP-2变异体。 主要表达NMDA或非NMDA受体的小脑颗粒细胞- 通道将被用来检验慢性激活的假说 NMDA受体诱导CaM亚型“类前脑”的表达 基伊。初步数据显示MAP-2蛋白表达 正常，在这两个表型上没有区别，而Cam Kii 亚单位mRNA的表达发生改变。在神经元成熟过程中， MAP-2的磷酸化状态及其表达变体在发育中的作用 调节类似于完整的大脑。第二系列实验将 检测转化细胞中MAP-2和CaM KII表达的调控因素 一种新的MAP-2信使核糖核酸调控模式的神经元培养 观察到的。这些实验最终将提供对 钙依赖性MAP-2磷酸化的复杂关系 蛋白激酶和突触功能。一种分子的组合 将使用生物、免疫化学和生化方法来 回答以下具体问题：小脑颗粒细胞 主要表达NMDA谷氨酸受体亚型表达a Cam Kii的“类似前脑”的异构体？做小脑颗粒细胞 主要表达非NMDA谷氨酸受体亚型表达a Cam Kii的“小脑样”亚型？有什么不同吗？ CaM KII在小脑颗粒细胞中的亚细胞分布 NMDA与非NMDA受体？MAP-2mRNA是如何被主动调控的 分裂NG108细胞？

Modification of synaptic neurotransmission at glutamatergic synapses and activation of Ca+2- dependent second messenger systems contribute to the processes of learning and memory, and the neuronal dysfunction observed following stroke and ischemia, focal epilepsies, and Alzheimer's disease. In addition, these systems play an important, yet poorly understood, role in neuronal development, regeneration, and mitosis. A multifunctional type II Ca+2 and calmodulin-dependent protein kinase (CaM KII) and a substrate for this kinase, the microtubule-associated protein MAP-2 have been implicated in these processes. In the present proposal, Primary neuronal cultures and transformed cells will be used to examine potentially important physiological factors that regulate expression and targeting of CaM KII isoforms and MAP-2 variants during neuronal development. Cerebellar granule cells expressing primarily NMDA or non-NMDA receptor- channels will be used to test the hypothesis that chronic activation of NMDA receptors induces expression of a "forebrain-like", isoform of CaM KII. Preliminary data indicate that MAP-2 protein expression appears normal and does not differ in these two phenotypes, whereas CaM KII subunit mRNA expression is altered. During neuronal maturation, the phosphorylation state and expressed variant of MAP-2 are developmentally regulated similar to intact brain. A second series of experiments will examine factors regulating expression of MAP-2 and CaM KII in transformed neuronal cultures where a novel mode of MAP-2 mRNA regulation has been observed. These experiments will ultimately provide insights into the complex relationship between phosphorylation of MAP-2 by Ca+2-dependent protein kinases and synaptic function. A combination of molecular biological, immunochemical and biochemical approaches will be used to answer the following specific questions: Do cerebellar granule cells expressing primarily the NMDA glutamate receptor subtype express a "forebrain-like" isoform of CaM KII? Do cerebellar granule cells expressing primarily the non-NMDA glutamate receptor subtype express a "cerebellar-like" isoform of CaM KII? Is there a difference in the subcellular distribution of CaM KII in cerebellar granule cells expressing NMDA versus non-NMDA receptors? How is MAP-2 mRNA regulated in actively dividing NG108 cells?