NEURONAL CA++ SEQUESTERING COMPARTMENTS--PROTECTIVE ROLE

这项拨款的主要研究重点是深入了解 细胞内钙离子隔离的结构与功能 神经元的隔间。这些隔间很有价值 因为最近积累的证据表明它们起到了保护作用 暴露于谷氨酸的神经元，谷氨酸是主要的兴奋性神经递质 在中枢神经系统和“兴奋性毒性”的原因。兴奋性毒性是一种 中风和创伤中神经元损伤的原因以及可能的 老年性神经退行性疾病中神经元损伤的介体， 例如亨廷顿氏症、肌萎缩侧索硬化症。 具体目标是：1.定位CaATPase的特定亚型 和钙网织蛋白2，由神经元钙离子隔离成分 大脑、培养的神经元和神经母细胞瘤细胞的隔室。2. 探讨CAATPase和CAATPase的生物合成途径(S) 紧随其后的是钙网硬蛋白，到达钙离子隔离隔室。3.至 研究细胞内钙离子升高药对大鼠心肌细胞内钙离子浓度的影响。 神经母细胞瘤细胞CaATPase和钙网硬蛋白的蛋白水平 培养的神经元。4.用cDNA转染人神经母细胞瘤细胞 CaATPase和钙网硬蛋白来确定这些蛋白是否具有保护性 对抗细胞内钙离子升高的毒性效应和5.使用 用细胞内染料直接测定神经元和 神经母细胞瘤细胞。我们将使用生化、形态和 分子技术在这些研究中的应用。

The principle research focus of this grant is to gain insight into the structure and function of the intracellular Ca2+ sequestering compartments of neurons. These compartments are of great interest because recent evidence has accumulated that they play a protective role in neurons exposed to glutamate, the major excitatory neurotransmitter in the CNS and the cause of "excitotoxicity". Excitotoxicity is a major cause of neuronal damage in stroke and trauma as well as a likely mediator of neuronal injury in age related neurodegenerative diseases, e.g. Huntington's, ALS. The specific aims are: 1. To localize the specific isoforms of CaATPase and calreticulin two, by components of neuronal Ca2+ sequestering compartments in brain and cultured neurons and neuroblastoma cells. 2. To investigate the biosynthetic route(s) by which the CaATPase and calreticulin follow to reach the Ca2+ sequestering compartments. 3. To study the effects of agents which raise cytosolic Ca2+ on the mRNA and protein level of CaATPase and calreticulin using neuroblastoma cells and cultured neurons. 4. To transfect neuroblastoma cells with cDNAs for CaATPase and calreticulin to determine if these proteins are protective against toxic effects of elevated cytosolic Ca2+ and 5. To employ intracellular dyes to directly measure cytosolic Ca2+ in neurons and neuroblastoma cells. We will employ biochemical, morphological and molecular techniques in these investigations.