GLUTAMATERGIC AND IP3 SIGNALING IN CNS AND PERIPHERY

肌醇磷脂信号系统是一种广泛存在的机制 用于传递神经递质、激素和生长的信号 影响细胞内过程的因素。此应用程序的重点是 与肌醇磷脂相关的两个新发现的研究 信号：(1)在高等动物物种中(豚鼠、兔、恒河猴 在人类神经母细胞瘤细胞中，锂含量增加 乙酰胆碱刺激的第二信使INS(1，4，5)P3。(2)在 在没有添加激动剂的情况下，锂会增加胞外谷氨酸 大脑皮层切片，通过激活NMDA受体， 提高INS(1，4，5)P3。该计划的目标是了解Li+如何 升高激动剂刺激的Ins(1，4，5)P3，Li+如何升高细胞外 谷氨酸，以及它的生理相关性和后果是什么 细胞信号。Li+对突触谷氨酸调节的影响 通过对切片、胶质细胞和细胞摄取谷氨酸的研究来探讨 突触体，PKC调节的胞吐作用(DAG水平和 选择性蛋白激酶C抑制剂)和谷氨酸释放的内源性抑制剂。 Li+对突触体释放谷氨酸的影响 突触前模型)将被确定。Li+链接之间的联系 谷氨酸和INS(1，4，5)P3的变化。将使用抑制剂进行探索 谷氨酸释放和NMDA受体激动剂。一般假设 Li+升高任何激活受体产生的Ins(1，4，5)P3 将测试通过G蛋白或通过增加的钙离子内流连接到PLC 通过测定Li+对大脑皮层脑片Ins(1，4，5)P3的影响 在大量合适的神经递质存在的情况下。在……里面 与谷氨酸灵长类中心的Terasawa博士合作 未麻醉和非应激恒河猴大脑皮层的释放 猴子将在活体内通过推拉灌流和 在存在和不存在锂的情况下的微透析。初步准备 对这一模型的研究非常有前途。其他产品的发布 神经递质也将被研究。锂对人体健康的影响 Ins(1，4，5)P3和Ins(1，3，4，5)P4在皮质下结构中的蓄积 将进一步研究，主要是在海马体。的影响 急性和慢性锂对人神经母细胞瘤各参数的影响 细胞，即Ins(1，4，5)P3和Ins(1，3，4，5)P4(慢性)，数量和 INS(1，4，5)P3受体DAG和胞内钙离子的亲和力 接受检查。这些效应也将在分化的细胞中进行研究， 它们更接近于成年神经元。其分子机制的研究进展 锂升高的Ins(1，4，5)P3在胰腺和Will中的研究效果最好 有待进一步调查；即，海拔是否因PLC增加而引起 Ins(1，4，5)P3 5-磷酸酶的活性或抑制，还是两者兼而有之？最后， NMDA-和INS(1，4，5)P3-受体可能下调 将对长期使用锂治疗的豚鼠进行研究。初步 数据显示，小鼠的INS(L，4，5)P3受体表达下调。这些 研究可能有助于阐明该疗法的分子机制。 锂的作用，从长远来看，这可能会带来医疗利益。

The phosphoinositide signaling system is a widespread mechanism for transducing signals from neurotransmitters, hormones, and growth factors to intracellular processes. The focus of this application is an investigation of two recent discoveries related to phosphoinositide signaling: (1) In higher animal species (guinea pig, rabbit, rhesus monkey) and in human neuroblastoma cells, lithium increases acetylcholine-stimulated second messenger Ins(1,4,5)P3. (2) in the absence of added agonist, lithium raises extracellular glutamate in cerebrocortical slices, which, via activation of the NMDA receptor, raises Ins(1,4,5)P3. The objectives of this proposal are to learn how Li+ elevates agonist-stimulated ins(1,4,5)P3, how Li+ raises extracellular glutamate, and what are the physiological relevance and consequences for cell signaling. The effect of Li+ on synaptiC glutamate regulation will be probed by studies of uptake of glutamate by slices, glia and synaptosomes, PKC-modulated exocytosis (levels of DAG and effect of selective PKC inhibitors) and endogenous inhibitors of glutamate release. The effect of Li+ on glutamate release from synaptosomes (an isolated presynaptic model) will be determined. The connection between Li+ linked changes in glutamate and Ins(1,4,5)P3. will be explored using inhibitors of glutamate release and NMDA receptor agonists. The general hypothesis that Li+ elevates Ins(1,4,5)P3 generated by any activated receptor coupled to PLC via G proteins or via increased Ca2+ influx will be tested by measuring the effect of Li+ on Ins(1,4,5)P3 in cerebrocortical slices in the presence of a wide selection of appropriate neurotransmitters. In collaboration with Dr. Ei Terasawa at the Primate Center, glutamate release in cerebral cortex of unanesthetized and unstressed rhesus monkeys will be measured in vivo by push-pull perfusion and by microdialysis in the presence and absence of lithium. preliminary studies in this model are very promising. The release of other neurotransmitters will also be studied. The effects of lithium on Ins(1,4,5)P3 and Ins(1,3,4,5)P4 accumulation in subcortical structures will be further explored, primarily in the hippocampus. The effects of acute and chronic lithium on various parameters in human neuroblastoma cells, i.e., Ins(1,4,5)P3 and Ins(1,3,4,5)P4 (chronic), number and affinities of the Ins(1,4,5)P3 receptor, DAG, and cytosolic Ca2+, will be examined. These effects will also be studied in differentiated cells, which more closely resemble adult neurons. The molecular mechanism of lithium-elevated Ins (1,4,5)P3 can best be studied in pancreas and will be further investigated; i.e., is the elevation due to increased PLC activity or inhibition of Ins(1,4,5)P3 5-phosphatase, or both? Lastly, possible down-regulation of the NMDA- and Ins(1,4,5)P3-receptors in guinea pigs chronically treated with lithium will be studied. Preliminary data suggest down-regulation of the Ins(l,4,5)P3 receptor in mice. These studies may throw light on the molecular mechanism of the therapeutic action of lithium, which in the long run may lead to medical benefit.