MECHANISMS OF TROPHIC MOLECULES DURING NEURONAL DEVELOPM

神经元发育过程中营养分子的机制

• 批准号: 2416286
• 负责人: Rae Nishi
• 金额: $ 27.35万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-04-01 至 2001-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2416286
• 关键词: Retroviridae; antisense nucleic acid; blocking antibody; bungarotoxins; cell death; choroid uvea; ciliary ganglion; developmental neurobiology; eye; eye pharmacology; gene induction /repression; growth factor receptors; inhibin; iris; laboratory mouse; laboratory rat; mixed tissue /cell culture; myotubes; neurons; neurotrophic factors; receptor expression; recombinant proteins; smooth muscle; somatostatin; uvea ciliary body; Retroviridae; antisense nucleic acid; blocking antibody; bungarotoxins; cell death; choroid uvea; ciliary ganglion; developmental neurobiology; eye; eye pharmacology; gene induction /repression; growth factor receptors; inhibin; iris; laboratory mouse; laboratory rat; mixed tissue /cell culture; myotubes; neurons; neurotrophic factors; receptor expression; recombinant proteins; smooth muscle; somatostatin; uvea ciliary body

The long term goal of this study is to understand the molecular and cellular mechanisms of target- neuron interactions during development. We will accomplish this goal by applying a variety of methods for manipulating gene and protein expression during chicken embryo development in vivo. These studies are expected to advance the understanding of the development and maintenance of the nervous system, providing insights into the processes of neural maintenance and neurodegenerative disease. Our studies will use the avian ciliary ganglion (CG) because of its simple neuronal composition, the accessibility of the neurons and their targets throughout development, and the coincidence of synapse formation, cell death, and transmitter expression within a well-defined window of development. The CG contains two populations of neurons: ciliary neurons that innervate iris and ciliary body and choroid neurons that innervate arterial smooth muscle in the choroid layer of the eye. We have identified three molecules by their actions on CG neurons in cell culture: growth promoting activity (GPA), which supports neuronal survival; activin A, which induces the expression of somatostatin; and follistatin, an inhibitor of activin. Our working model is that availability and levels of these three target-derived molecules regulate survival and neuropeptide phenotype of CG neurons. We will determine the validity of this model by testing the following hypotheses: l). Cell death in the CG during development is controlled by GPA expressed in the iris, ciliary body, and choroid layer; 2) Specific expression of somatostatin in choroid neurons of the CG is induced by activin expressed in the choroid layer, whereas follistatin blocks the effects of activin expressed in the iris/ciliary body. The specific aims are: l) to overexpress GPA in order to determine if CG neurons are rescued from cell death; 2) to use cell cultures of CG target tissues as well as Co-cultures to assay the effectiveness of GPA blocking agents; 3) to block GPA during embryonic development in vivo to determine if CG cell death is exacerbated; 4) to identify the mechanism by which alpha bungarotoxin increases CG neuron survival; 5) to quantify and characterize GPA receptor and receptor associated molecules during development; 6) to test the effectiveness of activin and follistatin blocking agents on co-cultures of CG neurons with choroid smooth muscle and iris/ciliary body; 7) to block to the action of activin in vivo to determine if somatostatin expression in choroid neurons is prevented; 8) to inject activin in the anterior chamber of the eye to induce somatostatin in ciliary neurons; 9) to block the effects of follistatin in the iris/ciliary body to determine if somatostatin is induced in ciliary neurons.

这项研究的长期目标是了解分子和 发育过程中靶神经元相互作用的细胞机制。我们 将通过应用多种方法来实现这一目标 在鸡胚胎发育过程中操纵基因和蛋白质表达 体内。期望这些研究能够提高对 神经系统的开发和维护，为 神经维持和神经退行性疾病的过程。我们的 研究将使用禽睫状神经节（CG），因为它的简单 神经元组成，神经元的可及性及其靶标 在整个发育中，以及突触形成的巧合，细胞 死亡和发射器表达在明确的窗口中 发展。 CG包含两个神经元群：纤毛神经元 支配虹膜和睫状体和脉络膜神经元，这些神经元 眼睛脉络膜中的动脉平滑肌。我们已经确定了 三个分子通过对细胞培养中CG神经元的作用的作用：生长 促进活动（GPA），支持神经元生存；激活素A， 诱导生长抑素的表达；和follistatin，an 激活素抑制剂。我们的工作模型是可用性和 这三个靶点分子调节生存和神经肽 CG神经元的表型。我们将通过 测试以下假设：L）。在CG中的细胞死亡 开发由虹膜，睫状体和 脉络层； 2）脉络膜神经元中生长抑素的特异性表达 CG的诱导素在脉络膜层中表达，而 Follistatin阻断虹膜/睫状体中表达的激活素的作用 身体。具体目的是：l）过表达GPA以确定 如果CG神经元从细胞死亡中救出； 2）使用CG的细胞培养物 靶组织以及共培养以测定GPA的有效性 阻止代理； 3）在体内阻止胚胎发育期间的GPA 确定CG细胞死亡是否加剧； 4）通过 哪种αBungarotoxin会增加CG神经元存活； 5）量化和 表征GPA受体和受体相关的分子 发展; 6）测试激活素和卵泡素的有效性 用脉络膜平滑肌阻止CG神经元共培养的剂 和虹膜/睫状体； 7）阻止活化蛋白在体内的作用 确定是否预防脉络膜神经元中的生长抑素表达； 8） 在眼前室内注入激活素以诱导 睫状神经元中的生长抑素； 9）阻止Follistatin在 虹膜/睫状体，确定睫状体中是否诱导生长抑素 神经元。