Central and Peripheral Actions of Nitric Oxide

一氧化氮的中枢和外周作用

• 批准号: 0342330
• 负责人: Barry Trimmer
• 金额: $ 34.33万
• 依托单位: Tufts University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-03-01 至 2008-02-29
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0342330&HistoricalAwards=false
• 关键词: Central; Peripheral; Actions; Nitric; Oxide

Nitric oxide (NO) is a small, highly reactive molecule produced in both plants and animals. It is used to protect tissues from infections and acts as a signaling molecule in the vasculature of vertebrates and the central nervous system (CNS) of most species. Because NO can dissolve in both cell membranes and the fluid around cells it is thought to spread widely from where it is produced. However, the range of NO signaling has not been rigorously defined and may vary between tissues and species. In the CNS an effective range of 200 microns would enable NO to control hundreds of neurons, but in peripheral tissues such as muscles this range could limit NO to a very local or even intracellular role. Hence, the immense differences in size, metabolic function and cellular architecture of NO target tissues raise important questions about the mechanisms of NO signaling at different locations. The proposed experiments take advantage of a well-characterized NO signaling system in an insect model system (larval Manduca sexta) to establish the functional roles of NO at central and peripheral sites. A key advantage of studies in this insect is that signaling can be manipulated in the intact, freely moving larva, or in tissues dissected from the insect and maintained alive for several hours under defined conditions. Pharmacological tools are available to block or supplement NO production and to block or increase its effectiveness. It is now also possible to alter the synthesis of key enzymes at the molecular level in intact animals using injections of double-stranded RNA. This treatment interferes with the expression of specific genes in the fully developed animal or in selected parts of the CNS. The results of preliminary physiological experiments, together with the anatomical distribution of enzymes that make or respond to NO (nitric oxide synthase, NOS; and soluble guanylyl cyclase, sGC; respectively), suggest that NO is involved in the control of feeding (chewing movements and foregut activity), the regulation of normal body wall tension and the direct cellular immune response. The neural and muscular activity will be recorded using electrodes and pressure sensors implanted in normal larvae and in larvae that are experimentally deficient in NOS or sGC. These actions of NO on motor patterns and neuromuscular physiology will also be examined pharmacologically in isolated tissues. The role of NO in the insect immune response will be tested by implanting fluorescent micro spheres into the body cavity and assessing the degree of encapsulation in normal and NO-deficient larvae. It is expected that feeding (and growth), gut movements and activity of body wall sensory neurons will all be strongly modulated by NO. The control of body turgor, motor coordination, and abiotic encapsulation are also potentially very important sites of action.The long-term goal of the proposed activity is to better understand how NO carries out its diverse functions. In particular, the results will help to define the specializations and limitations of NO signaling in very different tissue environments. Because NO is involved in the locomotion and feeding behavior of this herbivorous insect, the results could also impact the development of pest-specific antifeedants for crop protection.

一氧化氮(NO)是一种小的、高活性的分子，在植物和动物中都能产生。它被用来保护组织免受感染，并在脊椎动物的血管系统和大多数物种的中枢神经系统(CNS)中充当信号分子。因为NO可以溶解在细胞膜和细胞周围的液体中，所以它被认为是从它产生的地方广泛传播的。然而，NO信号的范围还没有得到严格的定义，而且可能在组织和物种之间有所不同。在中枢神经系统，200微米的有效范围将使NO能够控制数百个神经元，但在肌肉等外围组织中，这个范围可能会将NO限制在非常局部甚至细胞内的作用。因此，NO靶组织在大小、代谢功能和细胞结构上的巨大差异引发了关于不同位置NO信号转导机制的重要问题。这项实验利用昆虫模型系统(幼虫)中的NO信号系统来确定NO在中枢和外周位置的功能作用。对这种昆虫进行研究的一个关键优势是，信号可以在完整的、自由移动的幼虫中操纵，也可以在从昆虫身上解剖并在特定条件下存活数小时的组织中操纵。有药理工具可用来阻断或补充NO的产生，并阻断或增加其有效性。现在，通过注射双链RNA，也可以在分子水平上改变完整动物体内关键酶的合成。这种治疗会干扰发育完全的动物或中枢神经系统特定部分的特定基因的表达。初步的生理实验结果，以及产生或响应NO的酶的解剖分布(分别为一氧化氮合酶和可溶性鸟苷环酶，sGC)，表明NO参与控制摄食(咀嚼运动和前肠活动)、调节正常体壁张力和直接细胞免疫反应。神经和肌肉活动将使用植入正常幼虫和实验上缺乏一氧化氮合酶或sGC的幼虫的电极和压力传感器来记录。NO对运动模式和神经肌肉生理学的这些作用也将在分离的组织中进行药理学研究。NO在昆虫免疫反应中的作用将通过将荧光微球植入体腔并评估正常幼虫和NO缺乏幼虫的包裹程度来测试。预计摄食(和生长)、肠道运动和体壁感觉神经元的活动都将受到NO的强烈调控。控制身体膨胀、运动协调和非生物包裹也是潜在的非常重要的作用部位。拟议活动的长期目标是更好地了解NO如何执行其不同的功能。特别是，这些结果将有助于确定NO信号在非常不同的组织环境中的专门化和局限性。由于NO参与了这种草食性昆虫的运动和取食行为，因此这一结果也可能影响害虫特异性拒食剂的开发，以保护作物。