CELLULAR MECHANISMS OF NEUROENDOCRINE HORMONE SECRETION

描述(申请人摘要)：释放递质和激素 神经元在信息传递中起着至关重要的作用 邻近的细胞和远处的组织。揭示了控制 神经分泌对于我们理解广泛的 当发生这种情况时，会发生各种生理系统和疾病过程 传输中断。我们的长期目标是了解神经控制 多肽分泌及其对生理和行为的影响。的广泛目标 这项提议是为了揭示和研究潜在的细胞和分子事件。 调节神经激素(产卵激素，ELH)的合成和分泌 以海兔袋状细胞神经元为模型系统。我们现在能够 监测BAG细胞神经元膜兴奋性和ELH分泌 自由活动的动物，来自切除的神经组织中的袋状细胞神经元 在体外保持，并从单袋细胞培养神经元。这给了我们 一种相对独特的能力，可以在解决分子和 细胞生理问题以及解决系统和行为问题的问题 使用单一的神经内分泌系统。这有三个具体目标 求婚。第一个目标是确定细胞内钙的作用。 储存在调节ELH的分泌模式中。我们将使用 电生理学、钙成像和激素分泌的RIA测量 测定细胞内钙离子对电刺激的反应模式 后放电、细胞内钙调控对ELH的影响 分泌，以及控制ELH分泌的细胞内钙的来源 对后放电的反应。第二个目标是研究蛋白激酶A 蛋白激酶C(PKC)和钙调素激酶(CaM)在BAG细胞神经元中表达。我们计划 使用传统的放射性标记方法监测激酶活性，如 以及新开发的光学成像技术。此外，我们还将 研究CaM激酶II在调节ELH分泌中的作用。第三 具体目的是了解细胞和分子事件，这些事件介导了 后放电对ELH生物合成的刺激作用。我们会调查的 后放电对翻译路径上特定步骤的影响， 钙、PKA和PKC信号通路在调控中的作用 导致ELH生物合成改变的特定翻译事件。这个 这项工作的结果应该使我们能够开发一个详细的细胞模型 将膜兴奋与神经激素分泌和 神经激素的合成。

DESCRIPTION (applicant's abstract): Release of transmitters and hormones from neurons plays a critical role in transmission of information between neighboring cells and distant tissues. Revealing mechanisms that control neurosecretion is vital to our understanding of both homeostasis of a wide variety of physiological systems and disease processes that occur when this transmission breaks down. Our long-term goal is to understand neural control of peptide secretion and its impact on physiology and behavior. The broad aim of this proposal is to reveal and study underlying cellular and molecular events regulating synthesis and secretion of a neurohormone (egg laying hormone, ELH) using the bag cell neurons of Aplysia as a model system. We are now able to monitor membrane excitability and ELH secretion from bag cell neurons of freely-behaving animals, from bag cell neurons in excised neural tissue maintained in vitro, and from single bag cell neurons in culture. This gives us a relatively unique ability to move between studies addressing molecular and cell physiological issues and those addressing systems and behavioral questions using a single neuroendocrine system. There are three specific aims of this proposal. The first aim is to determine the role of calcium from intracellular stores in regulating the pattern of ELH secretion. We will use electrophysiology, calcium imaging, and RIA measurement of hormone secretion to determine the pattern of intracellular calcium in response to electrical afterdischarge, the effects of manipulating intracellular calcium on ELH secretion, and the source of intracellular calcium controlling ELH secretion in response to afterdischarge. The second aim is to investigate protein kinase A (PKA), protein kinase C (PKC), and CaM kinase in bag cell neurons. We plan to monitor kinase activities using both a traditional radiolabeling approach, as well as newly developed optical imaging techniques. In addition, we will investigate the role of CaM kinase II in regulating ELH secretion. The third specific aim is to understand cellular and molecular events that mediate the stimulatory effects of afterdischarge on ELH biosynthesis. We will investigate the effects of afterdischarge on specific steps along the translation pathway, and the roles of the calcium, PKA and PKC signaling pathways in regulating specific translational events leading to alterations in ELH biosynthesis. The outcome of this work should enable us to develop a detailed model of cellular mechanisms that link membrane excitation to neurohormone secretion and neurohormone synthesis.