Circuitry and Modulation of the Circardian Clock Network of D. melanogaster

在动物中，睡眠/活动节律所必需的计时发生在位于离散的 中枢神经系统(CNS)区域。值得注意的是，单个神经元生物钟可以维持 在没有来自环境或其他细胞的时间提示的情况下，分子和生理节律。尽管 这种细胞自主的计时，日常行为节律的编排依赖于时钟网络 神经元。我试图了解这些网络的电路属性，并检查确定 神经元在昼夜节律的控制中发挥作用。在我研究的Roo阶段！将使用解剖学上的， 遗传和实时成像技术1是在我的K99资助阶段开发的，目的是发现时间是如何保持的 以及它是如何被用来协调果蝇日常和季节性行为的变化 黑猩猩。此外，在活体成像、苍蝇遗传学和光控制方面的新技术 将开发神经元信号来研究飞行中生物钟网络的电路特性 大脑。为了了解神经元时钟网络的组织，以及 神经系统中的神经调节信号，我提出了以下特殊目标：1)发育 同时操作和观察神经元信号的实时成像方法 结合cAMP和Ca2-f，H基因编码传感器对细胞兴奋性的光遗传控制 神经钟网络的多肽/胺调节剂的识别。Ill)身份识别和 利用活体成像和靶向遗传技术表征时钟神经元输出的非时钟目标 操作方法，IV)对这些神经元内GPCR/cAMP/钙-t信号的遗传解剖。这个 由我的K99拨款资助的工作支持拟议研究的所有方面的可行性，以及 Roo提供的资源将使我能够在我的研究中使用先进的成像技术 神经元时钟网络。此外，这里提出的工作将涉及以下几个基本方面 神经生物学，为研究神经元回路和动物控制创造新的方法 行为。

In animals the timekeeping necessary for sleep/activity rhythms takes place within neurons located in discrete regions of the central nervous system (CNS). Remarkably, individual neuronal circadian clocks can maintain molecular and physiological rhythms in the absence of time-cues from the environment or other cells. Despite such cell-autonomous timekeeping, the orchestration of daily behavioral rhytiims depends on networks of clock neurons. I seek to understand the circuit properties of these networks and to examine the roles that identified neurons play in the control of circadian rhythms. During the ROO phase of my research! will use the anatomical, genetic, and live-imaging techniques 1 developed during the K99 phase of my grant to discover how time is kept within the brain and how it is used to orchestrate daily and seasonal changes in behavior in the fly Drosophila melanogaster. Furthermore, newly developed technologies in live imaging, fly genetics, and light-control of neuronal signaling will be developed to investigate the circuit properties of the circadian clock network in the fly brain. To understand the organization of the neuronal clock network, and fundamental questions of neuromodulatory signaling in the nervous system, I propose the following speciflc aims: 1) The development of live Imaging methods for the simultaneous manipulation and observation of neuronal signaling using optogenetic control of cell excitability in conjunction with genetically encoded sensors for cAMP and Ca2-f, H) The identiflcation of peptide/amine modulators of the neuronal clock network. Ill) The identiflcation and characterization of non-clock targets of clock neuron output using live imaging and targeted genetic manipulation approaches, IV) The genetic dissection of GPCR/cAMP/Ca2-t- signaling within these neurons. The work supported by my K99 grant supports the feasibility of all aspects of the proposed research and the resources made possible by the ROO will allow me to employ advanced imaging technologies in my study of the neuronal clock network. Furthermore, the work proposed here will address fundamental aspects of neurobiology and create new methodologies for the investigation of neuronal circuitry and the control of animal behavior.