Prokineticin 2 and Suprachiasmatic Circadian Output

描述（由申请人提供）：生理和行为的组织与重复的日常环境条件是一种适应，基本上发生在所有生物体。昼夜节律由三个部分调节：昼夜节律起搏器、输入机制和输出机制。在哺乳动物中，驱动昼夜节律的主起搏点位于下丘脑前部的视交叉上核（SCN）。环境光暗周期将SCN时钟带入24小时。SCN神经元的同步导致协调的昼夜节律输出，调节表达的节律。SCN内的分子钟机制的清晰视图已经出现。SCN生物钟的分子时钟由具有正和负元件的自动调节转录和翻译反馈环组成。类似地，光输入如何重置SCN时钟以与环境亮/暗周期同步的信号路径也正在出现。与SCN起搏器时钟和输入途径的分子机制相反，SCN昼夜节律起搏器发送定时信息以控制生理和行为节律的输出机制相对知之甚少。前动力蛋白2（PK 2），一种富含半胱氨酸的蛋白质，最近被证明是一种SCN输出分子，可传递昼夜运动节律。PK 2满足SCN生物钟的真实输出分子的预期标准：1）PK 2是分泌分子; 2）PK 2的转录受核心时钟基因调控，并且PK 2 mRNA在SCN中以高幅度振荡; 3）PK 2的产生响应于光夹带; 4）PK 2的受体在初级SCN输出靶区域中表达;和5）当内源性PK 2水平较低时，在主观夜间脑室内（ICV）施用PK 2，抑制高夜间轮跑活动。我们建议进一步研究PK 2信号在SCN生物钟输出机制中的作用。具体而言，将通过定量免疫细胞化学和/或放射免疫测定法研究细胞体、SCN神经元末端区域和脑脊液中PK 2蛋白的节律。将研究PK 2是否是介导SCN昼夜节律钟和光掩蔽输出的共同信号。PK 2的节奏输出从SCN响应突然转变的光/暗周期也将被调查。此外，将通过在大鼠中急性和慢性输注PK 2和PK 2拮抗剂来研究PK 2对SCN生物钟控制的运动和睡眠/觉醒节律的影响。此外，PK 2基因将在小鼠中被破坏，并将检查其对SCN控制的昼夜行为节律以及核心SCN昼夜循环的影响。最后，将通过遗传方法研究SCN PK 2输出途径。这些拟议的研究将有助于我们进一步了解PK 2信号转导在介导SCN昼夜节律钟输出定时信息中的机制，并可能对未来治疗一些昼夜节律紊乱如时差综合征、轮班工作综合征和慢性失眠症产生重大影响

DESCRIPTION (provided by applicant): Organization of physiology and behavior with recurring daily environmental conditions is an adaptation that occurs in essentially all living organisms. Circadian rhythms are regulated by three components: the circadian pacemaker, an input mechanism and an output mechanism. In mammals, the master pacemaker driving circadian rhythms resides in the suprachiasmatic nuclei (SCN) of the anterior hypothalamus. Environmental light-dark cycles entrain the SCN clock to the 24-hr day. Synchronization of SCN neurons leads to coordinated circadian outputs that regulate expressed rhythms. A clear view of molecular clock mechanisms within the SCN has emerged. The molecular clockwork of the SCN circadian clock consists of auto-regulatory transcriptional and translational feedback loops that have both positive and negative elements. Similarly, the signal pathway of how light input resets SCN clock to synchronize with the environmental light/dark cycle is also emerging. In contrast to molecular mechanisms of SCN pacemaker clockwork and input pathway, relatively little is known about the output mechanism by which the SCN circadian pacemaker sends timing information to control physiological and behavioral rhythms. Prokineticin 2 (PK2), a cysteine-rich protein, has recently been shown as a SCN output molecule that transmits the circadian locomotor rhythm. PK2 fulfill the criteria expected for a bona fide output molecule from SCN circadian clock: 1) PK2 is a secreted molecule; 2) The transcription of PK2 is regulated by core clock genes, and PK2 mRNA oscillates in the SCN with high magnitude; 3) The production of PK2 is responsive to light entrainment; 4) Receptor for PK2 is expressed in primary SCN output target areas; and 5) Intracerebroventricular (ICV) administration of PK2 at subjective night, when endogenous PK2 levels are low, suppressed high nocturnal wheel-running activity. We propose to further investigate the role of PK2 signaling in the output mechanism of the SCN circadian clock. Specifically, the rhythms of PK2 protein in the cell bodies, terminal areas of SCN neurons and cerebral spinal fluid will be investigated by quantitative immunocytochemistry and/or radioimmunoassay. Whether PK2 is the common signal that mediates the output of SCN circadian clock and light masking will be investigated. How the PK2 rhythmic output from the SCN responds to abrupt shifts of light/dark cycle will also be investigated. Moreover, the effects of PK2 on SCN circadian clock-controlled locomotor and sleep/wake rhythms will be investigated by acute and chronic infusion of PK2 and PK2 antagonist in rats. Furthermore, the PK2 gene will be disrupted in mice and its effect on SCN-controlled circadian behavioral rhythms as well as core SCN circadian loops will be examined. Finally, the SCN PK2 output pathway will be investigated by a genetic approach. These proposed studies should help us gain further insight into the mechanism of PK2 signaling in mediating the output of timing information from the SCN circadian crock, and could have a major impact on the future treatment of a number of circadian disorders such as jet-lag, shift work syndrome, and chronic insomnia