Cellular Basis of Circadian Rhythms in Mammals

描述（由申请人提供）：哺乳动物视交叉上核（SCN）是行为和生理中日常节律所需的昼夜节律起搏器。在体内和体外，SCN内的多个昼夜节律振荡器彼此同步以维持近24小时的节律。目前尚不清楚这种节律性是否是SCN神经元的特定群体所固有的，或者是什么机制耦合了它们的昼夜节律。此外，最近的分子证据表明，其他组织也可以作为昼夜节律振荡器，但其节律性的基础和在行为中的作用尚不清楚。拟议的研究通过利用长时间记录技术（多电极阵列和基因活性的生物发光报告）以及具有与昼夜节律计时相关的基因突变的小鼠和仓鼠的独特特性，直接解决了这些问题。第一个特定目标测试的假设，个别SCN神经元是自主的，昼夜节律的起搏器和起搏神经元是一个小的SCN神经元的子集。该策略是表征完全分离的SCN神经元的节律能力，然后，固定后，其神经化学物质的内容。 具体目标2测试SCN神经元通过血管活性肠多肽而不是快速突触通信彼此同步的假设。使用特定的拮抗剂，激动剂和基因敲除，这一目标补充了第一个目标，即确定起搏器和协调活动所需的信号。最近在许多哺乳动物组织中发现了假定的昼夜节律振荡器，这导致了昼夜节律系统是分层组织的假设。具体目标3和4将确定主嗅球（OB）昼夜节律的功能和分子基础，这是一个与SCN进行比较的昼夜节律振荡器模型。使用行为，解剖学和生理学测定的节奏在OB，这些目标将建立一个体内的作用，在OB的时钟。此外，他们将直接测试这样一个假设，即SCN和OB之间至少有一个参与昼夜节律计时的基因不同。这些实验将首次确定两个大脑区域的昼夜节律起搏器，协调其整体节奏的机制，以及它们在行为中发挥的独特作用。

DESCRIPTION (provided by applicant): The mammalian suprachiasmatic nucleus (SCN) is a circadian pacemaker required for daily rhythms in behavior and physiology. In vivo and in vitro, multiple circadian oscillators within the SCN synchronize to each other to sustain near 24-h rhythms. It is presently unclear if this rhythmicity is intrinsic to a specialized population of SCN neurons or what mechanisms couple their circadian rhythms. In addition, recent molecular evidence suggests that other tissues can act as circadian oscillators, but the bases for their rhythmicity and roles in behavior are unknown. The proposed studies directly address these issues by taking advantage of long-duration recording technologies--multielectrode arrays and bioluminescent reporters of gene activity--and the unique properties of mice and hamsters with mutations in genes involved in circadian timekeeping. The first Specific Aim tests the hypotheses that individual SCN neurons are autonomous, circadian pacemakers and that the pacemaking neurons are a small subset of SCN neurons. The strategy is to characterize the rhythmic ability of fully isolated SCN neurons and then, after fixation, their neurochemical content. Specific Aim 2 tests the hypothesis that SCN neurons synchronize to each other via vasoactive intestinal polypeptide and not fast synaptic communication. Using specific antagonists, agonists, and genetic knockouts, this aim complements the first aim in identifying pacemakers and the signals required for their coordinated activity. The recent discoveries of putative circadian oscillators in many mammalian tissues have led to the hypothesis that the circadian system is hierarchically organized. Specific Aims 3 and 4 will determine the function and molecular basis for circadian rhythms in the main olfactory bulb (OB), a model circadian oscillator to be compared to the SCN. Using behavioral, anatomical and physiological assays for rhythmicity in the OB, these aims will establish an in vivo role for the clock in the OB. Furthermore, they will directly test the hypothesis that at least one of the genes involved in circadian timekeeping differ between the SCN and OB. These experiments will, for the first time, identify circadian pacemakers in two brain areas, the mechanisms that coordinate their ensemble rhythms, and the distinct roles they play in behavior.