PHYSIOLOGICAL DISSECTION OF THE SCN

描述(申请人提供)：视交叉上核(SCN)是大脑时钟的轨迹。在细胞水平上，节律性是通过调节各种时钟基因的转录/翻译的细胞负反馈循环而产生的。虽然我们对基于细胞的振荡器的分子机制的了解在最近几年有了很大的进步，但这些振荡器在SCN组织水平上的组织还不太发达。一方面，有人提出所有的SCN细胞都是昼夜节律振荡器。另一方面，我们对仓鼠SCN的研究表明，基于周期mRNA(PER1，PER2，PER3)的表达，SCN内存在不同的功能区块；一个包含节律振荡器，另一个包含昼夜节律门控、光响应、非振荡细胞。该组织建议，振荡和夹带信息可以在SCN内的不同站点进行整合。这种观点将改写我们对相移、夹带、后效等现象的理解。我们建议利用小鼠基因组解码带来的进展，来解决SCN组织的问题。在基线研究中，我们描述了一个特征良好的小鼠品系的昼夜节律和昼夜节律，以及对相位推进和延迟光脉冲的反应(目标I)。初步研究表明，小鼠的SCN，就像仓鼠的SCN一样，由两种截然不同的细胞类型组成，基于它们的光反应和振荡特性。为了了解这个昼夜节律调控系统的组织结构，我们将使用带有GFP记者的转基因动物，这些转基因动物是SCN(目标II和III)的节律(PER1：：GFP)和光诱导(Calbindin：：GFP)间隔的标记。最后，我们将使用钙成像和电记录和刺激来表征SCN的微电路，以了解SCN的网络特性，使用GFP来表征已识别的(PER1：：GFP和Calb：：GFP)SCN细胞类型(目标IV)的反应。

DESCRIPTION (provided by applicant): The suprachiasmatic nuclei (SCN) are the locus of the brain clock. At the cellular level, rhythmicity is produced by regulation of a cell-based negative feedback loop of transcription/translation of various clock genes. While our understanding of the molecular mechanisms of cell based oscillators has advanced tremendously in recent years, the organization of these oscillators at the level of SCN tissue is less well developed. On the one hand, it has been proposed that all SCN cells are circadian oscillators. On the other hand, our work on hamster SCN, based on Period mRNA (Per1, Per2, Per3) expression, suggests that there are functionally distinct compartments within the SCN; one contains rhythmic oscillators and the other contains circadian-gated, light responsive, non-oscillating cells. This organization suggests that oscillating and entraining information might be integrated at different sites within the SCN. Such a view would rewrite our understanding of such phenomena as phase shifting, entrainment, after-effects, etc.We propose to take advantage of the advances presented by the decoding of the mouse genome, to address questions of SCN organization. In baseline studies, we characterize the circadian and diurnal rhythms, and responses to phase advancing and delaying light pulses in a well characterized mouse strain (Aim I). Pilot studies suggest that the mouse SCN, like that of the hamster is made up of two distinctly different cell types, based on their light responsiveness and oscillating properties. To understand the organization of this circadian regulatory system, we will use transgenic animals with GFP reporters that are markers for the rhythmic (Per1 ::GFP) and light-induced (calbindin::GFP) compartments of the SCN (Aim II and III). Finally, we will characterize the microcircuitry of the SCN using calcium imaging and electrical recording and stimulation to understand the network properties of the SCN, using GFP to characterize the responses of identified (Per1 ::GFP and the CalB::GFP) SCN cell types (Aim IV).