Investigating circadian communication within the brain and body

SCN内的振荡器如何相互作用，以及大脑的其他部分如何相互作用，仍然是一个定义模糊但关键的研究领域。SCN的不同区域投射到很大程度上重叠的大脑区域；然而，尚不清楚每个区域的输出路径传递的是冗余信号还是不同的信号。为了评估区域相互作用和不同SCN区域的功能贡献，我使用活体光周期操作分离了背侧和腹侧SCN的节律。为了跟踪SCN内单个神经元的相，我们使用了来自PER2：LUC小鼠的SCN切片的实时生物发光成像，这是一个转基因小鼠模型，其中单个细胞内的PER2产生使用萤火虫荧光素酶报告程序进行监测。在初步研究中，我使用超长的光周期(20小时光：4小时暗，LD 20：4)在体外将SCN重组为背侧和腹侧区域，表达第一个周期的解离节律。在随后的体外循环中，背侧和腹侧SCN之间的相位差减小，表明分离的区域在体外相互作用。我建议验证以下假设：1)体外时相关系随时间的变化取决于SCN区域之间的耦合；2)SCN背侧和腹侧SCN向下游组织传递功能上不同的定时信号。为了研究这些问题，我将使用多种组织类型的实时生物发光成像，成像数据的高级计算分析，药物操作，以及SCN靶点时钟基因表达的免疫组织化学。我的长期目标是了解SCN内的神经振荡器如何相互作用，形成能够调节行为和生理节奏的功能性起搏器。

How oscillators within the SCN interact with one another and the rest of the brain remains an ill-defined but critical area of research. Different regions of the SCN project to largely overlapping brain regions; however, it is unknown whether output pathways from each region convey redundant or distinct signals. To assess regional interactions and the functional contribution of different SCN regions, I have dissociated rhythms in the dorsal and ventral SCN using an in vivo photoperiodic manipulation. To track the phase of individual neurons within the SCN, we employ real-time bioluminescence imaging of SCN slices from the PER2:LUC mouse, a transgenic mouse model where a PER2 production within individual cells is monitored using a firefly luciferase reporter. In preliminary studies, I have used an ultra long photoperiod (20h light: 4h darkness, LD 20:4) to reorganize the SCN into dorsal and ventral regions that express dissociated rhythms on the first cycle in vitro. Over the subsequent cycles in vitro, the phase difference between dorsal and ventral SCN is reduced, suggesting that dissociated regions interact in vitro. I propose to test the hypotheses 1) that changes in phase relationships over time in vitro depend on coupling between SCN regions and 2) that dorsal and ventral SCN convey functionally distinct timing signals to downstream tissues. To investigate these questions, I will use real-time bioluminescence imaging of multiple tissue types, advanced computational analyses of the imaging data, pharmacological manipulations, and immunohistochemistry for clock gene expression in SCN targets. My long-term objective is to understand how neural oscillators within the SCN interact to form a functional pacemaker capable of regulating rhythms in behavior and physiology.