Subparaventricular zone pathways to circadian synchrony

描述（由申请人提供）：下丘脑的室旁亚区（SPZ）是同步和驱动行为和生理昼夜节律的电路的重要组成部分。SPZ接收来自视交叉上核（SCN）的大部分轴突输出，SCN是哺乳动物的昼夜节律起搏器，并进一步投射到大多数SCN的下游目标。对大鼠的生理和解剖学研究表明，SPZ是异质性的，由不同的子区域组成，以不同的方式调节特定的节律。具体来说，背侧SPZ病变，由室旁核（PVH）向腹侧延伸的区域定义，选择性地破坏体温（Tb）节律，对运动活动（LMA）或睡眠/觉醒节律的影响最小。相比之下，腹侧SPZ病变，由SCN向背侧和尾侧延伸的区域定义，严重破坏LMA和睡眠/觉醒节律，对Tb的影响较小。从大鼠SPZ的顺行追踪进一步表明，该区域由四个解剖学上和功能上不同的亚区组成：腹内侧和腹外侧SPZ （vmSPZ, vlSPZ）和背内侧和背外侧SPZ （dmSPZ, dlSPZ）。某些条件敲除小鼠的最新进展允许直接测试SPZ亚区在体内的神经递质特异性作用。我们试图更好地了解SPZ的组织和流出及其在小鼠昼夜节律系统中的作用。阐明这些途径可能揭示慢性昼夜节律紊乱改变行为并对整体健康产生不利影响的机制。目的1。为了评估SPZ的GABA能亚群在Tb、LMA和睡眠/觉醒的昼夜节律调节中的作用：SPZ主要由GABA能神经元组成，它们使用VGAT将GABA包装成囊泡。利用Cre-Lox技术，我们将注射一种病毒载体来删除SPZ亚区VGAT的表达，阻止这些细胞释放GABA。我们将记录小鼠在恒定黑暗下的自由运动，同时使用植入腹腔的生物遥测发射器测量Tb和LMA，并使用脑电图和肌电图记录睡眠/清醒。我们希望这些实验能够确定SPZ中哪些gaba能神经元群调节LMA、睡眠/觉醒和Tb的昼夜节律。目标2。为了描述gaba能产生的SPZ神经元的传出投射，并将Aim 1的生理模式与gaba能在特定终末场传递的缺失联系起来：目前还没有发表的关于小鼠SPZ传出的详细分析的报告。Aim 1中使用的载体也追踪来自转导神经元的传出投射，并将揭示介导这些节律的投射靶标，然而，它也追踪来自非gaba能神经元。为了阐明可能的影响，我们将使用在VGAT启动子下表达Cre的小鼠和表达Cre依赖示踪基因的载体来选择性地追踪SPZ gaba能投射。然后，我们将比较特定靶点的VGAT缺失与Aim 1中的生理反应，以确定调节特定昼夜节律的SPZ靶点。

DESCRIPTION (provided by applicant): The subparaventricular zone (SPZ) of the hypothalamus is an important component of the circuitry that synchronizes and drives circadian rhythms of behavior and physiology. The SPZ receives most of the axonal output from the suprachiasmatic nucleus (SCN), the mammalian circadian pacemaker, and further projects to most of the SCN's downstream targets. Physiological and anatomical studies in rats suggest the SPZ is heterogeneous and comprised of subregions differentially modulating particular rhythms. Specifically, dorsal SPZ lesions, defined by the region extending ventrally from the paraventricular nucleus (PVH), selectively disrupt body temperature (Tb) rhythms with minimal impact on locomotor activity (LMA) or sleep/wake rhythms. In contrast, ventral SPZ lesions, defined by the region extending dorsally and caudally from the SCN, profoundly disrupt LMA and sleep/wake rhythms with less effect on Tb. Anterograde tracing from the SPZ in rats further suggests this area is comprised of four anatomically, and perhaps functionally, distinct subregions: the ventromedial and ventrolateral SPZ (vmSPZ, vlSPZ) and the dorsomedial and dorsolateral SPZ (dmSPZ, dlSPZ). Recent developments of certain conditional knockout mice allow for the direct testing of the neurotransmitter-specific role of SPZ subregions in vivo. We seek to gain a better understanding of SPZ organization and outflow and its role in the circadian timing system in mice. Elucidating such pathways may reveal mechanisms by which chronic circadian disruption alters behavior and adversely affects overall health. Aim 1. To assess the role of GABAergic subpopulations of the SPZ in the modulation of circadian rhythms of Tb, LMA, and sleep/wake: The SPZ is primarily comprised of GABAergic neurons, which package GABA into vesicles using VGAT. Using Cre-Lox technology, we will inject a viral vector to delete VGAT expression in SPZ subregions, preventing these cells from releasing GABA. We will record freely moving mice in constant dark, while measuring Tb and LMA using biotelemetry transmitters implanted in the intraperitoneal cavity, and sleep/wake using EEG and EMG recordings. We expect these experiments to identify which populations of GABAergic neurons in the SPZ regulate circadian rhythms of LMA, sleep/wake, and Tb. Aim 2. To delineate efferent projections from GABAergic SPZ neurons and correlate the physiological patterns in Aim 1 with deletion of GABAergic transmission in specific terminal fields: There are no published reports of a detailed analysis of SPZ efferents in mice. The vector used in Aim 1 also traces the efferent projections from transduced neurons and will reveal the projection targets that mediate these rhythms, however, it also traces from non-GABAergic neurons. To clarify the projections likely underlying our effects, we will use mice expressing Cre under the VGAT promoter and a vector expressing the gene for a Cre-dependent tracer to selectively trace SPZ GABAergic projections. We will then compare VGAT deletion in specific targets with physiological responses in Aim 1 to determine SPZ targets regulating specific circadian rhythms.