Section on Light and Circadian Rhythms

2017财年开展的研究项目 1-视杆/视锥光感受器对非图像功能的贡献： 在2010年，我们发表了视杆细胞和黑视蛋白光转导对于信号光与昼夜节律光夹带是互补的（Altimus等人，Nature Neurosci 2010），并且最近我们显示了瞳孔光反射的相同结果（Keenan等人，eLife 2016）。我们现在已经开始确定视杆细胞和视锥细胞对包括急性体温调节在内的其他非表象行为的贡献。我们发现，光对体温调节影响的视网膜回路与昼夜光诱导的视网膜回路不同，我们正在通过使用NIMH正在建立的新行为设置来确定为什么存在这些差异。 2-定义驱动昼夜光诱导和相移的视网膜和大脑回路： 我们已经产生了转基因小鼠品系，以揭示ipRGC亚型和相应的大脑回路对内部生物钟与太阳日同步的贡献。我们有只携带SCN投射ipRGC的动物（Chen等人，Nature 2011）或缺乏投射到SCN的ipRGC（未发表）。这将使我们能够确定ipRGC的各个亚型对昼夜光夹带和相移的贡献。昼夜节律振荡器的相位可以被称为相移的急性光脉冲提前或延迟。我们令人兴奋的初步数据显示，不同的ipRGC群体控制昼夜节律振荡器中的相位延迟与相位提前。这一发现挑战了该领域目前的观点，即相位提前和相位延迟的基础是类似的机制，并且光对时钟具有简单的开/关效应。未来的研究将确定哪些ipRGC群体是进步与延迟所必需的，并绘制受光影响导致昼夜节律振荡器相位变化的大脑区域。 3-200个M1-Brn 3b阴性ipRGC的分子和功能表征： 我们最近做出了一个惊人的发现，ipRGC的亚群（200个M1-Brn 3b阴性，我们称之为昼夜光感受器）对于昼夜节律钟以及视觉的发育是至关重要的，尽管它们不投射到视觉中心（Chew等人，eLife 2017）。一个令人兴奋的假设是，这200个ipRGC（Chen et al.，Nature 2011）代表了一个进化的古老感光器类别，因为它们对几种不同的行为（光诱导，时钟和视觉的发展以及局部瞳孔光反射）有广泛的影响。因此，了解该群体相对于其他ipRGC和常规神经节细胞的分子和功能特化是至关重要的。因此，我们已经开始使用我们在NIMH的资源来检查这个人群的转录组和表观遗传标记。该项目将提供分子手柄，以了解200个M1 ipRGC相对于其他ipRGC和传统神经节细胞的个体发育和功能特化。 4-SCN-神经肽调节光介导的睡眠效应： SCN被认为主要通过与下游脑区域的直接传出连接来影响昼夜节律。SCN产生几种神经肽，包括血管活性肠肽（VIP）、胃泌素释放肽（Grp）、前动力素-2（Prok 2）和精氨酸加压素（Avp）。这些神经肽被认为同步SCN内的细胞振荡器。然而，它们是否能够吸引下游振荡器尚不清楚。研究SCN神经肽在控制下游振荡器中的功能的主要障碍是仍然具有SCN同步性但缺乏神经肽表达的动物模型的可用性。 我们已经产生了一个小鼠模型，缺乏主要SCN神经肽的表达，但在SCN保持同步细胞振荡。这使我们能够第一次专门讨论SCN神经肽在调节昼夜节律行为中的必要性和充分性。在与Seth Blackshaw（约翰霍普金斯医学院）的合作中，我们有条件地删除了LIM同源结构域转录因子Lhx 1，特别是在SCN中（Six 3-Cre; Lhx 1 lox/lox此后称为Lhx 1突变小鼠）。我们发现，Lhx 1突变体表现出显着下调的主要SCN神经肽，VIP，Grp，Avp和Prok 2的水平。值得注意的是，SCN的成熟、核心SCN时钟工作和时钟控制的基因表达仍然保留（Bedont等人，Cell Reports 2014）。Lhx 1突变小鼠在轮运行活动的昼夜输出行为中表现出适度的缺陷，但重要的是，我们观察到光对睡眠调节的严重缺陷（Bedont等人，Current Biology 2017）。我们将确定SCN神经肽在控制睡眠中的必要性。 5. Lhx 6神经元群体调节睡眠 在与Seth Blackshaw实验室的一次令人兴奋的合作中，我们发现了一个未定义的Lhx-6 GABA神经元群体，它们在调节睡眠的大脑中（Liu，Nature 2017）。 我们将继续开拓新的领域，了解来自环境的光信号如何调节对人类福祉至关重要的几种功能。

Research projects carried during the Fiscal Year 2017 1- Contribution of rod/cone photoreceptors to non-image functions: In 2010, we published that rods and melanopsin phototransduction are complementary for signaling light to circadian photoentrainment (Altimus et al., Nature Neurosci 2010) and recently we showed the same for the Pupillary light reflex (Keenan et al., eLife 2016). We have now begun to determine the contribution of rods and cones to other non-image behaviors including acute body temperature regulation. We found that the retinal circuits that underlie the effects of light on body temperature regulation are distinct from those for circadian photoentrainment and we are in the process of determining why these differences exist by using the new behavioral setup that is being built at NIMH. 2- Defining the retinal and brain circuits that drive circadian photoentrainment and phase shifts: We have generated genetically modified mouse lines to uncover the contribution of ipRGC subtypes and the corresponding brain circuits to the synchronization of the internal biological clock to the solar day. We have animals that either harbor only the SCN-projecting ipRGCs (Chen et al., Nature 2011) or lack the ipRGCs that project to the SCN (unpublished). This will allow us to determine the contribution of individual subtypes of ipRGCs to circadian photoentrainment and phase shifts. The phase of the circadian oscillator can be advanced or delayed by acute pulses of light, known as phase shifts. Our exciting preliminary data reveal that different populations of ipRGCs control phase delays versus phase advances in the circadian oscillator. This finding challenges the current view in the field that similar mechanisms underlie phase advances and phase delays, and that light has a simple ON/OFF effect on the clock. Future studies will determine which ipRGC populations are necessary for advances versus delays, and map the brain regions that are influenced by light to cause changes in the phase of the circadian oscillator. 3- Molecular and functional characterization of the 200 M1-Brn3b-negative ipRGCs: We recently made a startling discovery that a subpopulation of ipRGCs (200 M1-Brn3b-negative, which we called circadian photoreceptors) is critical for the development of the circadian clock as well as vision, although they do not project to visual centers (Chew et al., eLife 2017). An exciting hypothesis is that these 200 ipRGCs (Chen et al., Nature 2011) represent an evolutionary ancient photoreceptor class given their broad influence on several distinct behaviors (photoentrainment, development of the clock and vision as well as local pupillary light reflex). Therefore, it is critical to understand the molecular and functional specification of this population in relation to other ipRGCs and conventional ganglion cells. Thus, we have started to examine the transcriptome and epigenetic marks of this population using our resources at NIMH. This project will provide the molecular handles to understand the ontogeny and the functional specialization of the 200 M1 ipRGCs in relation to other ipRGCs and conventional ganglion cells. 4- SCN-neuropeptides in regulating light-mediated effects on sleep: The SCN is believed to influence circadian rhythms primarily by direct efferent connections with downstream brain regions. The SCN produces several neuropeptides including vasoactive intestinal peptide (VIP), gastrin releasing peptide (Grp), prokineticin-2 (Prok2), and arginine-vasopressin (Avp). These neuropeptides are thought to synchronize cellular oscillators within the SCN. However, whether they are capable of entraining downstream oscillators is unclear. A major obstacle to the study of SCN neuropeptide functions in controlling downstream oscillators has been the availability of animal models that still have SCN synchrony but lack neuropeptide expression. We have generated a mouse model that lacks the expression of major SCN neuropeptides but maintains synchronous cellular oscillations in the SCN. This allows us, for the first time, to specifically address the necessity and sufficiency of SCN neuropeptides in regulating circadian behaviors. In collaboration with Seth Blackshaw (Johns Hopkins School of Medicine), we conditionally deleted a LIM homeodomain transcription factor, Lhx1, specifically in the SCN (Six3-Cre;Lhx1lox/lox henceforth called Lhx1 mutant mice). We found that Lhx1-mutants exhibit a dramatic downregulation in the levels of the major SCN neuropeptides, Vip, Grp, Avp and Prok2. Notably, the maturation of SCN, the core SCN clock-work and clock-controlled gene expression are still preserved (Bedont et al., Cell Reports 2014). Lhx1-mutant mice exhibit modest deficits in the circadian output behavior of wheel running activity, but importantly, we observed profound deficits in sleep regulation by light (Bedont et al., Current Biology 2017). We will determine the necessity of the SCN neuropeptides in controlling sleep. 5. An Lhx6-neuronal population regulates sleep In an exciting collaboration with Seth Blackshaw's lab, we found an undefined population of Lhx-6 GABA neurons in the zona incerta regulate sleep (Liu, Nature 2017). Together, we will continue to break new ground about how light signaling from the environment regulates several functions that are essential for the well-being of humans.