CAREER: Biological Timing and Brain Circuits: Circadian influences on Prefrontal Cortex function

地球的自转提供了最显著的环境信号之一：昼夜节律（昼夜交替）。几乎所有陆生生物，从单细胞到多细胞物种，都拥有内部生物钟，使内部过程与环境同步。在哺乳动物（包括人类）中，大脑中的主生物钟与整个大脑和身体的其他生物钟同步，维持整个生物体的时间组织。随着昼夜节律的退化与许多负面健康影响相关，这些时钟的重要性在时间中断期间变得明显。然而，要充分了解“坏”时钟如何造成负面影响，了解它们在正常情况下如何促进最佳功能是必要的。这项研究的目的是了解生物钟如何促进大脑回路的正常功能，这些回路在决策、注意力和认知灵活性等复杂行为中很重要。一个小鼠模型被用来研究正常或中断的昼夜节律如何调节大脑中神经元的大小、形状和功能，以及这些变化如何影响认知。脑化学测量、成像、神经元三维重建和药理学的先进技术被用来帮助理解生物钟如何维持正常功能，以及生物钟被破坏如何导致负面结果。该奖项的一个组成部分是让农村和城市的本科生参与莫比乌斯科学中心和斯波坎儿童博物馆的推广活动，帮助提高人们对生物钟如何影响生理功能的认识，从最简单的生物体到我们自己的大脑。在了解视交叉上核（SCN）生物钟的细胞和分子功能方面取得了重大进展。然而，昼夜节律在复杂行为背后的大脑区域，如前额皮质（PFC）中的基本作用仍然是虚幻的。利用环境昼夜节律干扰作为工具，本研究确定了昼夜节律如何在行为、生理、结构和生化水平上调节正常PFC功能。这项研究建立在我们的发现之上，即昼夜节律紊乱会损害认知灵活性并导致PFC神经元萎缩。该项目的总体假设是，昼夜节律主要通过调节谷氨酸能信号来促进PFC的正常功能，因为谷氨酸对PFC的最佳功能至关重要。昼夜节律中断对PFC的影响是通过检查PFC介导的行为来确定的，并通过使用可植入的生物传感器来实时确定对细胞外PFC谷氨酸的影响。为了确定谷氨酸的因果作用，进行了PFC AMPA和NMDA信号的药理操作。使用共聚焦显微镜和PFC神经元的三维重建来研究神经形态和树突棘的昼夜变化，树突棘是兴奋信号的主要部位。生化研究确定了正常和异常的昼夜节律如何驱动谷氨酸受体的膜运输，为昼夜节律调节PFC功能提供了另一种底物。第三个也是最后一个实验目的是确定节律激素（皮质酮）的变化是否介导了这些影响，这将提供一种机制，通过这种机制，正常和中断的时间线索被传递到scn外的大脑区域。

The rotation of the Earth provides one of the most salient environmental signals: the circadian (daily) alternation of night and day. Nearly all terrestrial organisms, from single-celled to multi-cellular species, possess internal biological clocks allowing for synchronization of internal processes with the environment. In mammals (including humans), a master circadian clock in the brain synchronizes other clocks throughout the brain and body, maintaining temporal organization in the whole organism. The importance of these clocks becomes apparent during times of temporal disruption, with the degradation of circadian rhythms associated with numerous negative health effects. However, to fully understand how "broken" clocks cause negative effects, understanding how they promote optimal function under normal circumstances is necessary. The goal of this research is to understand how circadian clocks promote normal functioning of brain circuits important in complex behaviors like decision making, attention, and cognitive flexibility. A mouse model is used to investigate how normal or disrupted circadian rhythms regulate the size, shape, and function of neurons in the brain, and how these changes affect cognition. Advanced techniques in the measurement of brain chemistry, imaging, 3-dimensional reconstruction of neurons, and pharmacology is used to help understand how circadian clocks maintain normal function, and how disrupted clocks lead to negative outcomes. An integral component of this award engages rural and urban undergraduate students in outreach involving the Mobius Science Center and Children's Museum in Spokane, helping increase awareness of how biological clocks affect physiological function, from the simplest organisms, to our own brain.Significant inroads have been made in understanding the cellular and molecular function of the suprachiasmatic nucleus (SCN) circadian clock. However, the fundamental role of circadian rhythms in brain areas underlying complex behaviors, such as the prefrontal cortex (PFC), remains illusive. Using environmental circadian disruption as a tool, this research determines how circadian rhythms modulate normal PFC function at the behavioral, physiological, structural, and biochemical levels. This research builds upon our findings that circadian disruption impairs cognitive flexibility and causes atrophy of PFC neurons. The overarching hypothesis of this project is that circadian rhythms promote normal PFC function primarily through modulation of glutamatergic signaling, since glutamate is crucial for optimal PFC function. Circadian disruption effects on the PFC is determined by examining PFC mediated behaviors, and through the use of implantable biosensors to determine effects on extra-cellular PFC glutamate in real time. To establish a causal role for glutamate, pharmacological manipulation of PFC AMPA and NMDA signaling is undertaken. Confocal microscopy and 3-D reconstruction of PFC neurons is used to investigate circadian changes in neural morphology and dendritic spines, primary sites of excitatory signaling. Biochemical studies determine how normal and abnormal circadian rhythms drive membrane trafficking of glutamate receptors, providing another substrate on which circadian rhythms may act to modulate PFC function. The third and final experimental aim is to determine whether changes in a rhythmic hormone (corticosterone) mediates these effects, which would provide a mechanism by which normal and disrupted timing cues are relayed to extra-SCN brain regions.