睡眠觉醒周期中内侧前额叶皮层胞内钙信号活动变化及与视觉信息处理的关系

The prefrontal cortex, especially the medial prefrontal cortex (mPFC), is the most evolved brain regions highly associated with level of wakefulness and plays a critical role in many higher cognitive functions, such as working memory and attention. In brain, most depolarizing electrical signals are associated with Ca2+ influx. Such Ca2+ signals are essential for elementary forms of neuronal communication. In addition, Ca2+ signaling is obligatory for complex processes, such as the induction of memory- and learning- related forms of neuronal plasticity. Our previous findings support the role of arousal systems-innervating neural circuits of mPFC in working memory performance. While little is known about the characteristics and significance of intracellular Ca2+ in mPFC during information processing across sleep and wakefulness states in vivo. In this project, we will first use in vivo two photon calcium imaging technique to investigate the characteristics of intracellular Ca2+ in primary neurons, interneurons and astrocytes, especially, in dendrites and spines from I~VI in mPFC in sleep and wakefulness cycle. Then the activity of intracellular Ca2+ and different process of visual information will be recorded. And the correlation between activity of intracellular Ca2+ in mPFC and corresponding changes of visual information processing during prolonged wakefulness will be analyzed eventually. Multiple methods will be used frequently in our project, including optogenetic activation/inactivation, EEG and EMG recordings, and bolus loading Ca2+ indicators, if any possible. We expect to clarify the relationship between the activities of intracellular Ca2+ in mPFC and information processing, which will provide more evidence for better understanding the neural substrates of learning and memory as well as a theoretical basis for the treatment of cognitive impairment in prolonged wakefulness.

内侧前额叶皮层是与觉醒水平高度相关的脑区，在整合信号、参与多种认知活动的发生中具有重要的作用。研究指出，与海马相互形成神经回路振荡，内侧前额叶皮层神经元在获得、贮存信息中具有昼夜节律变化的特点。胞内钙信号在中枢系统内扮演着特殊的角色，对于许多复杂的神经细胞生物过程都是必不可少的。在前期研究内侧前额叶皮层神经回路昼夜活动特点的基础上，本研究拟采用在体双光子钙成像技术、光遗传学技术、在体电生理技术和分子生物技术，观察睡眠觉醒周期中，内侧前额叶皮层中 I~VI层神经元、胶质细胞，树突和树突棘钙信号活动规律，以及与视觉信息处理的相关性；不同觉醒水平下，细胞和树突（棘）钙信号的变化以及信息处理的改变。以期通过对内侧前额叶皮层睡眠觉醒周期胞内钙活动规律，及参与信息获得、编码、提取等学习记忆活动关系的阐述，为理解大脑皮层认知功能的神经生物基础及持续觉醒时认知功能受损治疗提供理论依据。

内侧前额叶皮层是与脑认知功能高度相关的脑区，尤其是在短期记忆的形成过程中发挥着重要的作用。内侧前额叶皮层等记忆相关脑区介导的空间学习记忆依赖良好的觉醒状态。本项目围绕皮层下促觉醒系统支配的内侧前额叶皮层等记忆相关脑区的活动特点，以及觉醒期神经回路与学习记忆相关的行为变化进行了探讨。采用神经束路追踪、免疫荧光技术，本研究发现向内侧前额叶皮层和内嗅皮层主要记忆脑区发出纤维投射的外侧下丘脑区促醒肽orexin神经元存在显著的空间分布特点。其中，投射到内侧前额叶皮层2个亚区边缘前区和边缘下区的orexin神经元胞体从下丘脑吻侧到尾侧均有分布，以中部数量最多，呈梭形分布特点，而投射到边缘前区的orexin神经元胞体在各个轴面上比投射到边缘下区的少。投射至内嗅皮层的orexin神经元胞体同样呈现出吻、尾两侧少、中间多的梭形分布规律，但数量显著少于投射到内侧前额叶皮层的orexin神经元数目。大部分投射到内侧前额叶皮层和内嗅皮层的orexin来源于不同亚群，少量的orexin神经元可同时投射到内嗅皮层和边缘下区亚区。行为学实验进一步发现，T迷宫任务中，orexin基因敲除小鼠正确率低于对照组，空间物体识别任务中，orexin敲除小鼠时间长于对照组，提示orexin空间工作记忆和空间识别能力将有所下降。除了提供背景兴奋水平外，促醒神经元orexin至内侧前额叶皮层和内嗅皮层的神经纤维投射参与空间工作记忆中的空间信息处理过程。前额叶皮层对持续觉醒产生疲劳非常敏感，本实验结果进一步发现，与睡眠期相比，前额叶皮层神经元在以觉醒为主的黑暗期具有较高的自噬活动，延长觉醒6小时后，自噬体蛋白水平表达显著增加，p-AMPK可能参与了觉醒期前叶皮层神经元的自噬活动。通过投射至内侧前额叶皮层等主要记忆相关脑区的皮层促觉醒系统的活动和学习记忆脑认知功能的探讨，睡眠觉醒期前额叶皮层神经元自噬周期性活动规律的阐述，为理解觉醒期脑认知功能的神经基础及认知障碍疾病的治疗提供了新思路。