CHOLINERGIC INFLUENCE ON CIRCADIAN FUNCTION

所有哺乳动物都有一个昼夜节律钟，它以近24小时为周期，控制着生物体的神经、荷尔蒙和代谢功能的日常振荡。位于大脑下丘脑的视交叉上核（SCN）存放着这个时钟。SCN与大脑的不同区域有神经连接。在这些连接中，突出的是来自大脑中负责睡眠和警报的区域，即基底前脑（BF）和脑干的胆碱能投射。关于SCN内的细胞内信号转导途径，已有相当多的工作。体外药理学研究表明，毒蕈碱乙酰胆碱受体（M1AChR）的m1亚型是介导胆碱能对昼夜节律系统影响的受体。最近，国际药理学联合会受体命名和药物分类委员会对用于暗示M1AChR参与胆碱能信号级联的药物的特异性提出了质疑。代替特定的药理学，必须采用其他方法来确定介导胆碱能昼夜节律信号的受体。在这项提议中，电生理、行为和解剖学研究将在缺乏M1AChR （M1KO）的小鼠中进行。首先，B6129PF1/J小鼠，一个与M1K0具有相似遗传背景的小鼠，将在体外和体内测试对胆碱能激动剂，carbachol的敏感性。其次，将测试M1KO对碳醇的敏感性是否降低。最后，将使用双标记荧光共聚焦显微镜来评估SCN内的胆碱能末端是否与含m1achr的细胞并置，并绘制其分布。

All mammals possess a circadian clock that governs daily oscillations with neural, hormonal, and metabolic functions for the organism with a near 24-h period. The suprachiasmatic nucleus (SCN) within the brain's hypothalamus houses this clock. The SCN has neural connections with various regions of the brain. Prominent among these connections are cholinergic projections from regions of the brain responsible for asleep and alerting, namely the basal forebrain (BF) and brainstem. A considerable body of work is available regarding intracellular signal transduction pathways within SCN. Pharmacological studies in vitro have implicated the M1-subtype of the muscarinic acetylcholine receptor (M1AChR) as the receptor mediating the effects of cholinergics on the circadian system. Recently, the International Union of Pharmacology Committee on Receptor Nomenclature and Drug Classification has questioned the specificity of the drugs used to implicate the M1AChR in the cholinergic signaling cascade. In lieu of specific pharmacology, other methods must be employed to confidently identify the receptor(s) mediating cholinergic circadian signaling. In this proposal, electrophysiological, behavioral and anatomical studies will be carried out in mice devoid of the M1AChR (M1KO). First, the B6129PF1/J mouse, a mouse of similar genetic background to the M1K0, will be tested for sensitivity to the cholinergic agonist, carbachol, both in vitro and in vivo. Second, the M1KO will be tested for decreased sensitivity to carbachol. Finally, double-label fluorescence confocal microscopy will be used to evaluate whether the cholinergic terminals within the SCN are juxtaposed to M1AChR-containing cells and to map their distribution.