外侧缰核介导抑郁大鼠睡眠障碍的作用和机制

Circadian rhythm disturbance has been shown to be associated with etiology of depression. The activity of lateral habenula (LHb) neurons has circadian rhythm characteristics, and a increased LHb neuron activity plays an important role in pathogenesis of depression. In addition, there are a close relationship with morphologically and functionally between tha labenula and the pineal gland which rhythmically secrets melatonin and regulates sleep-wake circadian rhythm. Therefore, based on our previous work, we will continue to target LHb in this proposed study. We plan to detect whether there is correlation between changes in LHb rhythmic gene expression and sleep disturbance caused by depression in rats though behavior test, EEG and EMG technique, and also determine the impact of depressive behavior and sleep disturbance by lesion of LHb or inhibition/activation of rhythmic gene expression.We will further testify LHb's influence on melatonin secretion of pineal gland by testing pineal melatonin, plasma metatonin, plasma cortisol and thyroid hormone in different time window, and finally to identify whether activity changed LHb neurons in depression model project the fiber to pineal gland and influence the function of pineal gland by using single cell recording in vivo, patch-clamp recording in vitro combined with retrograde tracing technique and immunofluorescence technique. Successful completion of this proposed study will reveal the function and mechanism of LHb in mediating depression-induced sleep disturbance based on the theory of circadian rhythms. . The expected outcome of this study will provide new evidence to help develop the therapeutic strategy in anti-depression treatment based on regulation of circadian rhythms.

昼夜节律紊乱与抑郁症发病有关，外侧缰核（LHb）神经元活动具有昼夜节律特征，并且其功能增强在抑郁症发病中起重要作用。此外，LHb也与通过节律性分泌褪黑素调节睡眠觉醒昼夜节律的关键部位松果体有纤维和功能联系。因此，本课题在我们原有工作基础上，继续选择缰核做为研究靶点，首先应用行为学、脑电肌电记录等方法，弄清抑郁伴有睡眠障碍大鼠与LHb节律基因表达变化是否相关，然后证明LHb损毁或影响节律基因表达对抑郁大鼠抑郁行为、睡眠障碍的影响；进一步通过松果体和血浆中不同时间点褪黑素含量、血浆皮质醇和甲状腺素含量测定，证明其作用可能与影响松果体褪黑素的分泌有关；最后应用整体电生理记录、膜片钳记录结合逆行纤维示踪和免疫荧光技术，明确抑郁大鼠LHb活动改变的神经元是否其纤维投射到松果体并影响松果体功能。以从昼夜节律角度揭示LHb介导抑郁大鼠睡眠障碍的作用和机制，为临床通过调整昼夜节律达到抑郁症治疗提供新依据。

抑郁症发病与昼夜节律失调密切相关，外侧缰核（LHb）活动具有昼夜节律特征，并且是抑郁症发病的关键部位，此外，LHb也与松果体有纤维联系。因此，本课题探讨LHb在抑郁大鼠睡眠障碍中的作用和机制。我们完成了计划中的大部分内容，并调整了研究计划中的个别内容。结果如下：.1.抑郁大鼠活动的昼夜节律性及LHb和松果体内节律基因的表达.（1）与正常大鼠比较，抑郁大鼠的糖水偏爱率、强迫游泳中的上窜时间在夜晚均明显降低，导致其两种行为的昼低夜高的振幅降低。.（2）正常与抑郁大鼠LHb内基因的表达均呈现出昼夜节律振荡。其中在抑郁鼠Bmal1、Per2及Cry1基因的表达在夜晚均明显低于正常鼠，导致其昼夜节律振幅降低；松果体内正常和抑郁大鼠节律基因表达均呈现节律振荡，其中Per1和per2基因表达在ZT22时显著高于对照组。这些节律基因表达的变化可能与抑郁大鼠行为的昼夜节律改变有关。.2. 在抑郁大鼠发病中LHb与松果体之间的功能联系改变.（1）松果体对LHb活动的影响：1）正常大鼠的活动表现明显的昼夜节律性，松果体摘除可诱导大鼠抑郁样行为，表现夜晚的不动时间增加，导致其昼夜节律震荡减弱；2）松果体摘除不影响LHb内节律基因表达的昼夜节律特征，但Per1 mRNA在夜晚的表达明显高于对照组；3）记录离体脑片LHb神经元放电，发现褪黑色素可以明显抑制LHb神经元放电频率。.（2）LHb对松果体的影响：1)LHb损毁可以使松果体组织的重量降低；2）LHb损毁能破坏松果体的超微结构，如突触带消失、线粒体肿胀、嵴断裂，固缩等变化。.3. LHb在睡眠稳态调节中的作用.我们观察到6小时睡眠剥夺可明显提高LHb内Per2mRNA表达；LHb 损毁可以缩短睡眠剥夺恢复期NREM睡眠，延长觉醒时间。.以上结果说明，抑郁大鼠表现昼夜节律性活动的改变，可能与LHb 和松果体内节律性基因表达的昼夜节律振幅降低有关；松果体摘除可以引起大鼠抑郁样行为，可能通过改变LHb内节律基因的表达，进而影响其功能实现；LHb参与睡眠稳态的调节与其Per2基因的表达变化有关。