【作者】 许光辉；

【导师】 李廷利；

【作者基本信息】 黑龙江中医药大学 ， 中药学， 2010， 博士

【摘要】 目的:以果蝇为模式生物,在基因水平探讨刺五加改善睡眠的作用机制。方法:本实验采用果蝇活动监测技术和夜间亮-暗交替刺激剥夺果蝇睡眠的方法,研究刺五加对正常果蝇和果蝇睡眠剥夺模型的睡眠-觉醒周期的影响。在此基础上,应用果蝇全基因组表达谱芯片技术研究刺五加对果蝇睡眠-觉醒相关基因表达的影响,并通过生物信息学中数据分析技术探讨刺五加改善睡眠的机制。结果:1、雌果蝇攀爬指数在1日龄到32日龄内稳定的保持在最高水平;雄果蝇攀爬指数在1日龄到22日龄内稳定的保持在最高水平;7日龄到17日龄的雌、雄果蝇的攀爬指数无显著性差异。2、雌果蝇的睡眠总时间和睡眠指数随日龄的变化, 2日龄到12日龄时呈增加趋势,12日龄到32日龄时呈降低趋势;雄果蝇的睡眠总时间和睡眠指数随日龄的变化,2日龄到22日龄时呈增加趋势。3、7日龄的雌、雄果蝇的睡眠总时间存在着显著差异(P<0.01),雌果蝇总睡眠时间明显少于雄果蝇(P<0.01);雌果蝇的睡眠主要集中夜间,而雄果蝇白天的睡眠时间和夜晚的睡眠时间无显著性差异。4、夜间亮-暗交替刺激均能显著降低雌、雄果蝇的睡眠时间(P<0.01)和睡眠指数(P<0.01)。5、0.125mg/ml的地西泮对雌、雄果蝇的亮-暗交替刺激睡眠剥夺模型的睡眠时间和睡眠指数有明显的恢复作用(P<0.05)。6、0.125%-1%的刺五加对均能显著增加雌、雄果蝇睡眠总时间(P<0.05)和雌果蝇的睡眠指数(P<0.05),0.25%的刺五加可极显著增加雌、雄果蝇睡眠总时间(P<0.01)和睡眠指数(P<0.01)。7、0.25%的刺五加给药4d-7d能极显著增加雌果蝇的睡眠总时间(P<0.01)和睡眠指数(P<0.01);0.25%的刺五加给药2d-7d能极显著增加雄果蝇的睡眠总时间(P<0.01),给药4d-7d能显著增加雄果蝇睡眠指数(P<0.05)。8、0.5%和1%的刺五加能显著增加雌果蝇睡眠剥夺模型的睡眠总时间(P<0.05),0.25%-1%的刺五加能显著增加雌果蝇睡眠剥夺模型的睡眠指数(P<0.05);刺五加对雄果蝇睡眠剥夺模型的睡眠总时间和睡眠指数均无显著性影响。9、1%的刺五加给药2d-7d能极显著增加雌果蝇睡眠剥夺模型睡眠时间(P<0.01),给药3d-7d能显著增加雌果蝇的睡眠指数(P<0.05)。10、1%的刺五加给药4天,可使果蝇头部194个已知功能基因差异表达,上调基因80个,下调基因114个;夜间亮-暗交替刺激,可使果蝇头部184个已知功能基因差异表达,上调基因83个,下调基因101个;1%的刺五加给药4天和夜间亮-暗交替刺激共同作用,可使果蝇头部685个已知功能基因差异表达,上调基因188个,下调基因497个。结论:1、果蝇攀爬指数随日龄增加逐渐降低。2、雌果蝇的睡眠总时间和睡眠指数随日龄的变化,在12日龄前呈增加趋势,之后呈降低趋势;雄果蝇的睡眠总时间和睡眠指数随日龄的变化,2日龄到22日龄时呈增加趋势。3、浓度为0.25%-1%刺五加可明显改善雌、雄果蝇的睡眠-觉醒周期;刺五加改善雌、雄果蝇睡眠-觉醒周期的最佳给药方案:0.25%的刺五加,给药时间≥4天。4、浓度为0.25%-2%刺五加可明显改善雌果蝇的睡眠剥夺模型的睡眠-觉醒周期,刺五加对雄果蝇睡眠剥夺模型睡眠-觉醒周期没有改善作用;刺五加改善雌果蝇睡眠剥夺模型睡眠-觉醒周期的最佳给药方案:1%的刺五加,给药时间≥3天;。5、刺五加改善果蝇睡眠的作用,是以dco、slo和DopR2等基因为主要参与者的多基因参与的过程、多基因共同调节的结果。更多还原

【Abstract】 Objective:The mechanism of Ci Wu Jia on improving sleep in drosophila was studied using the microarray of whole drosophila genome. Methods:In this study, the effects of Ci Wu Jia on improving sleep of normal flies and flies sleep-derprivation models was determined by using the Drosophila activity monitoring technology. The flies models of sleep-deprivation was maked by the alternative light-dark stimulating at night. On this basis, the acting mechanism was explored by using the microarray technology of whole drosophila and bioinformatics data analysis techniquesResults:1. The climbing index of female flies aged 1-day-old to 32-day-old maintained stability at the highest level; the climbing index of male flies aged 1-day-old to 22-day-old remain stability at the highest level; the climbing index female and male flies aged 7-day-old to 17-day-old were no significant difference.2. The sleeping time and sleep consolidation index of female flies was increasing with age from 2-day to 12-day, and decreasing with age from 12-day to 32-day; the sleeping time and sleep consolidation index of male flies was increasing with age from 2 day to 22 day.3. The sleeping time of female fruit flies total sleeping time was significantly less than the male fruit flies(p<0.01); female flies sleep mainly at night, and the sleeping time of male flies during the day and night was no significant difference.4,The alternative light-dark stimulation can reduce sleeping time and sleeping index of female and male flies significantly(p<0.01).5. The sleeping time and sleeping consolidation of sleep-deprivation models of female and male flies was recoveried by administering diazepam at the concentration of 0.125 mg/ml (P <0.05).6. CWJ can increase sleeping time of the male and female flies significantly (p<0.05) at the concentration of 0.125% to 1% , and the sleeping consolidation index was increased (p<0.05); CWJ can increase the sleeping time(p<0.01) and sleeping consolidation index(p<0.01) significantly at the concentration of 0.25%.7. CWJ of 0.25% can significantly increase the sleeping time (p<0.01) and sleeping index (p<0.01) of female flies administration 4d-7d; CWJ of 0.25% can significantly increase the sleeping time (p<0.01) and sleeping index (p<0.05) of male flies administration 4d-7d.8. CWJ of 0.5% and 1% can increase sleeping time of the sleep deprivation model of female flies(p<0.05), CWJ can increase sleeping index of the sleep deprivation model of female flies(P <0.05) significantly at the concentration of 0.25%-1%; the sleepin time and sleep index of the male flies sleep deprivation model were not significantly affected by CWJ.9. Administration of 2 to 7d of CWJ of 1% can significantly increase the female flies sleep deprivation sleep time (p<0.01), administration of 3d-7d can significantly increase the female flies sleep index (p<0.05)..10. CWJ(concentration of 1%, administration 4 days) can changed expression of 194 genes in the head of fly, 80 genes up-regulated, and 114 genes were down-regulated; night light-dark stimulus can changed expression of 184 genes in the head of fly, 83 genes up-regulated, and 101 genes were down-regulated; the interaction of CWJ(concentration of 1%, administration 4 days) and night light-dark stimulating can changed expression of 685 genes in the head of fly, 188 genes up-regulated, and 497 genes were down-regulated.Conclusion:1. The climbing index of flies decreased with increasing age.2. The sleeping time and sleeping index of female flies changes with age, before age 12 tended to increase, then decreasing; the sleeping time and sleeping index of male flies changes with age, 2day age to 22 days of age tended to increase.3. CWJ can improve male and female fruit flies sleep-wake cycle at the best dosage regimen: CWJ of 0.25%, delivery time≥4days.4. CWJ can improve sleep-wake cycle of sleep deprivation model of female fruit flies at the best dosage regimen: CWJ of 1%, delivery time≥3days; CWJ was no effect on sleep-wake cycle of male flies sleep deprivation model.5. CWJ improve the role of fruit flies sleep, is dco, slo and DopR2 major players such as genes involved in the process of multi-gene, multi-gene co-regulation results.更多还原