The Effect And Mechanism Of Ziziphi Spinosae Decoction Regulating SCN And VLPO System For Senile Insomnia Based On 5-HTR

Objective : Arrange pertinent literature of sleep deprivation and Ziziphi Spinosae Decoction combine the previous research foundation of our research group.Through the establishment of chronic sleep deprivation senile insomnia rats,explore the effect and mechanism of the SCN,VLPO system and their projection relation in senile sleep deprivation rats.Methods:Theoretical study: Review the pertinent literature of sleep deprivation and Ziziphi Spinosae Decoction,summarize the classicaltheoretical content and advancing front of scientific research.General understand the Circadian Rhythms of Sleep-wake and materials base for pharmacological,Research and discuss the Ziziphi Spinosae Decoction regulating sleep deprivationexperimental animalfrom the angle of mechanism.Experimental study: Select SPF SD senile insomnia rats,deprive them from REM sleep 21 d with self-made sleeping depriving cage,continuous administration 14 d,the detection indexas follows:1.The effect of myocardial cell apoptosis in chronic sleep deprivation senile insomnia rats,and expression of protein bcl-2,baxObserve rats' myocardial cell apoptosis with Tunel staining technique after chronic sleep deprivation,observe the change of rats' myocardial cell bcl-2,bax expression with immunohistochemical methods after chronic sleep deprivation.2.The effect of Ziziphi Spinosae Decoction on SCN system in chronic sleep deprivation senile insomnia ratsThe transmission electron microscope observe the morphologic changes in SCN.The change of c-fos protein expression is observed by immunohistochemical methods.Immunohistochemical methods and Realtime-PCR can observe vSPZ in SCN,n NOS in DMH,and the change of VIP,AVP neuron expression.3.The effect of Ziziphi Spinosae Decoction on VLPO system in chronic sleep deprivation senile insomnia ratsImmunohistochemical methods and immunofluorescence staining assaycan observe the change of galanin expression.Nissl's staining check the change of VLPO neuronal expression.Immunohistochemical methods and immunofluorescence staining assay observe the change of GABA expression,Realtime-PCR check the expression of hypothalamus GABAAR?1,?2 subunit.Check thecontent of 5-HT with fluorescence spectrophotometry,the change of hypothalamus 5-HT1 AR,5-HT2 AR mRNA expression with Realtime-PCR.Results:1.The effect of myocardial cell apoptosis in chronic sleep deprivation senile insomnia rats,and expression of protein bcl-2,baxThe result of apoptosis of myocardial cellsobserved by Tunel staining technique With respect to environmental control group,chronic sleep deprivation model group show obvious decrease of apoptotic cell number(P<0.05);and with respect to chronic sleep deprivation model group,Surazepam group and Ziziphi Spinosae Decoction show varying degrees ofdecrease of apoptotic cell number(P<0.05).The result of bcl-2,bax expression with immunohistochemical methodsWith respect to environmental control group,chronic sleep deprivation model group showrising trend of bcl-2,baxactivity(P<0.05);and with respect to chronic sleep deprivation model group,Surazepam group and Ziziphi Spinosae Decoction group show obvious decrease of bcl-2,bax activity(P<0.05).2.The effect of Suanzao-Rentang on SCN system in chronic sleep deprivation senile insomnia rats.The result of SCN shape with ultrastractural pathology With respect to environmental control group,chronic sleep deprivation model group show increase of Lipo in SCN neuron,obvious increase of swelling Golgivesicles.With respect to chronic sleep deprivation model group,Surazepam group and Ziziphi Spinosae Decoction group show relatively decrease and lessening of LIPO.Ziziphi Spinosae Decoction group is of the most remarkable change.With respect to environmental control group,chronic sleep deprivation model group show down-regulation of VIP,AVP mRNA expression;with respect to chronic sleep deprivation model group,Surazepam group and Ziziphi Spinosae Decoction group show up-regulation of VIP,AVP mRNA expression.The result of immunohistochemical methods With respect to environmental control group,chronic sleep deprivation model group show decrease of VIP,AVP expression,increase of nNos,c-fos expression(P<0.05);and with respect to environmental control group,Surazepam group and Ziziphi Spinosae Decoction group show increase of VIP,AVP expression,decrease of nNos,c-fos expression(P<0.05);3.The effect of Ziziphi Spinosae Decoction on VLPO system in chronic sleep deprivation senile insomnia rats.With respect to environmental control group,chronic sleep deprivation model group show down-regulation of 5-HT1 AmRNA,up-regulation of 5-HT2AmRNA;with respect to chronic sleep deprivation model group,Surazepam group and Ziziphi Spinosae Decoction group show up-regulation of 5-HT1 AmRNA,down-regulation of 5-HT2 AmRNA.With respect to environmental control group,chronic sleep deprivation model group show down-regulation of GABAAR?1,GABAAR?2expression level;and with respect to chronic sleep deprivation model group,Surazepam group and Ziziphi Spinosae Decoction group show up-regulation of GABAAR?1,GABAAR?2 expression level.With respect to environmental control group,chronic sleep deprivation model group showincrease of GABA,galanin expression(P<0.05);and with respect to chronic sleep deprivation model group,Surazepam group and Ziziphi Spinosae Decoction group show decrese of GABA,galanin expression in rat hypothalamic(P<0.05).The analysis of 5-HT content in rat hypothalamic checked with fluorescence spectrophotometryWith respect to environmental control group,chronic sleep deprivation model group showdecrease of 5-HT relative expression(P<0.05);and with respect to chronic sleep deprivation model group,Surazepam group and Ziziphi Spinosae Decoction group show varying degrees of 5-HT content(P<0.05).The result of Nissl's staining checking the neuronal expression of VLPO : The neuronal cellular morphology of VLPO in environmental controlgroup is normal,space closely,cyto-architectonics is fininshed,the nucleus is round or oval,nucleolus clear,abundant Nissl bodies can be seen in cytoplasm.After senile sleep deprivation,the dyeing condition of rats change obviously,space loosely,partial neuronal cell shrinkage,Nissl bodie decrease in cytoplasm.The result of GABA,galanin expression checked with immunofluorescence staining assay With respect to environmental control group,chronic sleep deprivation model group show decrease of GABA,galanin neuronal expression(P<0.05);and with respect to chronic sleep deprivation model group,Surazepam group and Ziziphi Spinosae Decoction group show increase of GABA,galanin neuronal expression(P<0.05).Conclusion:Ziziphi Spinosae Decoction can regulate sleep-wakefulness mechanismbyregulating SCN and VLPO system,effectively improve the senile chronicinsomnia.