Xiaoyaosan Regulatory Role Of Liver Stagnation And Spleen Deficiency Rats Hippocampus And Amygdala Bdnf

1 theoretical discussionLiver invading the spleen syndrome,a common type of depression patterns,is also known as "the wood overacting the earth".With good operability and repeatability,our group ensure the rat model subjected to chronic immobilization stress(CIS) was an ideal model for research on this syndrome.CIS simulates the pathological process of the liver failing to maintain the normal flow of QI and the treatment of Xiaoyao san promotes the functional recovery of liver governs smoothing qi flow(LGSQF).Since multiple systems relate to chronic stress,and different stressor,different individuals and different circumstances might lead to different results,it's so difficult for prevention and treatment to chronic stress diseases,whereas Chinese medicine,which with a concept of whole adjustment,treatment based on syndrome differentiation and individualized diagnosis,has shown greater superiority.Considerable interest in this paper has focused on the molecular mechanisms and regulation channel underly Xiaoyao san affording effective protection agaist this depression syndrome.The present project was supported by National Natural Science Foundation of China(NO.30672578).The topic chooses intercommunication and interaction of the two regions--the hippocampus and amygdala,it is to discuss distribution and expression of BDNF, observation of brain synaptic plasticity,regulating role of xiaoyaosan on BDNF when chronic immobilization stress,from the theory of modern stress,combined with research results in the stress theory.It is further explore human injury and the internal mechanism for the disease when chronic stress,and liver controlling dispersion has the leading position,and part Mechanism that xiaoyaosan eliminate or reduce the chronic stress,and also is the manifestation of function of LGSQF.This is not only conducive to the basic theories and clinical research of Chinese medicine and development,but it has great significance for prevention and treatment of many stress disease.There is now general consensus that antagonists of AMPA receptors,such as CNQX, appear to be much more effective than NMDA antagonists(MK-801) in preventing CA3 cell from atrophy even death following CIS.This experiment was designed to explore CIS rats' behaviors,Immunohistochemistry findings and RT-PCR results in hippocampal subregions CA1,CA3 and DG and amygdaloid subregion BLA after suppressing BDNF by microinjection of CNQX,to exploer the regulatory effect of Xiaoyao san.2 Experimental studyThis experiment is composed of four parts.Part 1,to establish an animal model of depression induced by CIS in accord with syndrome of liver invading the spleen in TCM. From the animal behavior,observion the signs and symptoms and detection the rate of urinary D-fructose excretion to confirmate the rat model of syndrome of liver invading the spleen by CIS.In part 2,3 and 4,from rats' behaviors,Immunohistochemistry findings and RT-PCR to explore the expression of BDNF in sudregion of hippocampus and amygdale.Rats were randomly divided into 5 groups,control group,model group, sham operation group,CNQX group and Xiaoyao san group.,15/group,5/cage.After 21d's immobilization,the last four group were operated in 22^th day.The sham operation group and Xiaoyao san group rats were microinjected 0.5ul normal saline in both amygdaloid subregions,and CNQX group rats were microinjected 0.5 ul CNQX solution. Except Xiaoyao san group rats were gavaged Xiaoyao san solution,of other groups were gavaged same volume normal saline.The model group rats didn't operated.Rats were observed behavior in the 25^th day and perfused,fixed and drawed meterials in the 26^th day.3 results and conclusion3.1 Part 1RESULTS:CIS group:The results revealed that subjected to CIS resulted in a decrease of behavioral activity,weight-gain,food intake,low rate of urinary D-fructose excretion, and weak ambulation,rising and decoration in the open field test especially during the period of 14th to 21st day.Xiaoyao san group:Xiaoyao san treatment significantly reversed the behavioral deficit of these depression model rats especially during the period of 14th to 21st day.CONCLUSION:With good operability and repeatability,the CIS rat model is an ideal model for research on syndrome of liver invading the spleen.Long term depression leads to liver invading the spleen,that is to say,results in syndrome of depression in combination with spleen deficiency.Xiaoyao san can regulate the behavior abnormality in CIS rats.3.2 Part 2RESULTS:Signs and symptoms of Liver invading the spleen syndrome were seen in model group rats;anxiety-related behaviors were shown in sham operation group; anxiety-related behaviors were controlled in CNQX group revealing antidepressant treatment of CNQX;rats of Xiaoyao san group resist the injury of CIS and operation and has little difference with control group.CONCLUSION:â‘ Suppression of both amygdaloid excitability by microinjection of CNQX,AMPA receptor antagonist,can relieve anxiety acting as an antidepressant drug.â‘¡Xiaoyao san can effectively regulate the rats' behavior abnormality.3.3 Part 3Immunohistochemistry findings:â‘ The positive area,MOD and IOD of BDNF were similar with the number of positive cells.â‘¡The neurons in CA3 were damaged more seriously than in CA1,which were same with in BLA,and in DG were damaged slighter than in CA3 and CA1.â‘¢The change trend of BDNF positive cells:in the CA1 region of hippocampus,compared with control group,sham operation group and CNQX group had statistical significance,the number of BDNF positive cells decreased.Compared with sham operation group,Xiaoyao san group had statistical significance,both between CNQX group and Xiaoyao san group.In the CA3 region of hippocampus,compared with control group,model group,sham operation group,and Xiaoyao san group had statistical significance,the numbers of BDNF positive cells decreased.In the DG region of hippocampus,compared with control group,model group,sham operation group,and Xiaoyao san group had statistical significance.Compared with model group,sham operation group,CNQX group and Xiaoyao san group had no statistical significance.,but the number of positive cells in sham operation group and CNQX group were less and more in Xiaoyao san group.Statistical analysis showed that no difference existed in sham operation group and model group,CNQX group and Xiaoyao san group,but Xiaoyao san group were more than sham operation group.In the region of BLA, compared with control group,model group and sham operation group had statistical significance,the cells decreased.Statistical analysis showed that difference existed in model group and CNQX group,and number of positive cells increased in CNQX group. And the situations were same in sham operation group and CNQX group.CONCLUSION:â‘ Statistical analysis showed that no difference existed in model group and sham operation group,which showed than surgical injury had little effects with the expression of BDNF.â‘¡Respectively compared with model group and sham operation group,the expressional tendency of BDNF was almost the same in CNQX and Xiaoyao san group in sudregion of hippocampus and BLA,which showed than Xiaoyao san can relieve effectively the stimulation of surgical injury and binding.Xiaoyao san may regulate the balance between the hippocampus and amygdale.This finding also revealed that CNQX inhibited excitation of basolateral amygdale by antagonizing basolateral amygdale AMPA receptors,which can reduce stimulation intensity in rengions of hippocampus.â‘¢In this experiment,the expressional tendency of BDNF decreased in rengions of hippocampus when chronic binding stress,especially in BLA obviously because BLA seriously damaged when excitation in chronic stress.Microinjection CNQX can inhibit BLA's overexcitation,so the expressional tendency of BDNF in basolateral amygdale didn't decreased significantly inverse increased.This finding revealed that CNQX may react treatment.3.4 Part 4RT-PCR:Results was respectively perfectly coincident with indexes of part 3 to ensure the correction of this experiment and this hypothesis.In all,the balance coordination between hippocampus and amygdala may be the key in transformation from adaption to damage in a state of stress,is one of the central mechanisms of LGSQF on regulating the stress,chronic binding stress effected the expressional tendency of BDNF in the hippocampus and amygdale.BDNF in the hippocampus in model group and sham operation group decreased,and increased after use Xiaoyao san,as soon as increased after microinjection CNQX.While in amygdale, the expressional tendency of BDNF decreased when chronic binding stress because BLA overexcited.Microinjection CNQX can inhibit BLA's overexcitation,so the expressional tendency of BDNF in basolateral amygdale didn't decreased significantly inverse increased.This finding revealed that CNQX may reduce stimulation intensity in rengions of hippocampus,which react treatment.This also explains that BDNF protection to nervous system is diminishing,which indicate that the loss of BDNF protection during stress is one of the key mechanism stress the protection of the central nervous system damage.It illustrate Xiaoyao san has a significant regulatory role to liver invading the spleen syndrome.This hypothesis may reveal the molecular mechanisms and regulation balance beteen hippocampus and amygdala on Xiaoyao san affording effective protection agaist Liver invading the spleen syndrome.