| Background and ObjectiveDepression is a common clinical emotional mental disorder with high prevalence,high recurrence rate,high suicide rate and it is difficult to cure.At present,the pathological mechanism of depression is still unclear,and there is a lack of effective treatment in the clinic.The JNK signaling pathway is a member of the mitogen-activated protein kinases(MAPK)family and is called c-jun amino terminal kinase(JNK).JNK plays an important role in cell growth,differentiation,apoptosis,stress response and the occurrence and development of various human diseases.Previous experimental studies have found that stress-induced activation of JNK signaling pathway is involved in the development and progression of depression.Acupuncture intervention can reduce the phosphorylation level of JNK,inhibit neuronal apoptosis,and improve depression-like behavior in rats.Based on the previous research,this study will establish a chronic stress model in rats by combining CUMS with chronic orphans,and inject JNK signaling pathway blocker SP600125 into the lateral ventricle to perform electroacupuncture intervention,with body weight,open field text and sucrose intake test.Evaluation of changes in behavioral indicators in rats;The difference of c-fos and hypothalamic c-jun,pituitary c-fos and serum c-fos and AP-1 expression in hippocampal CA3 area after electroacupuncture The expression of c-jun,c-fos and AP-1 is used as a symbol of the functions of hippocampus,hypothalamus and pituitary neurons.The purpose of this study is to investigate the changes of hippocampal,hypothalamic and pituitary neurons in chronic stress depression rats.Further explore the mechanism of acupuncture anti-depression effect,and provide a new scientific basis for the clinical application of acupuncture.Methods1 Experimental groupsA group of 64 male rats were randomly divided into eight groups:Blank control group(control group),model group,model+DMSO group(DMSO group),model+SP600125(SP group).Model + electroacupuncture group(electroacupuncture group),model +electroacupuncture+SP600125(electroacupuncture+SP group),model+fluoxetine group(fluoxetine group),model+fluoxetine+SP600125(Fluoxetine+SP group)2Modeling methodsIn addition to the control group,the other rats used the internationally accepted CUMS modeling method:seven unpredictable mild stimuli were applied.(Shock3。min,night light 12h,binding 2h,sandwich 3min,no food 24h,no water 24h,wet cushion 24h)continuous stimulus 21 days,Each stimulus is used three times on average.3 Intervention methodsElectroacupuncture intervention is carried out "Nei guan"(PC6)and "San yin jiao"(SP6)with no.30 one-inch needle with the depth of 0.3-0.5cm,and a handle connected to the HANS LH-202 electrometer with a frequency of 2hz and a current intensity of 0.6mA of 20 min,one time a day,lasts 21 days.Fluoxetine is mixed with distilled water into 0.2 mg/ml,and is fed with 1.8ml/kg,one time a day,lasts 21 days.4 Process methodsThe control group did not give any stimulus and was normally raised in groups.Each group was raised in a single cage and received chronic antibiotics for 21 days.At the same time,The electroacupuncture group interacts 30 minutes before the mold is made every day;Fluoxetine was given to the group 30 minutes before the stress every day;The DMSO group was given 10μl/Id DMSO side injection at 1h before stimulated;The SP group was given an injection of 10μl/1d block inhibitor SP600125 diluent liquid side vanadium at 1h prior to thorium.The electroacupuncture+SP group,the fluoxetine+sp group were given 10μl/I d block at 1h before stimulated.The inhibitor SP600125 dilution liquid side injection was given to the electroacupuncture intervention and the fluoxetine lavage 30 min before stimulated.5 Detection methodsBehavioral testing:Body weight,open field test and sucrose preference test were measured 1 day before the experiment and on the 21st day of the experiment to evaluate the behavioral changes of the model rats.Index detection:The expression of c-fos in hippocampal CA3 region was detected by immunohistochemistry.The expression of c-jun,pituitary c-fos,serum c-fos and AP-1 in hypothalamus were detected by ELISA.Result1.Behavioural changeOne day before the experiment,there was no statistically significant difference in body weight,open field test and sucrose preference test between the groups(P>0.05).On the 21st day of the experiment,compared with the control group,the body weight,open field test and sucrose preference test of the model group were significantly reduced(P<0.01),suggesting successful modeling;compared with the model group,the electroacupuncture group body weight,open field test and sucrose preference test increased significantly(P<0.01),the fluoxetine group increased the crossing level(P<0.05),and the body weight increased significantly(P<0.01),indicating electro acupuncture and fluoxetine.Intervention can improve the depression-like behavior of rats with depression,and the effect of electroacupuncture intervention is better than that of fluoxetine.There is no statistically significant difference between DMSO group,open field test and sucrose preference test(P>0.05),indicating lateral ventricle.The implantation of cannula had little effect on the behavior of rats;there was no significant difference in body weight and open field test between the operation group(DMSO group,SP group,electroacupuncture + SP group,fluoxetine + SP group)(P>0.05);Compared with the electroacupuncture group,the electroacupuncture + SP group body weight was significantly reduced(P<0.01),the open field test experiment was reduced(P<0.05),the sucrose preference test was not significantly different(P>0.05);Compared with the fluoxetine group,the body weight of the fluoxetine+SP group was significantly reduced(P<0.01),and there was no significant difference between the open field test experiment and the sucrose preference test(P>0.05).2.Expression level of molecules downstream of JNK signaling pathwayAfter 21 days of experiment,compared with the control group,the expression of c-fos,hypothalamic c-jun,pituitary c-fos,serum c-fos and AP-1 in the hippocampal CA3 area of the model group increased significantly(P<0.01).It indicated that chronic stress stimulation can activate the expression level of JNK-related downstream molecules;compared with the model group,the expression of c-fos in hippocampal CA3region of electroacupuncture group decreased(P<0.05),hypothalamic c-jun,pituitary c-fos,serum The expression of c-fos and AP-1 was significantly decreased(P<0.01).The expression of c-fos and pituitary c-fos in hippocampal CA3region of fluoxetine group decreased(P<0.05),hypothalamic c-jun,serum c-fos and The expression of AP-1 was significantly decreased(P<0.01),indicating that electroacupuncture and fluoxetine could reverse the expression of JNK-related downstream molecules to some extent.There was no significant difference in the expression of DMSO group(P>0.05).Compared with the DMSO group,the expression of c-jun and serum AP-1 in the hypothalamus of the SP group decreased(P<0.05),and the expression levels of the indicators in the electroacupuncture + SP group decreased significantly(P<0.01)· The expression of c-jun,serum c-fos and AP-1 in the hypothalamic group of fluoxetine + SP group was significantly decreased(P<0.01).Compared with the electroacupuncture group,there was no significant difference in the expression of the electroacupuncture+ SP group(P>0.05).Compared with fluoxetine group,there was no significant difference in c-fos,hypothalamic c-jun,pituitary c-fos and serum AP-1 expression in hippocampal CA3 area of fluoxetine+SP group(P>0.05),serum c-fos expression decreased(P<0.05).Conclusion1.Chronic mild unpredictable stress stimulation combined with orphaned resulted in abnormal behaviors such as weight loss,loss of pleasure and decreased behavior of the rats in the model rats.2.Electroacupuncture intervention can improve the depression-like behavior in rats,which is directly related to the down-regulation of c-fos,hypothalamic c-jun,pituitary c-fos and serum c-fos and AP-1 activities in hippocampal CA3area.3.Blocker SP600125 has an intervention effect on the expression of c-jun,c-fos and AP-1 in chronic stress depression rats;it has different effects on behavioral indicators.4.The electroacupuncture+SP group had a significant effect on the expression of c-fos in the chronic stress depression model than the fluoxetine + SP group.The blocking agent SP600125 and the electroacupuncture intervention method had a superposition effect on the inhibition of c-fos phosphorylation level.More synergistic.5.The effect of fluoxetine on the peripheral serum of chronic stress depression model was more obvious than that of electroacupuncture. |
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