The Research Of The Antidepressant Effects Of Acupuncture Treatment By Regulating The Expression Of Synaptic Relative Proteins And AMPA Receptors In Hippocampi Of Depression Model Rats

ObjectiveAMPA receptors is a post-synaptic excitatory neuron,which is also the target for both the important structure of the physiological and pathological and the antidepressant therapy of depression.In this study,we observed the antidepressant effects of acupuncture therapy for CUMS rats.We use behavioral experiments,western blot and immunofluorescence technique to observed the expression of synaptic relative proteins and AMPA receptors.Finally we discussed the mechanism of the antidepressant effects of acupuncture treatment in "glutamate-AMPA" synaptic trafficking.Experiment 1The anti-depressive effects of acupuncture therapy in rats induced by chronic unpredictable mild stress combined solitary feeding model:a viewpoint of behavior alterations.MethodsMale Sprague-Dawley（SD）rats（180-220g）from the SanYuanLi Experimental Animal Center of GuangZhou University of Chinese Medicine were kept under controlled environmental conditions（25℃,12h light/dark cycles,food and water freely available）.Before experimental manipulation,the animals were given a period of 1 week to adjust to the new surroundings,singly housed in perspex cages（14 x 14 x 24cm）.30 rats were randomly divided into 5 groups（n=6）:（1）Group A,the rats didn’ t received any manipulation except common feeding;（2）Group B,the rats received a consecutive 42 days of CUMS;（3）Group C,after undergoing a consecutive 21 days of CUMS,the rats didn’t received any manipulation except common feeding;（4）Group D,after undergoing a consecutive 21 days of CUMS,the rats received acupuncture treatment for three consecutive weeks;（5）Group E,the rats received a consecutive 42 days of CUMS,meanwhile received acupuncture treatment for three consecutive weeks beginning at the 22st day.The details of CUMS and acupuncture treatment are as follow.The SPT was performed three times,before the first beginning of experiment（day0）,the 21th day（day21）and 42th day（day42）the experiment,respectively.The open filed test was performed three times,the first day（dayl）,the 22th day（day22）,the 41th day（day）,respectively.The elevated plus maze test was performed three times,before the first beginning of experiment（day0）,the 20th day（day20）,the 40th day（day40）,respectively.The passive avoidance test was performed three times,the 2nd day（day2）,the 21th day（day21）,the 42th day（day42）,respectively.Result1.For the sucrose preference test（figure.1 A）,at the beginning of experimental procedure,there was no significant difference among the groups according to the physical state scores(F_4,25= 0.089,p>0.05).At the 21th day,stressed rats,including group B,group C,group D and group E,showed a significant decreasing of the state scores compared with group A(F_4,25=10.246,p<0.01).According to the 42th day(F_4,25=24.578,p<0.01),after CUMS for 42 days,group B showed a significant decreasing sucrose preference compared with group A（P<0.01）,which was also lower than group C（P<0.01）and group E（P<0.01）.Without CUMS since the 22th day,although the sucrose preference of group C showed an increasing at the 42th day than before,it still had a significant difference with group A（P<0.01）.After receiving 21 days acupuncture treatment,the sucrose preference of group D was improved obviously and showed no physical state difference with group A（P<0.01）,which was also higher than group B（P<0.01）,group C（P<0.01）and group E（P<0.01）.This test had a statistically significant in our experiment(F_4,25=16.303,p<0.01).2.According to the open filed test（figure.1 B and C）,at the first day,the crossings and rearings of rats showed no significant difference among the five groups(F（4,2,5）=0.025,p>0.05;F_4,25= 0.026,p>0.05;).At the 22th day,stressed rats,including group B,group C,group D and group E,showed a significant decreasing of the state scores each compared with group A（p<0.01）.At the 42th day(F_4,25= 5.35,p<0.01;F_4,25= 5.34,p<0.01),group D was close to group A,and this two group were higher than group B and group C（p<0.01;p<0.01）.Group E was higher than group B（p<0.01;p<0.01）.3.For the elevated plus maze test（figure.1D）,there was no statistically significant difference at the beginning of experimental procedure in our study F（4.25）=0.085,p>0.05).After CUMS for 20 days,the groups of stressed rats showed a significant decreasing of the state scores compared with group A(F_4,25= 10.246,p<0.01).At the 21th day,stressed rats,including group B,group C,group D and group E,showed a significant decreasing of the state scores compared with group A.When to the 40th day(F_4,25= 6.179,p<0.01),the ratio of time spent on the open arms to the time spent on the closed of group A was higher than any other groups(F_4,25= 34.946,p<0.01).However,the data showed no statistically significant difference between group A and group D（P>0.05）,group B and group C（P>0.05）,group D and group E（P>0.05）.The ratio of group D was also higher than group B（P<0.05）,group C（P>0.05）.4.For the passive avoidance test（figure.1E）,the latency of the rats showed no statistically significant difference at the 2nd day(F_4,25= 0.186,p<0.01).At the 21th day,the group A showed statistically significant difference with other stressed groups(F_4,25= 40.829,p<0.01).According to the 42th day,there had statistically significant difference among groups(F_4,25= 45.675,p<0.01).Group A was lower than any other groups（P<0.05）.Group D was lower than group B（P<0.01）,group C（P<0.01）,group E（P<0.01）.Group E showed statistically difference with group B（P<0.01）and group C（P<0.05）.Group B was higher than group C（P<0,01）.Experiment 2The research of the antidepressant effects of acupuncture treatrment by regulating the expression of synaptic relative proteins and AMPA receptors in hippocampi of rats induced by chronic unpredictable mild stress combined solitary feeding model.Methods30 male Sprague-Dawley（SD）rats（180-220g）from the SanYuanLi Experimental Animal Center of GuangZhou University of Chinese Medicinerats were randomly divided into 5 groups（n=6）:（1）Group A,the rats didn’ t received any manipulation except common feeding;（2）Group B,the rats received a consecutive 42 days of CUMS;（3）Group C,after undergoing a consecutive 21 days of CUMS,the rats received acupuncture treatment for three consecutive weeks;（4）Group D,after undergoing a consecutive 21 days of CUMS,the rats received Sham acupuncture treatment for three consecutive weeks;（5）Group E,after undergoing a consecutive 21 days of CUMS,the rats received acupuncture treatment for three consecutive weeks,and rats were injected NBQX into the paracele half an hour before the acupuncture treatment.We use western blot and immunofluorescence technique to observed the expression of synaptic relative proteins including CaMKⅡ、Stargazin、PSD-95、PICK1、p-GLUR1、p-GLUR2、GLUR1、GLUR2 and AMPA receptors,GluR1、GluR2.ResultAccording to the results of western blot experiments,we can analysis that compared with group A,the content of Strgazin showed an significant decreasing in group B and group E（P=0.000,P=0.000）.The content of Strgazin showed no significant changes compared with group A in group C（P=0.498）.Compared with group A,the content of Strgazin showed an significant increasing in group D（P=0.01）.Group B compared with group E,group D compared with group E,separately,showed no significant difference（P=0.061,P=0.227）.Compared with group A,the content of p-GLUR1 showed an significant decreasing in group B and group D and group E（P=0.000,P=0.00,P=0.000）.The content of p-GLUR1 showed no significant changes compared with group A in group C（P=0.056）.Compared with group B,the content of p-GLUR1 showed an significant increasing in group C and group D（P=0.000,P=0.01）.Group B compared with group E,showed no significant difference（P=0.235）.Group D compared with group E,showed significant difference（P=0.025）.Compared with group A,the content of p-GLUR2 showed an significant decreasing in group B and group D and group E（P=0.000,P=0.000,P=0.000）.The content of p-GLUR2 showed no significant changes compared with group A in group C（P=0.077）.Compared with group B,the content of p-GLUR2 showed an significant increasing in group C and group D（P=0.000,P=0.000）.Group B compared with group E,showed no significant difference（P=0.091）.Group D compared with group E,showed significant difference（P=0.012）.Compared with group A,the content of p-Camk Ⅱ showed an significant decreasing in group B and group D and group E（P=0.000,P=0.000,P=0.000）.The content of p-Camk Ⅱ showed no significant changes compared with group A in group C（P=P=0.332）.Compared with group B,the content of p-GLUR2 showed an significant increasing in group C and group D（P=0.000,P=0.20）.Group B compared with group E,showed no significant difference（P=0.210）.Group D compared with group E,showed no significant difference（P=0.242）.Compared with group A,the content of Pickl showed an significant decreasing in group B and group D and group C and group E（P=0.000,P=0.027,P=0.000,P=0.000）.Compared with group B,the content of Pickl showed an significant increasing in group C and group D（P=0.000,P=0.000）.Group B compared with group E,showed no significant difference（P=0.064）.Group D compared with group E,showed no significant difference（P=0.20）.Group C compared with group D,showed significant difference（P=0.002）.Compared with group A,the content of PSD95 showed an significant decreasing in group B and group D and group C and group E（P=0.000,P=0.008,P=0.000,P=0.000）.Compared with group B,the content of PSD95 showed an significant increasing in group C and group D（P=0.000,P=0.000）.Group B compared with group E,showed no significant difference（P=0.368）.Group C compared with group D,showed no significant difference（P=0.062）.Group D compared with group E,showed significant difference（P=0.001）.Compared with group A,the content of GLUR1 showed an significant decreasing in group B and group D and group C and group E（P=0.000,P=0.001,P=0.000,P=0.000）_Compared with group B,the content of GLUR1 showed an significant increasing in group C and group D（P=0.000,P=0.02）.Group B compared with group E,showed no significant difference（P=0.118）.Group C compared with group D,showed no significant difference（0.063）.Group D compared with group E,showed no significant difference（P=0.084）.Group C compared with group E,showed significant difference（P=0.001）.Compared with group A,the content of GLUR2 showed an significant decreasing in group B and group E（P=0.000,P=0.000）.Compared with group A,the content of GLUR2 showed no significant decreasing in group C and group D（P=0.671,P=0.087）.Compared with group B,the content of GLUR2 showed an significant increasing in group C and group D（P=0.000,P=0.02）.Group B compared with group E,showed no significant difference（P=0.382）.Group C compared with group D,showed no significant difference（P=0.189）.Group D compared with group E,showed no significant difference（P=0.001）.Group C compared with group E,showed significant difference（P=0.016）.According to the results of immunofluorescence technique,we can analysis that the expression of synaptic relative proteins AMPA receptors GluRl showed obviously in group A and group C,while GluR2 in group B and Group E.ConclusionOn the basis of Experiment 1 and Experiment 2 we can conclude that:（1）Acupuncture therapy can effectly repair the chronic unpredictable mild stress combined solitary feeding depression model rats’ behavior alterations.（2）Acupuncture therapy can effectly increase the expression of synaptic relative proteins including CaMKII、Stargazin、PSD-95、PICK1、p-GLUR1、p-GLUR2、GLUR1、GLUR2 in hippocampi of rats induced by chronic unpredictable mild stress combined solitary feeding model.（3）Acupuncture therapy can effectly increase the expression of synaptic AMPA receptors GluRl and GluR2 in hippocampi of rats induced by chronic unpredictable mild stress combined solitary feeding model.