Behavioral Effects Induced By Repetitive Transcranial Magnetic Stimulation In CUMS Animal Model Through Regulation Of Cannabinoid Receptor Type1

Objectives:This study was designed to explore whether alterations in cannabinoid type1receptor(CB1receptor) in the hippocampus might contribute to the antidepressant-like effectsinduced by rTMS treatment. We examined this question in a chronic unpredicted mildstress (CUMS) rat model of depression.Methods:In this study, all the experiments were performed in animals, and were divided intotwo parts. In the first part, we proposed to observe the behavioral effects and the change ofthe expression of hippocampal CB1receptor in CUMS model rats induced by rTMStreatment. In the second part, we intended to find out if AM251, a CB1receptor antagonist,can reverse the rTMS-induced antidepressant-like effects and hippocampusneuroprotective and neurogenesis.For the first part, the rats were randomly divided into four groups:(1) Sham group,the rats were kept in their home cages for the3-week period, and then subjected to Sham rTMS treatments for7days;(2) Sham+rTMS group, the rats were kept in their homecages for the3-week period, and then subjected to real rTMS treatments for7days;(3)CUMS group, the rats were subjected to CUMS for three weeks and then to sham rTMStreatments for7days;(4) CUMS+rTMS group, the rats were subjected to CUMS forthree weeks and then to real rTMS treatments for7days.For the second part, the rats were randomly divided into six groups:(1) Sham group,the rats were kept in their home cages for the3-week period, and then subjected to shamrTMS treatments for7days;(2) CUMS group, the rats were subjected to CUMS for threeweeks and then to sham rTMS treatments for7days;(3) CUMS+rTMS group, the ratswere subjected to CUMS for three weeks and then to real rTMS treatments for7days;(4)CUMS+rTMS+AM251group, the rats were administered with CUMS and AM251andreal rTMS treatments for7days. AM251was given30minutes before the o nset of rTMStreatment at1mg/kg intraperitoneally;(5) CUMS+AM251group, the rats wereadministered with CUMS and AM251and sham rTMS treatments for7days. AM251wasgiven30minutes before the onset of sham rTMS at1mg/kg intraperitoneally;(6) CUMS+rTMS+Vehicle group, the rats were administered with CUMS and0.6ml/kg Vehicle(DMSO: saline=1:4) and real rTMS treatments for7days.Behavioral testing (sucrose preference test, forced swimming test, open feld test)were used to assess the effects of CUMS and rTMS treatment; Western blot andimmunocytochemistry were used to measure the expressions of hippocampal CB1receptorand related neuroprotection and neurogenesis factors.Results：In the first part of study, CUMS paradigm induced alterations in depression-relatedbehaviors and the expression of hippocampal CB1receptor in rats; while rTMS treatmentinduced improvement of depressive behaviour and the expression of hippocampal CB1receptor in CUMS rats. The detail of the results was written as follow:1. Compared with Sham group,(1) there was significant decrease in sucrosepreference in depressive animal induced by CUMS (P <0.05);(2) CUMS group showed asignificant increase in the immobility time(P <0.01);(3) in open-field test, CUMS group showed significantly less horizontal distance (P <0.01) and less number of times enteredthe center field (P <0.01);(4) in CUMS group, less expression of CB1receptor wasobserved throughout the hippocampus (P <0.01).2. Compared with CUMS group,(1) treatment with rTMS significantly increasedsucrose preference (P <0.05);(2) rTMS treatment signifcantly reduced the immobilitytime (P <0.05);(3) in open-field test, administration of rTMS treatment induced anincrease in the total horizontal distance traveled of the open-field (P <0.05) and in thenumber of times entered the center field (P <0.05);(4) the expression of CB1receptor inhippocampus was significantly increased after the rTMS treatment (P <0.01).In the second part of the study, we found that the antidepressive-like behavioursinduced by rTMS was reversed by AM251, rTMS treatment promoted the hippocampalneurogenesis and neuroprotection in CUMS model rats, while AM251could attenuatedthe effects induced by rTMS treatment. The detail of the results was written as follow:1. rTMS treatment significantly improved the depression-associated behaviorscompared with that of the CUMS group (for instance: the sucrose preference, theimmobility time, the total horizontal distance traveled of the open-field and the number ofvisits to center field of the open-field, P <0.05). The CB1receptor antagonist AM251reversed the beneficial effect of rTMS treatment given30minutes before the onset ofrTMS treatment (for instance: the sucrose preference, the immobility time, the totalhorizontal distance traveled of the open-field and the number of visits to center field of theopen-field, P <0.05.(CUMS+rTMS+AM251vs. CUMS+rTMS). The depressive-likebehaviours of the CUMS+rTMS+vehicle group was similar to that of the CUMS+rTMS group (P>0.05). There were no statistical differences in the depressive-likebehaviours among CUMS, CUMS+rTMS+AM251and CUMS+AM251groups (P>0.05).2.(1) Relative to CUMS group, rTMS treatment signifcantly increased the numberof BrdU-positive cells in the dentate gyrus (P <0.01); increased the BDNF protein levelof the hippocampus (P <0.01); increased the Bcl-2protein level (P <0.05) and decreasedBax proteins (P <0.05);(2) Compared with CUMS+rTMS group, AM251administered before rTMS treatment decreased the number of BrdU-positive cells in the dentate gyrus(P <0.01); the BDNF protein levels were signifcantly decreased when administeredAM25130minutes before the onset of rTMS treatment; Both Bcl-2and Bax protein ofCUMS+rTMS group were reversed when administered AM25130minutes before theonset of rTMS treatment (Bcl-2, P <0.01; Bax, P <0.05);(3) There were no statisticaldifferences in the number of BrdU-positive cells, the expression level of BDNF and Bcl-2family protein between CUMS+rTMS group and CUMS+rTMS+vehicle group, andamong CUMS, CUMS+rTMS+AM251and CUMS+AM251groups.Conclusion：CUMS paradigm induced alterations in depression-related behaviors in rats like thedepression, and rTMS treatment could induced improvement of depressive behaviour inCUMS rats, while the antidepressive-like behaviours induced by rTMS was reversed byAM251, CB1receptor antagonist. These results indicated that rTMS treatment hasup-regulated the expression of CB1receptors, which promoted hippocampusneuroprotection and neurogenesis against depressive behaviour, suggesting a newmechanism of rTMS treatment to depressive illness.