Basolateral Amygdala Serotonin 2C Receptor Regulates Emotional Disorder Related Symptoms Induced By Chronic Methamphetamine Administration

Objective:Globally,methamphetamine（METH）is the second most abused drug,with psychotic symptoms being one of the most common adverse effects.Emotional disorders induced by METH abuse have been widely reported both in human and animal models;however,the mechanisms underlying such disorders have not yet been fully elucidated.This topic will use the mice METH chronic poisoning model to explore the molecular mechanism of emotional symptoms caused by long-term METH abuse.Methods:（1）Establish a METH chronic model in mice,intraperitoneal of METH solution（10 mg/kg body weight/times,1 time/day,continuous injection of 4 weeks）,forced swimming test,FST;tail suspension test,TST;sucrose preference test SPT and elevated plus maze,EPM.（2）Animal samples were collected:①the levels of TPH2 in the dorsal raphe nucleus（DRN）and 5-HT_2CR in the basolateral amygdala（BLA）were detected by immunofluorescence assay;②The levels of TPH2 mRNA in DRN and 5-HT_1AR、5-HT_2AR、5-HT2CR、5-HT₃R、5-HT4R mRNA in BLA were detected by qPCR.③Western blot was used to detect the levels of TPH2 in DRN and 5-HT2cR in BLA.④The levels of 5-HT and its metabolites in mice BLA were detected by HPLC.（3）Pharmacological methods were used to interven the activity of 5-HT2AR and 5-HT2cR.Three behavioral experiments,FST,SPT and EPM,were carried out after METH chronic poisoning model was established in mice.（4）Synthetic RNA laden virus down-regulates the expression level of 5-HT2cR.Three behavioral experiments,FST,SPT and EPM,were carried out after METH chronic poisoning model was established in mice.（5）Collecting animal samples:①The injection site of adeno-associated virus in BLA and the co-localization of 5-HT2cR and GABA neurons in amygdala were detected by immunofluorescence method;②The protein level of 5-HT2CR in mice BLA was detected by Western blot.③The micro inhibitory postsynaptic current（mIPSC）and action potential（AP）in the amygdala of mice were recorded by patch clamp.Results:（1）The immobility time of mice in FST and TST experiments increased after METH chronic poisoning.Decreased preference for sugared water;the time and frequency of entering the open arms of the elevated cross maze has been reduced.（2）The expression level of TPH2 in DRN decreased after METH chronic poisoning in mice.The levels of 5-HT and its metabolites in BLA decreased.The 5-HT1AR level decreased,the 5-HT2AR and 5-HT2cR levels increased,and the 5-HT3R and 5-HT4R levels did not change significantly.（3）Compared with METH mice without antagonist,the immobile time of mice in FST test decreased after 5-HT2cR antagonist injection.A return preference to sugared water;The time and frequency of entering the open arms of the elevated cross maze has increased.Compared with METH mice without antagonist,there was no significant change in the behavior of mice after 5-HT2AR antagonist injection.（4）Compared with METH mice without interference gene,the immobile time of mice in FST experiment decreased after the down-regulation of 5-HT2cR expression;A return preference to sugared water;The time and frequency of entering the open arms of the elevated cross maze has been increased.（5）After interfering with the expression of 5-HT2cR gene,glandular associated green fluorescence was observed in the amygdala region of mice.The expression level of 5-HT2CR was significantly decreased;The 5-HT2CR of BLA co-locates with GAD67.The frequency of mIPSC in BLA increased but the amplitude did not change significantly,and the number of AP decreased.Conclusion:The chronic effect of METH can reduce the expression of TPH2 in DRN,reduce the level of 5-HT transmitter released by DRN into BLA through neurons,up-regulate the expression of 5-HT2CR in BLA,thus inhibiting the inhibitory effect of GABAergic neurons on pyramidal neurons,enhancing the excitability of BLA,and finally leading to the occurrence of dysthymic disorder symptoms.