基于GABA能神经系统探究交泰丸对斑马鱼睡眠剥夺模型神经递质及受体的影响OA

Objective To observe the effects of Jiaotai Pills on related transmitters and receptors in the γ-aminobutyric acid(GABA)nervous system of sleep deprivation(SD)zebrafish.Methods 240 6-month-old wild-type AB line zebrafish were randomly divided into blank group,model group,Jiaotai Pill low-dose group(0.16 mg/mL),and Jiaotai Pill medium-dose group(0.47 mg/mL).Jiaotai Pill high-dose group(1.41 mg/mL),melatonin group(0.2 mg/mL).LED light(150 lux)light induction was used for 3 days to establish a zebrafish sleep deprivation model,and the administration group was in the model group.On top of that,drug treatment is added for 3 days.After 3 days,the zebrafish behavior tracking system was used to monitor the 24 h movement of zebrafish.T-maze was used to detect the learning and memory ability of zebrafish.HE staining method observes the morphological changes of zebrafish brain neurons.ELISA was used to determine contents of GABA,glutamic acid(Glu),and 5-hydroxytryptamine(5-HT)in brain tissue.Immunohistochemistry(IHC)method was used to detect the content of glutamic acid decarboxylase 67(GAD67)in brain tissue.Real-time polymerase chain reaction(Real-time PCR)and Western blot method were used to detect mRNA and protein expression levels of gamma-aminobutyric acid A receptor subtype α1(GABAARα1),metabotropic glutamate receptor 7(mGluR7),serotonin 1A receptor(5-HT1AR),GAD67,and γ-aminobutyric acid transporter isoform-1(GAT-1)in zebrafish brain tissue.Results Compared with the blank group,in the 24 h behavioral observation of the model group,the resting state count decreased significantly(P<0.01),the latency to first enter the nutrient-rich area(EC area)increased significantly(P<0.01).The morphology of brain neurons was irregular,the staining was fuzzy and uneven,the cytoplasm was more transparent and the staining was shallow.The GABA,5-HT content and the positive expression of GAD67 decreased significantly(P<0.01).The mRNA and protein expression of Glu and its receptor mGluR7 increased significantly(P<0.01).mRNA and protein expression levels of GABAARα1,5-HT1A,GAT-1 and GAD67 decreased significantly(P<0.01).Compared with model group,the content of Glu protein in the low dose group of Jiaotai Pill decreased significantly(P<0.01),while there was no significant difference in other above-mentioned indexes(P>0.05).The number of resting state in middle and high dose groups of Jiaotai Pill and melatonin group increased significantly(P<0.05 or P<0.01).The incubation period of first entry into the EC area decreased significantly(P<0.01).The morphology and cytoplasm transparency of neurons have been improved,and the damage has been alleviated,and the contents of GABA,5-HT and the positive expression of GAD67 increased significantly(P<0.05 or P<0.01).Glu content and its receptor mGluR7 mRNA relative expression level and protein expression level decreased significantly(P<0.01).mRNA and protein expression levels of GABAARα1,5-HT1A,GAT-1 and GAD67 increased significantly(P<0.05 or P<0.01).Conclusion Jiaotai Pill can regulate the neurotransmitter related to GABA in the brain of zebrafish with insomnia,inhibit the synthesis of Glu neurotransmitter,balance the levels of GABA and Glu,and then exert the sedative and hypnotic effect.