| Objective:To elucidate the intervention effects of Jiao-Tai-Wan(JTW)on p-chlorophenylalanine(PCPA)-induced insomnia in rats and the underlying mechanismMethod:Male SD rats were randomly normal group,model group and JTW group.The JTW group rats were given JTW water extraction by oral administration once a day and a total of five days,normal group and model group were given the same volume of saline.The experiment on the third day,the insomnia model was established by intraperitoneal injection of PCPA methyl ester hydrochloride(564 mg/kg,equivalent to 450 mg/kg)to the model group and the JTW group rats once,the normal group was given the same volume of saline by intraperitoneal injection.The locomotor activity of rats was observed and a proton nuclear magnetic resonance(~1H-NMR)based metabonomic approach was developed to profile insomnia-related metabolic perturbations in rat urine,serum and hippocampus.Orthogonal partial least-squares discriminant analysis was carried out on the metabolites,the variable importance projection and t-test were used to screen metabolites which significant difference between the normal group and model group.Moreover,the levels of neuroprotective factors such as brain-derived neurotrophic factor(BDNF),nerve growth factor(NGF);apoptotic factors such as B-cell lymphoma-2(Bcl-2),Bcl-2-Associated X(Bax);neurotransmitter such as glutamate(Glu),cholecystokinin 8(CCK-8);cytokines such as interleukin(IL)-2,IL-6 and glial fibrillary acidic protein(GFAP)in hypothalamus,hippocampus,prefrontal cortex and serum of rats were determined by using enzyme-linked immunosorbent assay(ELISA).Bradford method was used to measure the level of protein in tissue sample.Result:1.The JTW could significantly suppress the locomotor activity of insomnia model rats.2.(1)The levels of n-acetyl glycoprotein,acetone,pyruvic acid,succinic acid,alpha ketone glutaric acid,dimethylamine were remarkably reduced(P<0.05),dimethyl glycine,taurine,phenylacetyl glycine were remarkably increased in urine of insomnia rats in comparison with the normal group(P<0.05),compared with the model group,the levels of n-acetyl glycoprotein,pyruvic acid,succinic acid,alpha ketone glutaric acid,dimethylamine were remarkably increased(P<0.05),dimethyl glycine,taurine,phenylacetyl glycine were remarkably reduced in urine of insomnia rats by administration of JTW(P<0.05);(2)The levels of LDL and VLDL,unsaturated fatty acid,isoleucine,n-acetyl glycoprotein,glutamic acid,o-choline acetyl glycoprotein,choline,phosphoric acid,trimethylamine oxide were remarkably increased in serum of insomnia rats in comparison with the normal group(P<0.05),compared with the model group,the levels of LDL and VLDL,isoleucine,n-acetyl glycoprotein,glutamic acid,choline and choline phosphate were remarkably reduced in serum of insomnia rats by administration of JTW(P<0.05);(3)The levels of alanine,gamma-aminobutyric acid,inositol were remarkably reduced(P<0.05),n-acetyl aspartate,creatine/creatine phosphate,glycerin phosphoric acid choline,taurine were remarkably increased in hippocampus of insomnia rats in comparison with the normal group(P<0.05),compared with the model group,the levels of alanine,gamma-aminobutyric acid,inositol were remarkably increased(P<0.05),glycerol phosphate choline,taurine were remarkably reduced in serum of insomnia rats by administration of JTW(P<0.05).3.The levels of IL-2,IL-6,BDNF,NGF,CCK-8 in hypothalamus,hippocampal,prefrontal cortex and serum of rats with insomnia were remarkably reduced(P<0.05)meanwhile the levels of GFAP,Glu were significantly increased(P<0.05),We found the ratio of Bcl-2/Bax in hypothalamus,hippocampal and prefrontal cortex of model rats were remarkably reduced(P<0.05),but the ratio of Bcl-2/Bax in serum had no significant change of model group rats compared with the normal group.Compared with the model group,The levels of IL-2,IL-6,BDNF,NGF,CCK-8 in hypothalamus,hippocampal,prefrontal cortex and serum of rats with insomnia were remarkably increased(P<0.05),meanwhile the levels of GFAP,Glu were significantly reduced by administration of JTW(P<0.05).In addition,JTW could significantly increase the ratio of Bcl-2/Bax in hypothalamus,hippocampal and prefrontal cortex(P<0.05),but no obviously difference was found with the ratio of Bcl-2/Bax in serum compared with the insomnia model rats.Conclusion:1.The JTW could significantly suppress the locomotor activity of insomnia model rats.2.The current study demonstrated that the disturbed metabolic state associated with PCPA-induced insomnia in rat serum,hippocampus and urine could be restored by JTW,which was probably mediated metabolism of lipid,energy,amino acid,cholin and intestinal flora.3.The JTW exerted its intervention effects against PCPA-induced insomnia in rats,which was probably mediated by regulating the levels of nerve protection factor such as BDNF and NGF;apoptosis factors such as Bcl-2,Bax;neurotransmitter such as Glu,CCK-8;cytokines such as IL-2,IL-6 as well as suppressing the hyperplasia of astrocyte. |
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