Research On The Antidepressant Effects And Mechanisms Of Radix Polygalae

Depression is a common psychiatric disorder,characterized by anhedonia,loss of interest and the dysfunction in sleep and cognition,which impaired severely the physical and psychological health of the patients.Although there are many drugs for the treatment of depression,the low response rate and the obvious side effects of current antidepressants limited their applications in depression.Therefore,it is an urgent need to develop the effective medicine with less adverse effects.Radix Polygalae(RP)is the dried root of Polygala tenuifolia Willd.or Polygala sibirica L.,wildly used as a traditional Chinese herb medicine with excellent protective effect on central nervous system and could improve several neuropsychiatric diseases.The pathological mechanism of depression is far from clear for the complexity and heterogeneity of this disorder,and recently developed metabolomics has given strong support for researching the pathological basis of depression and the mechanism of typical antidepressants.Olfactory bulbectomy(OBX)-induced rat is an animal model applied early in depression,while changes in metabolites of OBX rats have not been revealed.Though the antidepressant action of RP has been explored preliminarily,researches in the antidepressant effect and mechanism of RP are not comprehensive.The present study firstly investigated the changes in the urine and hippocampus metabolite profiles based on metabolomics and the dysfuction related to energy regulation and autophagy in OBX rats.In addition,the antidepressant efficacy of RP was evaluated in behavior despair mice model,olfactory bulbectomy(OBX)rat model and chronic restraint stress(CRS)rat model.The antidepressant mechanism of RP was explored by several molecular biological techniques.The main contents of the present research are as follows:1.Study on the metabolite profiles and autophagy status of OBX ratsOBX model of rat was established by the surgery removal of bilateral olfactory bulbs.After a 14-day recovery,OBX rats were treated with fluoxetine(10 mg/kg)by gavage for another 14 days,and then the 24-h urine of rat was collected,followed with behavioral tests,including open field test,elevated plus maze(EPM)test,forced swimming test(FST),hyperemotionality test and Morris water maze(MWM)test.After these,the rats were sacrificed,and the prefrontal cortex(PFC)and hippocampus were harvested.The levels of metabolites related to tryptophan metabolism and neurotransmitters in the PFC were quantified by liquid chromatography-mass spectrometry.The metabolite profiles of urine and hippocampus were analyzed by UPLC-QTOF-MS-based metabolomics,and the protein expression involved in AMPK-mTOR pathway and autophagy were measured by Western blot in the hippocampus and PFC.As a result,OBX induced the significant hyperactivity and hyperemotionality in rats,and OBX rats displayed prolonged immobility time in FST and the impaired learn and memory ability in MWM test.Fluoxetine could alleviate the depressive-like behaviors,but had no effects on the deficiency in learn and memory of OBX rats.OBX caused marked increase in the levels of tryptophan,5-hydroxy indoleacetic acid(5-HIAA)and dopamine,and significant decrease in the levels of kynurenine and 5-hydroxytryptamine(5-HT)in the PFC of rats.Among of them,the abnormal changes of kynurenine,5-HT and 5-HIAA could be reversed by fluoxetine.The metabolite profiles of urine and hippocampus in OBX rats separated obviously from the Sham rats.Twenty-six and 16 potential biomarkers were identified in urine and hippocampus,respectively,which involved in the disturbance in amino acid metabolism,energy metabolism,gut bacteria metabolism,purine metabolism and ascorbate and aldaric acid metabolism.Fluoxetine could reverse partly the abnormal changes of metabolites in OBX rats.In addition,OBX caused the dysfunction in the AMPK-mTOR pathway(a key energy-sensing signal)and the inhibition of autophagy in the hippocampus and PFC of OBX rats,which could be normalized by fluoxetine treatment.2.Study on the antidepressant effects of RPSeveral behavioral tests were performed to measure the antidepressant activity of RP in the behavioral despair mice,OBX rats and CRS rats.RP(0.5 and 1 g/kg)treatment for 14 days remarkably decreased the immobility time of mice in FST.Twenty-eight-day treatment of RP(0.5 and 1 g/kg)could reverse the depressive-like behaviors of rats induced by OBX,displayed by the reduction of the open arm entry ratio and the movement distance in EPM test,the inhibition of the hyperlocomotion and the hyperemotionality,and decreasd immobility time in FST.In the depression model of rat induced by CRS,RP(0.5 and 1 g/kg)increased the index of sucrose preference and the movement distance of CRS rats in open field test,and decreased the feeding latency in novelty-suppressed feeding test and the immobility time in FST,but could’t ameliorate the reduced body weight and the anxiety-like behaviors of CRS rats in EPM test.3 Study on the antidepressant mechanism of RPSeveral molecular biological methods were used to uncover the antidepressant mechanisms of RP,including Western blot,immunofluorescence,transmission electron microscope and real-time quantitative polymerase chain reaction.As a consequence,RP(0.5 and 1 g/kg)could elevate the autophagy level in the cerebral cortex of behavioral despair mice and the PFC of OBX-and CRS-induced rats,displayed by increasing the protein expressions of LC3-II and beclinl,and decreasing the level of p62.The disturbed AMPK-mTOR pathway in the PFC of rats caused by OBX or CRS could be normalized by RP,and RP inhibited the excessive phosphorylation of Akt,mTOR and ULK1 and enhanced the expression of p-AMPK.Furthermore,the activated microglia and NLRP3 inflammasome in the PFC of OBX and CRS rats could be prevented by RP treatment.And the increased mRNA levels of several proinflammatory factors(NLRP3,IL-1 p,IL-6,IL-18 and TNF-α)in the PFC of OBX and CRS rats were significantly reduced by RP.In addition,RP could relieve the damage of astrocyte and neuron,and increase the protein levels of GFAP,NeuN and PSD 95.In conclusion,metabolomics study uncovered that OBX induced significantly changes in the urine and hippocampus metabolite profiles of rats,and identified biomarkers might be helpful to understand the pathological mechanism of depression and for the clinical diagnosis of depression.Meanwhile,we found that RP could relieve the depressive-like behaviors in behavior despair model,OBX model and CRS model of depression,and RP could correct the dysfunction in AMPK-mTOR pathway,enhance autophagy level,alleviate the neuro-inflammatory response and strengthen the neuroplasticity to exert powerful antidepressant potency.