Alternative Treatments For Major Depressive Disorder And Investigation Of Cerebral Neural Circuits

Major depressive disorder(MDD)is a neuropsychiatric disorder characterized by persistent despondent state accompanied by low self-esteem and a loss of interest or pleasure in normally enjoyable activities(anhedonia).It is a highly prevalent,multifactorial disorder with high disability and mortality rates.The psychosocial difficulties lead to serious physical,mental and socioeconomic consequences,such as impairments in social communication and vocational functioning.Many sufferers do not live independently.Half of patients attempt suicide at least once and up to 20%ultimately commit suicide.Almost all of them need long-term medication and so research into pharmacotherapy of MDD has evolved over the centuries.Some powerful antidepressants have made indelible contributions,such as selective-serotonin reuptake inhibitor(SSRIs),tricyclic antidepressant(TCA),and monoamine oxidase(MAO)inhibitors.However,some of them are of variable effectiveness and some exert undesirable side effects.Therefore,the development of alternative antidepressants is still the ideal aim pursued by researchers.This research is focused on the treatment of depression,the exploration of the neural mechanisms and the related neural circuits.The contents are introduced as follows:the effect of Ziziphi spinosae lily powder suspension against depression disorder;the effect of vanillin alleviating depressive-behaviors by olfactory pathway;the development of tissue transparent techniques;combination of the tissue transparent technique and virus conjugated fluorescent tracer to explore the olfactory pathway related to emotion modulation.1)The effect of ziziphi spinosae lily powder suspension on depression-like behaviors in rats.The aim of the present study was to evaluate the behavioral anti-depressant effects of a novel herbal compounds named ziziphi spinosae lily powder suspension,as well as to investigate its potential mechanisms.40 male SD rats were used to establish animal depression model by chronic unpredictable mild stress(CUMS).At last,6 rats were excluded from the experiment because of intolerance or insensitive to the CUMS procedure.Then all animals were randomized into three groups with 8 rats per group:the fluoxetine group,the lily jujube group and the control(untreated)group.Body weight and depression like behaviors were measured at each time point:before modeling,after modeling and after treatment.Depression-like behaviors including forced swimming test(FST),sucrose consumption test(SCT)and open field test(OFT)were performed.Changes in these parameters reflected the behavioral characteristic phenotypes of depression in rats.Additionally,in order to elucidate the possible underlying mechanisms,a quantitative analysis of serum serotonin(5-HT)in peripheral blood and brain 5-hydroxyindoleacetic acid(5-HIAA)were performed using ultrahigh-performance liquid chromatography(UPLC)and high-performance liquid chromatography(HPLC),respectively.Results:after treatment for 4w,ziziphi spinosae lily powder suspension obviously increased body weight of the animals(P<0.01).In the forced swimming test,ziziphi spinosae lily powder suspension obviously decreased immobility time(P<0.01)while increased swimming time(P<0.01).In the sucrose consumption test,ziziphi spinosae lily powder suspension obviously increased the sucrose consumption of the animals(P<0.01).In the open field test,ziziphi spinosae lily powder suspension obviously increased rearing movement(P<0.05)and decreased the number of fecal granule(P<0.05).All of the data showed that the herbal compounds ziziphi spinosae lily suspension could alleviate depressive symptoms in rat model of chronic depression.The underlying mechanisms may be related to the increase of serum 5-HT in peripheral blood and 5-HIAA in brain.2)Vanillin-induced amelioration of depression-like behaviors in rats by olfactory pathway.The beneficial effects of vanillin on depressive model rats were investigated using a combination of behavioral assessments and neurotransmitter measurements..Two models of depressive disorder were used to assess the effects of vanillin in rats.The chronic depressive disorder model was induced by chronic unpredictable mild stress(CUMS),and the acute depressive disorder model was induced by olfactory bulbectomy which disrupted the olfactory pathway.40 male SD rats were used to establish animal depression model by chronic unpredictable mild stress(CUMS).At last,10 rats were excluded from the experiment because of intolerance or insensitive to the CUMS procedure.Then all animals were randomized into three groups with 10 rats per group:the vanillin group,the fluoxetine group and the control(untreated)group.And another 20 rats were used to establish the acute depressive disorder model by olfactory bulbectomy which disrupted the olfactory pathway:the olfactory bulbectomy+vanillin group(OBX)and the sham group(Sham).Before and after chronic stress condition(or olfactory bulbetomy),and at the end of vanillin treatment,body weight,immobility time in the forced swimming test and sucrose consumption in the sucrose consumption test were measured.The quantitative changes in these assessments revealed the characteristic phenotypes of depression in rats.Brian neurotransmitters were measured using ultrahigh-performance liquid chromatography to investigate the possible mechanisms.Results:vanillin could obviously increase body weight of the animals(P<0.01),decrease the immobility time in the forced swimming test(P<0.01),and increase the sucrose consumption in the sucrose consumption test(P<0.01).All of the data showed that vanillin can relieve the depressive-like behaviors via olfactory pathway.The underlying mechanisms may be related to the modulation of monoamine neurotransmitters in brain,such as 5-HT and DA.3)Development of tissue transparent techniques.The tissue transparent technique is a new technology developed in recent years.It builds a bridge between the whole organisms and the cell,enabling the observation of 3D structure and detailed information at micro scales through intact organisms.It is an excellent tool to explore the olfactory neural circuit.At present,two families of the techniques are identified:organic solvents and aqueous solutions.However,each of them presented one or more following weak points,such as time-consuming,tedious,fluorescence quenching,tissue deformation,pungent odor and specialized equipment.Results:During our investigation of tissue transparent process,there are three tissue transparent protocols optimized.Each of them has its merits and drawbacks.A tissue clearing technique contains neither fructose nor ammonia water.It renders a whole mouse brain transparent with preferable effect by long time immersion.A rapid tissue clearing technique contains both fructose and ammonia water.It renders an adult mouse brain transparent by immersion in the clearing reagents for 12-15h.The characteristic of the method is rapid,simple,no obvious tissue deformation and compatible with fluorescence.A tissue clearing technique contains fructose but not ammonia water.Without complicated pretreatment,it renders whole organs transparent by incubation in the clearing reagents.It is demonstrated to preserve organ morphology and cellular membrane lipids.It is compatible with both fluorescent signals and macromolecular.Combined the tissue transparent protocol and specific dyes or antibodies,whole-organ staining and whole-organ immunostaining can be achieved.4)Investigation of olfactory circuit on brain scales.Neuroanatomist Papez suggested that the limbic system was the neural circuit of emotion regulation,namely the Papez neural circuit.The circuit includes visible papillae,hypothalamus;anterior thalamus,cingulate cortex,amygdala,septal area,fornix.After integrated in olfactory bulb,olfactory information will be transferred to the piriform cortex,entorhinal cortex,and so on.Part of them overlaps with the emotional center.Observation of olfactory projection at whole brain scale will offer better understanding of the antidepressant effect of vanillin.In this study,we combined the tissue transparent technique with whole organ staining and the tract tracing by fluorescent virus to investigate the relation of olfactory center and the emotional center.Results:combination of the tissue transparent technique and the whole organ staining would distinctly show the layers of olfactory bulb,the direction and the communication of the olfactory fibers.For example,it was the anterior commissure that communicated two sides of the olfactory system.The direction and route of the posterior limb of the anterior commissure,the fibers of mammillary body were also distinct on whole brain scales.Combination of the tissue transparent technique and the tract tracing by fluorescent virus verified the olfactory fibers,olfactory projection areas and their communications,such as olfactory tubercle,entrohinal cortex,hippocampus,and amygdala,which provided morphological basis for emotion regulation via olfactory pathway.