Studies On The Antidepressant-like Effect And Mechanisms Of Flavonoid Extracted From Apocynum Venetum Leaves

Depression is a prevalent psychiatric disorder which affects 21% of the world population. The symptoms of depression are intense feelings of sadness, loss of interest or pleasure, hopelessness, and despair, disturbed in sleeppatterns and appetite, as well as the inability to experience pleasure in usual activities, as its worst, can lead to suicide thoughts. The World Health Organization (WHO) report has predicted that major depression will become the second cause of illness induced disability by the year 2020. Consequently, more and more researchers have paid attention to pathogenesy and therapy of depression, however, the mechanism of antidepressants is still far from clearly understood. Indeed, studies have been proposed as possible mechanisms that depression is closely related to monoamine neurotransmitters, neurotrophic factors, signal transduction and other factors. The majority of Antidepressant drugs currently used has lead too much concern shortage, such as narrow-spectrum antidepressant, drug resistance, side effect and so on. From this perspective, we need to development new type of antidepressant drugs, while the naturally medications have higher antidepressant curative effect and fewer adverse effect. Apocynum venetum (AV) L. (Apocynaceae) is a traditional drug has been the subject of intensive research in China. In the present study, we extracted the major active component of Apocynum venetum-flavonoid, investigated its antidepressant effect, and corresponding mechanisms based on antidepressant-like effect.The present study will prescribe clinical application to provide the certain theories and experiments according to develop new natural mediciations.1. Evaluated the antidepressant-like effect of AV-extractIn this study, the classical animal models of depression including the tail suspention and force swimming of the mouse, and open-field test were applied to evaluate the antidepressant-like effect of AV-extract.The results showed that AV-extract showed a clear antidepressant-like effect in variety of animal models of depression.2. Influence of AV-extract on monoamine neurotransmitters and its metabolites in rat brain tissueThe LC-MS/MS detection was applied to investigate the variation of norepinephrine (NE), 5-hydroxytryptamine (5-HT), dopamine (DA) and its metabolites 5-hydroxyindole-3-acetic acid (5-HIAA),3,4-dihydroxyphenylacetic acid (DOPAC), and homovanillic acid (HVA) in rats brain tissue. The results showed that the concentration of NE,DA,HVA,DOPAC,5-HT,5-HIAA in depression rats'brain were degraded compared with the control group. While the content of NE,DA,HVA,DOPAC in AV-extract group were higher than depression group, the results were similar to fluoxetine group. However, the AV-extract have no effect on 5-HT/5-HIAA contrast to fluoxetine which can increased levels of 5-HT and/or 5-HIAA in rat brain regions.These results confirmed the neurotransmitter hypothesis of depression, as well as we found that AV-extract colud reversed the variation of NE,DA,HVA,DOPAC in rat brain tissue.3. Involvement of monoaminergic systems in the antidepressant-like effect of AV-extractTo further search for the neurotransmitter mechanism of antidepression of AV-extract, additionally, the monoaminergic mechanisms involved in the antidepressant-like effect of AV-extract in mice were also assessed. The anti-immobility effect of AV-extract (50 mg/kg, p.o.) was completely prevented by the pretreatment of mice with ketanserin (5 mg/kg, i.g., a serotonin 5-HT2A receptor antagonist), cyproheptadine (3 mg/kg, i.g., a serotonin 5-HT2 receptor antagonist), prazosin(1 mg/kg, i.g., anα1-adrenoceptor antagonist), yohimbine (1 mg/kg, i.g., anα2-adrenoceptor antagonist) or propranolol (2 mg/kg, i.g.,αβ-adrenoceptor antagonist) SCH23390 (0.05 mg/kg, i.g.. a dopamine D1 receptor antagonist) or sulpiride (50 mg/kg, i.g., a dopamine D2 receptor antagonist). On the other hand, the pretreatment of mice with WAY 100635 (0.1 mg/kg, s.c., a serotonin 5-HT1A receptor antagonist), did not block the antidepressant-like effect of AV-extract in the TST. Taken together, the data demonstrated that AV-extract produced an antidepressant-like effect that seems to be dependent on its interaction with the serotonergic (5-HT2A and 5-HT2 receptors), noradrenergic (α1-adrenoceptor, a2-adrenoceptor, aβ-adrenoceptor) and dopaminergic (D1 and D2 receptors) systems.4. The mechanism of neuroprotective effect of AV-extractNeuroprotective effect of AV extract was studied in corticosterone-induced neurotoxicity, using PC 12 cells as a suitable vitro model of depression to illustrate elementarily the pharmacologieal mechanism of antidepressant activity of AV extract. Cell viability was quantitated by 3-(4,5-Dimethylthiazol-2-y1)-2,5-diphenyltetrazolium bromide (MTT) assay. The release amount of lactic dehydrogenase (LDH) and intracellular Ca2+concentration was measured using kit, cell period change was tested by flow cytometry. The results showed that AV extract (25, 50 and 100μg/mL) increased the survival rate of PC 12 cells, especially in 100mg/kg arrived to 78%, but decreased LDH release and Ca2+concentration significantly (P<0.01), S period of the cells reduced significantly with the corresponding corticosteron-treated PC 12 cells. These results suggest AV extract could generate a neuroprotective effect on corticosterone-induced neurotoxicity in PC 12 cells, pointing to a possible action pathway of AV extract in vivo by decreasing decreased LDH release and Ca2+concentration to promote cell proliferation.5. Study on the expression of antidepressant effect associate-gene CREB and BDNF of AV-extractTaqman quantitative testing technique was used to test the expression of the BDNF and CREB of PC 12 cells in the presence of corticosterone (0.01 mmol·L-1) and flavonoid extract from Apocynum venetum leaves (25,50 and 100μg/ml) for 48 h. The result showed that the expression level of BDNF,CREB was reduced significantly after treatmented by corticosterone, but was elevated significantly by Apocynum venetum treatment which was up-regulated approximately 10 times corresponding to corticosterone treated. The result showed that the possible mechanism of Apocynum venetum maybe related to elevation the expression of BDNF,CREB through the signal path AC-cAMP-CREB.In conclusion, the study presented herein on the basis of evaluating the antidepressant effect of AV-extract demonstrated that the possible mechanism of these effect may be related to reduce the concentration of NE,DA,HVA,DOPAC, and for the first time the study investigated the antidepressant-like effect of AV-extract mediated by an interaction with the serotonergic (5-HT2A and 5-HT2 receptors), noradrenergic (α1-adrenoceptor,α2-adrenoceptor, aβ-adrenoceptor) and dopaminergic (D1 and D2 receptors)systems. Further works clarified that the antidepressant mechanism of neuroprotection may be achieved by decreasing LDH release and Ca2+ concentration to promote cell proliferation, and then speculated that the possible mechanism of Apocynum venetum antidepressant maybe related to elevating the expression of BDNF,CREB through the signal path AC-cAMP-CREB. The results presented here would be helpful to further study the pathogenesis of depression, and also provided a new idea for the use of natural anti-depressant medication to cure depression.