Establishment Of Chlorpyrifos-induced Animal Model Of Major Depression

Currently, considerable efforts are being made to improve the treatment andprevention of psychiatric disorders. Nevertheless, among these efforts, thepharmacological treatment still remains the most crucial one. Therefore, more andmore investigators realize the importance of understanding the following issues insearching for a better drugs of these disorders: the etiology of diseases, themechanism of action of currently available drugs, and the novel treatment targets andconcepts. It is beyond dispute that animal models could fulfill all these significantaspects.Depression, a psychiatric disease which characterized primarily by depressedmood and loss of interest or pleasure, is a major cause of disability with a lifetimeprevalence of about 15-20%. According to the World Health Organization, depressionis to reach second place as leading contributor to the global burden of disease by theyear 2020. Over the course of the past decades of years, animal models of depressionhave been largely developed owing to the constantly emerged etiological theories ofdepression, thus promoting the discovery and development of various effective drugs.However, the occurrences of false positives of negatives in these models arouse greatconcerns and controversies about the relevance and validities of animal models ofdepression, probably due to overemphasizing single endophenotypic of depressionrather than modeling the whole syndromes. Therefore, establishment of novel animalmodel of depression that comprising multiple endophenotypics of depression is ofgreat significance.Increasing evident has shown potential neurotoxic risks of usingorganophosphorothioate (OP) pesticide, especially on developing individuals.Chlorpyrifos (CPF), a OP pesticides, remains one of the most extensively usedinsecticides worldwide, due to its effective, cost-competitive broad spectrum ofactivity when compared with other alternative products. Despite of its agricultural usage being curtailed in United States and European Union in 2001, it is still served asan highly effective weapon against insects in agriculture and industry circumstancesamong the world. However, the association between emotional disorders and CPFexposure in humans is still a major environmental health issue, especially in children,a subgroup that highly susceptible to environmental hazards.The epidemiological studies on CPF shed partial light onto the possibilities ofmodeling depression symptom in rodents, thus we choose this chemical to inducebehavioural measures (including anhedonia-, anxiety- and depression- relatedparameters) that can be reflect relatively comprehensive depression-related symptoms,and we hypothesize that repeated exposure to CPF may induce depression behavioralalterations via its effect on neurotransmitter release, cytokine levels, signallingtransduction, as well as HPA axis function.Face, construct, and predictive validity were chosen to evaluate this model. In theface validity evaluation, we observed dramatic depression-related alterations invarious animal test of depression, manifested by increased behavioral despair state inforced swimming test, increased escape deficit in learned helplessness test, lost ofpleasure in anhedonia test, and increased approach-avoidance conflict innovelty-suppressed feeding test, while no evident effect on locomotor activity inopen-field activity test was observed.In the construct validity evaluation, several generally acknowledgedpathogeneses of major depression were chosen to explain these depression-relatedbehavioral alterations:⑴Monoamine hypothesis of major depression: levels of NE, DA, 5-HT and theirmetabolites were examined by HPLC-ECD assay. Our present results indicate thatthese neurotransmitter systems are affected dramatically by repeated exposure to CPF.In hippocampus, ANOVA analysis detected a significant decline of 5-HIAA (p<0.03),while NE and 5-HT not, however, in partial doses of CPF, an evident decrease effecton 5-HT was observed. In prefrontal cortex, both doses of CPF decrease the contentof 5-HT (p<0.001) and NE (p=0.050), while significantly increase that of 5-HIAA(p<0.008). In striatum, extraordinarily evident alteration of monoamineneurotransmitter and their metabolites was also observed. ANOVA analysis indicateda significant effect of CPF treatment on NE, DOPAC, DA, and 5-HIAA (p<0.0001, 0.003, 0.02, 0.008, respectively). Besides, the elevation of MAO-A and MAO-B invarious brain section as we observed indicated that CPF's effect on MAO maycontribute to the disturbance of neurotransmitters.⑵Cytokine hypothesis of major depression: alterations in levels ofpro-inflammatory cytokines (TNF-α, IL-1β, IL-6) and anti-inflammatory cytokineIL-4 in various brain sections were observed by ELISA. In hippocampus, ANOVAanalysis detected a elevated pro-inflammatory cytokines (TNF-α, IL-1β, IL-6) andreduced anti-inflammatory cytokine IL-4, while in prefrontal cortex and striatum, ageneral down-regulation effect on these cytokines were observed.⑶HPA axis hypothesis of major depression: three key hormones (CRF, ACTH,and CORT) in HPA axis were chosen to investigate the effect of repeated exposure tochlorpyrifos on HPA axis function. We observed an activation of HPA axis,manifested by elevation of levels of ACTH (p<0.05) and CORT (p<0.01) in serum.⑷Wnt signaling hypothesis of major depression: we primarily examineGKS-3β's regulation effect in this pathway as well as its related effectors, Wnt2 andβ-catenin, the upstream signaling molecules that regulating this kinase as well as itsdownstream targets. Thus brain levels of phosphorylated proteins of GSK-3βandβ-catenin, Wnt2, GSK-3β, andβ-catenin were determined using immunoblot assay.ANOVA analysis did not detected a significant effect of CPF on Wnt2, GSK-3βandβ-catenin, however, an extraordinarily elevation of phosphorylation of GSK-3βwasobserved, indicating another noncholinergic mechamisms that may contribute to thecellular response to neurotoxicity of CPF.In addition, we found AChE activity in prefrontal cortex, hippocampus, andstriatum were well below the 70 % threshold necessary for symptoms of cholinergichyperstimulation, excluding the possibilities of involvement of any cholinergicintoxication.Results from the above four generally acknowledged pathogeneses of majordepression, we confirm that the established animal model of major depression inducedby repeated exposure of CPF meet the standard of construct validity.In the predictive validity ecaluation, we used two kinds of anti-depressants,desipramine (DMI) and fluoxetine (FLU) to reverse the depressive-like behavioralalterations in forced-swimming test and novelty-suppressed feeding test, and found that our model is sensitive to chronic administration of DMI and FLU.In conclusion, by repeated exposure to CPF, we successfully induceddepressive-like behavioral alterations in adolescent male rats, and these ratsmanifested characteristics that consistent with what observed in clinical depressionpatients, moreover, these depressive-like behaviral can be reversed by chronicadministration of two kinds of anti-depressants including DMI and FLU. Therefore,this model can meet both face, construct, and predictive validity, indicating itpossibility as a potential animal model of major depression.