| Objectives Fenvalerate,one of the commonly used pyrethroid pesticides,is considered to be a potential environmental pollutant and endocrine disruptor with neurotoxicity.5-HT can regulate cell proliferation and differentiation as well as neuronal migration and synaptogenesis,and is considered to be a key signaling molecule during neural development.However,whether fenvalerate can affect 5-HT levels during embryonic development and have long-term effects on offspring neurobehavior remains unclear.Therefore,the purpose of this study was to investigate whether fenvalerate exposure during pregnancy leads to depression-like behavior in offspring,and to explore its effect on placental and brain-derived 5-HT synthesis and its mechanism.By establishing a model of fenvalerate exposure during pregnancy,this study explored the effects of 5-HT synthesis levels from different sources and provided a new explanation mechanism for its induced neurotoxicity.MethodsAn animal model of depression in offspring induced by fenvalerate exposure during pregnancy was established.In this study,20 mg/kg fenvalerate(1/10 of the LD50 of fenvalerate)was used as the maximum dose,and 2mg/kg,1/10 of the high dose,was selected as the low dose.Clean-grade 7-week-old ICR mice(male: 34-40g;Female: 25-29g)were purchased from Zhejiang Charles River.After one week of adaptive feeding,the mice were mated with the male mice at a ratio of 2:1.The Day of detection of vaginal plug was defined as Gestation Day 0(GD0).Pregnant mice were separated into three groups according to body weight S-type(control group,low-dose group,high-dose group).The control group received daily oral administration of corn oil,while the fenvalerate group was orally administered fenvalerate at doses of 2mg/kg and 20mg/kg,respectively.Body weight and food consumption were recorded.Five pregnant mice in each group were sacrificed on GD14 to obtain placenta and forebrain.The placenta,prefrontal cortex and midbrainObjectives Fenvalerate,one of the commonly used pyrethroid pesticides,is considered to be a potential environmental pollutant and endocrine disruptor with neurotoxicity.5-HT can regulate cell proliferation and differentiation as well as neuronal migration and synaptogenesis,and is considered to be a key signaling molecule during neural development.However,whether fenvalerate can affect 5-HT levels during embryonic development and have long-term effects on offspring neurobehavior remains unclear.Therefore,the purpose of this study was to investigate whether fenvalerate exposure during pregnancy leads to depression-like behavior in offspring,and to explore its effect on placental and brain-derived 5-HT synthesis and its mechanism.By establishing a model of fenvalerate exposure during pregnancy,this study explored the effects of 5-HT synthesis levels from different sources and provided a new explanation mechanism for its induced neurotoxicity.Methods An animal model of depression in offspring induced by fenvalerate exposure during pregnancy was established.In this study,20 mg/kg fenvalerate(1/10 of the LD50 of fenvalerate)was used as the maximum dose,and 2mg/kg,1/10 of the high dose,was selected as the low dose.Clean-grade 7-week-old ICR mice(male: 34-40g;Female: 25-29g)were purchased from Zhejiang Charles River.After one week of adaptive feeding,the mice were mated with the male mice at a ratio of 2:1.The Day of detection of vaginal plug was defined as Gestation Day 0(GD0).Pregnant mice were separated into three groups according to body weight S-type(control group,low-dose group,high-dose group).The control group received daily oral administration of corn oil,while the fenvalerate group was orally administered fenvalerate at doses of 2mg/kg and 20mg/kg,respectively.Body weight and food consumption were recorded.Five pregnant mice in each group were sacrificed on GD14 to obtain placenta and forebrain.The placenta,prefrontal cortex and midbrain were harvested at GD18.The remaining pregnant mice were delivered normally to produce offspring,the first Postnatal day was determined as PND1(postnatal day1),and each litter was adjusted to four females and four males within 12 hours after birth.One female and one male were randomly selected from each litter on PND60 for behavioral test,and the remaining offspring were sacrificed on PND70.The brains of the mice were immediately removed and fixed with paraformaldehyde and embedded in paraffin for immunohistochemical tests.Other brain samples were isolated from prefrontal tissue collected in separate cryotubes,snap-frozen in liquid nitrogen and stored at-80°C for further studies.All experiments were approved by the Society for Laboratory Animal Science,Anhui Medical University.ResultsBody weight and food intake of pregnant rats in the three groups did not differ significantly.However,pregnant rats in the high-dose group had a lower body weight than those in the control group,according to pairwise comparison using the least significant difference(LSD)test.Additionally,the body weight of adult male and female offspring in the high-dose group was lower than that of the control group.Depression-like behavior of adult offspring was evaluated using the sugar-water preference test,forced swimming test,and tail suspension test.The sucrose preference index of the high-dose group was significantly lower than that of the control group in the sucrose preference test.In the forced swimming test,male rats in the high-dose group had a higher immobility time than the control group.In the tail suspension test,both male and female mice in the high-dose group had a higher immobility time than those in the control group.The high-dose fenvalerate group had a significant reduction in PSD95 protein levels in adult male and female mice compared to the control group,but the Synapsin-1 protein level did not change.Immunolocalization of5-HT in placenta sections at GD14 showed that the positive cells were mainly located in the labyrinth layer,and the cytoplasm of cytotrophoblasts with large round nuclei and syncytiotrophoblasts with small oval nuclei were slightly stained.Quantitative analysis showed that there was no difference in 5-HT content between male and female placentas compared with the control group.Immunolocalization and quantification of 5-HT in GD18 placentas were consistent with the GD14 results.There was no significant difference in brain-derived 5-HT content in the forebrain at GD14 among all groups.Compared with the control group,the content of 5-HT in the PFC of GD18 fetal rats was decreased,and the difference in the content of 5-HT in the PFC of male fetal rats was statistically significant.In addition,the level of TPH2 protein was significantly downregulated in the fenvalerate exposed midbrain,although both showed a trend toward increased m RNA expression levels.In order to explore the mechanism of the down-regulation of TPH2 protein level in fetal rat brain induced by fenvalerate exposure,we used Western blot to detect the protein levels of p-e IF2α and GRP78 related to endoplasmic reticulum oxidative stress in fetal rat brain exposed to fenvalerate during pregnancy.The results showed that p-e IF2α and GRP78 protein levels were up-regulated in the fenvalerate exposure group compared with the control group.ConclusionFenvalerate exposure during pregnancy can increase depression-like behavior in offspring mice by down-regulating brain-derived 5-HT synthesis,resulting in synaptic damage in prefrontal cortex.The down-regulation of brain-derived 5-HT synthesis may be caused by down-regulating the expression of TPH2 protein by inducing endoplasmic reticulum stress. |
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