Effect Of Mouse Prenatal Chlorpyrifos On Interkinetic Nuclear Migration Of Neural Pregenitors And Hippocampus Cells

Recent studies reported the effect of prenatal Chlorpyrifos (CPF) on cell proliferation and differentiation of developmental neural progenitors, the mechanism is still unclear. Interkinetic nuclear migration(INM) is the migration of apical progenitors (APs) during cell proliferation cycle along the cortex apical-basal polarity. It is a hallmark event of neurogenesis and the basis of brain evlolution. The hippocampus is a key region involving learning, memory and behavior, the morphological structure changes are related with cognitive behavior anomaly. This study evaluates the effects of prenatal exposure CPF on INM and the embryo hippocampal organization.We first established the model of pregnant rat. The gestational mice was injected sub cutaneous ly with5mg/kg CPF on embryo days14(E14) or equivalent volume of dimethyl sulfoxide. The injections were administered every3h for total3times. At0.5,3.5or5.5h of first CPF exposure, animals were received5-bromo-2’-deoxyuridine (Brdu,50mg/kg) intraperitoneally, thus the S phase cells were labeled with Brdu, and formed marked embryo at different times. The animals were sacrificated by cervical dislocation at6.5h of the first exposure of CPF. The embryos were stripped, fixed and paraffin embeded, then stained with Brdu immunohistochemical assay. The Brdu labeled cells in ventricle area (ventricular Zone, VZ) and subvertricular Zone (SVZ) were calculated under a microscope, the percentage of labeling cells in each layer of VZ/SVZ areas were statisticalated, and the effects of instant exposure CPF to INM were evaluated.Next the gestational female mice were exposure to5mg/kg/d of CPF during embryo days7.5-11.5. E14animals were given Brdu (50mg/kg) intraperitoneally, at1,3,6or9hour of Brdu injection, the mice were sacrificated and the embryos were prepared as the above mentioned. The slices were observed under a microscope to determine whether CPF treatment cause delay effects on the INM.In order to assess the effects of prenatal CPF exposure on cytoarchitecture of the hippocampus, gestational mice were exposed to CPF through E7.5-11.5, the morphological samples were perfomed on E16. The embryos were prepared with continuous coronary slices and stained with nissl’s staining.3standard hippocampal slices were selected, the total cell number in hippocampus were calculated and analysed.The results of instant exposure CPF represented that CPF exposure slow the migration of Brdu-labeled nuclei toward VZ surface.1hr after Brdu administration, the positive cell nucleus increased8.3%in outer area of VZ, but reduced40.1%in the middle area of VZ (P<0.01);3hr after Brdu administration, the positive cell nucleus increased10.6%in outer area of VZ, increased61.1%in the middle area of VZ, but reduced82.0%in the inner area of VZ (P<0.01);6hr after Brdu administration, the positive cell nucleus increased25.1%in outer area of VZ, increased39.3%in the middle area of VZ, but reduced22.2%in inner area of VZ(P <0.01)The results of short-term exposure CPF represented that CPF elicited delay effects on the INM, which mean the toxicity effects did not recover after cessation of CPF exposure.1hr after Brdu administration, the positive cell nucleus increased6.5%in outer area of VZ, but reduced22.4%in the middle area of VZ (P<0.01);3hr after Brdu administration, the positive cell nucleus increased39.8%in middle area of VZ, but reduced25.9%in the inner area of VZ (P<0.01);6hr after Brdu administration, the positive cell nucleus increased15.1%in middle area of VZ, but reduced11.7%in inner area of VZ (P<0.01);9hr after Brdu administration, the positive cell nucleus decreased12.2%in outer area of VZ (P>0.05), reduced22.5%in the middle area of VZ (P<0.01), no differences in inner area of VZ, and increased95.3%in SVZ area (P<0.01)The morphological studies showed that the total cell number in hippocampus CA1and CA3sub-region did not have significant differences between CPF and control groups, but the total cell number in DG sub-region reduced in CPF group, mean reduced8.06%. Thus significantly difference was observed between control grou and CPF group in DG sub-region.In summary, these research results proved that prenatal CPF exposure in low concentration can cause subtle damage in mice VZ neural prenitors and embryo hippocampus architecture.