The Effect Of Doses Subthreshold For Overt Systemic Toxicity Of Chlorpyrifos On Mitochondrial Of Substantia Nigra In Newborn Rats

Our research group has found that the newborn SD rats were exposed in sub-threshold dose of chlorpyrifos, the microglia cells were activated until the rats'puberty, and dopaminergic neurons were injured in adolescence and adulthood. Many studies have found that the activated microglia cells would lead to the injury of dopaminergic neurons. Above all studies suggest us that in neonatal period of SD rats who were exposed in sub-threshold dose of chlorpyrifos may have potential risk factors in the injury of dopaminergic neurons. Our research group has also found that newborn SD rats exposed in sub-threshold dose of chlorpyrifos would have depression of SOD in brain homogenate, overexpression of MDA and brain protein carboxide, these tell us the oxidative stress of brain tissue maybe take part in Neurotoxic effect of chlorpyrifos.Oxidative stress is due to the overexpression of highly reactive molecules, such as ROS, which cannot be gotten rid of entirely. ROS are produced from mitochondria.When mitochondria are injured, ROS will be increased, but the overexpression of ROS also is harmfull to mitochondria itself.Objective To reserch the effect on mitochondria of substantial nigra in neonatal SD rats who were exposed in sub-threshold dose of chlorpyrifos (Chlorpyrifos, CPF).Methods There are 85 SD rats in their 9-10 days, were randomly assigned into groups of chlorpyrifos(group CPF, n=29) and control (n=56). The control groups were divided into the solvent DMSO control (DMSO group, n=29) and saline control (group NS, n=27). According to methods of our research group to establish animal model of neonatal SD rats exposed in sub-threshold dose of Clorpyrifos. After injected, animals were put to death at 24h,72h,6d.①To detect the expression of mitochondrial respiratory chain complex I subunit 1 (NADH dehydrogenase subunit 1,ND1) in substantial nigra by immunohistochemistry and Western Blotting.②To detect the expression of cytochrome C (Cytochrome C,Cyt C)by immunohistochemistry in substantia nigra by immunohistochemistry;③To observe mitochondrial ultrastructure in substantia nigra by electron microscopy though uranyl acetate, lead nitrate double staining;④To observe mitochondrial ultrastructure in substantia nigra by electron microscopy though immunohistochemical staining of tyrosine hydroxylase chlorpyrifos. All data are expressed as mean±standard deviation, and processed with SPSS18.0.Results1. The expression of mitochondrial respiratory chain complexⅠsubunits ND1 is changed, expression of ND1 in group chlorpyrifos is significantly increased, the difference was statistically significant (P<0.05); Between time points of the group chlorpyrifos,expression of ND1 is no significant difference, the difference is not statistically significant (P> 0.05).2.The expression of Cyt C mitochondrial respiratory chain is changed, at 72h,6d of group chlorpyrifos,the expression of Cyt C is increased significantly (P<0.05); At each time point of group chlorpyrifos, the expression of Cyt C is increased gradually, the difference was statistically significant (P<0.05).3. In electron microscope of uranyl acetate,lead nitrate double staining, dopaminergic neurons of group chlorpyrifos are edema, content of cytoplasmic are rare, and mitochondrial in opaminergic neurons are edema, vacuolation; But dopaminergic neurons of group dimethyl sulfoxide are not edema, content of cytoplasm are rich, there are more mitochondria in opaminergic neurons, and morphology of mitochondria are normal.4. In electron microscope of TH immunohistochemistry, the mitochondrial in positive cells (dopaminergic neurons) of group chlorpyrifos are edema, vacuolation, fewer in number; mitochondria of the group DMSO are normal, no edema and vacuolation and in larger quantities.Conclusion 1. Sub-threshold dose of chlorpyrifos can cause the expression of mitochondrial respiratory chain complexⅠsubunit 1 and cytochrome C in substantia nigra of neoborn SD rats increased. 2. Sub-threshold dose of chlorpyrifos can cause mitochondrial in substantia nigra dopaminergic neurons of neoborn SD rats edema, vacuolation, fewer in number.3. The change of mitochondrial structure and function maybe mechanisms of developmental toxicity of chlorpyrifos.