Dopaminergic System Damage In Mice Induced By Developmental Exposure To Paraquat

Paraquat, a non-selective herbicide, is widely used around the world. In recent years, the effects of PQ on central nervous system increasingly attract attention of researchers. It has been reported that chronic exposure to paraquat can cause injury of the dopaminergic nigrostriatal system. But it is still unclear whether developmental exposure to PQ also causes permanent changes in dopaminergic system as well as behavioral deficits in the adult animals.The present study, using C57BL/6J mice, was aimed to explore whether subacute developmental exposure to PQ alone causes permanent nigrostriatal DA system neurotoxicity. We also aimed to investigate the neurobehavioral modifications and the selective degeneration of nigrostriatal neurons.80 male C57BL/6J mice with age of 21 days were randomly divided into five groups, i.e. Saline (the control group),1.25mg/kg/d,2.5mg/kg/d,5mg/kg/d, 10mg/kg/d Paraquat treated groups. The chemical was given by gavage in the volume 0.1mL/10g body weight for 30 days. Morris water maze was used for measuring neurobehavioral changes. After the Morris water maze, the step-through passive-avoidance test was performed to assess aversive learning and memory in mice. Toluidine blue and H&E staining were used to assess general histopathological alterations of midbrain. The expression of dopamine-related genes of nigrostriatal was quantified by using RT-PCR analysis. The expressions of apoptosis-related proteins in the midbrain were measured by Western blot.No significant changes were observed in the general appearance and the weight of mice in all PQ treated groups when compared to mice in the saline treated control group. In addition, data of organ coefficients indicated that PQ exposure had no significant adverse on the organ coefficients of brain.1. Neurobehavioral effects of PQ in miceMorris water maze was performed after repeated exposure to PQ to assess the spatial learning and memory in mice. No significant difference of latencies to reach the hidden platform was observed among control and PQ-treated groups. PQ-treated groups failed to show selective search in the training quadrant, as measured by number of platform crossings. In addition, exposure to PQ did not significantly affect the total swimming distance in the training quadrant as compared to saline-treated mice. In summary, there was no evidence of effects of PQ., 10mg/kg/d for 30 days, on spatial learning and memory in mice.After the Morris water maze, the step-through passive-avoidance test was performed to assess aversive learning and memory in mice. The latency to leave the illuminated compartment was recorded. When compared to saline-treated mice, the 10mg/kg PQ-treated mice exhibited a better behavioral response in passive-avoidance test.2. Histopathological and Ultrastructure examinationsToluidine blue and H&E staining were used to assess general histopathological alterations of midbrain. Light microscopic examinations showed that the number of cells of substantia nigra was significantly lower in 10mg/kg PQ-treated mice than that in control group. However, there were no significant changes of the number of cells in other PQ dose-level treated groups.Transmission electron microscopic examinations showed that the 10mg/kg PQ-treated mice had marked swelling of mitochondria with disintegration of their cristae. Neurons have shrunken nucleus with corrugated nuclear membrane, swollen endoplasmic reticulum, multiple cytoplasmic vesicles and lipid droplets, which revealed that the neurons are in a state of apoptosis.3. Effects of PQ on gene expression in striatum of miceThe administration of PQ produced a significant alteration of expression of DAT and VMAT2 genes in nigrostriatal, while there were no significant changes of expression of D2R gene. Meanwhile, significant decrease of TH was found in 5mg/kg/d, 10mg/kg/d Paraquat treated groups. In addition, there were significant increase of COMT in 2.5mg/kg/d,5mg/kg/d, 10mg/kg/d Paraquat treated groups, while there was no change in the level of MAOB gene in all PQ-treated groups.4. Effects of PQ on apoptosis-related proteins in striatum of miceWestern blot analysis revealed that Caspase 9 expression in PQ-treated mice was at very low level and had no significant difference compared with the control group. Meanwhile, there were no PQ-related effects on the expressions of BAX in the midbrain.ConclusionThe present study showed that developmental paraquat exposure could alert the expression of dopamine-related gene of nigrostriatal and number of cells of substantia nigra. However, there were no significant changes in other aspects, such as neurobehavior function and general condition. The results were considered as "silent neurotoxicity" induced by developmental paraquat exposure, which could markedly enhance vulnerability to subsequent pesticide treatments. The research laid a foundation for more extensive work on the development of neurotoxical mechanism as well as risk assessment of paraquat. In conclusion, the results provided the orientation for the further study of silent neurotoxicity induced by PQ. Further studies will be needed with lower dose of PQ to confirm these results and demonstrate the "silent neurotoxicity" of PQ.