孕期亚硝酸盐暴露对子鼠视皮质突触改变的影响

Synapse loss in visual cortex of mouse after prenatal nitrite exposure

目的建立亚硝酸盐暴露模型,探讨孕期亚硝酸盐暴露对子鼠视皮质突触数量的影响及其改变机制。方法采用免疫荧光染色法观察对照组和模型组(低剂量组与高剂量组)子鼠出生后0d(P0)、P7、P14、P30的的视皮质突触素(SYN)和γ突触核蛋白(γ-synuclein)的改变,利用Western blotting和水迷宫实验对SYN蛋白的表达进行进一步验证。结果横向比较(剂量)发现,亚硝酸盐暴露组SYN减少(P<0.05)而γ-synuclein阳性细胞增多(P<0.05),且与亚硝酸盐存在剂量依赖关系(P<0.05)。纵向比较(年龄)发现,P0、P7、P14、P30 SYN表达随着年龄增长而增多(P<0.05),γ-synuclein表达减少(P<0.05),剂量(MS=2.00,F=55.22,P<0.05)与年龄(MS=1.11,F=30.68,P<0.05)之间存在交互作用且剂量的影响作用更大。Western blotting和水迷宫实验结果与免疫荧光统计结果一致。结论孕期亚硝酸盐暴露会影响子代视皮质突触的发育,减少突触数量,且对其影响与亚硝酸盐暴露剂量存在依赖关系,而γ突触核蛋白的过度表达可能是突触丢失的主要原因。

Objective To understand the nitrite＇ s toxicity to the quantitative alternations of synapses and explore its possible mechanism in mouse visual cortex using prenatal nitrite exposure（PNE） models. Methods The pregnant dams were exposure to nitrite from E0 （embryonic day 0） to parturition and divided into control group, moderate dose group （60mg/kg） and high dose groups （120mg/kg）. Pups with different treatments at P0, P7, P14 and P30 were harvested. Synaptophysin expression and γ-synucein expression in visual cortex were visualized with immunocytochemistry technique and Western blotting. Synaptophysin positive puncta and γ-synuclein positive neurons were measured, and their correlation was tested with statistical analysis. Results The pups＇ learning and memory reduced after nitrite exposure. The synaptophysin positive buttons reduced after prenatal nitrate exposure with dose dependency （ P 〈 0.05 ） , and the γ- synuclein positive cells increased with dose dependency （ P 〈 0. 05） as well. There was negative correlation between synaptophysin expression and γ-synuclein expression. Western blotting supported the results from immunofluorescent labeling. Conclusion Prenatal nitrite exposure can induce the synapse loss with dose dependency and long-term effect, and the abnormal aggregation of γ-synuclein may contribute to the synaptic loss.