超声重复辐照孕鼠对子代海马NMDA受体NR1、NR2B亚单位表达及其突触结构的影响

Effects of repeated ultrasound on expression of NR1,NR2B subunit of NMDA receptor and synaptic structure in prenatal rats

目的 明确不同剂量超声重复辐照孕鼠对仔鼠海马N-甲基-D-天门冬氨酸（NMDA）受体NR1、NR2B亚单位表达及突触结构的影响,探讨超声辐射对仔鼠学习记忆功能影响的分子机制.方法 用不同剂量超声于孕6 d、12 d、18 d重复辐照孕大鼠0 min、4 min、20 min （分别为对照组、4 min组及20 min组）,每组8只,应用Morris水迷宫检测2月龄仔鼠的空间学习记忆能力,通过HE染色观察海马CA1区锥体细胞形态学变化,免疫组化法和Western印迹法检测大鼠海马组织NR1、NR2B蛋白表达,电镜观察突触的超微结构.结果 （1）4 min组大鼠第1~4天逃避潜伏期短于对照组（P〈0.05）,20 min组第1～4天逃避潜伏期长于对照组（P〈0.01）;4 min组原站台象限停留时间长于对照组（P〈0.01）、穿越原站台次数多于对照组（P〈0.05）;20 min组原站台象限停留时间短于对照组（P〈0.01）、穿越原站台次数少于对照组（P〈0.01）;（2）4 min组海马CA1区单位面积锥体细胞密度高于对照组（P〈0.05）,20 min组锥体细胞密度低于对照组（P〈0.01）;（3）4 min组海马CA1区NR1、NR2B表达增强,Western印迹显示海马总NR1、NR2B表达高于对照组（P〈0.05）,20 min组海马CA1区NR1、NR2B表达减弱,Western印迹显示海马总NR1、NR2B表达低于对照组（P〈0.05）;（4）20 min组突触结构受损,突触数密度、面密度降低.结论 大剂量超声多次辐照孕鼠,仔鼠海马区NMDA受体NR1、NR2B亚单位表达减少,突触结构受损,使仔鼠学习记忆能力下降;小剂量超声辐照组仔鼠海马区NMDA受体NR1、NR2B亚单位表达增强,突触结构正常,仔鼠学习记忆能力提高.

Objective To explore the effects of different doses of repeated ultrasound on the expressions of NR1 and NR2B subunits of NMDA receptor and synaptic structure so as to understand the molecular mechanisms of ultrasound on learning and memory in prenatal rats. Methods The pregnant rats were randomly divided into 3 groups （n = 8, each） and exposed to uhrasound for 0, 4 and 20 minutes at Days 6, 12 and 18 of gestation respectively. Their offspring were tested by Morris water maze （MWM） for evaluating the learning and memory abilities at an age of 2 months. The hippocampal pyramidal cell was observed by hematoxylin and eosin staining, the expressions of NR1 and NR2B by immunohistochemistry and Western blot and the morphologies of synapse in CA1 by electron microscopy. Results （ 1 ） In the water maze test, compared with the control group, the escape latency of 4 min group was significantly shorter （P 〈 0. 05） while that of 20 min group significantly longer in the 1 -4 d latency （P 〈0. 01 ）. The time in platform quadrant （TQ） and the frequency of crossing through original platform increased in 4 min group （P 〈0. 01 ）. However, they decreased in the 20 min group （P 〈0.01 ） ; （2） in 4 rain group, the density of hippocampal CAI pyramidal cells per unit area was higher than that of the control group （P 〈 0. 05）. Sparse pyramidal cells and low density were observed in the 20 min group （ P 〈 0. 01 ） ; （ 3 ） compared with the control group, the expressions of total NR1, NR2B proteins in hippocampal regions significantly increased in 4 rain group and both significantly decreased in 20 min （P 〈0. 05） ; （4） the structure of synapses became damaged in 20min group with decreased number density and surface density. Conduslon High doses of repeated ultrasound may decrease the ability of learning and memory through a down-regulated expression of NR1 and NR2B subunits in hippocampal regions and damaging synaptic structure. However, low doses of repeated ultrasound may promote the expressions of NR1 and NR2B, maintain synaptie structure and further enhance cognitive ability in prenatal rats.