大鼠肠道神经元NR1表达与急性束缚应激所致内脏高敏感的关系

目的 研究急性束缚应激所致内脏高敏感大鼠肠道神经元N-甲基-D-天冬氨酸(NMDA)受体NRI亚基的表达变化.方法 30只雄性SD大鼠按照随机表分为3组:对照组、急性应激组和急性应激+MK-801组各10只.以急性束缚应激的方法建立内脏高敏感模型,以NMDA受体拮抗剂MK-801干预.记录大鼠结直肠扩张压力下腹外斜肌肌电图以评估内脏感觉功能,比较不同组问的差异.采用RT-PCR和Western印迹法检测回盲部、近端结肠和远端结肠NR1的mRNA和蛋白质水平的表达.结果 (1)急性应激组在40、60、80 men Hg(1 mm Hg=0.133 kPa)压力下行直、结肠扩张时,腹外斜肌放电次数均多于对照组(次:925±217比188±31、1480±347比510±68、1732±344比765±103,均P<0.01)和急性应激+MK-801组(次:210±47、525±97、841±156,均P<0.05);后2组差异均无统计学意义(均P>0.05).(2)急性应激组回盲部、近端结肠和远端结肠NRI的mRNA表达水平均高于对照组(A值:1.57±0.20比0.68±0.10、2.00±0.20比0.87±0.19、1.36±0.17比0.74±0.15,均P<0.01)和急性应激+MK-801组(A值:0.84±0.13、0.91±0.16、0.79±0.13,均P<0.05);后2组差异均无统计学意义(均P>0.05).(3)急性应激组回盲部、近端结肠和远端结肠NRI的蛋白表达水平均高于对照组(A值:1.69±0.20比0.54±0.11、1.41±0.12比0.71士0.06、1.63±0.15比0.71±0.07,均P=0.000)和急性应激+MK-801组(A值0.75±0.09、0.70±0.11、0.63±0.11,均P=0.000);后2组差异均无统计学意义(均P>0.05).结论 NMDA受体介导了急性束缚应激导致的内脏敏感性增高.

Relationship between NR1 subunit of N-methyl-D-aspartate receptor and visceral hypersensitivity caused by acute restraint stress: experiment with rats

Objective To investigate the response of NRI subunit of N-methyl-D-aspartate (NMDA)receptor to restraint stress in intestinal tract.Methods Thirty Sprague-Dawley rats were randomly divided into 3 equal groups:acute stress group undergoing intrapeitoneal injection of normal saline (NS),acute stress+MK-801 group,undergoing intrapeitoneal injection of MK-801,a NMDA receptor inhibitor.and then acute restraint for 1 h,30 min after the injection.Control group underwent intrapeitoneal injection of NS without acute restraint to be used as.Then all rats underwent eolorectal distension(CRD)at the pressure levels of 20.40,60,and 80 nun Hg respectively for 10 s 3 times with an interval of40 s so as to establish visceral hypersensitivity models.Electremyography was performed at the extemal oblique muscle of abdomen to record the frequency of discharge to evaluate the viseers]sensitivity.Then tlle rats weTe killed and specimens of ileocecum and the proximal and distal ends of colon were obtained.RT-PCR and Westem blotting were used to detect the mRNA and protein expression of NMDA receptor NR1 subunit in intestinal tract.Results (1)The frequencies of discharge of external oblique muscle of abdomen responding to CRD of the acute stress group at the pressure levels of 40.60.and 80 mm Hg were 925±217,1480±347,and 1732±344 respectively,all significantly higher than those of the control group(188±31,510±68,and 765±103,all P<0.01)and the acute stress+MK-801 group(210±47,525±97,84l±156,all P<0.05).But no significantly different from the acute stress+MK-801 group and the control group(all P>0.05).(2)The A valuese of the mRNA expression of NR1 in the ileocecal junction,proximal colon end,and distal colon end of the acute stress group were 1.57±0.20.2.00±0.20,and 1.36±0.17respectively,all significantly higher than those of the control group(0.68±0.10,0.87±0.19,and 0.74±0.15,all P<0.01)and the acute stress+MK-801 group(0.84±0.13,0.91±0.16,0.79±0.13,all P<0.05).But no significantly difierent from the acute stress+MK-801 group and the contml group(allP>0.05).(3)The A vaiues of the protein expression of NR1 in the ileocecal junction,proximal colon end.and distal colon end of the acute stress group were 1.69±0.20,1.41±0.12,and 1.63±0.15respectively.all significantly higher than those of the control group(0.54±0.11,O.71±0.06,0.71±0.07.all P=0.000)and the acute+MK-801 group(0.75±0.09,0.70±0.11,0.63±0.11,all P=0.000).But no signiilcanfly different from the acute stress+MK-801 group and the control group(all P>0.05).Conclusion NMDA receptor plays an important role in the visceral sensitivity induced by acute restraint stress.