新生大鼠反复惊厥对脑内γ-氨基丁酸A受体α1和β2亚单位表达的长期影响

目的 研究新生期大鼠反复惊厥对脑内γ-氨基丁酸（GABA）A受体（GABAAR）α1和β2亚单位表达的长期影响及成年期记忆功能和惊厥阈的长期改变。方法 生后7d的SD大鼠随机分成两组,每组16只,惊厥组每日吸入三氟乙醚诱导惊厥发作1次,每次持续30min,连续6d;对照组同样操作但不吸入三氟乙醚。两组大鼠于生后61～65d行Morris水迷宫实验,于生后75d时通过腹腔注射戊四唑测定大鼠的惊厥阈。随即处死大鼠,分别采用免疫组化和逆转录PCR方法观察大鼠大脑皮层及海马GABAARα1和B2亚单位表达的变化。结果 生后64d的寻找平台时间（82424ms±35622ms）明显长于对照组（40712ms±29467ms,P=0．001）。在生后65d,惊厥组大鼠120s内穿越目标次数（1．2次±0．9次）明显少于对照组（3．1次±1．3次,P〈0．01）。惊厥组大鼠注射PTZ后发生惊厥的潜伏期（1487s±662s）与对照组（1841s±648s）差异无统计学意义（P=0．133）。惊厥组大鼠GABAARα1亚单位免疫化学累积吸光度在顶叶及海马CA1、CA2和CA4区明显低于对照组（均P〈0．05）;惊厥组大鼠GABAARβ2亚单位免疫化学累积光密度在丘脑区和海马CA1～4区明显低于对照组（均P〈0．05）。惊厥组大鼠海马α1和β2亚单位mRNA的表达明显少于对照组（均P〈0．05）。结论 新生大鼠反复惊厥可造成脑内GABAARα1和陀亚单位表达的长期改变,这种改变可能与新生期反复惊厥导致的成年期记忆障碍相关。

Long-term effects of recurrent seizures in neonate period on gamma-aminobutyric acid A receptor alpha1 and beta2 subunits expression in adult brain: experiment with rats

OBJECTIVE: To investigate the long-term effects of recurrent seizures in neonate period on the expression of gamma-aminobutyric acid A receptor (GABAAR) alpha1 and beta2 subunits in brain and spatial memory and seizure susceptibility in adult period. METHODS: Thirty-two 7-day-old SD rats were randomly divided into 2 equal groups: seizure group, inhaling flurothyl to induce seizure daily for 6 days, and control group. On days 61 - 65 after birth Morris water maze test was used to record the escape latency. On day 75 after birth pentylenetetrazol (PTZ) was injected intraperitoneally to induce seizure so as to record the latency. Then the rats were killed to take their brains, 8 in each group used to undergo immunohistochemistry to examine the protein expression of the GABAAR alpha1 and beta2 subunits, and the other 8 in each group used to examine the mRNA expression of the GABAAR alpha1 and beta2 subunits in the brains using RT-PCR. RESULTS: On day 64 the escape latency of the seizure group was 82,424 ms +/- 35,622 ms, significantly longer than that of the control group (40,712 ms +/- 29,468 ms, P = 0.001). On day 75 the frequency of crossing target within 120 s in the water maze of the seizure group was 1.2 times +/- 0.9 times, significantly less than that of the control group (3.1 times +/- 1.3 times, P < 0.01). The seizure latency after the PTZ injection of the seizure group was (1487 +/- 662) s, not significantly different from that of the control group (1841 s +/- 648 s, P = 0.133). In comparison with the control group the accumulated optical density (AOD) of GABAAR alpha1 subunit protein immunoactivity in the parietal cortex, and hippocampal CA1-2 and CA4 regions of the seizure group decreased significantly (all P < 0.05), and was not significantly different in the frontal cortex, dentate gyrus, and hippocampal CA3 region (all P > 0.05). In comparison with the control group the accumulated optical density (AOD) of GABAAR beta2 subunit protein immunoactivity in the thalamus, and hippocampal CA1-4 regions of the seizure group decreased significantly (all P < 0.05), and was not significantly different in the frontal cortex and parietal cortex (both P > 0.05). In comparison with the control group the mRNA expression of GABAAR alpha1 subunit and the mRNA expression of GABAAR beta2 subunit of the seizure group were significantly lower in the hippocampus (both P < 0.05) and not significantly different in the cerebral cortex (both P > 0.05). CONCLUSION: Recurrent seizures in neonate period modify the expression of GABAAR alpha1 and beta2 subunits in the cerebral cortex and hippocampus in adult period which may be related to cognitive deficit.