新生大鼠反复惊厥对脑内γ-氨基丁酸B1受体表达的影响

Effects of neonatal recurrent seizures on γ-aminobutyric acid B1 receptor expression in the rat brain

摘要:

目的：发育期脑损伤能通过改变神经递质受体表达造成脑功能长期障碍,该研究通过观察新生期反复惊厥对大鼠脑内γ-氨基丁酸(γ-aminobutyricacid,GABA)B1受体(GABAB1R)表达的影响及成年期记忆功能和惊厥阈的长期改变,探讨其间可能存在的相关性。方法：生后7d(P7)新生SD大鼠48只,随机分入惊厥组和对照组,每组24只,惊厥组仔鼠通过吸入三氟乙醚诱导惊厥,每天1次,连续6d。于反复惊厥结束后7d,每组各随机处死12只大鼠,应用RT-PCR及免疫组织化学方法检测大鼠大脑皮层及海马的GABAB1RmRNA及蛋白表达的近期改变。剩余大鼠于P61~P64行Morris水迷宫实验,检测大鼠的学习记忆功能。于P75时给予大鼠腹腔注射戊四唑测定大鼠的惊厥阈后,即刻处死大鼠取脑,用RT-PCR及免疫组织化学方法检测大鼠大脑皮层及海马的GABAB1RmRNA及蛋白表达的长期变化。结果：①在反复惊厥后7d及P75时,惊厥组大鼠大脑皮层中GABAB1RmRNA及蛋白的表达较对照组均显著下调(P<0.05);②惊厥结束后7d时,惊厥组大鼠海马区中GABAB1RmRNA表达较对照组无明显改变(P>0.05),但在齿状核GABAB1R蛋白表达明显降低(P<0.001);P75时惊厥组大鼠海马区GABAB1RmRNA及其在齿状核GABAB1R蛋白表达较对照组差异无显著性(P>0.05);③Morris水迷宫实验显示惊厥组大鼠在P64的寻找平台时间为98533.8±27205.4ms较正常对照组的46723.3±40666.5ms明显延长(t=3.66,P<0.05);④惊厥组大鼠注射戊四唑后发生惊厥的潜伏期为1415.1±428.5s与对照组1156.0±308.9s比较差异无显著性(t=1.70,P>0.05)。结论：新生期大鼠反复惊厥后,大脑皮层和海马GABAB1R表达持续减少,可能参与新生期惊厥后脑损伤的病理过程,与新生期反复惊厥导致的成年期记忆障碍相关。

Abstract:

OBJECTIVE: This study investigated the effects of flurothyl-induced neonatal recurrent seizures on γ-aminobutyric acid B1 receptor (GABAB1R) expression in neonatal and adult rat brain, and explored the possible relationship between the alterations of GABAB1R in mature brain and the changes of spatial memory and seizure susceptibility in adult rats. METHODS: Forty-eight postnatal day (P) 7 Sprague-Dawley rats were randomly assigned into two groups: Control and Seizure group (n=24 each). Seizures were induced by inhalant flurothyl daily for six consecuive days in rat pups from the Seizure group. Twelve rats selected randomly in each group were sacrificed on the 7th day after the last seizure for detecting the expressions of GABAB1R mRNA and protein in cerebral cortex and hippocampus by reverse transcription-polymerase chain reaction (RT-PCR) and immuno-histochemistry method. The spatial memory was tested by using the Morris water maze task during P61 to P64 and the seizure threshold was measured at P75 following intraperitoneal injection of pentylenetetrazol ( PTZ ) in the remaining rats. The rats were then sacrificed for detecting the expressions of GABAB1R mRNA and protein in cerebral cortex and hippocampus. RESULTS: The expressions of GABAB1R mRNA and protein in the cerebral cortex on the 7th day after the last seizure and at P75 decreased significantly in the Seizure group when compared with the Control group (P<0.05). The GABAB1R protein expression in the dentate gyrus on the 7th day after the last seizure in the Seizure group was significantly lower than that in the Control group (P<0.05), but the GABAB1R mRNA expression in the hippocampus was not different from that in the Control group. There were no significant differences in the expressions of GABAB1R mRNA and protein in the hippocampus between the two groups at P75. The escape latencies in water maze of the rats in the Seizure group at P64 were significantly longer than those in the Control group (98 533.8± 27 205.4 ms vs 46 723.3±40 666.5 ms; P<0.05). There were no differences in the seizure threshold between the two groups. CONCLUSIONS: The expressions of GABAB1R mRNA and protein in the cerebral cortex and hippocampus of neonatal rats with recurrent seizures decreased significantly, suggesting the changes of GABAB1R may be related to acute brain injury following neonatal recurrent seizures and the memory deficit in adult rats caused by neonatal recurrent seizures.