戊四氮致痫幼鼠颞叶和海马区Cx43、P38的表达及天麻素干预研究
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摘要
目的在建立戊四氮点燃发育期大鼠致痫模型的基础上,观察各组大鼠行为学改变、颞叶与海马区Cx43、P38表达变化及天麻素的干预研究。
方法将50只21日龄Wistar发育期大鼠(50-70g)随机分为5组,分别为空白对照组、戊四氮组、天麻素大剂量组、天麻素小剂量组及丙戊酸钠组,每组10只。根据Racine分级法,观察大鼠行为学改变并记录大鼠痫性发作级别和次数,在点燃4周后常规方法心脏灌注、固定及取脑组织,采用免疫组织化学方法显示各组动物切片Cx43、P38表达的情况。
结果(1)在实验2周时,戊四氮组发作率为100%,天麻素小剂量组、大剂量组及丙戊酸钠组发作率均低于戊四氮组(P=0.03、0.003、0.003);4周后,天麻素小剂量组、大剂量组发作率与戊四氮组比较(P_a＞0.05),差异没有统计学意义,但丙戊酸钠组明显低于戊四氮组(P＜0.05),其差异具有统计学意义;4周后,戊四氮组点燃率为100%,天麻素大小剂量组及丙戊酸钠组点燃率均低于戊四氮组(P＜0.01)。(2)戊四氮点燃四周后,实验各组大鼠痫性发作级别比较存在显著性差异(χ~2=35.83,P＜0.01),戊四氮组发作级别最高(MeanRank=44.60),其次是天麻素小剂量组(Mean Rank=34.90)、天麻素大剂量组(Mean Rank=21.45)、丙戊酸钠组(Mean Rank=19.55)。(3)戊四氮点燃四周后,在各组实验大鼠颞叶皮质及海马区,戊四氮组Cx43表达量较其它组多(P_a＜0.01),差异具有显著性统计学意义,且天麻素大剂量组的表达量小于天麻素小剂量组(q=2.811,2.647 P_a＜0.05),而大剂量组与丙戊酸钠组比较(q=1.203,1.116P_a＞0.05),其差异没有统计学意义。(4)戊四氮点燃四周后,在各组实验大鼠颞叶皮质及海马区,戊四氮组P38表达量较其它组多(P_a＜0.01),差异具有显著性统计学意义,且天麻素大剂量组的表达量小于天麻素小剂量组(q=2.705,2.351P_a＜0.05),而大剂量组与丙戊酸钠组比较(q=1.194,1.031 P_a＞0.05),其差异没有统计学意义。
结论(1)Cx43、P38表达增加可能是PTZ致痫产生的原因。(2)天麻素能够降低致痫大鼠惊厥的易感性及大鼠颞叶和海马区Cx43表达,抑制了异常缝隙连接的形成,达到抗癫痫形成。(3)戊四氮反复点燃可导致发育期大鼠颞叶和海马区苔藓纤维发芽和突触重建的形成;天麻素具有明显对抗癫痫形成的作用,并且可通过降低P38的表达,抑制突触重建的形成,间接控制癲痫发作。
Objective On the basis of the establishment of pentylenetetrazole ignited immature rat model induced seizures,to observe the behavior change and the expression of Cx43and P38 in temporal lobe and hippocampus of each group of rats and its intervention study of Gastrodin.
Methods Fifty growth period Wistar rats(50-70g)were randomly divided into five groups,each 10,there were control group,pentylenetetrazole group,gastrodin high-dose group,gastrodin low-dose group and valproate sodium group.According to racine classification scheme for observing the behavior changes and record seizures rank and frequency of experimental rats.To adopt conventional method to perfuse cordis,fix and extract the brain,and the immunohistochemistry was used to detect Cx43 and P38 expression in the temporal lobe and hippocampus of the brain after 4 weeks.
Results(1)After two weeks of the experiment,attack rate was 100%in pentylenetetrazole group,attack rates in gastrodin low-dose group,high-dose group and valproate sodium group were all lower than PTZ group(P=0.03,0.003, 0.003);After 4 weeks,attack rate in gastrodin low-dose group and high-dose group were compared with PTZ group(P_α＞0.05),the difference was not statistically significant,but valproate sodium group was lower than that of PTZ group(P＜0.05),the difference were statistically significant;After 4 weeks,ignited rate in PTZ group was 100%,ignited rate in high-dose group and valproate sodium group were lower than PTZ group(P＜0.01).(2)Four weeks later,there was significant difference in comparison among seizures level in each group of experimental rats (χ~2=35.83,P＜0.01),attack level was the highest in PTZ group(Mean Rank= 44.60),followed by gastrodin low-dose group(Mean Rank=34.90),gastrodin high-dose group(Mean Rank=21.45)and valproate sodium group(Mean Rank= 19.55).(3)Four weeks later,rats who were ignited by pentylenetetrazole in all groups, Cx43 expression in temporal lobe cortex and the hippocampus of PTZ group was more than other groups(P_α＜0.01),the difference was statistically significant,and Cx43 expression in gastrodin high-dose group was less than gastrodin low-dose group (q=2.811,2.647 P_α＜0.05),but Cx43 expression in high-dose group compared with valproate sodium group(q=1.203,1.116 P_α＞0.05),there was no statistically significant.(4)Four weeks later,rats who were ignited by pentylenetetrazole in all groups,P38 expression in temporal lobe cortex and the hippocampus of PTZ group was more than other groups(P_α＜0.01),the difference was statistically significant,and P38 expression in gastrodin high-dose group was less than gastrodin low-dose group(q=2.705,2.351 P_α＜0.05),but P38 expression in high-dose group was compared with valproate sodium group(q=1.194,1.031 P_α＞0.05),there was no statistically significant.
Conclusion(1)The expression of P38 and Cx43 was increased which may be the cause of PTZ induced epilepsy.(2)Gastrodin could reduce the susceptibility of epileptic seizures and inhibited Cx43 expression of temporal lobe and,hippocampus, inhibited the formation of abnormal gap junction and achieved antiepileptic formation.(3)There was mossy fiber sprouting and the formation of synaptic reconstruction in temporal lobe and hippocampus of the growth period rats who were repeatedly ignited by pentylenetetrazole.Gastrodin have the role to the formation of antiepileptic obviously,and through decreasing the expression of P38,which can inhibit the formation of synaptic reconstruction and control seizures indirectly.
方法将50只21日龄Wistar发育期大鼠(50-70g)随机分为5组,分别为空白对照组、戊四氮组、天麻素大剂量组、天麻素小剂量组及丙戊酸钠组,每组10只。根据Racine分级法,观察大鼠行为学改变并记录大鼠痫性发作级别和次数,在点燃4周后常规方法心脏灌注、固定及取脑组织,采用免疫组织化学方法显示各组动物切片Cx43、P38表达的情况。
结果(1)在实验2周时,戊四氮组发作率为100%,天麻素小剂量组、大剂量组及丙戊酸钠组发作率均低于戊四氮组(P=0.03、0.003、0.003);4周后,天麻素小剂量组、大剂量组发作率与戊四氮组比较(P_a＞0.05),差异没有统计学意义,但丙戊酸钠组明显低于戊四氮组(P＜0.05),其差异具有统计学意义;4周后,戊四氮组点燃率为100%,天麻素大小剂量组及丙戊酸钠组点燃率均低于戊四氮组(P＜0.01)。(2)戊四氮点燃四周后,实验各组大鼠痫性发作级别比较存在显著性差异(χ~2=35.83,P＜0.01),戊四氮组发作级别最高(MeanRank=44.60),其次是天麻素小剂量组(Mean Rank=34.90)、天麻素大剂量组(Mean Rank=21.45)、丙戊酸钠组(Mean Rank=19.55)。(3)戊四氮点燃四周后,在各组实验大鼠颞叶皮质及海马区,戊四氮组Cx43表达量较其它组多(P_a＜0.01),差异具有显著性统计学意义,且天麻素大剂量组的表达量小于天麻素小剂量组(q=2.811,2.647 P_a＜0.05),而大剂量组与丙戊酸钠组比较(q=1.203,1.116P_a＞0.05),其差异没有统计学意义。(4)戊四氮点燃四周后,在各组实验大鼠颞叶皮质及海马区,戊四氮组P38表达量较其它组多(P_a＜0.01),差异具有显著性统计学意义,且天麻素大剂量组的表达量小于天麻素小剂量组(q=2.705,2.351P_a＜0.05),而大剂量组与丙戊酸钠组比较(q=1.194,1.031 P_a＞0.05),其差异没有统计学意义。
结论(1)Cx43、P38表达增加可能是PTZ致痫产生的原因。(2)天麻素能够降低致痫大鼠惊厥的易感性及大鼠颞叶和海马区Cx43表达,抑制了异常缝隙连接的形成,达到抗癫痫形成。(3)戊四氮反复点燃可导致发育期大鼠颞叶和海马区苔藓纤维发芽和突触重建的形成;天麻素具有明显对抗癫痫形成的作用,并且可通过降低P38的表达,抑制突触重建的形成,间接控制癲痫发作。
Objective On the basis of the establishment of pentylenetetrazole ignited immature rat model induced seizures,to observe the behavior change and the expression of Cx43and P38 in temporal lobe and hippocampus of each group of rats and its intervention study of Gastrodin.
Methods Fifty growth period Wistar rats(50-70g)were randomly divided into five groups,each 10,there were control group,pentylenetetrazole group,gastrodin high-dose group,gastrodin low-dose group and valproate sodium group.According to racine classification scheme for observing the behavior changes and record seizures rank and frequency of experimental rats.To adopt conventional method to perfuse cordis,fix and extract the brain,and the immunohistochemistry was used to detect Cx43 and P38 expression in the temporal lobe and hippocampus of the brain after 4 weeks.
Results(1)After two weeks of the experiment,attack rate was 100%in pentylenetetrazole group,attack rates in gastrodin low-dose group,high-dose group and valproate sodium group were all lower than PTZ group(P=0.03,0.003, 0.003);After 4 weeks,attack rate in gastrodin low-dose group and high-dose group were compared with PTZ group(P_α＞0.05),the difference was not statistically significant,but valproate sodium group was lower than that of PTZ group(P＜0.05),the difference were statistically significant;After 4 weeks,ignited rate in PTZ group was 100%,ignited rate in high-dose group and valproate sodium group were lower than PTZ group(P＜0.01).(2)Four weeks later,there was significant difference in comparison among seizures level in each group of experimental rats (χ~2=35.83,P＜0.01),attack level was the highest in PTZ group(Mean Rank= 44.60),followed by gastrodin low-dose group(Mean Rank=34.90),gastrodin high-dose group(Mean Rank=21.45)and valproate sodium group(Mean Rank= 19.55).(3)Four weeks later,rats who were ignited by pentylenetetrazole in all groups, Cx43 expression in temporal lobe cortex and the hippocampus of PTZ group was more than other groups(P_α＜0.01),the difference was statistically significant,and Cx43 expression in gastrodin high-dose group was less than gastrodin low-dose group (q=2.811,2.647 P_α＜0.05),but Cx43 expression in high-dose group compared with valproate sodium group(q=1.203,1.116 P_α＞0.05),there was no statistically significant.(4)Four weeks later,rats who were ignited by pentylenetetrazole in all groups,P38 expression in temporal lobe cortex and the hippocampus of PTZ group was more than other groups(P_α＜0.01),the difference was statistically significant,and P38 expression in gastrodin high-dose group was less than gastrodin low-dose group(q=2.705,2.351 P_α＜0.05),but P38 expression in high-dose group was compared with valproate sodium group(q=1.194,1.031 P_α＞0.05),there was no statistically significant.
Conclusion(1)The expression of P38 and Cx43 was increased which may be the cause of PTZ induced epilepsy.(2)Gastrodin could reduce the susceptibility of epileptic seizures and inhibited Cx43 expression of temporal lobe and,hippocampus, inhibited the formation of abnormal gap junction and achieved antiepileptic formation.(3)There was mossy fiber sprouting and the formation of synaptic reconstruction in temporal lobe and hippocampus of the growth period rats who were repeatedly ignited by pentylenetetrazole.Gastrodin have the role to the formation of antiepileptic obviously,and through decreasing the expression of P38,which can inhibit the formation of synaptic reconstruction and control seizures indirectly.
引文
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