An approach for anti-sense therapy based on the establishment of an animal model for the epileptogenic cortical malformations

基于致痫性皮质畸形动物模型的反义治疗方法

• 批准号: 12557077
• 负责人: FUKUDA Atsuo
• 金额: $ 7.3万
• 依托单位: HAMAMATSU UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-12557077/
• 关键词: focal cortical dysplasia; cortical freeze-lesion; epilepsy; Cl^-homeostasis; KCC2; NKCC1; GABA; cell migration; クロライドイオン; 免疫沈降法; yeast two-hybrid法; グリシン受容体; クロライドトランスポーター; カハールレチウス細胞; グリシン; 神経幹細胞; Cl^-トランスポーター; ClC; NKCC; KCC

Focal cortical malformations as a result of cell migration disorder are often associated with intractable epilepsy. By using the rats underwent neonatal cortical freeze-lesion (FL), as a model for human polymicrogyria, we studied [Cl^-]_i and the GABA/glycine effects during formation of the microgyrus. GABA/glycine-induced depolarizations and Ca^<2+> influxes were observed in the upper part of FL in which [Cl^-]_i were increased. Consistently, mRNA expression of NKCC1 (accumulates Cl^-) was up-regulated whereas that of KCC2 (extrudes Cl^-) was downregulated. Above characteristics are similar to the known characteristics of migrating cortical plate cells. Thus, cortical plate cells near by FL might regain or preserve the immature characteristics of Cl^-homeostasis and GABA/glycine actions, so that they could migrate into the FL to form microgyrus expressing hyperexcitability later on.We also investigated NKCC1 and KCC2 expressions in the human focal cortical dysplasia (FCD) tissue, characterized by disorganized lamination and cytomegalic dysplastic cells, from patients of intractable epilepsy. KCC2 mRNA and protein were decreased in small-sized dysplastic neurons as compared to non-dysplastic neurons in histologicaly normal portion. These results suggest that decreases in KCC2 expression in small-sized dysplastic neurons might change their Cl^-homeostasis, so that impairment of GABAergic actions could occur in FCD being involved in epileptogenesis.

细胞迁移障碍导致的局灶性皮质畸形通常与顽固性癫痫有关。通过使用新生儿皮质冷冻损伤（FL）大鼠作为人类多小脑回模型，我们研究了[Cl^-]_i和GABA/甘氨酸在小脑回形成过程中的作用。在FL上部观察到GABA/甘氨酸诱导的去极化和Ca^2+流入，其中[Cl^-]_i增加。一致地，NKCC1（积累 Cl^-）的 mRNA 表达上调，而 KCC2（排出 Cl^-）的 mRNA 表达下调。上述特征与已知的迁移皮质板细胞的特征相似。因此，靠近 FL 的皮质板细胞可能会恢复或保留 Cl^-稳态和 GABA/甘氨酸作用的不成熟特征，以便它们能够迁移到 FL 中形成表达超兴奋性的小回。我们还研究了 NKCC1 和 KCC2 在 FL 中的表达。来自顽固性癫痫患者的人类局灶性皮质发育不良 (FCD) 组织，其特征是层压紊乱和巨细胞发育不良细胞。与组织学正常部分中的非发育不良神经元相比，小尺寸发育不良神经元中的 KCC2 mRNA 和蛋白质减少。这些结果表明，小型发育不良神经元中 KCC2 表达的减少可能会改变其 Cl^-稳态，因此参与癫痫发生的 FCD 可能会损害 GABA 能作用。

项目成果

Shimizu-Okabe, C. et al.: "Layer-specific expression of Cl^-transporters and differential [Cl^-]_i in newborn rat cortex."Neuroreport. 13. 2433-2437 (2002)

Shimizu-Okabe, C. 等人：“新生大鼠皮层中 Cl^-转运蛋白的层特异性表达和差异 [Cl^-]_i”。Neuroreport。

Ikeda, M. et al.: "Differential development of cation-chloride cotransporters and Cl^- homeostasis contributes to differential GABAergic actions between developing rat visual cortex and dorsal lateral geniculate nucleus."Brain Res.. 984. 149-159 (2003)

Ikeda, M. 等人：“阳离子-氯化物协同转运蛋白和 Cl^- 稳态的差异发育有助于发育中的大鼠视觉皮层和背外侧膝状核之间的 GABA 能作用的差异。”Brain Res.. 984. 149-159 (2003)

Ueno, T. et al.: "Diversity of neuron-specific K^+-Cl^- cotransporter expression and inhibitory postsynaptic potential depression in rat motoneurons."J.Biol.Chem.. 277. 4945-4950 (2002)

Ueno, T. 等人：“大鼠运动神经元中神经元特异性 K^-Cl^- 协同转运蛋白表达的多样性和抑制性突触后电位抑制。”J.Biol.Chem.. 277. 4945-4950 (2002)

Okabe, A. et al.: "Amygdala kindling induces upregulation of mRNA for NKCC1,a Na^+,K^+,-2Cl^- cotransporter, in the rat piriform cortex."Neurosci.Res.. 44. 225-229 (2002)

Okabe, A. 等人：“杏仁核点燃诱导大鼠梨状皮层中 NKCC1（一种 Na^ 、K^ 、-2Cl^- 协同转运蛋白） mRNA 的上调。”Neurosci.Res.. 44. 225-229 (2002)

Farkas, I. et al.: "CD59 blocks not only the insertion of C9 into MAC but inhibits ion channel formation of homologous C5b-8 as well as C5b-9."J.Physiol.. 539. 537-545 (2002)

Farkas, I. 等人：“CD59 不仅阻止 C9 插入 MAC，而且抑制同源 C5b-8 和 C5b-9 的离子通道形成。”J.Physiol.. 539. 537-545 (2002)