Krembil Research Institute, University Health Network, 60 Leonard Avenue, Toronto, Ontario M5T 0S8, Canada.
Krembil Research Institute, University Health Network, 60 Leonard Avenue, Toronto, Ontario M5T 0S8, Canada.
Exp Neurol. 2020 Sep;331:113373. doi: 10.1016/j.expneurol.2020.113373. Epub 2020 Jun 2.
Neurofibromatosis type 1 (NF1) is associated with higher rates of epilepsy compared to the general population. Some NF1 patients with epilepsy do not have intracranial lesions, suggesting the genetic mutation itself may contribute to higher rates of epilepsy in these patients. We have recently demonstrated increased seizure susceptibility in the Nf1 mouse, but it is unknown whether this model displays altered epileptogenicity, as has been reported in patients with NF1. The aim of this study was to determine whether the Nf1 mouse is more susceptible to electrical kindling-induced epileptogenesis.
Young male or female adult Nf1 or Nf1 (wild-type; WT) mice were implanted with electrodes for neocortical or hippocampal kindling paradigms. Neocortical kindling was performed for 40 stimulation sessions followed by baseline EEG monitoring to detect possible SRSs. Hippocampal kindling was performed with a modified extended kindling paradigm, completed to a maximum of 80 sessions to try to induce spontaneous repetitive seizures (SRSs). Western blot assays were performed in naïve and kindled mice to compare levels of Akt and MAPK (ERK1/2), proteins downstream of the NF1 mutation.
The average initial neocortical after-discharge threshold (ADT) was significantly lower in the Nf1 group, which also required fewer stimulations to reach stage 5 seizure, had greater average seizure severity across all kindling sessions, had a greater number of convulsive seizures, and had a faster progression of after-discharge duration and Racine score during kindling. No WT mice exhibited SRS after neocortical kindling, versus 33% of Nf1 mice. The average initial hippocampal ADT was not significantly different between the WT and Nf1 groups, nor was there a difference in the number of stimulations required to reach the kindled state. The WT group had a significantly higher average seizure severity across all kindling sessions as compared with the Nf1 mice. The WT group also had faster progression of the Racine seizure score over the kindling sessions, mainly due to a faster increase in seizures severity early during the kindling process. However, SRSs were seen in 50% of Nf1 mice after modified extended kindling and in no WT mice. Western blots showed hippocampal kindling increased the ratio of phosphorylated/total Akt in both the WT and Nf1 mice, while neocortical kindling led to increased ratios of phosphorylated/total Akt and MAPK in Nf1 mice only.
We have demonstrated for the first time an increased rate of epileptogenesis in an animal model of NF1 with no known macroscopic/neoplastic brain lesions. This work provides evidence for the genetic mutation itself playing a role in seizures and epilepsy in patients with NF1, and supports the use of the Nf1 mouse model in future mechanistic studies.
与普通人群相比，1 型神经纤维瘤病（NF1）患者癫痫发作的发生率更高。一些患有癫痫的 NF1 患者没有颅内病变，这表明遗传突变本身可能导致这些患者癫痫发作的发生率更高。我们最近证明了 Nf1 小鼠的癫痫易感性增加，但尚不清楚该模型是否表现出与 NF1 患者中报道的那样改变的致痫性。本研究的目的是确定 Nf1 小鼠是否更容易发生电点燃诱导的癫痫发生。
年轻的雄性或雌性成年 Nf1 或 Nf1（野生型；WT）小鼠被植入用于新皮质或海马点燃范式的电极。进行新皮质点燃 40 个刺激疗程，然后进行基线 EEG 监测以检测可能的 SRS。海马点燃采用改良的扩展点燃范式进行，完成 80 个疗程以尝试诱导自发性重复发作（SRS）。在未点燃和点燃的小鼠中进行 Western blot 检测，以比较 Akt 和 MAPK（ERK1/2）的水平，这是 NF1 突变的下游蛋白。
Nf1 组的平均初始新皮质后放电阈值（ADT）明显较低，达到 5 级发作所需的刺激次数更少，在所有点燃疗程中的平均发作严重程度更大，有更多的强直发作，以及在点燃过程中放电持续时间和 Racine 评分的更快进展。没有 WT 小鼠在新皮质点燃后出现 SRS，而 Nf1 小鼠中有 33%出现。WT 组和 Nf1 组的平均初始海马 ADT 无显著差异，达到点燃状态所需的刺激次数也无差异。WT 组在所有点燃疗程中的平均发作严重程度均显著高于 Nf1 小鼠。WT 组的 Racine 发作评分在点燃疗程中的进展也更快，主要是由于在点燃过程早期发作严重程度的快速增加。然而，在改良的扩展点燃后，有 50%的 Nf1 小鼠出现 SRS，而没有 WT 小鼠出现。Western blot 显示，海马点燃增加了 WT 和 Nf1 小鼠中磷酸化/总 Akt 的比值，而新皮质点燃仅导致 Nf1 小鼠中磷酸化/总 Akt 和 MAPK 的比值增加。
我们首次证明了 NF1 动物模型中癫痫发生的发生率增加，而该模型没有已知的宏观/肿瘤性脑病变。这项工作为遗传突变本身在 NF1 患者的发作和癫痫中发挥作用提供了证据，并支持在未来的机制研究中使用 Nf1 小鼠模型。
