Overexpressing wild-type γ2 subunits rescued the seizure phenotype in Gabrg2(+/Q390X) Dravet syndrome mice.
Overexpressing wild-type γ2 subunits rescued the seizure phenotype in Gabrg2(+/Q390X) Dravet syndrome mice.
复制标题
DOI：
10.1111/epi.13810
复制
发表时间：
2017-08
期刊：
影响因子：
5.6
通讯作者：
MacDonald RL
中科院分区：
文献类型：
--
作者：
Huang X;Zhou C;Tian M;Kang JQ;Shen W;Verdier K;Pimenta A;MacDonald RL
突变型γ-氨基丁酸A型（GABAA）受体γ2（Q390 X）亚基（成熟肽中的Q351 X）与癫痫性脑病、Dravet综合征和癫痫综合征遗传性癫痫伴热性惊厥（GEFS+）相关。该突变产生了一个提前终止密码子，导致稳定的截短和错误折叠的γ2亚基的翻译，该亚基在神经元中积累，形成细胞内聚集体，破坏γ2亚基掺入GABAA受体，并影响配对α和β亚基的运输。杂合子Gabrg 2 +/Q390 X基因敲入（KI）小鼠的皮层抑制减少，EEG上的棘波放电，对惊厥药物戊四氮（PTZ）的癫痫发作阈值降低，自发性全身强直阵挛性癫痫发作。在这项主要研究的证据中，我们试图使用γ2亚基基因（GABRG 2）替代疗法来挽救KI小鼠的这些缺陷。我们通过将Gabrg 2 +/Q390 X KI小鼠与过表达HA（血凝素）标记的人γ 2 HA亚基的细菌人工染色体（BAC）转基因小鼠杂交，引入GABRG 2等位基因，并通过Western印迹和免疫组织化学染色比较GABAA受体亚基表达，通过监测PTZ注射后小鼠行为来比较癫痫发作阈值，通过切片记录来比较丘脑皮质抑制和网络振荡。与KI小鼠相比，携带突变等位基因和转基因的成年小鼠具有增加的野生型γ2和配对α1和β2/3亚基，增加的从第VI层皮层神经元记录的微型抑制性突触后电流（mIPSC）幅度，减少的丘脑皮层网络振荡和更高的PTZ癫痫发作阈值。基于这些结果，我们认为，癫痫突变体γ2（Q390 X）亚基引起的遗传性癫痫综合征中的癫痫发作具有显性负效应，可以通过野生型γ2亚基的过表达来挽救。
The mutant γ-aminobutyric acid type A (GABAA) receptor γ2(Q390X) subunit, (Q351X in the mature peptide) has been associated with the epileptic encephalopathy, Dravet syndrome, and the epilepsy syndrome genetic epilepsy with febrile seizures plus (GEFS+). The mutation generates a premature stop codon that results in translation of a stable truncated and misfolded γ2 subunit that accumulates in neurons, forms intracellular aggregates, disrupts incorporation of γ2 subunits into GABAA receptors and affects trafficking of partnering α and β subunits. Heterozygous Gabrg2+/Q390X knock-in (KI) mice had reduced cortical inhibition, spike wave discharges on EEG, a lower seizure threshold to the convulsant drug pentylenetetrazol (PTZ) and spontaneous generalized tonic-clonic seizures. In this proof of principal study, we attempted to rescue these deficits in KI mice using a γ2 subunit gene (GABRG2) replacement therapy. We introduced the GABRG2 allele by crossing Gabrg2+/Q390X KI mice with bacterial artificial chromosome (BAC) transgenic mice overexpressing HA (hemagglutinin) tagged human γ2HA subunits and compared GABAA receptor subunit expression by western blot and immunohistochemical staining, seizure threshold by monitoring mouse behavior after PTZ-injection, and thalamocortical inhibition and network oscillation by slice recording. Compared to KI mice, adult mice carrying both mutant allele and transgene had increased wild-type γ2 and partnering α1 and β2/3 subunits, increased miniature inhibitory postsynaptic current (mIPSC) amplitudes recorded from layer VI cortical neurons, reduced thalamocortical network oscillations, and higher PTZ seizure threshold. Based on these results we suggest that seizures in a genetic epilepsy syndrome caused by epilepsy mutant γ2(Q390X) subunits with dominant negative effects could be rescued potentially by overexpression of wild-type γ2 subunits.
登录
查看更多内容
影响因子：
5.6
作者：
Blumenfeld, H
通讯作者：
Blumenfeld, H
影响因子：
2.5
作者：
Salin, PA;Prince, DA
通讯作者：
Prince, DA
影响因子：
25
作者：
Kang, Jing-Qiong;Shen, Wangzhen;Zhou, Chengwen;Xu, Dong;Macdonald, Robert L.
通讯作者：
Macdonald, Robert L.
影响因子：
16.2
作者：
Beenhakker, Mark P.;Huguenard, John R.
通讯作者：
Huguenard, John R.
影响因子：
14.5
作者：
Marini, C;Harkin, LA;Berkovic, SF
通讯作者：
Berkovic, SF