AAV vector-mediated overexpression of CB1 cannabinoid receptor in pyramidal neurons of the hippocampus protects against seizure-induced excitoxicity.
AAV vector-mediated overexpression of CB1 cannabinoid receptor in pyramidal neurons of the hippocampus protects against seizure-induced excitoxicity.
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DOI：
10.1371/journal.pone.0015707
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发表时间：
2010-12-21
期刊：
影响因子：
3.7
通讯作者：
Klugmann M
中科院分区：
文献类型：
--
作者：
Guggenhuber S;Monory K;Lutz B;Klugmann M
CB1 大麻素受体是大脑中最丰富的 G 蛋白偶联受体，也是神经元兴奋性的关键调节因子。有强有力的证据表明，谷氨酸能海马神经元上的 CB1 受体有助于减轻小鼠和人类的癫痫样发作。因此，我们假设通过实验增加主要神经元中的 CB1 基因剂量将对红藻氨酸 (KA) 诱导的海马发病机制产生治疗作用。在这里，我们表明，病毒介导的海马锥体细胞和苔藓细胞中 CB1 受体的条件性过度表达具有神经保护作用，并能减轻小鼠急性 KA 癫痫模型中的惊厥。我们引入了重组腺相关病毒 (AAV) 基因组，该基因组具有侧翼为 loxP 位点的短终止元件，可在转录水平上高效减弱转基因表达。 Cre 重组酶的存在对于报告蛋白或 CB1 受体的体外和体内表达是严格必需的。将 AAV 立体定向递送至驱动小鼠 NEX-cre 的背侧海马后，转基因 CB1 受体免疫反应性靶向谷氨酸能神经元。 AAV 处理的 Cre-小鼠海马裂解物中 CB1 受体蛋白水平增加，同时大麻素诱导的 G 蛋白激活增强。与 AAV 治疗的对照动物相比，CB1 受体过表达者中 KA 诱导的癫痫发作严重程度和死亡率降低。在 AAV 处理的 Cre 转基因细胞中，海马 CA3 区的神经元损伤特别不存在，但在两个治疗组的整个皮质区域中都很明显。我们的数据提供了进一步的证据，证明锥体海马神经元中 CB1 信号传导的增加可以防止过度兴奋性网络活动的不利影响。
The CB1 cannabinoid receptor is the most abundant G-protein coupled receptor in the brain and a key regulator of neuronal excitability. There is strong evidence that CB1 receptor on glutamatergic hippocampal neurons is beneficial to alleviate epileptiform seizures in mouse and man. Therefore, we hypothesized that experimentally increased CB1 gene dosage in principal neurons would have therapeutic effects in kainic acid (KA)-induced hippocampal pathogenesis. Here, we show that virus-mediated conditional overexpression of CB1 receptor in pyramidal and mossy cells of the hippocampus is neuroprotective and moderates convulsions in the acute KA seizure model in mice. We introduce a recombinant adeno-associated virus (AAV) genome with a short stop element flanked by loxP sites, for highly efficient attenuation of transgene expression on the transcriptional level. The presence of Cre-recombinase is strictly necessary for expression of reporter proteins or CB1 receptor in vitro and in vivo. Transgenic CB1 receptor immunoreactivity is targeted to glutamatergic neurons after stereotaxic delivery of AAV to the dorsal hippocampus of the driver mice NEX-cre. Increased CB1 receptor protein levels in hippocampal lysates of AAV-treated Cre-mice is paralleled by enhanced cannabinoid-induced G-protein activation. KA-induced seizure severity and mortality is reduced in CB1 receptor overexpressors compared with AAV-treated control animals. Neuronal damage in the hippocampal CA3 field is specifically absent from AAV-treated Cre-transgenics, but evident throughout cortical areas of both treatment groups. Our data provide further evidence for a role of increased CB1 signaling in pyramidal hippocampal neurons as a safeguard against the adverse effects of excessive excitatory network activity.
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