Center for Genetics and Developmental Systems Biology, Department of Obstetrics & Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China.
Key Laboratory of Mental Health of the Ministry of Education, Guangdong-Hong Kong-Macao Greater Bay Area Center for Brain Science and Brain-Inspired Intelligence, Guangdong Key Laboratory of Psychiatric Disorders, Collaborative Innovation Center for Brain Science, Department of Neurobiology, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, China.
Theranostics. 2021 Sep 7;11(19):9296-9310. doi: 10.7150/thno.60531. eCollection 2021.
Mutations in serotonin pathway genes, especially the serotonergic receptor subunit gene , are associated with autism. However, the association of deficiency with autism and the underlying mechanisms remain unknown. The knockout (KO) mice were generated using transcription activator-like effector nuclease technology. Various behavior tests, including social interaction, social approach task, olfactory habituation/dishabituation, self-grooming, novel object recognition, contextual fear conditioning, elevated plus maze, open field and seizure susceptibility, were performed to assess the phenotypes. Transcriptome sequencing was carried out to search for molecular network and pathways underlying the phenotypes. Electrophysiological recordings, immunoblotting, immunofluorescence staining, immunoprecipitation, and quantitative real-time PCR were performed to verify the potential mechanisms. The N-methyl-D-aspartate receptor (NMDAR) antagonist memantine was used to treat the KO mice for rescuing the phenotypes. The KO mouse model showed three phenotypic domains: autistic-like behaviors (including impaired social behavior, cognitive deficits, and increased repetitive self-grooming), impaired memory, and attenuated susceptibility to pentylenetetrazol-induced seizures. We observed enhanced action potential-driven γ-aminobutyric acid-ergic (GABAergic) transmission in pyramidal neurons and decreased excitatory/inhibitory (E/I) ratio using the patch-clamp recording. Transcriptome sequencing on the hippocampus revealed the converged pathways of the dysregulated molecular networks underlying three phenotypic domains with upregulation of NMDAR. We speculated that KO promotes an increase in GABA release through NMDAR upregulation. The electrophysiological recordings on hippocampal parvalbumin-positive (PV) interneuron revealed increased NMDAR current and NMDAR-dependent excitability. The NMDAR antagonist memantine could rescue GABAergic transmission in the hippocampus and ameliorate autistic-like behaviors of the KO mice. Our data indicated that upregulation of the NMDAR in PV interneurons may play a critical role in regulating GABAergic input to pyramidal neurons and maybe involve in the pathogenesis of autism associated with deficiency. Therefore, we suggest that the NMDAR system could be considered potential therapeutic target for autism.
5-羟色胺（serotonin）途径基因，特别是 5-羟色胺能受体亚基基因的突变，与自闭症有关。然而， 缺乏与自闭症的关联及其潜在机制尚不清楚。使用转录激活因子样效应物核酸酶（transcription activator-like effector nuclease，TALEN）技术生成 敲除（knockout，KO）小鼠。进行各种行为测试，包括社交互动、社交接近任务、嗅觉习惯化/脱敏、自我梳理、新物体识别、情境恐惧条件反射、高架十字迷宫、旷场和癫痫易感性，以评估表型。进行转录组测序以搜索潜在的分子网络和途径。进行电生理记录、免疫印迹、免疫荧光染色、免疫沉淀和定量实时聚合酶链反应（polymerase chain reaction，PCR）以验证潜在机制。使用 N-甲基-D-天冬氨酸受体（N-methyl-D-aspartate receptor，NMDAR）拮抗剂美金刚（memantine）治疗 KO 小鼠以挽救表型。 KO 小鼠模型表现出三个表型领域：自闭症样行为（包括社交行为受损、认知缺陷和自我梳理增加）、记忆受损和戊四氮（pentylenetetrazol，PTZ）诱导的癫痫发作易感性降低。使用膜片钳记录观察到锥体神经元中动作电位驱动的γ-氨基丁酸能（GABAergic）传递增强，兴奋性/抑制性（excitatory/inhibitory，E/I）比值降低。对海马进行转录组测序揭示了三个表型领域中失调分子网络的趋同途径，其中 NMDAR 上调。我们推测 KO 通过上调 NMDAR 促进 GABA 释放增加。对海马 parvalbumin 阳性（parvalbumin-positive，PV）中间神经元的电生理记录显示 NMDAR 电流和 NMDAR 依赖性兴奋性增加。NMDAR 拮抗剂美金刚可挽救 KO 小鼠海马中的 GABA 能传递并改善其自闭症样行为。 我们的数据表明，PV 中间神经元中 NMDAR 的上调可能在调节 GABA 能传入到锥体神经元中起关键作用，并且可能与 缺乏相关的自闭症发病机制有关。因此，我们认为 NMDAR 系统可能是自闭症的潜在治疗靶点。
