SH3 and multiple ankyrin repeat domains protein 3 (SHANK3) monogenic mutations or deficiency leads to excessive stereotypic behavior and impaired sociability, which frequently occur in autism cases. To date, the underlying mechanisms by which Shank3 mutation or deletion causes autism and the part of the brain in which Shank3 mutation leads to the autistic phenotypes are understudied. The hypothalamus is associated with stereotypic behavior and sociability. p38α, a mediator of inflammatory responses in the brain, has been postulated as a potential gene for certain cases of autism occurrence. However, it is unclear whether hypothalamus and p38α are involved in the development of autism caused by Shank3 mutations or deficiency. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis and immunoblotting were used to assess alternated signaling pathways in the hypothalamus of Shank3 knockout (Shank3−/−) mice. Home-Cage real-time monitoring test was performed to record stereotypic behavior and three-chamber test was used to monitor the sociability of mice. Adeno-associated viruses 9 (AAV9) were used to express p38α in the arcuate nucleus (ARC) or agouti-related peptide (AgRP) neurons. D176A and F327S mutations expressed constitutively active p38α. T180A and Y182F mutations expressed inactive p38α. We found that Shank3 controls stereotypic behavior and sociability by regulating p38α activity in AgRP neurons. Phosphorylated p38 level in hypothalamus is significantly enhanced in Shank3−/− mice. Consistently, overexpression of p38α in ARC or AgRP neurons elicits excessive stereotypic behavior and impairs sociability in wild-type (WT) mice. Notably, activated p38α in AgRP neurons increases stereotypic behavior and impairs sociability. Conversely, inactivated p38α in AgRP neurons significantly ameliorates autistic behaviors of Shank3−/− mice. In contrast, activated p38α in pro-opiomelanocortin (POMC) neurons does not affect stereotypic behavior and sociability in mice. We demonstrated that SHANK3 regulates the phosphorylated p38 level in the hypothalamus and inactivated p38α in AgRP neurons significantly ameliorates autistic behaviors of Shank3−/− mice. However, we did not clarify the biochemical mechanism of SHANK3 inhibiting p38α in AgRP neurons. These results demonstrate that the Shank3 deficiency caused autistic-like behaviors by activating p38α signaling in AgRP neurons, suggesting that p38α signaling in AgRP neurons is a potential therapeutic target for Shank3 mutant-related autism.

中文翻译：

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Shank3缺陷通过激活下丘脑AgRP神经元中的p38α引发自闭症样行为

SH3 和多个锚蛋白重复结构域蛋白 3 (SHANK3) 单基因突变或缺陷会导致过度刻板行为和社交能力受损，这在自闭症病例中经常发生。迄今为止，Shank3 突变或缺失导致自闭症的潜在机制以及 Shank3 突变导致自闭症表型的大脑部分尚未得到充分研究。下丘脑与刻板行为和社交能力有关。 p38α 是大脑炎症反应的介质，被认为是某些自闭症发生的潜在基因。然而，尚不清楚下丘脑和p38α是否参与Shank3突变或缺陷引起的自闭症的发生。京都基因和基因组百科全书 (KEGG) 通路分析和免疫印迹用于评估 Shank3 敲除 (Shank3−/−) 小鼠下丘脑的交替信号通路。采用家笼实时监测试验记录小鼠的刻板行为，采用三室试验监测小鼠的社交能力。腺相关病毒 9 (AAV9) 用于在弓状核 (ARC) 或刺鼠相关肽 (AgRP) 神经元中表达 p38α。 D176A 和 F327S 突变表达组成型活性 p38α。 T180A 和 Y182F 突变表达无活性的 p38α。我们发现 Shank3 通过调节 AgRP 神经元中的 p38α 活性来控制刻板行为和社交能力。 Shank3−/− 小鼠下丘脑的磷酸化 p38 水平显着增强。一致地，ARC 或 AgRP 神经元中 p38α 的过度表达会引发过度的刻板行为并损害野生型 (WT) 小鼠的社交能力。值得注意的是，AgRP 神经元中激活的 p38α 会增加刻板行为并损害社交能力。相反，AgRP 神经元中失活的 p38α 显着改善了 Shank3−/− 小鼠的自闭症行为。相比之下，阿片黑皮质素原 (POMC) 神经元中激活的 p38α 不会影响小鼠的刻板行为和社交能力。我们证明 SHANK3 调节下丘脑中磷酸化的 p38 水平，AgRP 神经元中失活的 p38α 显着改善 Shank3−/− 小鼠的自闭症行为。然而，我们并没有阐明 SHANK3 抑制 AgRP 神经元中 p38α 的生化机制。这些结果表明，Shank3 缺陷通过激活 AgRP 神经元中的 p38α 信号传导导致类似自闭症的行为，这表明 AgRP 神经元中的 p38α 信号传导是 Shank3 突变相关自闭症的潜在治疗靶点。