Sex-specific mechanisms of cortical circuit dysfunction in a mouse ASD model

尽管自闭症谱系障碍 (ASD) 多见于男性，但自闭症谱系障碍 (ASD) 的行为表现以及神经解剖学特征 与 ASD 相关的变化和脑功能障碍在男性和女性之间有所不同。鲜为人知的是 自闭症谱系障碍基因如何影响女性大脑。雌激素，通过膜结合雌激素受体 α (ERα) 发挥作用， 选择性地在雌性神经元中与 Gq 偶联的第 1 组 mGluR、mGluR1 和 mGluR5 相互作用并激活 在不同的大脑区域中调节信号传导、神经生理学和行为。 mGluR 的功能改变， 主要是 mGluR5，与 ASD 小鼠模型的病理生理学密切相关，尤其是 Fragile X 综合症，但许多其他综合症和 mGluR5 拮抗剂正在 FXS 儿童的临床试验中。我们是第一个 在 FXS 小鼠模型 Fmr1 KO 中发现 mGluR5 功能过度活跃，并显示了这一结果 mGluR5 复合物异常； mGluR5 与其突触后支架蛋白 Homer 分离，该蛋白在 转，导致组成型 mGluR5 活性、下游效应器信号异常以及与疾病相关 表型，例如电路过度兴奋。尽管有 mGluR5 在 ASD 病理生理学中的证据， 已知雌激素对 mGluR1/5 的性别依赖性调节，但与 ASD 相关的基因如何与雌激素相互作用却知之甚少。 雌激素影响 mGluR1/5 功能以及对 ASD 相关神经生理学和行为的影响。 我们发现了性别特异性、mGluR5依赖性的感觉新皮质回路功能障碍 因 Pten 缺失（磷酸酶和张力蛋白同源物缺失）而产生的 ASD 小鼠模型 10 号染色体），PI3K/mTORC1 通路的抑制因子。具体来说，我们观察到过度兴奋 两种不同 PTEN 缺失模型的女性中的新皮质回路振荡，称为 UP 状态；一个胚胎， 海马和第 5 层新皮质神经元中 Pten 的敲除，以及种系 Pten 杂合小鼠 (Pten-het)，这是人类 Pten 相关 ASD 的遗传有效模型。过度兴奋的 UP 状态 女性 Pten 模型通过 mGluR5 或 ERα 的急性拮抗作用进行纠正，表明 mGluR5 过度活跃- ERα信号传导。增强 mGluR5-ERα 活性的候选分子底物是观察到的 mGluR5-ERα 活性增加 女性 Pten-het 皮质中的 mGluR5-ERα 和 mGluR5-Homer 复合物减少。我们假设一个 Pten 缺失的雌性神经元中 mGluR5 与 ERa 和 Homer 相互作用的不平衡导致 增强 mGluR5 信号对雌激素的反应，作用于 ERα，并具有组成性活性 mGluR5 会导致过度兴奋的皮质振荡和 ASD 相关行为。我们建议 以下旨在检验这一假设： 目标 1：确定 Pten 和 ERα 的性别依赖性相互作用 皮质回路功能障碍和 mGluR5 复合物调节。目标 2：检查 mGluR5 的性别特异性效应 和 ERα 对 Pten 删除的第 5 层神经元的内在和突触特性的影响。目标 3：确定性别特异性， Pten 缺失模型中脑电图显示 mGluR5 和 ERα 依赖性体内皮质振荡。目标 4. 确定 女性特有的、Pten 依赖性皮质回路过度兴奋在 ASD 相关行为中的作用。

Although ASD is more diagnosed in males, the behavioral manifestations of ASD, as well as the neuroanatomical changes and brain dysfunction associated with ASD, differ between males and females. There is little known of how ASD genes impact the female brain. Estrogen, acting through membrane bound estrogen receptor α (ERα), interacts with and activates Gq-coupled Group 1 mGluRs, mGluR1 and mGluR5, selectively in female neurons in diverse brain regions to regulate signaling, neurophysiology, and behavior. Altered functioning of mGluRs, primarily mGluR5, is strongly implicated in the pathophysiology of ASD mouse models, most notably Fragile X Syndrome, but many others, and mGluR5 antagonists are in clinical trials of FXS children. We were the first to discover hyperactivity of mGluR5 function in the FXS mouse model, Fmr1 KO, and show this results from an abnormal mGluR5 complex; mGluR5 is dissociated from its postsynaptic scaffolding protein, Homer, which, in turn, leads to constitutive mGluR5 activity, abnormal signaling to downstream effectors and disease relevant phenotypes, such as circuit hyperexcitability. Despite the evidence for mGluR5 in ASD pathophysiology, the known sex-dependent regulation of mGluR1/5 by estrogen, little is known of how ASD-linked genes interact with estrogen to affect mGluR1/5 function and the consequences on ASD-relevant neurophysiology and behavior. We have discovered a sex-specific, mGluR5 -dependent dysfunction of sensory neocortical circuits in a mouse model of ASD that results from deletion of Pten (Phosphatase and tensin homolog deleted on chromosome 10), a suppressor of the PI3K/mTORC1 pathway. Specifically, we observe hyperexcitable neocortical circuit oscillations, termed UP states, in females of two distinct PTEN deletion models; an embryonic, knockout of Pten in hippocampus and layer 5 neocortical neurons, and a germline Pten heterozygous mouse (Pten-het), which is a genetically valid model for Pten-related ASD in humans. Hyperexcitable UP states in female Pten models are corrected by acute antagonism of mGluR5 or ERα, indicative of hyperactive mGluR5- ERα signaling. A candidate molecular substrate for enhanced mGluR5-ERα activity is the observed increase in mGluR5-ERα, and decreased mGluR5-Homer, complexes in female Pten-het cortex. We hypothesize that an imbalance in mGluR5 interactions with ERa and Homer in Pten deleted female neurons results in enhanced mGluR5-signaling in response to estrogen, acting on ERα, as well as constitutively active mGluR5 which leads to hyperexcitable cortical oscillations and ASD-relevant behaviors. We propose the following aims to test this hypothesis: Aim 1: Determine the sex-dependent interactions of Pten and ERα in cortical circuit dysfunction and mGluR5 complex regulation. Aim 2: Examine the sex-specific effects of mGluR5 and ERα on intrinsic and synaptic properties of Pten-deleted layer 5 neurons. Aim 3: Determine the sex-specific, mGluR5- and ERα-dependent in vivo cortical oscillations with EEG in Pten deletion models. Aim 4. Determine the role of female-specific, Pten-dependent cortical circuit hyperexcitability in ASD-relevant behaviors.