Imaging dendritic spine abnormalities and circuit defects in fragile X mice.

描述(申请人提供)：脆性X综合征(FXS)是最常见的遗传性智力障碍形式，也是导致自闭症的最常见的单基因原因。对FXS的动物模型Fmr1基因敲除(KO)小鼠的研究发现，大脑中存在两个主要缺陷。第一个是树突棘的结构异常，树突棘是皮层兴奋性突触的主要接受者，第二个是突触和经验依赖性可塑性的功能异常。利用活体双光子显微镜，我们和其他人发现Fmr1KO小鼠皮质锥体神经元树突棘的稳定和成熟存在发育延迟，这可能是FXS最早的突触缺陷之一。现在，我们将测试这一假设，即由感觉体验引发的电路重构与脊柱动力学和大小密切相关，从而协调Fmr1 KO小鼠的结构和功能表型。我们还将用电子显微镜在超微结构水平上研究突触的完整性，以及皮质发育过程中轴突和轴突的动态，以确定它们在突变小鼠中是否也发生了变化。此外，利用体内双光子钙成像和电生理学记录完整回路中神经元的活动，我们发现Fmr1KO小鼠的锥体神经元表现出异常高的放电频率和同步性，这可以解释这些小鼠学习障碍和低惊厥阈值的原因。在这里，我们将测试这种网络超兴奋性转化为感觉诱发活动问题的假设，并将调查KO小鼠中这些电路水平的问题是否可以通过影响脑干神经调节和抑制通路的药物来挽救。实验设计采用了尖端的活体成像技术，并试图解决FXS中的重要知识空白和有争议的问题。由于树突棘异常和脆性X智力低下蛋白调控的许多信号通路也与其他神经发育障碍有关，我们认为我们独特的突触到回路的方法具有非常高的意义，可能对许多类型的自闭症和智能障碍具有广泛的重要性。

DESCRIPTION (provided by applicant): Fragile X syndrome (FXS) is the most common inherited form of intellectual impairment and the most common single gene cause of autism. Research in Fmr1 knockout (KO) mice, an animal model of FXS, has identified two major defects in the brain. The first is a structural abnormality in dendritic spines, the major recipiens of excitatory synapses in the cortex, and the second is a functional abnormality in synaptic and experience- dependent plasticity. Using in vivo two-photon microscopy, we and others have identified a developmental delay in the stabilization and maturation of dendritic spines of cortica pyramidal neurons in Fmr1 KO mice, which may be one of the earliest synaptic defects in FXS. Now, we will test the hypothesis that circuit remodeling triggered by sensory experience is intimately tied to the spine dynamics and size, thereby reconciling the structural and functional phenotypes of Fmr1 KO mice. We will also investigate synapse integrity at the ultrastructural level with electron microscopy, as well as the dynamics of axons and their boutons during cortical development, in order to ascertain whether they are also altered in mutant mice. In addition, using in vivo two-photon calcium imaging and electrophysiology to record neuronal activity in intact circuits, we have shown that pyramidal neurons in Fmr1 KO mice show abnormally high firing rates and synchrony, which could explain the deficits in learning and low seizure threshold in these mice. Here, we will test the hypothesis that this network hyperexcitability translates into problems with sensory-evoked activity and we will investigate whether these circuit-level problems in KO mice can be rescued with drugs that affect brainstem neuromodulation and inhibitory pathways. The experimental design employs cutting edge in vivo imaging techniques and seeks to address important knowledge gaps and controversial issues in FXS. Because dendritic spine abnormalities and many of the signaling pathways regulated by the fragile X mental retardation protein are also implicated in other neurodevelopmental disorders, we believe that our unique synapse-to- circuit approach has a very high significance and is likely to be of broad importance to many types of autism and mental impairment.