Early mechanisms mediating the organization of the axon initial segment impact the formation of axo-axonic synapses

项目摘要 大脑发育是一个复杂的过程，需要大量细胞的产生和分化， 细胞类型，以及连接神经回路的空间和时间精度的额外挑战 这些细胞。神经回路的不正确形成会导致对大脑活动模式的控制受损， 被广泛认为是导致许多儿童神经系统疾病的原因。多方面的研究表明 抑制性GABA能回路尤其有助于神经发育障碍的病理生理学， 然而，这些疾病仍然没有得到很好的治疗。该项目的重点是GABA能突触形成的- 在一个枝形细胞和一个称为轴突起始段（AIS）的专门神经元隔室之间，形成 轴-轴突触枝形细胞对神经活动模式施加强大的控制， 在AIS处抑制大群主细胞，这影响它们产生电刺激的可能性。 信号了轴-轴突触的特征是含有α2亚单位和collybistin的GABAA受体， 我们最近发现的一种特殊的相互作用伙伴。我们开发了一种小鼠模型， GABAA受体α2亚基（Gabra 2 -1）中的置换突变减少了与大肠杆菌的相互作用， bistin，导致轴突-轴突突触数量减少和发育过程中的自发性癫痫发作。对 在此前提下，我们假设α2亚基有助于AIS的组织，以促进 轴-轴突触的形成。在这个提议中，我们将研究轴-轴突触的发育， AIS组织使用免疫组织化学和扩张显微镜，以及电路功能障碍使用 皮层场记录我们将研究这些变化如何与Gabra 2 -1中的症状发作相关。 小鼠，并评估新的方法，以取代失去控制，通过使用一种新的光激活阳离子通道 针对AIS。与人类健康的相关性：拟议的项目预计将产生详细的信息 关于AIS在发育过程中是如何组织的，以及两种典型的轴突-轴突突触的形成， 和病理条件，提供有关大脑发育和翻译见解的基础知识， 治疗神经发育障碍该建议还提供了一种主动操纵AIS的新方法， 以其控制大脑皮层活动模式的潜力而闻名。结果：这项工作将完全由 由UNLV的本科生和研究生组成，UNLV是美国最多样化的校园之一。因此，在本发明中， 该项目将为生物医学劳动力的多样化提供有用的培训机会， 并改善我们机构的研究环境。

PROJECT SUMMARY Brain development is a complex process that requires the production and differentiation of numerous cells and cell types, with the additional challenge of spatial and temporal precision in the neural circuitry that connects these cells. Improper formation of neural circuitry leads to impaired control over brain activity patterns and is broadly thought to contribute to a number of childhood neurological disorders. Multiple lines of research suggest that inhibitory GABAergic circuitry in particular contributes to the pathophysiology of neurodevelopmental disor- ders, yet these disorders remain poorly treated. This project is focused on the GABAergic synapse formed be- tween a chandelier cell and a specialized neuronal compartment called the axon initial segment (AIS), forming an axo-axonic synapse. Chandelier cells exert powerful control over neural activity patterns exerting shunting inhibition at the AIS of large groups of principal cells, which impacts their probability of generating an electrical signal. Axo-axonic synapses are characterized by GABAA receptors containing the α2 subunit and collybistin, a specific interacting partner we have recently identified. We have developed and characterized a mouse model with a substitution mutation in the GABAA receptor α2 subunit (Gabra2-1) that diminishes interaction with colly- bistin, causing reduced numbers of axo-axonic synapses and spontaneous seizures during development. On this premise, we hypothesize that the α2 subunit contributes to the organization of the AIS in order to facilitate the formation of axo-axonic synapses. In this proposal we will examine axo-axonic synapse development and AIS organization using immunohistochemistry and expansion microscopy, as well as circuit dysfunction using cortical field recordings. We will examine how these changes correlate with symptom onset in the Gabra2-1 mouse and also evaluate novel methods to replace lost control by using a novel light activated cation channel targeted to the AIS. Relevance to human health: The proposed project is expected to yield detailed information about how the AIS organizes during development, as well as the formation of axo-axonic synapses in both typical and pathological conditions, providing both basic knowledge about brain development and translational insights for neurodevelopmental disorders. This proposal also offers a novel means of actively manipulating the AIS, a site known for its potential to control cortical activity patterns. Outcomes: This work will be conducted entirely by undergraduate and graduate students at UNLV, which is one of the nation’s most diverse campuses. Thus, this project will provide instrumental training opportunities for the diversification of the biomedical workforce, as well as enhance the research environment of our institution.