Regional and trans-regional heterogeneity of astrocyte morphology as a functional determinant of synaptic astrocyte-neuron interactions in the hippocampus

突触传递及其可塑性是由神经元和星形胶质细胞之间在突触水平上的快速反向信号交换调节的。这种星形细胞-神经元信号通常包括扩散性信号。因此，其效率可能取决于神经元结构和突触周围星形胶质细胞过程之间的空间接近程度，从而取决于星形胶质细胞的形态。因此，星形胶质细胞形态的异质性应该对突触传递具有重要意义。事实上，我们可以证明啮齿类动物CA1辐射层星形胶质细胞的结构异质性对CA3-CA1突触的突触前释放有重要影响，CA3-CA1突触是细胞神经生理学的标准模型系统。利用双光子激发荧光显微镜和新型电生理方法的结合，我们可以揭示星形胶质细胞结构复杂性的变化通过星形胶质细胞谷氨酸受体决定突触前短期可塑性，可能是通过星形胶质细胞过程对突触的不同覆盖。在这个项目中，我们将在这一发现的基础上，首先揭示星形胶质细胞形态的区域细胞间异质性如何决定谷氨酸能CA3-CA1突触和gaba能连接的局部覆盖。为此，我们建立了扩展显微镜。ExM使我们能够超分辨突触和突触周围星形胶质细胞结构之间的结构关系，这是衍射限制显微镜无法完全可视化的。其次，我们将把我们的研究扩展到CA1区域的邻近层，在那里星形胶质细胞显示不同的形态，并覆盖相同CA1锥体细胞神经元的其他树突结构域及其传入的兴奋连接。通过将成像和电生理技术与ExM相结合，我们将揭示星形胶质细胞形态的跨区域异质性如何决定单一神经元细胞类型接收的兴奋性突触的局部特性。第三，我们将建立星形胶质细胞形态及其细胞间和亚细胞特化如何决定星形胶质细胞对突触活动的反应。在这里，我们将重点关注星形胶质细胞过程与谷氨酸能突触的接近性如何决定突触刺激导致局部星形胶质细胞Ca2+瞬变的可能性，以及它如何影响它们的大小和潜在的信号级联。这些功能实验将再次得到使用ExM的结构分析的支持。第四，我们将探索人类组织样本中星形胶质细胞结构及其异质性与突触星形胶质细胞-神经元相互作用之间的关系。总之，我们将确定星形胶质细胞的区域和跨区域结构异质性和特化如何决定星形胶质细胞过程的局部突触覆盖以及星形胶质细胞-神经元相互作用的功能相关性。

Synaptic transmission and its plasticity are modulated by fast reciprocal signal exchange between neurons and astrocytes on the synaptic level. This astrocyte-neuron signalling often involves diffusible signals. Therefore, its efficiency is likely to depend on the spatial proximity between neuronal structures and perisynaptic astrocyte processes and thus on astrocyte morphology. As a consequence, the heterogeneity of astrocyte morphology should have important implications for synaptic transmission. Indeed, we could demonstrate that the structural heterogeneity of astrocytes in the rodent CA1 stratum radiatum has important consequences for presynaptic release at the CA3-CA1 synapse, a standard model system in cellular neurophysiology. Using combinations of two-photon excitation fluorescence microscopy and novel electrophysiological approaches we could reveal that variations of the structural complexity of astrocytes determines presynaptic short-term plasticity via astrocytic glutamate receptors, likely through differential coverage of synapses by astrocyte processes. In this project, we will now build on this finding by first uncovering how regional intercellular heterogeneity of astrocyte morphology determines the local coverage of glutamatergic CA3-CA1 synapses and also GABAergic connections. To this end, we have established expansion microscopy. ExM enables us to super-resolve the structural relationship between synapses and perisynaptic astrocyte structures, which cannot be fully visualized by diffraction-limited microscopy. Second, we will extend our investigation to adjacent layers of the CA1 region, where astrocytes display different morphologies and cover other dendritic domains of the same CA1 pyramidal cell neurons and their incoming excitatory connections. By combining imaging and electrophysiological techniques with ExM we will reveal how trans-regional heterogeneity of astrocyte morphology determines the local properties of excitatory synapses received by a single neuronal cell type. Third, we will establish how astrocyte morphology and its intercellular and subcellular specialisation determines the responsiveness of astrocyte to synaptic activity. Here we will focus on how the proximity of astrocyte processes to glutamatergic synapses determines the likelihood that a synaptic stimulus results in a local astrocytic Ca2+ transients and how it affects their magnitude and underlying signalling cascades. These functional experiments will again be supported by structural analysis using ExM. Fourth, we will explore the relationship between astrocyte structure, its heterogeneity and synaptic astrocyte-neuron interactions in human tissue samples. In summary, we will establish how regional and trans-regional structural heterogeneity and specialisation of astrocytes determine local synaptic coverage by astrocyte processes and the functional relevance for astrocyte-neuron interactions.