CRCNS US-German Research Proposal: Quantitative and Computational Dissection of Glutamatergic Crosstalk at Tripartite Synapses

CRCNS美国-德国研究提案：定量和计算剖析 三体突触的谷氨酸能串扰 (1)克里斯汀·R·罗斯，德国杜塞尔多夫海因里希·海涅大学 (2)克里斯蒂安·亨内伯格，德国波恩大学 (3)加尼姆·乌拉，美国佛罗里达州坦帕市南佛罗里达大学 项目说明 1导言和背景 化学突触的传递是信息传递的中心机制 在神经元之间转移。突触连接，如谷氨酸能兴奋 突触通常被认为是点对点连接，并被建模为点对点连接。然而，还有 大量证据表明，当不同的谷氨酸能突触之间发生串扰时 突触前释放的谷氨酸不仅由其直接的突触后伙伴感受到 但也受同一神经元和其他神经元附近的突触影响[4]。值得注意的是，这种现象 所谓的“谷氨酸溢出”不仅定义了给定突触的输入专一性 连接及其与相邻突触的串扰，但也参与和控制 依赖活性的塑性[1，7，8]。 谷氨酸从其释放部位逃逸的容易程度以及它扩散到组织中的程度 取决于细胞外间隙(ECS)AS的形态和分子特性 以及谷氨酸清除的效果，这主要取决于星形胶质细胞的摄取 [11，12]。我们和其他人已经证明，突触周围摄取谷氨酸的功效是通过 星形胶质细胞在脑区之间表现出显著的异质性，重要的是，可以 大脑区域内不同突触之间的差异非常大[3，7，8]。这在一定程度上 因为突触周围星形细胞突起对突触的形态覆盖 在不同的突触之间可以有很大的差异[14]。此外，Henneberger实验室已经 最近的研究表明，突触覆盖率越高，局部疗效越好。 谷氨酸摄取[3]。我们还展示了，除了异质性之外， 星形胶质细胞谷氨酸摄取和PAPs形态均受神经元可塑性控制 [1]。此外，谷氨酸的摄取受转运体的化学计量比控制，这意味着 谷氨酸分子进入星形胶质细胞，利用三者共同运输获得的能量 Na+和一个质子沿着电化学梯度下降，同时也输出一个K+[12]。 虽然向内导向的Na+梯度是谷氨酸摄取的主要驱动力，但最近 Rose实验室和其他人的研究表明，谷氨酸能活动导致局部或全球的Na+ 星形胶质细胞中的瞬变([Na+]A)[15]。事实上，在小鼠的海马体中，星形细胞的Na+信号 主要是由于谷氨酸转运体本身的活性，降解了 Na+梯度，从而暂时削弱负反馈环路中的摄取能力 [15-17]。在新皮质，谷氨酸能突触活动还导致显著的Na+ 通过NMDA受体流入，促进星形胶质细胞的Na+梯度[18]。 因此，人们越来越认识到，星形细胞对谷氨酸的摄取既不是静态的，也不是 制服。首先，它在功能上依赖于传输的离子的梯度 随着突触传递的动态变化[12]。其次，它是塑料的，因为它的结构 PAP在几分钟时间尺度上的重塑深刻改变突触周围谷氨酸扩散 [1]。因此，新出现的假设是谷氨酸溢出的程度和， 因此，大多数脑区的突触串扰是动态调控的。 星形胶质细胞的水平。此外，由于一个星形细胞可以接触数千个 各种神经元的突触，它有可能局部控制许多 突触。在这种情况下，星形胶质细胞或其亚细胞结构域可以协调 不同神经元上许多谷氨酸能突触之间的串扰。因此，星形胶质细胞 他们的论文将谷氨酸能突触传递的空间保真度设置为 结果深刻地控制着神经元的信号交换。 到目前为止，这些重要的假设在很大程度上还没有得到检验。我们将通过以下方式填补这一空白 结合定量荧光成像、星形细胞操作和预测 计算机建模。这将通过研究突触周围星形细胞Na+来实现 谷氨酸吸收的主要驱动力--梯度及其局部控制机制 1

CRCNS US-German Research Proposal: Quantitative and computational dissection of glutamatergic crosstalk at tripartite synapses (1) Christine R Rose, Heinrich Heine University, Düsseldorf, Germany (2) Christian Henneberger, University of Bonn, Germany (3) Ghanim Ullah, University of South Florida, Tampa, FL, USA Project Description 1 Introduction and Background Transmission at chemical synapses is the central mechanism by which information is transferred between neurons. Synaptic connections such as glutamatergic excitatory synapses are often perceived and modeled as point-to-point connections. However, there is substantial evidence that crosstalk between various glutamatergic synapses can occur when the presynaptically released glutamate is sensed not only by its direct postsynaptic partner but also by nearby synapses of the same and other neurons [4]. Notably, this phenomenon termed “glutamate spillover” not only defines the input-specificity of a given synaptic connection and its crosstalk to neighboring synapses, but is also involved in and controlled by activity-dependent plasticity [1, 7, 8]. How easily glutamate escapes from its release site and how far it spreads into the tissue depends on the morphological and molecular properties of the extracellular space (ECS) as well as on the efficacy of glutamate clearance, which primarily depends on astrocytic uptake [11, 12]. We and others have shown that the efficacy of perisynaptic glutamate uptake by astrocytes displays a remarkable heterogeneity between brain regions and, importantly, can vary drastically from one synapse to the next within a brain region [3, 7, 8]. This is in part because the morphological coverage of synapses by perisynaptic astrocyte processes (PAPs) can differ strongly between individual synapses [14]. Moreover, the Henneberger lab has recently shown that higher synaptic coverage by PAPs correlates with a higher local efficacy of glutamate uptake [3]. We have also demonstrated that in addition to being heterogeneous, astrocytic glutamate uptake and PAPs morphology both are controlled by neuronal plasticity [1]. Moreover, glutamate uptake is governed by the transporters’ stoichiometry, importing one glutamate molecule into the astrocyte by using the energy gained from co-transporting three Na+ and one proton down the electrochemical gradients, whilst also exporting one K+ [12]. While the inwardly-directed Na+ gradient is the main driving force for glutamate uptake, recent work by Rose lab and others have shown that glutamatergic activity causes local or global Na+ transients in astrocytes ([Na+]A) [15]. In the mouse hippocampus, astrocytic Na+ signals in fact arise predominately due to the activity of glutamate transporters themselves, degrading the Na+ gradient and thereby transiently weakening uptake capacity in a negative feedback-loop [15-17]. In the neocortex, glutamatergic synaptic activity in addition results in prominent Na+ influx through NMDA receptors, boosting astrocyte Na+ gradients [18]. Thus, it is increasingly appreciated that astrocytic glutamate uptake is neither static nor uniform. First, it is functionally dependent on the gradients of the transported ions which dynamically change with synaptic transmission [12]. Second, it is plastic because structural remodeling of PAPs on time scales of minutes profoundly alters perisynaptic glutamate spread [1]. Therefore, the emerging hypothesis is that the degree of glutamate spillover and, therefore, synaptic crosstalk in most brain regions are dynamically regulated and controlled at the level of the astrocytes. Furthermore, since a single astrocyte can contact thousands of synapses of various neurons, it has the potential to locally control the crosstalk of many synapses. In such a scenario, an astrocyte, or a subcellular domain of it, can coordinate crosstalk between many glutamatergic synapses on different neurons. Thereby, astrocytes and their PAPs set the spatial fidelity of glutamatergic synaptic transmission and as a consequence profoundly control neuronal signal exchange. So far, these important hypotheses remain largely untested. We will fill this gap by combining quantitative fluorescence imaging, astrocytic manipulations, and predictive computer modelling. This will be accomplished by investigating perisynaptic astrocytic Na+ gradients, the main driving force of glutamate uptake, and local mechanisms controlling them 1