Ca Release Pathways as Integrators of Synaptic Signals

描述（申请人提供）：第二信使信号的时空模式和通路之间的相互作用对突触信息的整合和处理以及调节突触连接的强度至关重要。本提案的总体目标是了解钙（Ca）信号在小脑浦肯野神经元突触功能中的作用。我们的重点是肌醇1,4,5-三磷酸受体（IP3Rs）和红嘌呤受体（RyRs）产生的钙信号的特性和功能，它们介导钙从细胞内储存的释放。我们的假设是Ca释放通路作为突触活动的整合者，并且这种特性是在被称为小脑长期抑制（LTD）的突触可塑性的诱导形式中联想性和突触特异性的细胞基础。我之前的研究证实浦肯野神经元中通过IP3敏感通路从细胞内储存的钙释放集中参与了LTD的诱导，IP3受体的钙释放受IP3和胞质钙的动态调节，这表明该通路在突触信号整合中起着巧合探测器的作用。其他研究也暗示一氧化氮（NO）是LTD的另一个关键决定因素。这里提出的研究有两个主要目标。第一个目标是了解钙在浦肯野细胞树突释放的时空动态是如何被钙信号通路和其他涉及LTD的第二信使级联之间的相互作用所调节的。第二个目标是确定这些相互作用对诱导LTD的功能后果。在这些实验中，我们将结合高速共聚焦显微钙测量、电生理学、固定化合物的局部光解和急性小脑切片信号通路的药理操作。我们首先验证了IP3Rs和RyRs对突触Ca信号的放大在LTD诱导过程中介导突触输入之间的关联。然后，我们研究了Ca释放和NO在LTD诱导和空间扩散中的关系，特别关注NO通过增强IP3Rs和RyRs的敏感性来调节LTD的模型。这些研究的结果将阐明突触信号通路如何促进LTD的诱导，并深入了解生化计算调节神经元之间通信和塑造单个神经元信号处理能力的基本机制。这一信息对于理解小脑对运动功能的控制以及导致共济失调和其他运动问题的小脑功能障碍是很重要的。

DESCRIPTION (provided by applicant): The spatiotemporal patterns of second messenger signals and interactions among pathways are central to the integration and processing of synaptic information and to regulating the strength of synaptic connections. The general goal of this proposal is to understand the role of calcium (Ca) signaling in the synaptic function of cerebellar Purkinje neurons. Our focus is on the properties and function of Ca signals produced by inositol 1,4,5-trisphosphate receptors (IP3Rs) and ryanodine receptors (RyRs), which mediate Ca release from intracellular stores. Our hypothesis is that the Ca release pathways function as integrators of synaptic activity, and that this property is the cellular basis for associativity and synapse specificity during the induction of a form of synaptic plasticity known as cerebellar long-term depression (LTD). My previous studies established that Ca release from intracellular stores via the IP3-sensitive pathway in Purkinje neurons is centrally involved in the induction of LTD and that Ca release by IP3 receptors is dynamically regulated by both IP3 and cytosolic Ca in a way that suggests this pathway acts as a coincidence detector for the integration of synaptic signals. Other studies have also implicated nitric oxide (NO) as another key determinant of LTD. The studies proposed here have two primary objectives. The first goal is to understand how the spatial and temporal dynamics of Ca release in Purkinje cell dendrites are regulated by the interplay between Ca signaling pathways and other second messenger cascades involved in LTD. The second goal is to determine the functional consequences of these interactions for the induction of LTD. For these experiments, we will use a combination of high-speed confocal microscopic Ca measurements, electrophysiology, localized photolysis of caged compounds, and pharmacological manipulations of signaling pathways in acute cerebellar slices. We first test the idea that amplification of synaptic Ca signals by IP3Rs and RyRs mediates associativity between synaptic inputs during the induction of LTD. We then examine the relationship between Ca release and NO in the induction and spatial spread of LTD, with particular focus on a model in which NO regulates LTD by enhancing the sensitivity of IP3Rs and RyRs. The results of these studies should clarify how synaptic signaling pathways contribute to the induction of LTD, and give insight into fundamental mechanisms by which biochemical computation regulates communication between neurons and shapes the signal processing capabilities of individual neurons. This information is important for understanding the control of motor function by the cerebellum and the cerebellar dysfunction that underlies ataxia and other motor problems.