Circuit Specializations of Cerebellar Molecular Layer Interneurons

项目摘要 小脑控制依赖于预测的行为，包括运动学习、眼球运动、 平衡和认知-情感功能。小脑皮质将苔藓纤维(Mf)的输入转化为 浦肯野细胞(PC)输出。至关重要的是要描绘出这个电路的元素以及它们之间的联系 电路特性对功能和疾病的影响。为了提高我们对小脑变化的理解 这一提议试图描绘出大脑中最丰富的中间神经元类型的电路特性。 小脑，分子层中间神经元(MLI)。MLIS抑制PC和其他MLI以控制 小脑皮层。然而，单核RNA测序最近发现了两种不同的分子类型 MLIS，MLI类型1(MLI1)和MLI类型2(MLI2)。有趣的是，MLI1表达连接蛋白36，而MLI2表达连接蛋白36 不。这表明MLI1可能是缝隙连接，这可能促进 同步触发，使MLI1非常适合于控制PC输出的时序。的电路特性 MLI1和MLI2未知。在这里，我将使用切片电生理学和 小鼠的连续电子显微镜(EM)重建。这项建议的第一个目标是确定 在脑片上使用配对的全细胞记录来输出MLI1和MLI2。首先，我将描述MLI1和 MLI2突触连接到PC。我还将描述MLI1和MLI2对触觉的贡献 抑制，由MLI专门化附近的大细胞外信号引起，称为Pinceaux 环绕PC轴突初始节段。最后，我将描述突触连接和电耦合 在MLI1和MLI2之间。这项提议的第二个目的是描述超微结构回路的特征。 MLI1和MLI2的特化并生成MLI1和MLI2突触和触觉的综合图谱 使用大规模EM重建的连接性。电生理学的互补途径 表征和EM重建将提供功能特性和 解剖学上的连接。初步研究表明，MLI1和MLI2以不同的细胞为靶点，其中MLI1 主要抑制PC，而MLI2主要抑制其他MLI。根据这些初步研究，我的 工作模式是MLI1非常适合于控制PC输出的时序，而MLI2促进PC输出 通过解除对PC的抑制而产生兴奋性。拟议工作的完成将确定这些电路的特性 元素，如果初步结果得到确认，将有必要修改 小脑皮层。这些实验还将导致未来的研究，将确定体内的激发 MLI1和MLI2的性质，并确定MLI1和MLI2如何促进加工、学习和 行为。这些研究有望阐明小脑计算的机制，这些机制与 神经紊乱和神经精神疾病。

Project Summary The cerebellum controls behaviors that depend on prediction, including motor learning, eye movements, balance, and cognitive-affective functions. The cerebellar cortex transforms mossy fiber (MF) inputs into Purkinje cell (PC) outputs. It is vital to delineate the elements of this circuit and their connectivity to relate circuit properties to function and disease. To improve our understanding of transformations in the cerebellar cortex, this proposal seeks to delineate the circuit properties of the most abundant type of interneuron in the cerebellum, the molecular layer interneuron (MLI). MLIs inhibit PCs and other MLIs to control the output of the cerebellar cortex. However, single nucleus RNA sequencing recently identified two molecularly distinct types of MLIs, MLI type 1 (MLI1) and MLI type 2 (MLI2). Intriguingly, MLI1s express connexin 36, whereas MLI2s do not. This suggests that MLI1s might be gap junction coupled with each other, which could promote synchronous firing and make MLI1s well-suited to controlling the timing of PC outputs. The circuit properties of MLI1 and MLI2 are not known. Here I will clarify MLI1 and MLI2 connectivity using slice electrophysiology and serial electron microscopy (EM) reconstructions in mice. The first aim of this proposal is to determine the outputs of MLI1 and MLI2 using paired whole-cell recordings in brain slice. First, I will characterize MLI1 and MLI2 synaptic connections onto PCs. I will also characterize the contributions of MLI1 and MLI2 to ephaptic inhibition, which arises from large extracellular signals near MLI specializations known as pinceaux that surround PC axon initial segments. Lastly, I will characterize synaptic connections and electrical coupling between MLI1 and MLI2. The second aim of this proposal is to characterize the ultrastructural circuit specializations of MLI1 and MLI2 and generate a comprehensive map of MLI1 and MLI2 synaptic and ephaptic connectivity using large-scale EM reconstructions. The complementary approaches of electrophysiological characterization and EM reconstructions will provide a complete picture of the functional properties and the anatomical connectivity. Preliminary findings suggest that MLI1 and MLI2 target different cells, with MLI1s mainly inhibiting PCs, whereas MLI2s primarily inhibit other MLIs. Based on these preliminary studies, my working model is that MLI1s are well suited to controlling the timing of PC outputs, whereas MLI2s promote PC excitability by disinhibiting PCs. Completion of the proposed work will define the properties of these circuit elements, and if preliminary results are confirmed, it will be necessary to revise the circuit diagram of the cerebellar cortex. These experiments will also lead to future studies that will determine the in vivo firing properties of MLI1 and MLI2 and determine how MLI1 and MLI2 contribute to processing, learning and behavior. These studies promise to shed light on mechanisms of cerebellar computations that are relevant to neurological disorders and neuropsychiatric diseases.