Voltage-gated ion channels and synaptic integration by cerebellar Purkinje cells

描述(申请人提供)：电压门控通道在小脑浦肯野细胞形成突触整合中的作用鲜为人知。由于许多表现小脑症状的衰弱遗传性疾病，包括发作性共济失调1型(EA1)和2型(EA2)，以及脊髓小脑型共济失调6型(SCA6)，都是这些膜蛋白的障碍，因此对它们的功能特性和相互作用的研究是至关重要的。这项研究计划的总体目标是确定选择的电压门控电导对浦肯野细胞兴奋性突触输入整合的贡献。浦肯野细胞整合了大量兴奋性和抑制性输入的汇聚，是小脑皮质的唯一输出。因此，它们是小脑的主要计算单位，最终实现适当的运动协调、姿势和平衡。实验室先前的工作证实，浦肯野细胞以其刺激后最大放电速率线性编码其突触前谷氨酸能颗粒细胞输入的强度。考虑到浦肯野细胞表现出由各种树枝状电压门控电导带来的活跃的树枝状钙尖峰，这是令人惊讶的，这种电导本身就是非线性的。浦肯野细胞中电压门控通道的活动是如何设计的，以允许颗粒细胞输入的线性求和？我们提出了两个目标来开始回答这个问题：1)非侵入性地评估选择的电压门控离子通道对颗粒细胞输入-浦肯野细胞输出函数的贡献；2)检验活性树突电导平衡的假设，以实现颗粒细胞输入-浦肯野细胞输出的线性关系。我们将通过将细胞外和全细胞膜片钳电生理学、谷氨酸去化和药理学相结合的实验来实现这一点，并将其应用于大鼠小脑切片制备。这些研究不仅对理解运动的神经控制和共济失调的机制至关重要，而且有助于从总体上阐明神经元的计算过程。

DESCRIPTION (provided by applicant): The role of voltage-gated channels in shaping synaptic integration in cerebellar Purkinje cells is little understood. Examination of their functional properties and interactions is of critical interest since many debilitating hereditary diseases presenting cerebellar symptoms, including episodic ataxia types 1 (EA1) and 2 (EA2), and spinocerebellar ataxia type 6 (SCA6), are disorders of these membrane proteins. The overall goal of this research plan is to define the contributions of select voltage-gated conductances to the integration of excitatory synaptic inputs by Purkinje cells. Purkinje cells integrate a massive convergence of excitatory and inhibitory inputs and are the sole output of the cerebellar cortex. They are thus the main computational units of the cerebellum, ultimately enabling proper motor coordination, posture, and balance. Prior work in the lab established that Purkinje cells linearly encode the strength of their presynaptic glutamatergic granule cell inputs in their maximum post-stimulus firing rate. This was surprising considering that Purkinje cells exhibit active dendritic calcium spikes brought about by various dendritic voltage-gated conductances, which are inherently nonlinear. How is the activity of voltage-gated channels in Purkinje cells choreographed to allow for the linear summation of their granule cell inputs? We propose two aims to begin answering this question: 1) To noninvasively assess the contribution of select voltage-gated ion channels to the granule cell input-Purkinje cell output function; and 2) To test the hypothesis that active dendritic conductances are balanced to achieve the linear granule cell input-Purkinje cell output relationship. We will accomplish this through experiments combining extracellular and whole-cell patch clamp electrophysiology, glutamate uncaging, and pharmacology applied to a rat cerebellar slice preparation. These investigations are not only crucial to understanding neural control of movement and the mechanisms underlying ataxia, but also will help elucidate neuronal computational processes in general.