The cellular basis of Episodic Ataxia/Myokemia type1

描述(由申请人提供)：本提案的总体目标是确定电压门控钾通道亚单位Kv1.1中的错义突变如何改变小脑输出并导致发作性共济失调1型(EA1)。V408A是人类EA1突变之一，导致Kv1.1通道处于不稳定的开放状态。我们已经建立了EA1的V408A小鼠模型，并发现它概括了疾病的关键方面。小脑浦肯野细胞(PC)表现出一种内在产生的紧张性放电模式，这种模式被它们的抑制性和兴奋性输入所改变。小脑篮细胞(BCs)在其突触前终末表达Kv1.1，是抑制PC的主要来源。EA1小鼠中的PC从BCS获得更多的GABA能传递。我发现，GABA能音的这种增加降低了PC放电的间歇放电的精确度，这对运动控制至关重要。PC发射精度缺陷是其他小脑性共济失调的特征。这种差异被GABAAR拮抗剂印防已毒素(PTX)抵消，表明GABA能张力增加在疾病模型中的重要性。为此，我建议研究V408A增加从BCS到PC的GABA能传递的细胞机制。我将结合双光子钙成像和电生理学来确定V408A是否增加了BCS大型突触前终末的动作电位侵袭。我将评估Kv1.1功能降低增加动作电位诱发事件释放概率的可能性。我还将使用BC动作电位和PC iPSCs的配对记录来确定AP诱导的GABA释放的失败率。实现这一目标将有助于我们理解K+通道在突触前控制神经递质释放中的生理作用，以及它参与EA1的病理生理过程。拟议的研究将对小脑性共济失调的细胞机制产生新的见解，并可能适用于其他共济失调。

DESCRIPTION (provided by applicant): The overall goal of this proposal is to determine how a missense mutation in the voltage-gated potassium channel subunit, Kv1.1 alters cerebellar out-put and results in Episodic Ataxia type 1 (EA1). V408A is one of the human EA1 mutations, and results in Kv1.1 channels with an unstable open state. We have generated a V408A mouse model of EA1, and find that it recapitulates the key aspects of the disease. Cerebellar Purkinje Cells (PCs) exhibit an intrinsically generated, tonic firing pattern that is modified by their inhibitory and excitatory inputs. Cerebellar Basket Cells (BCs) express Kv1.1 in their presynaptic terminals, and are the main source of inhibition to PCs. PCs in EA1 mice receive increased GABAergic transmission from BCs. I have found that this increase in GABAergic tone reduces the precision of the interspike interval of PC firing, which is critical for motor control. This PC firing precision deficit is a characteristic of other cerebellar ataxias. This difference is equalized by the GABAAR antagonist picrotoxin (PTX), indicating the importance of increased GABAergic tone in the disease model. For this reason, I propose to investigate the cellular mechanism by which V408A increases GABAergic transmission from BCs to PCs. I will use a combination of 2-photon Ca2+ imaging and electrophysiology to determine whether V408A increases action potential invasion throughout the large presynaptic terminals of BCs. I will assess the possibility that reduced Kv1.1 function increases probability of release for action potential-evoked events. I will additionally use paired-recordings of BC action potentials and PC IPSCs to determine percent failure of AP-induced GABA release. Accomplishing this aim will contribute to our understanding of the physiological role of K+ channels in the presynaptic control of neurotransmitter release, as well as its involvement in the pathophysiology of EA1. The proposed studies will engender novel insights into the cellular mechanisms responsible for a cerebellar ataxia, and may be applicable to additional ataxias.