Does a Common Mechanism Trigger Attacks in Episodic Ataxia Type 2?

 描述(申请人提供)：发作性共济失调2型(EA2)是一种神经性疾病，由编码P/Q型电压门控钙通道(CAV2.1)孔形成α-1a亚单位的CACNA1a基因突变引起。EA2患者表现出由不同应激源引发的严重共济失调发作，如咖啡因和酒精消费以及身体或情感压力。我们实验室以前的工作表明，啮齿动物EA2模型中的运动攻击，摇摇欲坠，是由于浦肯野细胞(PC)不稳定的突然放电驱动小脑的异常输出。摇摇欲坠的老鼠是EA2的成熟啮齿动物模型。它们携带CACNA1A基因的自发突变，导致Cav2.1通道不起作用，并表现出类似于EA2患者的应激、咖啡因和酒精诱导的运动攻击。有趣的是，所有这三种触发因素都会导致这些动物PC的放电模式发生相同的变化，这表明了一种共同的汇聚机制。个人计算机本质上是活跃的，在体内以每秒50个尖峰的速度表现出紧张的激发。此激发模式部分受 Cav2.1通道与小电导钙激活钾通道(SK2)紧密耦合，有助于调节这些细胞的内在活性。已有研究表明，阻断SK2通道可促进野生型PC的爆发性放电，而在摇摇欲坠的小鼠小脑中激活SK2通道可恢复PC的规律性，并减轻咖啡因或应激引起的攻击。鉴于通过Cav2.1通道的摇摇欲坠的小鼠的钙电流已经减弱，并且SK2通道被依赖于Cav2.1的钙离子所控制，SK2活性的进一步降低可能是观察到的PC在攻击期间爆发的原因。事实上，我们实验室的初步数据表明，所有三个触发因素都集中在SK2活性的调节上。有趣的是，这种调节似乎依赖于小脑中去甲肾上腺素(NE)水平的增加，这种现象已知发生在对压力、咖啡因和酒精的反应中。因此，我们的工作假设是，所有三个触发器在功能上收敛，通过NE依赖机制降低PCSSK2通道的开放概率。我在这项提议中的主要目标是通过干扰SK2门控并询问它将如何影响攻击来进一步描述在摇摇欲坠的小鼠中驱动攻击的机制。我计划通过将电生理技术与分子生物学和行为测试相结合来实现这一点。这种强大的方法将使我能够将分子机制、生理功能和行为联系起来。

 DESCRIPTION (provided by applicant): Episodic ataxia type 2 (EA2) is a neurological disorder caused by mutations in the CACNA1A gene encoding for the pore forming α-1a subunit of the P/Q type voltage gated calcium channel (Cav2.1). EA2 patients exhibit episodic attacks of severe ataxia triggered by different stressors such as, caffeine and alcohol consumption and physical or emotional stress. Previous work in our lab has shown that the motor attacks in a rodent EA2 model, tottering, are due to an aberrant output from the cerebellum driven by erratic burst firing of Purkinje cells (PC). The tottering mouse is a well-established rodent model of EA2. They carry a spontaneous mutation in the CACNA1A gene rendering the Cav2.1 channel non-functional and exhibit motor attacks induced by stress, caffeine and alcohol similar to EA2 patients. Interestingly, all three triggers result in the same change in the firing pattern of PCs n these animals, suggesting a common converging mechanism. PCs are intrinsically active and in vivo exhibit tonic firing at 50 spikes per second. This firing pattern is partially governed by the tight coupling of Cav2.1 channels to the small conductance calcium activated potassium channels (SK2) that help regulate the intrinsic activity of these cells. It has been shown that blocking SK2 channels promotes burst firing of wild type PCs, whereas activating them in the cerebellum of tottering mice restores PC regularity and alleviates attacks induced by caffeine or stress. Give that the calcium current through Cav2.1 channels in tottering mice is already diminished and that SK2 channels are gated by Cav2.1 dependent calcium, a further reduction in SK2 activity could be the cause for the observed bursting of PCs during attacks. Indeed, preliminary data from our lab suggests that all three triggers converge on the modulation of SK2 activity. Interestingly, this modulation seems to be dependent on increases in norepinephrine (NE) levels in the cerebellum, a phenomenon known to occur in response to stress, caffeine and alcohol. Thus, our working hypothesis is that all three triggers functionally converge to reduce open probability of PCs SK2 channels via a NE dependent mechanism. My main goal in this proposal is to further delineate the mechanism driving the attacks in tottering mice by perturbing SK2 gating and asking how it will affect attacks. I plan to achieve this by combining electrophysiological techniques together with molecular biology and behavioral tests. This powerful approach will allow me to link molecular mechanisms, physiological function and behavior.