Afterhyperpolarization in dopamine neurons, H2O2 and KATP channels

描述：黑质致密部(SNC)中的多巴胺(DA)神经元在基底节介导的运动功能中起着至关重要的作用。事实上，黑质多巴胺神经元的缺失会导致帕金森氏症的运动障碍。然而，这些神经元在正常运动控制中的作用还知之甚少。对帕金森氏病潜在机制的最终理解和治疗策略的发展在很大程度上取决于对DA神经元特性的理解。控制神经元放电模式的最关键的机制之一是单个动作电位或动作电位爆发后的延长后超极化(AHP)。在DA神经元中，AHP有助于调节自发和诱发放电的频率和模式，并部分通过APamin敏感的钙激活的K+通道来调节。然而，此外，一种对apamin不敏感的成分是由一种尚未确定的钙不敏感通道介导的。我们以前的研究表明，内源性过氧化氢(H_2O_2)对黑质多巴胺能神经元的膜特性和放电频率有调节作用，这种调节是通过ATP敏感K+(KATP)通道实现的。在DA神经元中，由去极化电流注入诱导的突发式放电伴随着过氧化氢等活性氧物种的产生增加，以及AHP的延长。令人兴奋的是，这项应用中的新数据显示，过氧化氢代谢酶过氧化氢酶和选择性KATP通道阻滞剂格列本脲显著降低DA神经元AHP的幅度和持续时间，表明H_2O_2和KATP通道都参与了这一过程。因此，拟议的研究将测试这些因素在DA神经元AHP中的参与。我将使用膜片钳记录结合H_2O_2荧光成像技术对豚鼠中脑切片中的SNc DA神经元进行成像。将采用电流钳制模式研究AHP的幅值和持续时间，并采用混合钳制记录来评估AHP的尾流。目标1中的实验将解决DA神经元中的AHP是否具有过氧化氢敏感成分的问题，并评估其对阿帕明的敏感性。目标2将评估H_2O_2敏感的KATP通道在黑质多巴胺神经元AHP中的作用。总体而言，该项目将为H_2O_2和K_(ATP)通道对AHP进而DA神经元活性的动态调节提供独特的见解。公共卫生相关性这个项目的重点是测试一个关于黑质致密部(SNC)多巴胺能神经元活动调节的新假设。这些细胞在基底节介导的功能中起着至关重要的作用。事实上，黑质多巴胺神经元的缺失会导致帕金森氏症患者的运动障碍。因此，这项研究可能会导致新的抗帕金森病药物的开发。因此，它与NINDS直接相关，NINDS支持专注于发现神经退行性疾病机制的项目。

DESCRIPTION: Dopamine (DA) neurons in the substantia nigra pars compacta (SNc) play a crucial role in basal- ganglia mediated motor function. Indeed loss of SNc DA neurons leads to the motor deficits in Parkinson's disease. However, the role of these neurons in the normal control of movement is poorly understood. Eventual understanding of mechanisms underlying Parkinson's disease and development of treatment strategies largely depends on understanding DA neuron properties. One of the most critical mechanisms that controls neuronal firing pattern is the prolonged afterhyperpolarization (AHP) that follows a single action potential or burst of action potentials. In DA neurons the AHP contributes to the regulation of the frequency and pattern of spontaneous and evoked discharges and is mediated in part by apamin-sensitive Ca2+-activated K+ channels. In addition, however, an apamin-insensitive component is mediated by an as yet unidentified Ca2+- insensitive channel. Our previous studies showed that membrane properties and firing frequency of DA neurons in the SNc are modulated by endogenous hydrogen peroxide (H2O2) and this modulation occurs via ATP-sensitive K+ (KATP) channels. In DA neurons burst-like firing induced by depolarizing current injection is accompanied by an increase in the production of reactive oxygen species, like H2O2, as well as a prolonged AHP. Excitingly, new data presented in this application shows that the H2O2-metabolyzing enzyme, catalase, and the selective KATP channel blocker, glibenclamide, significantly decrease the amplitude and duration of the AHP in DA neurons, suggesting an involvement of both H2O2 and KATP channels. Proposed studies will therefore test the involvement of these factors in the AHP in DA neurons. I will use patch-clamp recording coupled with fluorescence imaging of H2O2 in SNc DA neurons in guinea pig midbrain slices. Current-clamp mode will be used to study AHP amplitude and duration and hybrid-clamp recording to assess the AHP tail current. Experiments in Aim 1 will address the question whether the AHP in DA neurons has an H2O2- sensitive component and assess its sensitivity to apamin. The involvement of H2O2-sensitive KATP channels in the AHP in SNc DA neuron will be assessed in Aim 2. Overall, this project will provide unique insights into dynamic regulation of AHP and consequently DA neuron activity by H2O2 and KATP channels. PUBLIC HEALTH RELEVANCE This project is focused on testing a novel hypothesis concerning the regulation of the activity of dopaminergic neurons in the substantia nigra pars compacta (SNc). These cells play a crucial role in basal ganglia mediated function. Indeed loss of SNc dopamine neurons leads to motor deficits in Parkinson's disease. This study could therefore potentially lead to the development of novel antiparkinsonian drugs. Thus, it is of direct relevance to NINDS which supports projects focused on the discovery of mechanisms of neurodegenerative disorders.