Afterhyperpolarization in dopamine neurons, H2O2 and KATP channels

描述：黑质致密部（SNc）中的多巴胺（DA）神经元在基底神经节介导的运动功能中起着至关重要的作用。SNc DA神经元的缺失确实会导致帕金森病的运动缺陷。然而，这些神经元在正常运动控制中的作用尚不清楚。对帕金森病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一个最关键的机制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有助于调节自发放电和诱发放电的频率和模式，并部分由阿帕胺敏感的Ca2+激活的K+通道介导。此外，然而，阿帕胺不敏感成分是由一个尚未确定的Ca2+不敏感通道介导的。我们之前的研究表明，SNc中DA神经元的膜性质和放电频率受到内源性过氧化氢（H2O2）的调节，这种调节通过atp敏感的K+ (KATP)通道发生。在DA神经元中，去极化电流注入诱导的爆发样放电伴随着活性氧（如H2O2）产生的增加以及AHP的延长。令人兴奋的是，本研究中提出的新数据表明，H2O2代谢酶过氧化氢酶和选择性KATP通道阻滞剂格列本脲可显著降低DA神经元AHP的振幅和持续时间，这表明H2O2和KATP通道都参与其中。因此，拟议的研究将测试这些因素在DA神经元AHP中的参与。我将采用膜片钳记录结合荧光成像的方法对豚鼠中脑切片SNc DA神经元中的H2O2进行成像。电流箝位模式用于研究AHP振幅和持续时间，混合箝位记录用于评估AHP尾电流。Aim 1的实验将解决DA神经元AHP是否具有H2O2敏感成分的问题，并评估其对apamin的敏感性。对h2o2敏感的KATP通道在SNc DA神经元AHP中的参与将在Aim 2中进行评估。总的来说，该项目将为H2O2和KATP通道对AHP和DA神经元活性的动态调节提供独特的见解。本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。这些细胞在基底神经节介导的功能中起关键作用。事实上，SNc多巴胺神经元的缺失导致帕金森病的运动缺陷。因此，这项研究可能会导致新型抗帕金森药物的开发。因此，它与NINDS直接相关，NINDS支持的项目侧重于发现神经退行性疾病的机制。

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.