Modulation of Neuronal Ion Channels by 2nd Messengers

描述（由申请人提供）：离子通道电流是大多数生物体中电活动的基本单位。在我们的神经系统中，K 通道和 Ca2 通道至关重要，它们的调节提供了一种直接控制神经元兴奋性和突触神经递质释放的方法。这些调节对于基本神经功能至关重要，对它们的理解应该有助于针对涉及大脑、神经和肌肉的一系列疾病的新的医疗干预模式。我们主要关注作为多个神经元 K 电流基础的 Kv7（KCNQ/M 型）K 通道家族和 Cav2.2（N 型）Ca2 通道。特别是，我们试图阐明作用于这些类型离子通道的几种调节途径的分子机制。我们使用异源系统（其中通道、受体和信号分子的 cDNA 克隆在哺乳动物组织培养细胞中表达）和初级交感神经元的制剂。我们研究的两条途径存在于与 G 蛋白 Gq/11 家族偶联的共同受体中：磷脂酶 C 的激活和磷脂酰肌醇 4,5-二磷酸 (PIP2) 的水解。 Kv7 和 Cav2.2 通道均受 PIP2 调节，在本项目中，我们将研究 PIP2 如何与通道相互作用、相互作用的位点以及介导门控调节的通道蛋白部分。刺激某些物质会增加细胞内 Ca2 并与钙调蛋白 (CaM) 协同作用于 Kv7 通道。我们将进一步研究Ca2/CaM作用的分子机制。其他 Gq/11 偶联受体不会升高细胞内 Ca2+，而是通过消耗 PIP2 膜发挥作用。我们将使用多种方法来探讨受体作用的这种特异性。所使用的工具包括膜片钳电生理学、分子生物学、生物化学和先进的成像技术。我们研究的分子和信号通路与人类健康和疾病具有广泛的相关性。 Kv7通道在神经元兴奋性调节中起主导作用，其调节可能是情绪状态、记忆和身体器官调节变化的基础。 Kv7 通道功能失调会导致特定的癫痫综合征。 Cav2.2 通道的调节调节神经递质的释放，神经递质是神经元之间突触的基本信号。因此，我们的研究应该为开发各种神经疾病治疗干预的新模式提供基础。简单总结：该项目研究如何调节神经细胞中介导电流的离子通道。我们使用生物物理和分子工具在分子和单个活细胞水平上研究这些信号。我们试图了解人类思想、情感和行为的复杂现象背后的神经系统调节。我们的发现可能有助于缓解许多情绪、运动和意识疾病，即神经功能障碍。

DESCRIPTION (provided by applicant): Ion channel currents are the fundamental units of electrical activity in most organisms. In our nervous system, K+ and Ca2+ channels are critical, and their regulation provides a way to directly control neuronal excitability and release of neurotransmitter at synapses. The modulations are crucial to basic nervous function and their understanding should contribute to novel modes of medical interventions for a range of disorders involving the brain, nerves and muscles. We focus primarily on the family of Kv7 (KCNQ/M-type) K+ channels that underlies several neuronal K+ currents and on Cav2.2 (N-type) Ca2+ channels. In particular, we seek to elucidate the molecular mechanisms of several modulatory pathways that act on these types of ion channels. We use both a heterologous system in which cDNA clones of the channels, receptors and signaling molecules are expressed in mammalian tissue-culture cells, and a preparation of primary sympathetic neurons. Two pathways that we study have in common receptors coupled to the Gq/11 family of G proteins, the activation of phospholipase C, and hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2). Both Kv7 and Cav2.2 channels have emerged as being regulated by PIP2, and in this project we will investigate how PIP2 interacts with the channels, the site(s) of the interaction, and the parts of the channel proteins that mediate the regulation of gating. Stimulation of certain raises intracellular Ca2+ and acts on Kv7 channels in concert with calmodulin (CaM). We will further investigate the molecular mechanism by which Ca2+/CaM acts. Other Gq/11-coupled receptors do not raise intracellular Ca2+ and act by depleting the membrane of PIP2. Using a variety of approaches, we will probe this specificity in receptor action. The tools to be used include patch-clamp electrophysiology, molecular biology, biochemistry, and advanced imaging techniques. The molecules and signaling pathways that we study have broad relevance to human health and disease. Kv7 channels play a dominant role in regulating excitability of neurons, and their regulation likely underlies changes in emotional state, memory and regulation of body organs. Dysfunctional Kv7 channels cause specific epileptic syndromes. The modulation of Cav2.2 channels regulates release of neurotransmitter which is the basic signal at synapses between neurons. Thus, our research should provide the basis for the development of novel modes of therapeutic intervention for a variety of nervous diseases. Lay summary: This project studies how ion channels, which mediate the electricity in nerve cells, are regulated. We investigate these signals at the level of the molecule and the individual living cell using biophysical and molecular tools. We seek to understand this regulation of the nervous system that underlies the complex phenomena of human thought, emotion and behavior. Our findings may help to alleviate the many diseases of mood, motion and consciousness that are disorders of nervous function.