Modulation of Olfactory Bulb Neuron Current Properties

描述（由申请人提供）：设计的研究是对嗅球颗粒和二尖瓣细胞中钾电流调制的多学科分析。本研究的广泛和长期目标是阐明神经营养因子和生长因子如何利用离子通道作为磷酸化的底物，从而引起突触功效的短期和长期可塑性变化，或帮助嗅球神经回路的建立。了解控制这些转导级联的一般原理和离子通道的参与将提供蛋白质激酶和蛋白质磷酸酶如何促进特定神经元疾病（如阿尔茨海默氏症）的发病或严重程度的信息，或者这些酶的不受控制的信号传导如何导致细胞增殖失控和癌症和糖尿病等疾病。由于嗅觉系统中神经元独特的营养和再生能力，神经调节剂的持续表达除了在生长和分化中的作用外，还可以改变电兴奋性的模式。本提案的具体目的是利用膜片钳电生理学表征嗅球中受体连接的酪氨酸磷酸化信号是如何被感官体验、模式电刺激和营养因子输注改变的。利用克隆的嗅球钾通道Kv1.3作为平行模型，结合激酶诱导酪氨酸磷酸化、共免疫沉淀和分子诱变的生化测量，将阐明离子通道如何通过在SH2、SH3、PDZ和PTB结构域的离散蛋白-蛋白相互作用与激酶和衔接蛋白形成分子支架的机制细节。Kv1.3通道、胰岛素受体激酶和TrkB激酶的基因靶向缺失将为酪氨酸磷酸化信号在嗅觉和中枢神经系统中的神经调节中的作用提供机制细节，如用敲除小鼠品系进行的功能丧失实验（行为、生化、电生理）所定义的那样。该建议将为通过构建蛋白质-蛋白质与离子通道的相互作用来整合信号分子提供新的重要信息。因此，离子通道功能的调节将取决于特定神经元中表达的信号蛋白的数量，这一背景可能会随着感觉体验或电模式而改变。

DESCRIPTION (provided by applicant): The designed research is a multidisciplinary analysis of the modulation of potassium currents in granule and mitral cells of the olfactory bulb. The broad, long-term objective of this research is to elucidate how neurotrophins and growth factors can utilize ion channels as substrates for phosphorylation to give rise to short-term and long-term plastic changes in synaptic efficacy or to aid in the establishment of neural circuits in the olfactory bulb. Understanding the general principles governing these transduction cascades and the involvement of ion channels will provide information of how protein kinases and protein phosphatases contribute to the onset or severity of specific neuronal diseases, such as Alzheimer's, or how uncontrolled signaling of these enzymes leads to deregulated cell proliferation and diseases such as cancer and diabetes. Because of the unique trophic and regenerative capacity of neurons in the olfactory system, continual expression of neuromodulators could alter patterns of electrical excitability in addition to their well-studied roles in growth and differentiation. The specific aims of this proposal are to characterize using patch-clamp electrophysiology how receptor-linked tyrosine phosphorylation signaling in the olfactory bulb is altered by sensory experience, patterned electrical stimulation, and trophic factor infusion. By utilizing the cloned, olfactory bulb potassium channel Kv1.3 as a parallel model, combined biochemical measurement of kinase-induced tyrosine phosphorylation, co-immunoprecipitation, and molecular mutagenesis will elucidate the mechanistic details of how ion channels form molecular scaffolds with kinases and adaptor proteins through discrete protein-protein interactions at SH2, SH3, PDZ, and PTB domains. Gene-targeted deletions in Kv1.3 channel, insulin receptor kinase, and TrkB kinase will provide mechanistic details for the role for tyrosine phosphorylation signaling in olfaction and for neuromodulation in the CNS in general, as defined by loss of function experiments (behavioral, biochemical, electrophysiological) using knock-out mice strains. The proposal will provide new important information regarding the integration of signaling molecules by construction of protein-protein interactions with ion channels. Modulation of ion channel function would thus be dependent upon the repertoire of signaling proteins expressed in a given neuron, a background that could change with sensory experience or electrical patterning.