Direct nicotinic excitation of layer 5 neocortical pyramidal neurons

描述（由申请人提供）：新皮层在许多高级功能中起着核心作用，如感官信息的解释，语言的理解和随意运动的控制。这些过程中的许多都是由基底前脑复合体（主要是基底核）中的神经元的胆碱能驱动所形成的。 这一通路的功能障碍导致许多涉及新皮层的行为缺陷，并与抑郁症、帕金森病和阿尔茨海默病等临床疾病有关。 我们的长期目标是了解来自基底核的ACh如此深刻地影响皮质功能的细胞和网络机制。ACh作用于在新皮层中广泛表达的毒蕈碱型和烟碱型ACh受体（mAChR和nAChR）。新皮质中的锥体神经元表达这两种受体类型，但锥体神经元中nAChR的生理功能尚不清楚。 在这个建议中，我们描述了锥体神经元的nAChRs的功能。我们直接评估ACh，从基底核轴突释放，影响运动皮层锥体神经元，使用的技术组合，包括光遗传学，病毒工具，转基因小鼠，免疫细胞化学，细胞电生理学，双光子显微镜和电子显微镜。我们在基底核的胆碱能神经元和新皮层的轴突中表达通道视紫红质-2。在初步实验中，胆碱能轴突的激活通过nAChR去极化和促进第5层锥体神经元的尖峰，使我们假设ACh促进信息通过新皮层网络从上行兴奋性输入（例如，从丘脑）传递到靶结构（例如，脊髓中的运动回路）。 在这 我们将确定ACh对锥体神经元的影响是否支持这一假设。我们将研究nAChR影响尖峰和nAChR受体亚单位参与的机制（具体目标1），确定激活的nAChR位于锥体神经元树突树内的位置（具体目标2），并确定这些突触后nAChR是否介导了其他层和新皮层区域锥体神经元中ACh的作用（具体目标3）。 我们的结果将揭示ACh调节锥体神经元兴奋性的新机制。据我所知，我们的研究还将提供第一个证据，证明乙酰胆碱具有层特异性作用。由此产生的假设有可能改变我们对基底核胆碱能通路改变新皮层网络功能的方式的理解，因此可能对帕金森病和阿尔茨海默病等衰弱性疾病具有重要意义。

DESCRIPTION (provided by applicant): The neocortex plays a central role in many higher-order functions such as the interpretation of sensory information, comprehension of language and control of voluntary movements. Many of these processes are shaped by ascending cholinergic drive from neurons in the basal forebrain complex, principally nucleus basalis. Dysfunction of this pathway leads to deficits in many behaviors that involve neocortex and has been tied to clinical conditions such as depression, Parkinson's disease and Alzheimer's disease. Our long-term aim is to understand the cellular and network mechanisms by which ACh from nucleus basalis so profoundly influences cortical function. ACh acts at muscarinic and nicotinic ACh receptors (mAChRs and nAChRs), which are widely expressed in neocortex. Pyramidal neurons in neocortex express both of these receptor types, but the physiological functions of nAChRs in pyramidal neurons are unknown. In this proposal we describe the functions of nAChRs on pyramidal neurons. We directly assess how ACh, released by axons from nucleus basalis, affects pyramidal neurons in motor cortex, using a combination of techniques, including optogenetics, viral tools, genetically-modified mice, immuno-cytochemistry, cellular electrophysiology, two-photon microscopy and electron microscopy. We express channelrhodopsin-2 in cholinergic neurons in nucleus basalis and their axons in neocortex. In preliminary experiments, activation of cholinergic axons depolarized and promoted spiking of layer 5 pyramidal neurons via nAChRs, leading us to hypothesize that ACh facilitates the transfer of information through neocortical networks from ascending excitatory inputs, e.g. from thalamus, to target structures, e.g. motor circuits in the spinal cord. In this proposal we will determine whether the effects of ACh on pyramidal neurons support this hypothesis. We will investigate the mechanisms by which nAChRs affect spiking and the nAChR receptor subunits involved (specific aim 1), determine where in the activated nAChRs are located within the dendritic trees of pyramidal neurons (specific aim 2), and determine whether these postsynaptic nAChRs mediated the effects of ACh in pyramidal neurons in other layers and neocortical areas (specific aim3). Our results will reveal a new mechanism by which ACh modulates the excitability of pyramidal neurons. Our studies will also provide the first evidence, to my knowledge, that ACh has layer-specific effects. The resulting hypothesis has the potential to transform our understanding of the manner in which the cholinergic pathway from nucleus basalis changes network function in neocortex and may therefore have important implications for debilitating conditions such as Parkinson's disease and Alzheimer's disease.