AGRP NEURONS. NMDARs, Spines, Source of Excitatory Input and Downstream Effectors

描述（由申请人提供）：大脑调节进食行为和代谢，但负责这一过程的神经回路在很大程度上是未知的。大脑的复杂性以及在清醒的行为动物中操纵它的需要代表了重大的技术挑战。采用新的创新办法可能会大大加快进展。最近，我们和其他人，使用药物遗传学和光遗传学技术，发现AgRP神经元驱动强烈的摄食行为。考虑到这一点，现在至关重要的是确定控制AgRP神经元活动的传入信号（激素和神经输入）以及引起AgRP神经元驱动进食的效应器（GABA，NPY，AgRP和下游神经回路）。值得注意的是，很少有人注意到传入（上游）神经输入调节AgRP神经元。这是不幸的，因为a）AgRP神经元的神经控制缺陷可能对复杂的进食障碍（从厌食症到肥胖症）起重要作用，以及B）AgRP神经元的激素调节，一个例子是生长激素释放肽，可能通过调节这些输入起作用。最近，在初步研究中，我们发现NMDA受体（NMDARs），兴奋性突触可塑性的关键调节因子，在调节AgRP神经元的活动和进食中起着关键作用;另一方面，POMC神经元上的NMDARs几乎没有或没有发挥作用。与此一致，AgRP神经元具有丰富的树突棘（突触后特化，其中大多数兴奋性突触存在，并且NMDAR在其中操作以控制可塑性）; POMC神经元相反，缺乏棘。最后，我们发现AgRP神经元的快速激活需要NMDAR，并且涉及树突棘发生，并且很可能增加兴奋性突触发生。因此，NMDAR介导的可塑性的兴奋性输入到AgRP神经元起着关键的，以前未知的作用，在调节AgRP神经元的活动，从而喂养行为。本申请将追求上述发现的关键含义。在目标1中，使用先进的双光子显微镜，我们将机械地调查“监管”的兴奋性输入可塑性，通过NMDAR/棘，AgRP神经元。目的2：利用Cre依赖的单突触狂犬病标测和视紫红质辅助的回路标测，我们将识别和评估向AgRP神经元发送神经元兴奋性输入的传入神经元的功能。在目标3中，使用无法通过GABA、NPY和/或AgRP发出信号的AgRP神经元的药物遗传学激活以及AgRP神经末梢的解剖选择性光遗传学激活，我们将鉴定AgRP神经元的下游效应子（递质和回路）。

DESCRIPTION (provided by applicant): The brain regulates feeding behavior and metabolism however the neurocircuitry responsible for this is largely unknown. The complexity of the brain and the need to manipulate it in awake, behaving animals, represent significant technical challenges. The application of new, innovative approaches is likely to greatly accelerate progress. Recently, we and others, using pharmacogenetic and optogenetic technology, have discovered that AgRP neurons drive intense feeding behavior. With this in mind, it is now critical to determine the afferent signals (hormones and neural inputs) controlling AgRP neuron activity as well as the effectors (GABA, NPY, AgRP, and downstream neural circuits) that bring about AgRP neuron-driven feeding. Remarkably, little attention has been paid to afferent (upstream) neural inputs regulating AgRP neurons. This is unfortunate because a) defective neural control of AgRP neurons could contribute importantly to complex eating disorders (ranging from anorexia to obesity), and b) the hormonal regulation of AgRP neurons, one example being ghrelin, likely works by modulating these inputs. Recently, in Preliminary Studies, we discovered that NMDA receptors (NMDARs), key regulators of excitatory synaptic plasticity, play a critical role in regulating AgRP neuron activity and feeding; NMDARs on POMC neurons, on the other hand, play little or no role. Consistent with this, AgRP neurons have abundant dendritic spines (postsynaptic specializations where most excitatory synapses reside and within which NMDARs operate to control plasticity); POMC neurons, in contrast, lack spines. Finally, we have found that fasting-activation of AgRP neurons require NMDARs and involves dendritic spinogenesis, and very likely increased excitatory synaptogenesis. Thus, NMDAR-mediated plasticity of excitatory input to AgRP neurons plays a key, previously unknown role in regulating AgRP neuron activity and consequently feeding behavior. The present application will pursue key implications of the above-mentioned findings. In Aim 1, using advanced 2-photon microscopy, we will mechanistically investigate "regulatory" plasticity of excitatory inputs, via NMDARs/spines, to AgRP neurons. In Aim 2, using Cre-dependent, Monosynaptic Rabies Mapping and also Channelrhodopsin-Assisted Circuit Mapping, we will identify and assess function of afferent neurons sending glutamatergic input to AgRP neurons. In Aim 3, using pharmacogenetic activation of AgRP neurons unable to signal via GABA, NPY and/or AgRP and also anatomic-selective, optogenetic activation of AgRP nerve terminals, we will identify the downstream effectors (transmitters and circuits) of AgRP neurons.