Circuit-level control of forelimb motor behaviour and plasticity following injury

中枢神经系统（CNS）的主要输出是通过从大脑投射到脊髓的下行运动通路来控制运动，以协调运动神经元的活动。它们的重要性在脊髓损伤的情况下是非常明显的，因为它们的破坏会导致严重的瘫痪。然而，它们在运动行为中的具体作用和介导损伤后运动系统恢复的能力仍不清楚。* 在这里，我们正在开展一项新的研究计划，旨在利用化学遗传学和个体遗传学技术了解脊髓损伤后的神经可塑性和功能补偿。* 具体来说，我们提出了一系列使用最近开发的技术的初步实验，该技术允许体内特定神经回路的非侵入性和瞬时沉默，称为设计师药物独家激活的设计师受体（DREADDs）（PMID：17360345）。DREADD是生物工程G蛋白偶联受体，可以激活或沉默神经元，以响应不激活体内其他受体的配体。 这些配体可以通过腹膜内注射（短效）或通过饮用水（长效）非侵入性地施用。DREADD受体可以以两种方式之一靶向特定的神经元群体。首先，可以施用编码DREADD的病毒，使得通常仅病毒注射区域内的细胞被感染。第二，通过使用cre依赖性构建体，可以通过采用双重病毒方法靶向特定的解剖学投射，其中一种cre依赖性病毒被施用在神经元的细胞体处，并且第二种编码cre的病毒被施用用于逆行感染神经元的轴突/突触末端（PMID：26889809）。我们已经成功地使用了这种方法（PMID：27053214）。*在本项目中，我们将使用沉默DREADD hM 4Di来特异性沉默涉及前肢运动控制的下行运动通路。这项资助的第一个目标是阐明皮质脊髓，红核脊髓，和延髓网状脊髓腹侧（MdV）的运动行为功能在未受伤的成年小鼠中枢神经系统的途径的作用。第二个目的是阐明红核脊髓和MdV通路在锥体水平皮质脊髓束损伤后介导恢复的可塑性。 我们将评估给予CNO以沉默这些特定神经元群体之前和之后小鼠中单个食物颗粒到达、梯子穿越、旋转棒和握力的恢复，并在同一动物中进行比较，使得每只动物充当其自身对照。 这将揭示特定神经元投射系统在执行特定任务中的功能参与，无论是在正常未受伤状态下还是在不完全损伤后。

The major output of the central nervous system (CNS) is to control movement via descending motor pathways that project from the brain to the spinal cord to orchestrate motoneuron activities. Their importance is strikingly obvious in cases of spinal cord injury, when severe paralysis can ensue because of their disruption. However, their specific roles in motor behaviour and ability to mediate motor system recovery following injury remain unclear. ***Here, we are embarking on a new research program aiming at an understanding of neural plasticity and functional compensation after spinal cord injury using chemogenetic and ontogenetic technologies. ***Specifically, we propose a series of initial experiments using a recently developed technology that allows non-invasive and transient silencing of specific neural circuits in vivo called Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) (PMID:17360345). DREADDs are bioengineered G-protein coupled receptors that can activate or silence neurons in response to ligands that do not activate other receptors within the body. These ligands can be administered non-invasively via intraperitoneal injection (short acting) or through drinking water (long acting). DREADD receptors can be targeted to specific neuronal populations in one of two ways. First, viruses encoding DREADDs can be administered such that only cells within the area of viral injection is generally infected. Second, by using cre-dependent constructs, specific anatomical projections can be targeted by employing a dual virus approach, wherein one cre-dependent virus is administered at the neuron's cell body and the second, cre-encoding virus is administered for retrograde infection at the neuron's axon/synaptic terminal (PMID: 26889809). We have used this approach successfully (PMID: 27053214).***In the present project we will use the silencing DREADD hM4Di to specifically silence descending motor pathways implicated in forelimb motor control. The first objective of this grant is to elucidate the role of corticospinal, rubrospinal, and medullary reticulospinal ventral (MdV) pathways in motor behavioural function in the uninjured adult mouse CNS. The second objective is to elucidate the plasticity of rubrospinal and MdV pathways in mediating recovery following corticospinal tract injury at the level of the pyramid. We will assess the recovery of single food pellet reaching, ladder crossing, rotarod and grip strength in mice before and following the administration of CNO to silence these specific neuronal populations and make comparisons within the same animal such that each animal acts as its own control. This will reveal the functional involvement of specific neuronal projection systems in the execution of specific tasks, either in the normal uninjured state or after incomplete injury.