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路径来控制运动，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非常明显，当脊髓损伤时，它们的破坏可能导致严重的瘫痪。然而，它们在运动行为中的具体作用以及在损伤后调节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。*具体地说，我们提出了一系列初步实验，使用最近开发的技术，允许非侵入性和瞬时沉默体内的特定神经电路，称为由设计药物(DREADD)独占激活的设计受体(DREADD)(PMID：17360345)。DREADD是一种生物工程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腔注射(短效)或通过饮用水(长效)非侵入性地给药。DREADD受体可以通过两种方式中的一种靶向特定的神经元群体。首先，可以对编码DREADD的病毒进行管理，使其通常只感染病毒注射区域内的细胞。其次，通过使用cre依赖的构建物，可以通过采用双重病毒方法来靶向特定的解剖投射，其中一个cre依赖的病毒被注射在神经元的细胞体，第二个cre编码的病毒被注射在神经元的轴突/突触末端逆行感染(pID：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 hM4Di来专门沉默与前肢运动控制有关的下行运动通路。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)通路在未受损伤的成年小鼠中枢神经系统运动行为功能中的作用。第二个目的是阐明红核脊髓和MDV通路在调节锥体水平的皮质脊髓束损伤后恢复中的可塑性。我们将评估在给予CNO之前和之后小鼠的单一食物颗粒到达、梯子穿越、旋转杆和握力的恢复，以使这些特定的神经元群体安静下来，并在同一动物内进行比较，使每种动物作为自己的对照。这将揭示特定神经元投射系统在执行特定任务时的功能参与，无论是在正常的未损伤状态下，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.