Plasticity of spinal L3 propriospinal neurons in urination recovery after thoracic SCI

胸部 SCI 后排尿恢复中脊髓 L3 本体脊髓神经元的可塑性

• 批准号: 10575973
• 负责人: Lingxiao Deng
• 金额: $ 43.59万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-22 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10575973
• 关键词: Acute; Adult; Affect; Axon; Bladder; Bladder Control; Brain Stem; Cells; Chronic; Commissure; Cytomegalovirus; Data; Dorsal; Electric Stimulation; Electromyography; Electrophysiology (science); Female; Frequencies; Functional disorder; Future; Injury; Interneurons; Kidney Failure; Label; Lead; Mediating; Methods; Micturition Reflex; Modeling; Morbidity - disease rate; Motor Neurons; Names; Neuronal Plasticity; Neurons; Neurostimulation procedures of spinal cord tissue; Neurotransmitters; Paralysed; Pathway interactions; Pattern; Phenotype; Physiological; Pilot Projects; Population; Rattus; Recombinant adeno-associated virus (rAAV); Recovery; Recovery of Function; Recurrence; Reflex action; Relaxation; Role; Spinal; Spinal Cord; Spinal Cord transection injury; Spinal cord injury; Spinal cord injury patients; Synapses; Synaptic Transmission; Techniques; Therapeutic; Thoracic spinal cord structure; Urethral sphincter; Urination; Urine; Viral; Viral Vector; Virus; deprivation; effective therapy; improved; mad itch virus; male; mortality; nerve supply; neural circuit; optogenetics; partial recovery; patient population; postsynaptic; postsynaptic neurons; recombinant virus; relating to nervous system; spinal reflex; tetanospasmin; urinary

Abstract Bladder function recovery is a top priority among the SCI patient population, yet so far there are few effective therapies. One major reason is the limited understanding of intraspinal mechanisms for both spontaneous plasticity and for therapeutic functional recovery after SCI. A transection SCI deprives supraspinal control of the external urethral sphincter (EUS) which paralyzes its relaxation function and leads to urine retention. Partial recovery of EUS relaxation and voiding occurs spontaneously only if the neural circuits between lumbar 3 (L3) and lumbar 6 (L6) spinal cord level remain intact. Voiding function can be further improved by electrical stimulation of the L3 spinal cord in a chronic SCI model. However, the cellular mechanism underneath is unclear. In our preliminary study, we applied an anterograde trans-monosynaptic AAV1-GFP-Cre virus and found that a population of L3 propriospinal neurons (PSNs) projected to the L6 dorsal commissure (DCM) and synapsed with EUS-related interneurons (INs). We named these PSNs as PSNsL3-L6 DCM. Optogenetic stimulation of these PSNsL3-L6 DCM induced field potential of EUS Motoneurons (MNEUS) as well as EUS electromyography (EMGEUS) potential. These results indicate a functional connection between PSNsL3-L6 DCM and the EUS. We hypothesize that PSNsL3-L6 DCM participate in a spinal circuit to modulate EUS relaxation and mediate the spontaneous recovery of the EUS relaxation function after complete thoracic SCI. To evaluate the physiological function of PSNsL3-L6 DCM, we will inject recombinant viruses to transduce those neurons and express channelrhodopsin for optogenetic stimulation or hM4di for chemogenetic inhibition of neuronal function. We will evaluate whether manipulation of these neurons affects EUS relaxation and voiding function by bladder cystometry, EMGEUS, and field potential recording of DCM and MNEUS at L6. We will also combine anterograde and retrograde transsynaptic viral tracing and immunostaining to characterize the neurotransmitter phenotype of these PSNsL3-L6 DCM and their postsynaptic neurons. We will then further explore whether a T8 transection SCI will enhance synaptic connections between PSNsL3-L6 DCM and their target cells to mediate the spontaneous functional recovery of the EUS. We will use similar methods as above to verify whether more synaptic connections are established in this circuit at 4 weeks after SCI when the spinal reflex for EUS relaxation spontaneously recovers. Furthermore, chronic inhibition of PSNsL3-L6 DCM will be made immediately after SCI for 4 weeks to determine the causal contribution of neural plasticity to the functional recovery of EUS relaxation and voiding function. This study will be the first to explore the role of PSNs in EUS control in both intact and injured spinal cords, thus extending our understanding of cellular mechanisms of EUS control. This will provide a fundamental basis for some promising future therapies such as electrical stimulation of the spinal cord as a means of recovering bladder function.

抽象的 膀胱功能恢复是SCI患者人群中的重中之重，但到目前为止，有效的有效很少 疗法。一个主要原因之一是对两种自发的主要内部机制的理解有限 可塑性和科学后的治疗功能恢复。 Transection Sci剥夺了对 外部尿道括约肌（EUS）瘫痪其松弛功能并导致尿液保留。部分的 EUS松弛和空隙的恢复是自发发生的，仅当腰3（L3）之间的神经回路 和腰椎6（L6）脊髓水平保持完整。电气可以通过电气进一步提高功能 在慢性SCI模型中刺激L3脊髓。但是，下面的细胞机制尚不清楚。 在我们的初步研究中，我们采用了同类反式抗突触AAV1-GFP-CRE病毒，发现A 投影到L6背侧连合（DCM）的L3前脊髓神经元（PSN）的种群 EUS相关的中间神经元（INS）。我们将这些PSN命名为PSNSL3-L6 DCM。这些这些 PSNSL3-L6 DCM诱导EUS运动神经元（MNEU）以及EUS肌电图（EMGEUS）的场电位 潜在的。这些结果表明PSNSL3-L6 DCM与EUS之间的功能连接。我们假设 PSNSL3-L6 DCM参与脊柱回路以调节EUS松弛并自发介导 完整的胸科SCI后，EUS松弛功能的恢复。评估生理功能 PSNSL3-L6 DCM的of，我们将注入重组病毒来转导这些神经元并表达通道旋转蛋白 用于光遗传学刺激或HM4DI用于化学遗传学抑制神经元功能。我们将评估是否 这些神经元的操纵会影响EUS的松弛和膀胱囊肿，Emgeus和 DCM和MNEUS在L6处的现场潜在记录。我们还将结合顺行和逆行透射性突触 病毒追踪和免疫染色以表征这些PSNSL3-L6 DCM及其的神经递质表型 突触后神经元。然后，我们将进一步探索T8横断SCI是否会增强突触 PSNSL3-L6 DCM与其目标细胞之间的连接，以介导自发的功能恢复 EUS。我们将使用上述类似的方法来验证此中是否建立了更多的突触连接 SCI后4周回路，当时EUS松弛的脊柱反射自发恢复。此外， 慢性抑制PSNSL3-L6 DCM将在SCI之后立即进行4周后，以确定因果关系 神经可塑性对EUS松弛和空隙功能的功能恢复的贡献。这项研究会 成为第一个探索PSN在EUS控制中的作用在完整和受伤的脊髓中的作用，从而扩展了我们的 了解EUS控制的细胞机制。这将为某些有希望的人提供基本的基础 未来的疗法，例如对脊髓的电刺激作为恢复膀胱功能的一种手段。