Promoting targeted reinnervation of phrenic motor neurons and restoration of respiratory function using cell-specific expression of BDNF after cervical spinal cord injury

颈脊髓损伤后利用 BDNF 的细胞特异性表达促进膈运动神经元的定向神经支配和呼吸功能的恢复

• 批准号: 9917847
• 负责人: Brittany Charsar
• 金额: $ 5.05万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-05-01 至 2021-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9917847
• 关键词: Address; Axon; Brain Stem; Brain-Derived Neurotrophic Factor; Breathing; Cell Nucleus; Cells; Cervical; Cervical spinal cord injury; Cervical spinal cord structure; Contralateral; Cues; Denervation; Dependovirus; Down-Regulation; Drug Receptors; Electromyography; FRAP1 gene; Family; Forelimb; Functional disorder; Growth; Growth Factor; Human; Injections; Injury; Innovative Therapy; Intrinsic drive; Ipsilateral; Label; Limb structure; Modeling; Monitor; Motor; Motor Neurons; Natural regeneration; Nervous System Trauma; Neuraxis; Neurons; PTEN gene; Pain; Paralysed; Pathway interactions; Peptides; Periodicity; Phenotype; Population; Rattus; Recovery; Recovery of Function; Respiration Disorders; Respiratory Center; Respiratory Diaphragm; Respiratory physiology; Rest; Rodent Model; Serotyping; Signal Transduction; Source; Spinal; Spinal Cord; Spinal cord injury; Spinal cord injury patients; Synapses; Testing; Therapeutic; Therapeutic Intervention; Time; Tracer; Trauma; adeno-associated viral vector; axon growth; axon guidance; axon regeneration; base; central nervous system injury; designer receptors exclusively activated by designer drugs; functional restoration; in vivo; in vivo evaluation; inhibitor/antagonist; injured; member; minimally invasive; neural circuit; neurotrophic factor; novel strategies; overexpression; painful neuropathy; postsynaptic neurons; reconstruction; reinnervation; repaired; respiratory; restoration; vector

PROJECT SUMMARY We are testing a novel strategy to regenerate damaged descending bulbospinal respiratory axons and reinnervate phrenic motor neurons (PhMN) after cervical spinal cord injury (SCI) in rats. SCI is caused by trauma to the spinal cord, and more than half of all cases occur in the cervical region, leading to breathing compromise by damaging circuits involved in respiratory control. Restoration of functional deficits caused by SCI is limited due to the low intrinsic drive of neurons to regenerate axons and a lack of guidance cues to signal growing axons to appropriate targets, among others. The C3-C5 mid-cervical spinal cord levels house the PhMNs, which are responsible for diaphragm activation. PhMNs are predominately mono-synaptically innervated by supraspinal respiratory neurons located in a brainstem nucleus called the rostral Ventral Respiratory Group (rVRG). We are seeking to reverse respiratory dysfunction after SCI by restoring the crucial circuit controlling PhMNs, and thus diaphragm activation. Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family of growth factors, promotes axonal growth and acts as a guidance cue. Phosphatase and tensin homolog (PTEN) is a negative regulator of mammalian target of rapamycin (mTOR), which is responsible for pro-growth pathways including axon growth. Downregulation of PTEN has been shown to induce axon regeneration. We aim to promote targeted reinnervation of PhMNs and restore diaphragm function by systemically inhibiting PTEN with antagonist peptides to induce axon growth through the injury, followed by BDNF overexpression selectively in PhMNs via an adeno-associated virus (AAV) to direct growing axons. In Aim 1, we will determine whether providing a PhMN-specific source of the axon guidance molecule, BDNF, promotes targeted PhMN reinnervation by rVRG axons following cervical SCI. We will assess rVRG axons using an AAV vector expressing a dual anterograde/trans-synaptic tracer, examining regrowth and collateral sprouting, and identify synaptic reconnection with spared PhMNs by post-synaptic accumulation of the trans-synaptic marker. In Aim 2, we will determine whether rVRG-PhMN circuit re-connectivity promotes diaphragmatic recovery after cervical SCI. We will assess the ability of rVRG-PhMN reconnection to restore diaphragm function by testing in vivo diaphragm activation via electromyography (EMG). Excitingly, we can distinguish between modes of recovery, i.e. ipsilateral regrowth versus contralateral sprouting, by selectively silencing unilateral rVRG neurons with inhibitory Designer Receptor Exclusively Activated by Designer Drugs (DREADDs) and recording any subsequent changes in EMGs. BDNF overexpression is associated with neuropathic pain and abnormal motor function. We will test for unintended consequences of BDNF, including pain phenotypes and motor gains/deficits. We aim to use the strategy proposed here to reconnect motor neurons responsible for diaphragm activation with respiratory centers in the medulla to restore respiratory function following disruption after cervical SCI. The potential therapeutic benefits being explored will have profound implications for SCI patients suffering respiratory dysfunction.

项目总结