Transmitter Repletion: Key to Phrenic-Diaphragm Function

发射器补充：膈隔膜功能的关键

• 批准号: 6608784
• 负责人: ERIK VAN LUNTEREN
• 金额: $ 25.8万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-08 至 2006-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6608784
• 关键词: 2,4 dinitrophenol; acetylcholine; calcium flux; diaphragm; disease /disorder model; electrophysiology; fluorescent dye /probe; hypercapnia; hypothermia; hypoxia; laboratory rat; myasthenia gravis; neuromuscular function; neuromuscular junction; neuromuscular transmission; neurotransmitter transport; phrenic nerve; potassium channel; respiratory function; vesicle /vacuole

DESCRIPTION (provided by applicant): Failure of phrenic nerve-diaphragm neuromuscular transmission leads to hypercapnic respiratory failure. This occurs not only in overtly diseased neuromuscular junctions (eg. myasthenia gravis, botulism), but also in normal junctions subjected to high intensity activation during mechanical loading by lung disease (eg. COPD) or during exposure to systemic factors such as hypoxia and hypothermia. Neurotransmission requires sufficient prejunctional release of acetylcholine (ACh) to ensure muscle contraction. During repetitive activation, ACh release diminishes, which when severe leads to transmission failure. Restoration of ACh available for release depends on two separate but interrelated processes: recycling of transmitter from the synaptic cleft, and repletion of the immediately releaseable vesicle pool from one or more reserve pools. Respiratory muscles are active continuously, so that transmitter replenishment needs to be sufficiently robust to ensure that a constant supply of ACh is available for release. The overall objective of this proposal is to further examine the role of transmitter replenishment, and the factors which regulate replenishment, in determining the integrity of transmission in respiratory neuromuscular junctions. The specific hypotheses to be tested are as follows. 1) The rapidity of, and time available for, ACh replenishment are critical determinants of transmission at the phrenic-diaphragm neuromuscular junction, especially in diseased neuromuscular junctions. 2) ACh replenishment is hastened by high frequency stimulation, an accommodation to the adverse effects of high frequency activation on release and depletion. Furthermore, the acceleration of replenishment is mediated by elevated presynaptic [Ca+ +]. 3) Hypoxia and hypothermia impair neurotransmission to a large extent by slowing transmitter replenishment, rather than primarily by a direct inhibition of transmitter release. 4) Presynaptic K+ channels regulate not only Ach release but also transmitter replenishment, providing two mechanisms of improving neurotransmission by pharmacologic manipulation of K- channel conductances. Neuromuscular transmission will be assessed using a combination of force measurements to quantify the neuromuscular component of fatigue, electrophysiological recording to determine ACh release and recovery from transmitter rundown, and optical approaches using fluorescent styry1 dyes ( FM1-43, FM2-10) to assess vesicle pool dynamics. These studies may lead to novel therapeutic approaches to respiratory muscle impairment and resulting hypercapnic respiratory failure for conditions which produce neuromuscular junction dysfunction.

描述（由申请人提供）：膈神经-膈神经肌肉传递失败导致高碳酸血症性呼吸衰竭。这不仅发生在明显病变的神经肌肉连接处(如：重症肌无力，肉毒杆菌中毒)，但在肺部疾病引起的机械负荷期间受到高强度激活的正常连接(例如：慢性阻塞性肺病)或暴露于全身因素，如缺氧和体温过低。神经传递需要充分的突触前释放乙酰胆碱（ACh）以确保肌肉收缩。在重复激活过程中，乙酰胆碱释放减少，严重时导致传输失败。可释放乙酰胆碱的恢复取决于两个独立但相互关联的过程：突触间隙的递质再循环，以及一个或多个储备池中立即释放的囊泡池的补充。呼吸肌持续活动，因此递质补充需要足够稳健，以确保持续供应的乙酰胆碱可用于释放。本提案的总体目标是进一步研究递质补充的作用，以及调节补充的因素，以确定呼吸神经肌肉连接中传递的完整性。需要检验的具体假设如下：1)乙酰胆碱补充的速度和可用时间是膈-膈神经肌肉连接处，特别是病变神经肌肉连接处传递的关键决定因素。2)高频刺激可加速乙酰胆碱的补充，缓解了高频激活对乙酰胆碱释放和消耗的不利影响。此外，补充的加速是由突触前[Ca+ +]升高介导的。3)低氧和低温在很大程度上通过减缓递质补充而损害神经传递，而不是主要通过直接抑制递质释放。4)突触前K+通道不仅调节乙酰胆碱释放，还调节递质补充，提供了通过药物调控K通道传导改善神经传递的两种机制。神经肌肉传递将通过力测量来量化疲劳的神经肌肉成分，电生理记录来确定乙酰胆碱的释放和从递质衰竭中恢复，以及使用荧光styry1染料（FM1-43, FM2-10）来评估囊泡池动力学的光学方法来评估。这些研究可能会导致新的治疗方法，以呼吸肌肉损伤和导致高碳酸血症性呼吸衰竭的条件下，产生神经肌肉连接功能障碍。