Transmitter Repletion: Key to Phrenic-Diaphragm Function

发射器补充：膈隔膜功能的关键

• 批准号: 6736977
• 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/6736977
• 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-隔膜神经肌肉传递的失败导致高碳酸血症性呼吸衰竭。这种情况不仅发生在明显患病的神经肌肉接头（例如重症肌无力、肉毒杆菌中毒）中，而且也发生在肺部疾病（例如慢性阻塞性肺病）机械负荷期间或暴露于缺氧和体温过低等全身因素期间受到高强度激活的正常接头中。神经传递需要足够的乙酰胆碱 (ACh) 交界前释放以确保肌肉收缩。在重复激活过程中，乙酰胆碱释放减少，严重时会导致传输失败。可释放的乙酰胆碱的恢复取决于两个独立但相互关联的过程：从突触间隙回收递质，以及从一个或多个储备池补充可立即释放的囊泡池。呼吸肌持续活跃，因此递质补充需要足够强大，以确保有持续的乙酰胆碱供应可供释放。该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接头传输的完整性。具体待检验的假设如下。 1) 乙酰胆碱补充的速度和可用时间是膈膈神经肌肉接头处传输的关键决定因素，特别是在患病的神经肌肉接头处。 2) 高频刺激可加速乙酰胆碱的补充，以适应高频激活对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）缺氧和低温在很大程度上通过减慢递质补充来损害神经传递，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 Ach 释放，还调节递质补充，提供两种通过药物操纵 K- 通道电导来改善神经传递的机制。将使用力测量组合来评估神经肌肉传递，以量化疲劳的神经肌肉成分，电生理记录以确定乙酰胆碱释放和递质耗尽的恢复，以及使用荧光 styry1 染料（FM1-43、FM2-10）的光学方法来评估囊泡池动态。这些研究可能会带来针对呼吸肌损伤和导致神经肌肉接头功能障碍的高碳酸血症呼吸衰竭的新治疗方法。