Neurogenic Calcium Signals in Small Arteries

小动脉中的神经源性钙信号

• 批准号: 6728132
• 负责人: Withrow Gil Wier
• 金额: $ 29.22万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-02-01 至 2008-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6728132
• 关键词: action potentials; adenosine triphosphate; artery; calcitonin gene related peptide; calcium flux; calcium indicator; confocal scanning microscopy; genetically modified animals; laboratory mouse; laboratory rat; membrane potentials; neuromuscular transmission; neuropeptide Y; neuropeptide receptor; norepinephrine; purinergic receptor; receptor binding; receptor expression; vascular smooth muscle nervous control

DESCRIPTION (provided by applicant): The research will focus on the roles and mechanisms of neurogenic Ca 2+ signals in autonomic nervous system control of arterial diameter, particularly the Ca2+ signals generated in smooth muscle cells (SMC) by the concerted action of the sympathetic 'triad' of co-transmitters, ATP, norepinephfine (NE) and neuropeptide Y (NPY). Although components of neurogenic vasoconstriction are attributable to ATP, NE and NPY, the underlying Ca 2+ signals in SMC are not known. Preliminary confocal imaging of Ca 2+ within the SMC of intact pressurized rat mesenteric small arteries during low frequency nerve stimulation has revealed novel Ca2+ transients, putatively associated with excitatory junction potentials (EJPs) and provisionally called "junctional Ca 2+ transients" (jCaTs). With higher frequency stimulation, Ca 2+ 'flashes' (may be due to SMC action potentials) and Ca 2+waves are generated. Stimulation of perivascular sensory nerves elicits vasodilation; preliminary results implicate an increase in the frequency of Ca 2+ sparks. Specific Experimental goals include: 1) Test the hypothesis that jCaTs arise from Ca 2+entering SMCs through P2X1 receptors, which are activated by the ATP contained in one or a few quanta of transmitter, 2) Test the hypothesis that neurally released ATP elicits Ca 2+ influx, but not Ca 2+ waves, 3) Test the hypothesis that neurally released NE elicits Ca 2+ waves that function to activate contraction, 4) Test the hypothesis that neurally released NPY activates the Y1 receptor on SMCS to modulate the effects of NE by changing the frequency of Ca 2+ waves, and 5) Test the hypothesis that calcitonin gene related peptide (CGRP) released from sensory nerves decrease Ca 2+ spark frequency; thus causing vasodilation. Mesenteric small arteries from rats and mice (including a P2Xz-receptor deficient transgenic mouse) will be loaded with fluo-4 and mounted on a confocal myograph for simultaneous Ca2+ imaging, recording of isometric force, and measurement of membrane potential. A real-time confocal microscope will be used to provide 2D images fast enough to resolve jCaTs, flashes and waves. The research will test a comprehensive hypothesis on sympathetic neuromuscular transmission and the control of smooth muscle contraction by the autonomic nervous system: Neurally released ATP activates P2X1 receptors. The resulting Ca 2+ influx activates a small contraction, generates EJPs and is visualized as jCaTs. EJPs may summate to trigger action potentials, visualized as Ca 2+ 'flashes' Neurally released NE and NPY (as metabotropic or G protein-lined receptors) increase the frequency of Ca 2+ waves, which activate strong contraction. By visualizing novel, physiological neurogenic Ca 2+ signals and contraction of intact arteries, the research seeks to provide new information on the concerted action of neurotransmitters in controlling arterial function.

描述（由申请人提供）：该研究将重点关注神经源性 Ca 2+ 信号在自主神经系统控制动脉直径中的作用和机制，特别是通过共递质 ATP、去甲肾上腺素 (NE) 和神经肽 Y (NPY) 交感“三联体”的协同作用在平滑肌细胞 (SMC) 中产生的 Ca2+ 信号。尽管神经源性血管收缩的成分可归因于 ATP、NE 和 NPY，但 SMC 中潜在的 Ca 2+ 信号尚不清楚。在低频神经刺激过程中，对完整加压大鼠肠系膜小动脉的 SMC 内的 Ca 2+ 进行初步共聚焦成像，发现了新的 Ca2+ 瞬变，推测与兴奋性交界电位 (EJP) 相关，暂时称为“交界 Ca 2+ 瞬变”(jCaTs)。通过较高频率的刺激，Ca 2+ “闪烁”（可能是由于 SMC 动作电位）并产生 Ca 2+ 波。刺激血管周围感觉神经引起血管舒张；初步结果表明Ca 2+ 火花的频率增加。具体实验目标包括： 1) 检验 jCaT 是由 Ca 2+ 通过 P2X1 受体进入 SMC 产生的假设，P2X1 受体由一个或几个量的递质中包含的 ATP 激活，2) 检验神经释放的 ATP 引发 Ca 2+ 流入，而不是 Ca 2+ 波的假设，3) 检验神经释放的 NE 引发可激活收缩的 Ca 2+ 波的假设， 4）测试神经释放的NPY激活SMCS上的Y1受体以通过改变Ca 2+ 波的频率来调节NE的作用的假设，以及5）测试感觉神经释放的降钙素基因相关肽（CGRP）降低Ca 2+ 火花频率的假设；从而引起血管舒张。来自大鼠和小鼠（包括 P2Xz 受体缺陷的转基因小鼠）的肠系膜小动脉将装载 Fluo-4 并安装在共焦肌动描记器上，用于同步 Ca2+ 成像、记录等长力和测量膜电位。实时共焦显微镜将用于提供足够快的 2D 图像，以解析 jCaT、闪光和波。该研究将测试关于交感神经肌肉传递和自主神经系统控制平滑肌收缩的综合假设：神经释放的 ATP 激活 P2X1 受体。由此产生的 Ca 2+ 流入激活小幅收缩，产生 EJP 并可视化为 jCaT。 EJP 可能会触发动作电位，表现为 Ca 2+“闪烁”。神经释放的 NE 和 NPY（作为代谢型或 G 蛋白排列的受体）增加 Ca 2+ 波的频率，从而激活强烈的收缩。通过可视化新颖的生理神经源性 Ca 2+ 信号和完整动脉的收缩，该研究旨在提供有关神经递质在控制动脉功能中的协同作用的新信息。