Department of Kampo Diagnostics, Institute of Natural Medicine, University of Toyama, Toyama, Japan.
Department of Molecular Pharmacology and Physiology, Morsani College of Medicine, University of South Florida, Tampa, Florida.
Am J Physiol Heart Circ Physiol. 2019 Sep 1;317(3):H561-H574. doi: 10.1152/ajpheart.00564.2018. Epub 2019 Jul 5.
In the current study, the potential contributions of ryanodine receptors (RyRs) to intrinsic pumping and responsiveness to substance P (SP) were investigated in isolated rat mesenteric collecting lymphatic vessels. Responses to SP were characterized in lymphatic vessels in the absence or presence of pretreatment with nifedipine to block L-type Ca channels, caffeine to block normal release and uptake of Ca from the sarcoplasmic reticulum, ryanodine to block all RyR isoforms, or dantrolene to more selectively block RyR1 and RyR3. RyR expression and localization in lymphatics was also assessed by quantitative PCR and immunofluorescence confocal microscopy. The results show that SP normally elicits a significant increase in contraction frequency and a decrease in end-diastolic diameter. In the presence of nifedipine, phasic contractions stop, yet subsequent SP treatment still elicits a strong tonic contraction. Caffeine treatment gradually relaxes lymphatics, causing a loss of phasic contractions, and prevents subsequent SP-induced tonic contraction. Ryanodine also gradually diminishes phasic contractions but without causing vessel relaxation and significantly inhibits the SP-induced tonic contraction. Dantrolene treatment did not significantly impair lymphatic contractions nor the response to SP. The mRNA for all RyR isoforms is detectable in isolated lymphatics. RyR2 and RyR3 proteins are found predominantly in the collecting lymphatic smooth muscle layer. Collectively, the data suggest that SP-induced tonic contraction requires both extracellular Ca plus Ca release from internal stores and that RyRs play a role in the normal contractions and responsiveness to SP of rat mesenteric collecting lymphatics. The mechanisms that govern contractions of lymphatic vessels remain unclear. Tonic contraction of lymphatic vessels caused by substance P was blocked by caffeine, which prevents normal uptake and release of Ca from internal stores, but not nifedipine, which blocks L-type channel-mediated Ca entry. Ryanodine, which also disrupts normal sarcoplasmic reticulum Ca release and reuptake, significantly inhibited substance P-induced tonic contraction. Ryanodine receptors 2 and 3 were detected within the smooth muscle layer of collecting lymphatic vessels.
在当前的研究中，研究人员调查了兰尼碱受体 (RyR) 对固有泵送和对 P 物质 (SP) 反应的潜在贡献，方法是在分离的大鼠肠系膜收集淋巴管中进行。在没有或存在硝苯地平预处理以阻断 L 型钙通道、咖啡因以阻断内质网中 Ca 的正常释放和摄取、兰尼碱以阻断所有 RyR 同工型或丹曲林以更选择性地阻断 RyR1 和 RyR3 的情况下，对淋巴管中的 SP 反应进行了特征描述。通过定量 PCR 和免疫荧光共聚焦显微镜评估了淋巴管中 RyR 的表达和定位。结果表明，SP 通常会引起收缩频率的显著增加和舒张末期直径的减小。在硝苯地平存在的情况下，相位收缩停止，但随后的 SP 处理仍会引起强烈的紧张收缩。咖啡因处理逐渐使淋巴管松弛，导致相位收缩丧失，并防止随后的 SP 诱导的紧张收缩。兰尼碱也逐渐减弱相位收缩，但不会引起血管松弛，并显著抑制 SP 诱导的紧张收缩。丹曲林处理不会显著损害淋巴管收缩或对 SP 的反应。所有 RyR 同工型的 mRNA 在分离的淋巴管中均可检测到。RyR2 和 RyR3 蛋白主要存在于收集淋巴管平滑肌层中。总的来说，这些数据表明 SP 诱导的紧张收缩需要细胞外 Ca 加上来自内部储存库的 Ca 释放，并且 RyRs 在大鼠肠系膜收集淋巴管的正常收缩和对 SP 的反应中发挥作用。调节淋巴管收缩的机制尚不清楚。由 P 物质引起的淋巴管紧张收缩被咖啡因阻断，咖啡因可防止内质网中 Ca 的正常摄取和释放，但不能被硝苯地平阻断，硝苯地平可阻断 L 型通道介导的 Ca 进入。兰尼碱也破坏了内质网 Ca 释放和再摄取的正常功能，显著抑制了 P 物质诱导的紧张收缩。RyR2 和 RyR3 在收集淋巴管的平滑肌层中被检测到。
