Name: measurement of cytosolic ca2+ in isolated contractile lymphatics
Uploaded: 2011-12-08T13:00:00
Description: Watch this Scientific Journal Video about Measurement of Cytosolic Ca2+ in Isolated Contractile Lymphatics at JoVE.com

<p class="jove_content"这项工作是由美国国立卫生研究院授予P20RR018766和酒精研究从ABMRF /基金会授予的支持。</p

1。动物<ol><li>机构动物护理和使用委员会在路易斯安那州立大学健康科学中心的所有程序批准，并按照国家卫生研究所（NIH出版物，编号85-12，1996年修订）的指引执行。雄性SD大鼠（查尔斯河实验室，体重270-350克）被安置在一个控制温度（22℃）和控制的照明（12:12 h光照明暗周期）环境。抵达后，将大鼠提交了为期一周的驯化期间，提供了标准的大鼠议员（2018 Teklad全球18％的蛋白质鼠类饮食，...

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一个新颖的方法组合研究内在的淋巴管抽水。能够同时测量变化的[Ca 2 +] i和抽水淋巴管的直径将允许的 Ca 2 +依赖和 Ca 2 +敏的信号通路在整体管收缩淋巴循环的机制。相对贡献的研究淋巴管收缩周期由过音叠加的阶段性收缩。数据表明，每一个阶段性的收缩与中的[Ca 2 +] i的瞬态上升，这表明一个核心作用?...

<table border="1"><tbody><tr><td> 1。林格5倍股 </td></tr><tr><td化学</td><td> 公司 </td><td> 产品编号 </td><td> 金额 </td></tr><tr><td>氯化钠</td><td>电解二氧化锰</td><td> SX0420 - 3 </td><td> 35摹</td></tr><tr><td>氯化钾</td><td> JT贝克</td><td> 3040 </td><td> 1.75摹</td></tr><tr><td>氯化钙</td><td>西格玛</td><td> C - 3881 </td><td> 1.47摹</td></tr><tr><td>硫酸镁</td><td>西格玛</td><td> M - 9397 </td><td> 1.44摹</td></tr><tr><td>无菌过滤水</td><td> N / A </td><td> N / A </td><td>置于1升</td></tr><tr><td>进入蒸压瓶和商店的无菌过滤器，在4 ° C </td></tr><tr><td></td><td></td><td></td><td></td></tr><tr><td> 2。 MOPS缓冲 </td><td></td><td></td><td></td></tr><tr><td> 化学 </td><td> 公司 </TD&gt; <td> 产品编号 </td><td> 金额 </td></tr><tr><td> MOPS </td><td>西格玛</td><td> M3183 </td><td> 125.6摹</td></tr><tr><td>无菌过滤水</td><td> N / A </td><td> N / A </td><td>置于1升</td></tr><tr><td>进入蒸压瓶和商店的无菌过滤器，在4 ° C </td></tr><tr><td></td><td></td><td></td><td></td></tr><tr><td> 3。白蛋白的生理盐溶液（APSS） </td></tr><tr><td> 化学 </td><td> 公司 </td><td> 产品编号 </td><td> 金额 </td></tr><tr><td>林格股票（5X） </td><td> N / A </td><td> N / A </td><td> 200毫升</td></tr><tr><td> MOPS缓冲</td><td> N / A </td><td> N / A </td><td> 5毫升</td></tr><tr><td>磷酸钠</td><td>西格玛</td><td> S - 9638 </td><td> 0.168摹</td></tr><tr><td>丙酮酸钠</td><td>西格玛</td><td> P5280 </td><td> 0.22摹</td></tr><tr><td> EDTA钠SAL吨</td><td>西格玛</td><td> ED2SS </td><td> 0.0074摹</td></tr><tr><td>血糖</td><td>西格玛</td><td> G7528 </td><td> 0.901摹</td></tr><tr><td>白蛋白，牛</td><td> USB </td><td> 10856 </td><td> 10克</td></tr><tr><td>无菌过滤水</td><td> N / A </td><td> N / A </td><td>置于1升</td></tr><tr><td>调整pH至7.4，在37 °，然后放入高压灭菌瓶和储存在4℃的无菌过滤° C。 </td></tr></tbody></table> 表1具体所用试剂。所有存储在4 ° C。

measurement of cytosolic ca2+ in isolated contractile lymphatics

淋巴管组成一个多功能的交通运输系统保持流体的动态平衡，血脂中央流通，作为一个潜在的有害抗原的监控系统，优化粘膜免疫和适应性免疫反应1。从组织间液进入盲端的初始淋巴管，然后是对运输压力梯度较大的收集淋巴管形成淋巴。每个收集淋巴是由一系列称为lymphangions段，二尖瓣阀门，防止回流隔开。每个lymphangion拥有一个收缩的周期，推动反压力梯度向中央循环 2淋巴结。这种阶段性的收缩模式类似心动周期，收缩压和舒张压阶段，并具有较低的收缩频率4。此外，淋巴管平滑肌产生提示音并显示生肌的收缩和扩张，我ñ回应管腔压力的增加和减少，分别为5。因此，建议同时支持淋巴管的阶段性和补品收缩的分子机制的混合体。 平滑肌收缩一般是受细胞质的Ca 2 +浓度（内[Ca2 +] i的），加上灵敏度的Ca 2 +，在周围的细胞6环境变化的响应收缩元素。的[Ca 2 +] 我是运动的Ca 2 +通过质膜配体或电压门控钙离子通道的释放和吸收的Ca 2 +从内部商店相结合确定。胞质的Ca 2 +结合到钙调蛋白和酶的激活，如肌球蛋白轻链（MLC）的激酶（MLCK），这反过来又磷酸化MLC的肌动蛋白-肌球蛋白介导的收缩 8 。然而，这种通路的敏感性钙<sup&gt; 2 +可调节的MLC磷酸酶（MLCP）9。 MLCP活性受Rho激酶（ROCK）和肌球蛋白磷酸酶抑制蛋白CPI - 17。 在这里，我们目前超过隔离，淋巴管灌注时间的方法，以评估变化的[Ca 2 +] 我为了研究的Ca 2 +依赖和Ca 2 +敏淋巴管平滑肌收缩机制。采用离体大鼠肠系膜淋巴管收集，我们研究的[Ca 2 +]拉伸引起的变化我和收缩活动。孤立淋巴模型提供了优势的压力，流量，和化学成分的沐浴液，可以严格控制。的[Ca 2 +] 我是通过加载比例的Ca 2 +结合染料FURA - 2淋巴确定。这些研究将提供新的方法来学习不同的分子机制，规范阶段性的更广泛的问题进补收缩与淋巴管平滑肌的收缩。

Watch this Scientific Journal Video about Measurement of Cytosolic Ca2+ in Isolated Contractile Lymphatics at JoVE.com

Measurement of Cytosolic Ca2+ in Isolated Contractile Lymphatics

我们介绍的方法来评价细胞质CA<sup&gt; 2 +</sup&gt;孤立淋巴集中在研究CA<sup&gt; 2 +</sup&gt;依赖和Ca<sup&gt; 2 +</sup&gt;敏淋巴管平滑肌收缩机制。

胞内钙的测量 2 +</sup在隔离的收缩淋巴管

Lymphatic vessels comprise a multifunctional transport system that maintains fluid homeostasis, delivers lipids to the central circulation, and acts as a ...

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Immunology and Infection

Measurement of Cytosolic Ca2+ in Isolated Contractile Lymphatics

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