Name: standardized rat coronary ring preparation and real-time recording of dynamic tension changes along vessel diameter
Uploaded: 2022-06-16T14:50:00
Description: Watch this Scientific Journal Video about Standardized Rat Coronary Ring Preparation and Real-Time Recording of Dynamic Tension Changes Along Vessel Diameter at JoVE.com

这项工作得到了四川省科技计划重点研发项目（2022YFS0438）、国家自然科学基金（82104533）、中国博士后科学基金（2020M683273）和四川省科技厅（2021YJ0175）的支持。

该动物方案由成都中医药大学管理委员会审查批准（备案号：2021-11）。雄性Sprague Dawley（SD）大鼠（260-300g，8-10周龄）用于本研究。这些老鼠被关在动物室里，在实验期间可以自由饮水和进食。
1. 溶液制备
<ol><li>通过溶解118 mM的NaCl，4.7 mM的K +，2.5 mM的CaCl2，1.2 mM的KH2PO4，1.2 mM的MgCl2∙6H2O，25 mM的NaHCO3，11 m...

<ol>
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冠状动脉微循环紊乱涉及广泛的CAD患者，已逐渐得到认可，并关注充分心肌灌注的基础。考虑到突发性冠心病和心血管疾病的严重并发症，及时药物预防和治疗对于CAD17的临床个体非常重要。不可避免地，冠状动脉解剖结构的保密性及其生理结构的复杂性严重制约了对CAD18、19、20、21、22<...

冠状动脉疾病（CAD）已被广泛认为和关注为一种典型和具有代表性的心血管疾病，是发达国家和发展中国家的主要死因1，2。作为正常心脏生理功能的血液和氧气供应途径，循环血液通过心肌表面的两个主要冠状动脉和血血管网络3，4进入并滋养心脏。冠状动脉中的胆固醇和脂肪沉积物切断了心脏的血液供应和血管系统的剧烈炎症反应，导致动脉粥样硬化，稳定型心绞痛，不稳定型心绞痛，心肌梗塞或心源性猝死5，6。响应冠状动脉的病理性狭窄，代偿性加速生理心跳通过增加左心室7的输出来满足心脏本身或身体重要器官的血液供应。如果长时间的冠状动脉狭窄不能及时缓解，心脏的某些区域可能会出现广泛的新血管8。目前，CAD的临床治疗往往采用药物溶栓或手术机械溶栓和外源性仿生血管旁路，用药频繁，手术残疾很大9.因此，冠状动脉功能研究生理活动仍是心血管疾病10项的急需突破口。
除了无线遥测系统，没有可用的技术手段来检测冠状动脉生理活动，它可以动态记录 体内 冠状动脉压力，血管张力，血氧饱和度和pH值11。因此，考虑到冠状动脉的质地隐秘性和复杂性，冠状动脉的准确识别和分离无疑是探索CAD 体外4多种机制的最佳选择。
一系列多肌图系统，特别是线材显微照片微血管张力检测仪（见 材料表），是一种非常成熟的市场化设备，用于记录小血管、淋巴管和支气管的 体外 组织张力变化，具有高精度和连续动态记录12的特点。该系统已被广泛用于记录直径为60μm至10mm的腔体结构的 体外 组织张力特性。线材显微照片平台的连续加热特性在很大程度上抵消了对不利外部环境的刺激。同时，气体混合物的恒定输入和pH值使我们能够在相似的生理状态下获得更准确的血管张力数据13。然而，考虑到大鼠冠状动脉解剖定位的复杂性（图1），其分离一直令人困惑，并限制了其对多样化心血管疾病和药物开发的机制的探索。因此，本方案详细介绍了大鼠冠状动脉的解剖位置和分离过程，随后在钢丝显微照片14的平台上进行张力测量。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Apigenin</td>
			<td>Sangon Biotech Co., Ltd., Shanghai, China</td>
			<td>150731</td>
			<td></td>
		</tr>
		<tr>
			<td>CaCl2</td>
			<td>Sangon Biotech Co., Ltd., Shanghai, China</td>
			<td>A501330</td>
			<td></td>
		</tr>
		<tr>
			<td>D-glucose</td>
			<td>Sangon Biotech Co., Ltd., Shanghai, China</td>
			<td>A610219</td>
			<td></td>
		</tr>
		<tr>
			<td>HEPES</td>
			<td>Xiya Reagent Co., Ltd., Shandong, China</td>
			<td>S3872</td>
			<td></td>
		</tr>
		<tr>
			<td>KCl</td>
			<td>Sangon Biotech Co., Ltd., Shanghai, China</td>
			<td>A100395</td>
			<td></td>
		</tr>
		<tr>
			<td>KH2PO4</td>
			<td>Sangon Biotech Co., Ltd., Shanghai, China</td>
			<td>A100781</td>
			<td></td>
		</tr>
		<tr>
			<td>LabChart Professional version 8.3&#160;</td>
			<td>ADInstruments, Australia</td>
			<td>&#8212;</td>
			<td></td>
		</tr>
		<tr>
			<td>MgCl2&#183;6H2O</td>
			<td>Sangon Biotech Co., Ltd., Shanghai, China</td>
			<td>A100288</td>
			<td></td>
		</tr>
		<tr>
			<td>Multi myograph system&#160;</td>
			<td>Danish Myo Technology, Aarhus, Denmark</td>
			<td>620M</td>
			<td></td>
		</tr>
		<tr>
			<td>NaCl</td>
			<td>Sangon Biotech Co., Ltd., Shanghai, China</td>
			<td>A100241</td>
			<td></td>
		</tr>
		<tr>
			<td>NaHCO3</td>
			<td>Sangon Biotech Co., Ltd., Shanghai, China</td>
			<td>A100865</td>
			<td></td>
		</tr>
		<tr>
			<td>Steel wires</td>
			<td>Danish Myo Technology, Aarhus, Denmark</td>
			<td>400447</td>
			<td></td>
		</tr>
		<tr>
			<td>U46619</td>
			<td>Sigma, USA</td>
			<td>D8174</td>
			<td></td>
		</tr>
	</tbody>
</table>

standardized rat coronary ring preparation and real-time recording of dynamic tension changes along vessel diameter

冠状动脉疾病（CAD）作为心血管系统疾病的重要事件，被广泛认为是动脉粥样硬化、心肌梗塞、心绞痛等严重威胁全世界人民生命和健康的罪魁祸首。然而，如何记录孤立血管的动态生物力学特征，长期以来一直困扰着人们。同时，精确定位和分离冠状动脉以测量 体外 动态血管张力变化已成为CAD药物开发的趋势。本方案描述了大鼠冠状动脉的宏观鉴定和微观分离。使用已建立的多肌图系统监测冠状动脉环沿血管直径的收缩和扩张功能。冠状动脉环张力测量的标准化和编程方案，从采样到数据采集，极大地提高了实验数据的可重复性，保证了生理、病理和药物干预后血管张力记录的真实性。

解剖学上定位，大鼠冠状动脉分布并隐藏在心肌组织深处不容易识别。通过比较人（图1A）和大鼠的冠状动脉（图1B），根据 图2中的采样过程进行大鼠冠状动脉的快速准确分离。在光学显微镜下从前部精确定位右耳廓，肺动脉和顶点后，沿着 图2A所示的实心黑线解剖心肌。冠状动脉脑室间分支的约5毫米明显...

Watch this Scientific Journal Video about Standardized Rat Coronary Ring Preparation and Real-Time Recording of Dynamic Tension Changes Along Vessel Diameter at JoVE.com

Standardized Rat Coronary Ring Preparation and Real-Time Recording of Dynamic Tension Changes Along Vessel Diameter

本方案描述了用于测量大鼠冠状动脉血管反应性的线肌图技术。

标准化大鼠冠状动脉环制备及实时记录血管直径动态张力变化

As a key event of cardiovascular system diseases, coronary artery disease (CAD) has been widely regarded as the main culprit of atherosclerosis, ...

该方案详细介绍了大鼠冠状动脉的定位，分离和肌肉环固定。为各种心血管疾病的机制、探索和治疗提供了一种新的方法。该系统适用于和稳定记录体外动脉张力的动态变化，读数从60微米到10毫米。我们建议新手首先应准确了解每个血管的解剖位置，并且可以从稍大的血管开始，例如胸动脉和肠系膜动脉。解离并取出心脏后，首先用医用塑料镊子轻轻挤压，从所有心腔排出残留的血液。将预处理的心脏快速置于含有95%氧气和5%二氧化碳饱和生理盐溶液的培养皿中，在4摄氏度下，pH值为7.40。为了准确识别冠状动脉的解剖位置，根据示意图在光学显微镜下调整孤立心脏的姿势。用手术剪刀和镊子从肺动脉根部沿心室间隔切除左心室和右心室腔。为了将左右冠状动脉与心肌组织分离，请在光学解剖显微镜下解剖右心室，以彻底暴露右冠状动脉分支。然后通过顺时针旋转心脏组织45度来识别左冠状动脉的位置。去除周围粘稠的心肌组织后，明确辨别脉动的左右冠状动脉。立即分离中间的冠状动脉，并在四摄氏度下完全浸入生理盐溶液中。通过用解剖学剪刀垂直切割分离的动脉来记录不同刺激下的血管张力，从而获得约两毫米的动脉环。准备两根两根两厘米的不锈钢丝，预浸泡在饱和95%氧气和5%二氧化碳的四摄氏度生理盐溶液中。在光学解剖显微镜下，将两根导线沿着血管的方向平行穿过动脉环，并在血管腔的两端暴露相等长度的导线。用前后钢丝将动脉环固定在充满95%氧气和5%二氧化碳的冒泡生理盐溶液的线显微照片的浴中。旋转水平螺钉旋钮以获得适当的前后间距，使两根导线水平，动脉环处于自然松弛状态。在恒温装置上安装DMT浴槽后，打开数据采集软件，确保记录到相应的路径信号。设置参数。通过沿血管直径施加合理的张力，实现动脉环的最佳初始张力。此时，将显示的血管张力值设置为零。之后，通过旋转浴的螺旋轴对动脉环施加三毫牛顿脉冲刺激。单击LCD面板将其设置为零毫牛顿并保持一小时。在37摄氏度pH 7.40的氧饱和生理盐溶液缓冲液中孵育一小时后，在张力控制面板上再次设置张力值为零毫牛顿。使用线肌图技术执行冠状动脉环的收缩活性，并通过用60毫摩尔钾离子溶液刺激每个10分钟，在三个单独的操作中进行验证。每次刺激后，用氧饱和生理盐溶液冲洗浴，直到血管张力恢复到其初始状态。加入0.3微摩尔U46619到浴中，观察10分钟。加入一个微摩尔芹菜素并观察血管效应。当扩张达到平台时，加入下一个浓度。向60毫摩尔钾预缩溶液中加入一微摩尔芹菜素，观察血管效应。当扩张达到平台时，加入下一个浓度。向静息血管环中加入28毫摩尔钾离子。当收缩达到平台时，加入下一个浓度。将0.01微摩尔U46619加入静息血管环。当收缩达到平台时，加入下一个浓度。在对动脉环施加初始三毫牛顿张力后，通过平行施加60毫摩尔钾离子浓度三次，其张力超过两毫牛顿。因此，上述程序产生了具有出色生理活性的孤立冠状动脉环。将累积钾离子浓度或U46619加入到DMT620M的浴中，导致体外血管张力的浓度依赖性增加。当血管收缩效应达到平台时，加入下一个浓度的钾离子或U46619。对于由钾离子和U46619收缩的孤立冠状动脉环，测试药物芹菜素以令人惊讶的浓度依赖性方式引起血管舒张。心脏从肺动脉根部沿着心室间隔解剖，暴露并分离两毫米冠状动脉肌环。在此程序之后，我们可以记录视网膜微血管，肾功能，肾动脉和大脑中动脉的血管张力。从60微米到10毫米的区域动态。

Research

JoVE Journal

Medicine

X-ray Dose Reduction through Adaptive Exposure in Fluoroscopic Imaging

通过自适应曝光的减少剂量X射线透视影像

X-ray fluoroscopy is widely used for image guidance during cardiac intervention. However, radiation dose in these procedures can be high, and this is a ...

科研

医学

Retrograde Perfusion and Filling of Mouse Coronary Vasculature as Preparation for Micro Computed Tomography Imaging

微型计算机断层扫描成像制备小鼠冠状动脉血管逆行灌注和灌装

Visualization of the vasculature is becoming increasingly important for understanding many different disease states. While several techniques exist for ...

Real-time Digital Imaging of Leukocyte-endothelial Interaction in Ischemia-reperfusion Injury (IRI) of the Rat Cremaster Muscle

Real-time Digital Imaging of Leukocyte-endothelial Interaction in Ischemia-reperfusion Injury IRI of the Rat Cremaster Muscle

实时数字成像大鼠提睾肌缺血再灌注损伤（IRI）的白细胞 - 内皮细胞相互作用

Ischemia-reperfusion injury (IRI) has been implicated in a large array of pathological conditions such as cerebral stroke, myocardial infarction, ...

2-Vessel Occlusion/Hypotension: A Rat Model of Global Brain Ischemia

2血管闭塞/低血压：全脑缺血模型大鼠

Cardiac arrest followed by resuscitation often results in dramatic brain damage caused by ischemia and subsequent reperfusion of the brain. Global brain ...

Movement Retraining using Real-time Feedback of Performance

的运动再培训采用实时反馈的性能

Any modification of movement - especially movement patterns that have been honed over a number of years - requires re-organization of the neuromuscular ...

Cerebrospinal Fluid MicroRNA Profiling Using Quantitative Real Time PCR

脑脊液的microRNA分析采用实时定量PCR技术

MicroRNAs (miRNAs) constitute a potent layer of gene regulation by guiding RISC to target sites located on mRNAs and, consequently, by modulating their ...

Chemotherapy-induced Vascular Toxicity - Real-time In vivo Imaging of Vessel Impairment

Chemotherapy-induced Vascular Toxicity - Real-time In vivo Imaging of Vessel Impairment

化疗引起的血管毒性 - 实时<em&gt;在体内</em&gt;船舶减值成像

Certain classes of chemotherapies may exert acute vascular changes that may progress into long-term conditions that may predispose the patient to an ...

Real-time Cytotoxicity Assays in Human Whole Blood

实时的细胞毒性分析在人全血

A live cell-based whole blood cytotoxicity assay (WCA) that allows access to temporal information of the overall cell cytotoxicity is developed with ...

Cavernous Nerve Stimulation and Recording of Intracavernous Pressure in a Rat

大鼠海绵状神经刺激与海绵压力记录

The stimulation of the cavernous nerve (CN) and measurement of intracavernous pressure (ICP) have been used extensively to test and evaluate therapies for ...

Real-time Pressure-volume Analysis of Acute Myocardial Infarction in Mice

小鼠急性心肌梗死的实时压力容积分析

Acute myocardial infarction can lead to acute heart failure and cardiogenic shock. The evaluation of hemodynamics is critical for the evaluation of any ...