笔者想从NHLBI（K08 HL102066和R01 HL114832）承认资金。

执行任何在本协议中所描述的过程之前，获得由当地机构动物护理和使用委员会的批准。 1.安装试验装置 注意:在麻醉动物进行该程序和数据的质量成比例提供给动物的麻醉剂支持体的质量。该第一部分将详细介绍必要的设备和程序，以提供麻醉的小鼠的同时执行该协议。 <ol><li>选择麻醉协议。吸入麻醉剂有用于执行光伏回路分析许多有益的性能，但也有一些可注射的协议已被使用为好。查看选择的麻醉剂政权以获取更多信息的讨论。 </li><li>固定压缩氧气罐到手术台或壁手术部位附近。 </li><li>如果使用吸入麻醉剂，用蒸发器，以保证适当的剂量。每年校准汽化器，以确保他们所提供麻醉气体的适当剂量。连接蒸发器到一个流量计，它允许在该气体进入麻醉剂回路中的速率的控制。设定在0.5 - 1.0升/分。 </li><li>使用的歧管，以允许麻醉气体的，以1定向流量）的吸气室，2）的掩模，以及3）的呼吸机。清除麻醉气体是非常重要的，并应在一个活动的系统来执行，要么通风口进入通风橱（或其它类似建筑物基础设施），或通过旨在除去麻醉气体的罐。 注:与当地的职业卫生官员检查，以确保符合当地的法规。 </li><li>通过使用加热垫和/或加热灯保持体温。持续监测体温用直肠体温计。这将允许对主动增加或加热减小，以确保一个生理体温（ͭ6;血液动力数据的采集期间37℃）。 </li><li>流体提供支持，以抵消失血昏迷体积损失。 <ol><li>抽出1 ml等于25％白蛋白和在注射器加入1.5毫升的0.9％NaCl的制备白蛋白的10％溶液中的0.9％NaCl。 </li><li>准备低残留量血管内导管。 <ol><li>用钳子把美眉0.5英寸30号针头的塑料轮毂。用持针器，把握针并取出枢纽。刮使用止血钳针头剩余的粘合剂。插入针的钝端插入微孔管的长度。使用长度小于20英寸的管道。 </li></ol></li><li>使用注射泵，允许精确卷交付。 </li></ol></li><li>确保高品质的光伏数据的收集适当的通风。有多种可供购买鼠标的呼吸机。压力控制呼吸机提供吸入剂所需要的封闭环境nesthetic气体和提供在手术过程中的通风的更好的控制。 <ol><li>确保吸气压力限制为&lt;15 cm H2O的，以防止气压伤。设置通气机呼吸周期的35％时，提供吸气脉冲。在4的一级使用呼气末正压（PEEP） - 5 cm H2O的将大大提高鼠标的通风，防止肺不张及配套的气体交换。 </li><li>限制死腔远端的通气电路中的Y型接头。这是至关重要的，因为鼠标的潮气量是非常小的和新鲜的空气灵感交付任何死角减去。 </li><li>通过削减尖端关20号留置血管内导管的创建一个鼠标大小的气管（ET）管。这提供了更容易地插入一锥形点。放置的切断端插入麻醉剂电路的Y接头。 </li><li>使用软管在呼吸电路。在管道的任何结构的内存将创建具有拉气管插管出鼠标的气道的潜在的外部力量。 </li><li>让鼠标确定通过使用抑制内源性呼吸动力的最低利率的呼吸速率。开始以每分钟约60次呼吸的相对缓慢的呼吸速率。 注意:使用适当的通风鼠标应该很少的努力呼吸。然而，如果通风不足，CO 2的血液中的积聚将启动由鼠标呼吸努力。如果发生此种情况，增加呼吸频率是增加肺泡通气一个直接的方式。通常有必要增加响应于海拔与β-肾上腺素能受体刺激相关的心脏的工作负荷的呼吸速率。 </li><li>一旦呼吸电路制备，压力由堵塞气管的前端（ET）的管测试系统用手指。确保≈10CMH 2 O的气道压力这个试验前应在每个步骤进行。 </li></ol></li></ol> 2.手术方法<ol><li>使用小镊子和剪刀等设备从表1中 。所有仪表是相当小的，以允许在放大手术区域容易使用。使用外科立体显微镜，为外科手术过程的若干方面提供足够的放大。 </li><li>使用烧灼的手术过程中最大限度地止血。 注:有几个大类烧灼的。 Thermocautery加热将通过肌肉组织切割和止血薄金属元素。这些系统最初相对便宜;然而需要注意的是，导线的提示是脆弱和相对昂贵的是重要的。电灼系统更昂贵的最初购买，但提示是很坚固的，并且不会需要被更换ð。 </li><li> 诱导和手术准备 <ol><li>获得体重鼠标。 </li><li>鼠标放置成填充有5％异氟醚的感应腔室。 注意:鼠标不能比在此环境中的活几分钟。只有45 - 60秒都需要鼠标失去翻正反射（努力当放置在其背面或侧面翻转）。 </li><li>一旦翻正反射丧失，异氟烷浓度降低至2％，并打开麻醉气体到面罩。 </li><li>很快鼠标转移到手术台，并与面罩内的鼻子将其放置在背卧。 </li><li>如果使用电，用生理盐水浸湿纱布电连接鼠标电灼系统的接地垫。 </li><li>安全与手术磁带四肢。此胶带提供即使当湿粘合性能。 </li><li>将监测芯体直肠测温器温度。用胶带固定。 </li><li>应用脱毛鼠标的颈部和胸部。等待2 - 3分钟为脱毛工作，然后再从用棉签这些区域和/或实验室擦拭除去皮毛。 注意:不要求手术野的悬垂，因为这是一个非生存的方法，但可能希望限制外科医生暴露于烧灼接地垫，如果使用的话。 </li><li>一旦鼠标准备向手术，通过执行脚趾捏评估小鼠的外科平面。一旦保证适当的麻醉深度的小鼠是准备用于第一切口。 </li></ol></li><li>获得通过口头插管或气管切开气道的控制权。气管切开术是一种方便的方法是相对简单执行。 注意:在本过程描述所有方向和方位将相对于外科医生。 <ol><li>请在Le切口从中线的≈5毫米右延伸胸骨切迹的VEL到≈左中线5毫米。 </li><li>使沿第一切口的右边缘延伸的第二切口，吻侧延伸到一个水平≈2毫米尾椎下颌骨的末尾。 </li><li>使第三切口，从切口第二的喙端延伸过左中线向≈为5mm。缩回所得皮瓣向左以暴露下层组织。 </li><li>在钝性分离中线分开腮腺及颌下唾液腺。这将暴露底层肌肉组织覆盖气管。 </li><li>直截了当地分开以暴露气管右和左sternohyoideus肌肉。 </li><li>通过一个≈10厘米长3-0丝线缝合气管下，注意不包括食道。 </li><li>确定位置的气管切开术:仅仅尾椎喉部有第一气管环之前的差距，这是一个想法L存储单元进行气管切开术。 </li><li>调整所述歧管提供麻醉气体的呼吸机和打开呼吸机。 </li><li>通过用手指堵塞气管（ET）的管的尖端检查是否泄漏。确保≈10厘米水柱的气道压力。 </li><li>使用20号针头作为手术刀，切开气管。使切口比较宽，因为ET管将填补大部分气管腔。 </li><li>快速移动，取出面膜，小心地将ET管插入气管。不要强迫它作为组织是很脆弱的，并通过气管破壁可能导致气胸。 注意:立即在插入时，胸部应短途旅行变得明显。 </li><li>固定通气电路用胶带，以防止在ET管被拉出。 </li><li>在3-0缝线绑一个上手结，形成ET管周围的密封。 注意:此时胸部excursi附件应该是显而易见的。如果没有它通常是ET管的，这是问题的气管内的定位。拉气管插管，并尝试重新定位，着眼于气管为导向的方向。 </li></ol></li><li> 颈静脉网站的制备 <ol><li>缩回左唾液腺喙-侧向暴露颈外静脉。 </li><li>平分瘦肌肉（sternomastoideus），包括与钝性分离静脉。这将暴露颈静脉的外表面。 </li><li>仔细理清组织的主要部分，但作为静脉壁非常薄必须谨慎使用。一旦这项工作完成，覆盖唾液腺颈静脉后保存它插管。 </li></ol></li><li> 开胸手术;进入胸膜间隙而不损害心脏或肺 <ol><li>除去多覆盖延伸的右边缘胸部皮肤原始皮肤切口至剑突的水平，然后在整个中线到左中线≈1.5厘米。 注:应通过外部乳腺血管，其可以是出血显著源切割时作出。切割之前烧灼这些船只将在很大程度上防止这种出血。 </li><li>使用钝性分离，收回皮瓣横向暴露底层肌肉。 </li><li>隔离在右侧邻近使用血管扩张镊子胸骨尾侧方面胸大肌的插入。烧灼并切断肌肉。 </li><li>通过沿其附着到胸骨胸大肌切断。烧灼切割边缘，以确保止血。 </li><li>下一个破坏在同一侧的背阔肌，这是一个大张肌的覆盖鼠标的外侧面。烧灼和切这块肌肉，然后缩回切口处头侧。这可能需要一些生硬解剖。 注意:肋现在显而易见和心脏也可能在某些小鼠品系可见。在大多数的小鼠，在第四肋间尾半进入胸腔将提供对心脏很方便。第四肋间是第二个最尾的空间。 </li><li>要进入胸部，使用一对锋利的钳通过肋间肌肉层认真剖析了下来。 </li><li>一旦胸腔空间已经打开，小心地插入钝头血管扩张器。使用容器扩张器提供胸壁温和向上的力量，用钝端弹簧剪刀小心地切开肋间肌肉的剩余部分。 </li><li>一是横向切割，小心不要切断肺叶下方。接下来内侧延长切口，但停留3 - 中线4毫米横向避免乳内动脉。 注:乳内血管胸骨平行并且可导致显著失血如果切意外。 </li><li>仔细烧灼肋间肌的切割边缘，用小棉签向上滚动的组织，以便与切割组织烧灼接触而不与肺部或心脏接触。 </li><li>将盐水浸湿的棉花小朝通过中线指向切口倾斜涂药。提供一个温和向上的牵引，从底层结构拉胸壁路程。开始在切口的内侧边缘烧灼胸壁和结束左侧中线≈1厘米外侧。 </li><li>推进涂抹到左侧，使其始终保持在烧灼冰山下。 </li><li>一旦组织彻底烧灼，用剪刀通过胸骨经过精心切割的。心脏的顶点应该是在这一点上清晰可见。 </li><li>使用钝性分离，扰乱心包，并确定尾腔静脉。 </li><li>检查是否有出血的迹象，现在烧灼它。一旦所有出血已得到解决，小心地取出任何立体裁剪，烧灼接地垫，和生理盐水浸湿的纱布。 </li></ol></li><li> 放置颈静脉导管 <ol><li>连接在步骤1.6.2至通过在30号针头小心滑动管含有10％白蛋白的注射器所作的导管。 </li><li>开始通过导管注入的白蛋白。 </li><li>定向导管使得针在于在其自身的颈静脉，用针斜面向上。如果有必要，用针持有人在管旋转针头斜面移动到正确的方向。 </li><li>当导管完全冲洗停止输液。 </li><li>抓住用钳子导管针。用另一只手，用组织钳缩回唾液腺，以允许颈静脉的可视化。轻轻牵引周围远端颈组织静脉产生对血管壁的张力。使用方法浅的角度，仔细针头插入静脉。超前的3针针尖 - 4 mm产品进入血管。 </li><li>释放导管针固定的导管用一块胶带之前，这将限制针的任何运动一次释放。 </li><li>释放针并轻轻向后拉对针筒柱塞以确认该导管是在容器的腔，在该行可视化的血液。 </li><li>一旦定位正确固定导管与手术胶水在针附着到底层唾液腺。 </li><li>计算的总体积要输注。如果有不显著失血5微升/ g体重的体积是足够的。如果有显著失血注入6 - 6.5微升/ g的可能需要。设置流率，使得整个输液将在10个完整的 - 15分钟。 </li></ol></li><li> PV导管放置在左心室<ol><li>在上述外科手术过程中，放置光伏导管在含有生理盐水溶液的注射器，以允许它达到平衡。 </li><li>之前，为了安置，将注射器和导管旁边的鼠标。与在大致相同的高度的心脏导管尖端，零压力读数。 </li><li>用盐水浸泡小棉签，机动的心脏，让顶点的可视化。 </li><li>用25号针头，使刺切口尽可能靠近顶点尽可能的中心。 </li><li>以下在除去针，迅速通过切口插入导管。它并不需要多大的力来插入导管，所以推进导管入脑室时表现出克制。偶尔需要执行附加的刺切口。如果这是必要的，尽量靠近的初始位置执行后续切口，以尽量减少对心脏的损伤。 不E:一旦导管被推进到心室的最终位置是非常重要的。心室压力跟​​踪将通过低舒张压（&lt;10毫米汞柱）和高收缩压是明显的（&gt;在此点80毫米汞柱）。理想情况下，该导管将与刚刚脑室内外部电极心室居中。在导管位置的微调可通过观察光伏环路数据，寻找一个箱形追踪与侧面之间≈90°的角度来执行。 </li></ol></li></ol> 3.程序细节 注意:一旦导管就位简要稳定期（10 - 15分钟）是必要的，让动物从一些急性手术应激中恢复，并允许时间为流体的输注。在此之后的稳定期的实际协议就可以开始。 <ol><li>一旦光伏导管放置和手术操作已经停止，调低异氟醚≈1％作为麻醉的深刻手术飞机的需求正在减少。 <ol><li>在此期间，仔细监测鼠标以确保麻醉的适当电平被维持。仔细评估任何运动;呼吸肌的运动表明通气不足的电平，并可以通过提高呼吸的呼吸速率来解决。四肢或晶须的抽动运动迹象表明，老鼠越来越太轻，需要更多的麻醉剂。 注意:有各种各样的，可以结合该协议被用来治疗排列。许多这些治疗需要的药物输注。它有效地管理死空间体积是必不可少的。溶液交换机可以通过从一个注射泵向其他的针滑动的导管来实现。不久，早期输液结束前这么做是允许的导管被装与下一个的药物。有必要知道导管管道内的体积，以确定溶液开关的定时。引入在该行的小气泡的允许确定输液开关的精确定时;该气泡注入静脉循环的耐受性良好。 </li></ol></li><li>改变通过降低负荷和后负荷增加心脏的负荷条件。 <ol><li>通过阻断静脉回流到心脏前负荷降低。在此制备，可视化和随着从膜片到心脏通过阻塞尾部腔静脉。平滑且相对快速地执行该闭塞，持续不超过2 - 3秒。 2秒 - 通过执行一个温柔的腹部压迫持续1瞬时增加左室后负荷。 </li><li>在这些变化在心脏负荷，暂停呼吸，消除呼吸机引入的任何神器。 </li></ol></li><li>使用校准第体积信号Ë电导导管。这些过程是不必要的，使用导纳技术导管。 <ol><li>实验方案后注入5 - 10微升高渗盐水（20％NaCl）中的计算出并联电导。 </li><li>由取出导管和从左心室采血到肝素化的注射器采集血液。将此血液进入已知体积的比色杯中，并使用导管来测量电导。 </li><li>使用比色皿电导措施的电导信号转换成体积和并联电导必要定义由导管测量的绝对体积。 </li></ol></li><li>一旦该协议完成后，通过除去心脏下列腔静脉和主动脉附件的横断安乐死鼠标。 </li></ol>

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The author has nothing to disclose.

有在此过程中三个关键步骤:1）气管导管和适当的通风，2）颈静脉导管的安置，以及3）在左心室光伏导管的正确放置的位置。确定适当的呼吸频率提供通气支持的重要组成部分。清醒小鼠一般保持快速浅呼吸肺泡通气。一般情况下，通风老鼠将有更大的潮气量。从而较慢的呼吸速率是必需的。这很重要，因为太少的通风会导致呼吸性酸中毒和太多的通风会导致呼吸性碱中毒，这两个条件，这将改...

心血管疾病仍然是世界各地的1死亡率和发病率的一个显著原因。心脏的疾病，目前在开发新的治疗特别困难的挑战。在遗传学的进展提供了识别潜在的遗传贡献者多种心脏疾病发展的可能性。心血管系统的综合性质，要求这些遗传目标的完整的动物模型中进行验证。鼠标的遗传灵活性和低住房成本已经把它带到了最前沿的一个特定基因的生理作用的评估。鼠标的小尺寸提出了心脏功能评估一些独特的挑战。有几种方式，可以提供关于心脏功能的信息，但只有心室压力和体积的同时测量允许压力容积（PV）心室功能的循环分析。光伏全循环流心脏功能进行分析独立于其连接到脉管的;在确定特定遗传元件的功能作用的重要因素。 压力-体积环的评估已经用于实验和临床上多年和广泛的文献存在对这些数据的分析台2,3。光伏循环技术的鼠标适配一直小鼠心脏生理4-6的理解的重要进展。基于导管的光伏循环技术对夫妇一个压力传感器和采用电导估计心室容积。心室体积是通过检查在由导管产生的电场的变化来确定。心室为圆柱体，它的高度由在导管和半径电极之间的距离所限定的本方法的模型是由电场的传导通过血液中计算心室7-9。由导管测定的电导信号有两个分量。第一种是通过血液中的导通;这个变化与心室的体积，并构成用于确定心室体积的初级信号。第二成分通过，并沿着心室的壁由传导引起的。这就是所谓的并联电导，并且必须以确定的绝对心室容积中移除。有对压力-体积数据的在研究实验室的收集和用于计算和删除并行电导的方法两种市售系统在它们6,10,11之间的主要区别。电导导管需要高渗盐水的平行电导的计算的注入。此注入瞬时改变心室血液的导电性，而壁的导电性保持不变。从这个数据，可以判断从血液起源和什么电导信号的分量来自于室壁。这种方法假设平行电导不会在心动周期中发生变化。准入方法依赖于电场的相位变化来评估心室壁的整体音量信号的贡献。此方法依赖于多种用于血液和心肌确定最终体积的电导率预定常数，不过在心动周期中，使平行电导的连续的措施。这两个系统的提供左心室体积的良好估计和它们之间的差异不太可能是生理显著。心室和其他假设的圆柱模型渲染不一样准确其他方式这些基于导管的方法，但是这种数据设置在一个节拍逐节拍的基础是对心功能的负载独立措施的评价是至关重要的。 这里所概述的程序是在我的实验室中使用，并已进行了大量的研究探讨营养不良病12-18的基本病理生理机制提供的数据。下面概述的过程是一个可以被用于获得光伏环两个数据之一。虽然许多原则适用于任何一种方法，该协议将专注于开胸顶的方法;一个胸外协议已在别处19,20详细说明。而程序将详细描述的那样，重要的首要原则是与要么心脏或肺部损害最小暴露心脏。在整个协议的是要记住，这是一个非生存过程，并且具有一个心脏良好曝光为导管的正确放置极为重要的是非常重要的。

<table border="0" cellpadding="0" cellspacing="0" width="523">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Dumont 5/45 (2)</td>
			<td style="width:127px;">Fine Science Tools</td>
			<td style="width:119px;">11251-33</td>
			<td style="width:63px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Vessel Dilating Forceps</td>
			<td style="width:127px;">Fine Science Tools</td>
			<td style="width:119px;">18153-11</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Castroviejo Micro Dissecting Spring Scissor</td>
			<td style="width:127px;">Roboz Instruments</td>
			<td style="width:119px;">RS-5668</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Octogon Forceps - Serrated/Curved</td>
			<td style="width:127px;">Fine Science Tools</td>
			<td style="width:119px;">11041-08</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Octogon Forceps - Serrated/Straight</td>
			<td style="width:127px;">Fine Science Tools</td>
			<td style="width:119px;">11040-08</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Dissector Scissors- Heavy Blade</td>
			<td style="width:127px;">Fine Science Tools</td>
			<td style="width:119px;">14082-09</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Transpore Surgical Tape</td>
			<td style="width:127px;">3M</td>
			<td style="width:119px;">1527-1</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">3-0 Silk Suture</td>
			<td style="width:127px;">Fine Science Tools</td>
			<td style="width:119px;">18020-30</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">TOPO Ventilator</td>
			<td style="width:127px;">Kent Scientific</td>
			<td style="width:119px;">TOPO</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Martin ME 102 Electrosurgical Unit</td>
			<td style="width:127px;">Harvard Apparatus</td>
			<td style="width:119px;">PY2 72-2484</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Syringe Pump</td>
			<td style="width:127px;">Lucca Technologies</td>
			<td style="width:119px;">GenieTouch</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Stereomicroscope with boom stand</td>
			<td style="width:127px;">Nikon</td>
			<td style="width:119px;">SMZ-800N</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Thermocouple Thermometer</td>
			<td style="width:127px;">Cole Parmer</td>
			<td style="width:119px;">EW-91100-40</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">T/Pump Warm Water Recirculator</td>
			<td style="width:127px;">Kent Scientific</td>
			<td style="width:119px;">TP-700</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">ADVantage Pressure-Volume System</td>
			<td style="width:127px;">Transonic</td>
			<td style="width:119px;">ADV500</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Data Acquision and Analysis</td>
			<td style="width:127px;">DSI</td>
			<td style="width:119px;">Ponemah ACQ-16</td>
			<td></td>
		</tr>
	</tbody>
</table>

measuring pressure volume loops in the mouse

理解的原因和心脏疾病的进展呈现给生物医学界一个显著挑战。鼠标的遗传灵活性提供了很大的潜力在分子水平上，以探索心脏功能。鼠标的体积小巧确实提供的问候一些挑战进行详细的心脏表型。小型化和技术进步等已在小鼠心脏评估可能的方法很多。在这些中，压力和体积的数据的同时集合提供心脏功能的详细的图片是不通过任何其他方式。此处压力 - 容积环数据的收集的详细过程进行说明。包括的是的测量和误差的潜在来源的基本原则的讨论。麻醉管理和手术方法非常详细，因为他们是获得高质量的血流动力学测量两个关键的讨论秒。血流动力学协议开发和数据分析的相关方面的原则也被解决。

按照惯例，体积绘制在X轴和压力对Y轴如图1中。从对体积绘制压力产生应类似于一个矩形的压力-容积环，代表在压力（等容的变化，即在垂直的边缘，当两个二尖瓣和主动脉阀关闭）。底部水平代表心室通过二尖瓣和上部水平部填充代表心室通过主动脉瓣排空。在90健康的野生型小鼠左心室压力- 110毫米汞柱，预计8000的最大DP / DT - 12000毫?...

Watch this Scientific Journal Video about Measuring Pressure Volume Loops in the Mouse at JoVE.com

Measuring Pressure Volume Loops in the Mouse

该原稿描述压力 - 体积数据从小鼠收集的详细协议。

测量压力容量环中的鼠标

Understanding the causes and progression of heart disease presents a significant challenge to the biomedical community. The genetic flexibility of the ...

measuring-pressure-volume-loops-in-the-mouse

Universidad de Cartagena UDEC

Research

JoVE Journal

Biology

16.2K 次观看.  University of Minnesota. 该原稿描述压力 - 体积数据从小鼠收集的详细协议。 

视频: Measuring Pressure Volume Loops in the Mouse

Measuring Blood Pressure in Mice using Volume Pressure Recording, a Tail-cuff Method

The CODA 8-Channel High Throughput Non-Invasive Blood Pressure system measures the blood pressure in up to 8 mice or rats simultaneously. The CODA tail-cuff system uses Volume Pressure Recording (VPR) to measure the blood pressure by determining the tail blood volume. A specially designed differential pressure transducer and an occlusion tail-cuff measure the total blood volume in the tail without the need to obtain the individual pulse signal. Special attention is afforded to the length of the occlusion cuff in order to derive the most accurate blood pressure readings. VPR can easily obtain readings on dark-skinned rodents, such as C57BL6 mice and is MRI compatible. The CODA system provides you with measurements of six (6) different blood pressure parameters; systolic and diastolic blood pressure, heart rate, mean blood pressure, tail blood flow, and tail blood volume. Measurements can be made on either awake or anesthetized mice or rats. The CODA system includes a controller, laptop computer, software, cuffs, animal holders, infrared warming pads, and an infrared thermometer. There are seven different holder sizes for mice as small as 8 grams to rats as large as 900 grams.

The CODA 8-Channel High Throughput Non-Invasive Blood Pressure system measures the blood pressure in up to 8 mice or rats simultaneously. This tail-cuff system uses Volume Pressure Recording (VPR) to measure the blood pressure by determining the tail blood volume.

使用卷压力记录，一条尾巴袖套法小鼠的血压测量

The CODA 8-Channel High Throughput Non-Invasive Blood Pressure system measures the blood pressure in up to 8 mice or rats simultaneously. The CODA ...

科研

生物学

Catheterization of Intestinal Loops in Ruminants

The intestine is a complex structure that is involved not only in absorption of nutrients, but also acts as a barrier between the individual and the outside world. As such, the intestine plays a pivotal role in immunosurveillance and protection from enteric pathogens. Investigating intestinal physiology and immunology commonly employs 'intestinal loops' as an experimental model. The majority of these loop models are non-recovery surgical procedures that study short-term (&lt;24 hr) changes in the intestine (1-3). We previously created a recovery intestinal loop model to specifically measure long-term (&lt;6 mo) immunological changes in the intestine of sheep following exposure to vaccines, adjuvants, and viruses (4). This procedure localized treatments to a specific 'loop', allowing us to sample this area of the intestine. A significant drawback of this method is the single window of opportunity to administer treatments (i.e. at the time of surgery). Furthermore, samples of both the intestinal mucosa and luminal contents can only be taken at the termination of the project. Other salient limitations of the above model are that the surgical manipulation and requisite post-operative measures (e.g. administration of antibiotics and analgesics) can directly affect the treatment itself and/or alter immune function, thereby confounding results. Therefore, we modified our intestinal loop model by inserting long-term catheters into the loops. Sheep recover fully from the procedure, and are unaffected by the exteriorized catheters. Notably, the establishment of catheters in loops allows us to introduce multiple treatments over an extended interval, following recovery from surgery and clearance of drugs administered during surgery and the post-operative period.

We describe a novel surgical method for catheterizing 'intestinal loops' within the ileum of sheep. Once animals have recovered from surgery and have cleared antibiotics and analgesics, multiple treatments can be deposited directly in loops via the catheters.

导尿在反刍动物的肠袢

The intestine is a complex structure that is involved not only in absorption of nutrients, but also acts as a barrier between the individual and the ...

In vivo Imaging and Therapeutic Treatments in an Orthotopic Mouse Model of Ovarian Cancer

Human cancer and response to therapy is better represented in orthotopic animal models. This paper describes the development of an orthotopic mouse model of ovarian cancer, treatment of cancer via oral delivery of drugs, and monitoring of tumor cell behavior in response to drug treatment in real time using in vivo imaging system. In this orthotopic model, ovarian tumor cells expressing luciferase are applied topically by injecting them directly into the mouse bursa where each ovary is enclosed. Upon injection of D-luciferin, a substrate of firefly luciferase, luciferase-expressing cells generate bioluminescence signals. This signal is detected by the in vivo imaging system and allows for a non-invasive means of monitoring tumor growth, distribution, and regression in individual animals. Drug administration via oral gavage allows for a maximum dosing volume of 10 mL/kg body weight to be delivered directly to the stomach and closely resembles delivery of drugs in clinical treatments. Therefore, techniques described here, development of an orthotopic mouse model of ovarian cancer, oral delivery of drugs, and in vivo imaging, are useful for better understanding of human ovarian cancer and treatment and will improve targeting this disease.

In vivo Imaging and Therapeutic Treatments in an Orthotopic Mouse Model of Ovarian Cancer

Orthotopic animal models of ovarian cancer replicate better human disease and therefore enhance our understanding of cancer progression and tumor response to therapy. A mouse model receives an intrabursal injection of luciferase-expressing ovarian tumor cells. Treatment is administered via oral gavage. Tumor growth is monitored by in vivo imaging system.

活体成像和治疗卵巢癌的原位小鼠模型

Human cancer and response to therapy is better represented in orthotopic animal models.  This paper describes the development of an orthotopic mouse model ...

Measuring Caenorhabditis elegans Life Span in 96 Well Microtiter Plates

Lifespan is a biological process regulated by several genetic pathways. One strategy to investigate the biology of aging is to study animals that harbor mutations in components of age-regulatory pathways. If these mutations perturb the function of the age-regulatory pathway and therefore alter the lifespan of the entire organism, they provide important mechanistic insights1-3.
Another strategy to investigate the regulation of lifespan is to use small molecules to perturb age-regulatory pathways. To date, a number of molecules are known to extend lifespan in various model organisms and are used as tools to study the biology of aging4-16. The number of molecules identified thus far is small compared to the genetic "toolset" that is available to study the biology of aging.
Caenorhabditis elegans is one of the principle models used to study aging because of its excellent genetics and short lifespan of three weeks. More recently, C.elegans has emerged as a model organism for phenotype based drug screens5,7,16-20 because of its small size and its ability to grow in microtiter plates.
Here we present an assay to measure C.elegans lifespan in 96 well microtiter plates. The assay was developed and successfully used to screen large libraries for molecules that extend C.elegans lifespan7. The reliability of the assay was evaluated in multiple tests: first, by measuring the lifespan of wild type animals grown at different temperatures; second, by measuring the lifespan of mutants with altered lifespans; third, by measuring changes in lifespan in response to different concentrations of the antidepressant Mirtazepine. Mirtazepine has previously been shown to extend lifespan in C.elegans7. The results of these tests show that the assay is able to replicate previous findings from other assays and is quantitative. The microtiter format also makes this lifespan assay compatible with automated liquid handling systems and allows integration into automated platforms.

Measuring Caenorhabditis elegans Life Span in 96 Well Microtiter Plates

In this protocol we present a method to measure Caenorhabditis elegans lifespan in 96 well microtiter plates.

测量线虫寿命在96孔板

Lifespan is a biological process regulated by several genetic pathways. One strategy to investigate the biology of aging is to study animals that harbor ...

Generation of Mice Derived from Induced Pluripotent Stem Cells

The production of induced pluripotent stem cells (iPSCs) from somatic cells provides a means to create valuable tools for basic research and may also produce a source of patient-matched cells for regenerative therapies. iPSCs may be generated using multiple protocols and derived from multiple cell sources. Once generated, iPSCs are tested using a variety of assays including immunostaining for pluripotency markers, generation of three germ layers in embryoid bodies and teratomas, comparisons of gene expression with embryonic stem cells (ESCs) and production of chimeric mice with or without germline contribution2. Importantly, iPSC lines that pass these tests still vary in their capacity to produce different differentiated cell types2. This has made it difficult to establish which iPSC derivation protocols, donor cell sources or selection methods are most useful for different applications.
The most stringent test of whether a stem cell line has sufficient developmental potential to generate all tissues required for survival of an organism (termed full pluripotency) is tetraploid embryo complementation (TEC)3-5. Technically, TEC involves electrofusion of two-cell embryos to generate tetraploid (4n) one-cell embryos that can be cultured in vitro to the blastocyst stage6. Diploid (2n) pluripotent stem cells (e.g. ESCs or iPSCs) are then injected into the blastocoel cavity of the tetraploid blastocyst and transferred to a recipient female for gestation (see Figure 1). The tetraploid component of the complemented embryo contributes almost exclusively to the extraembryonic tissues (placenta, yolk sac), whereas the diploid cells constitute the embryo proper, resulting in a fetus derived entirely from the injected stem cell line.
Recently, we reported the derivation of iPSC lines that reproducibly generate adult mice via TEC1. These iPSC lines give rise to viable pups with efficiencies of 5-13%, which is comparable to ESCs3,4,7 and higher than that reported for most other iPSC lines8-12. These reports show that direct reprogramming can produce fully pluripotent iPSCs that match ESCs in their developmental potential and efficiency of generating pups in TEC tests. At present, it is not clear what distinguishes between fully pluripotent iPSCs and less potent lines13-15. Nor is it clear which reprogramming methods will produce these lines with the highest efficiency. Here we describe one method that produces fully pluripotent iPSCs and "all- iPSC" mice, which may be helpful for investigators wishing to compare the pluripotency of iPSC lines or establish the equivalence of different reprogramming methods.

Generating induced pluripotent stem cell (iPSC) lines produces lines of differing developmental potential even when they pass standard tests for pluripotency. Here we describe a protocol to produce mice derived entirely from iPSCs, which defines the iPSC lines as possessing full pluripotency1.

诱导多能干细胞来源于小鼠的生成

The production of induced pluripotent stem cells (iPSCs) from somatic cells provides a means to create valuable tools for basic research and may also ...

Measuring the Osmotic Water Permeability Coefficient (Pf) of Spherical Cells: Isolated Plant Protoplasts as an Example

Studying AQP regulation mechanisms is crucial for the understanding of water relations at both the cellular and the whole plant levels. Presented here is a simple and very efficient method for the determination of the osmotic water permeability coefficient (Pf) in plant protoplasts, applicable in principle also to other spherical cells such as frog oocytes. The first step of the assay is the isolation of protoplasts from the plant tissue of interest by enzymatic digestion into a chamber with an appropriate isotonic solution. The second step consists of an osmotic challenge assay: protoplasts immobilized on the bottom of the chamber are submitted to a constant perfusion starting with an isotonic solution and followed by a hypotonic solution. The cell swelling is video recorded. In the third step, the images are processed offline to yield volume changes, and the time course of the volume changes is correlated with the time course of the change in osmolarity of the chamber perfusion medium, using a curve fitting procedure written in Matlab (the &#8216;PfFit&#8217;), to yield Pf.

Measuring the Osmotic Water Permeability Coefficient Pf of Spherical Cells: Isolated Plant Protoplasts as an Example

Measuring the osmotic water permeability coefficient (Pf) of cells can help understand the regulatory mechanisms of aquaporins (AQPs). Pf determination in spherical plant cell protoplasts presented here involves protoplasts isolation and numerical analysis of their initial rate of volume change as a result of an osmotic challenge during constant bath perfusion.

测量渗透水渗透系数（P<sub&gt; F</sub&gt;）球形细胞:隔离植物原生质体为例。

Studying AQP regulation mechanisms is crucial for the understanding of water relations at both the cellular and the whole plant levels. Presented here is ...

Measuring Glutathione-induced Feeding Response in Hydra

Hydra is among the most primitive organisms possessing a nervous system and chemosensation for detecting reduced glutathione (GSH) for capturing the prey. The movement of prey organisms causes mechanosensory discharge of the stinging cells called nematocysts from hydra, which are inserted into the prey. The feeding response in hydra, which includes curling of the tentacles to bring the prey towards the mouth, opening of the mouth and consequent engulfing of the prey, is triggered by GSH present in the fluid released from the injured prey. To be able to identify the molecular mechanism of the feeding response in hydra which is unknown to date, it is necessary to establish an assay to measure the feeding response. Here, we describe a simple method for the quantitation of the feeding response in which the distance between the apical end of the tentacle and mouth of hydra is measured and the ratio of such distance before and after the addition of GSH is determined. The ratio, called the relative tentacle spread, was found to give a measure of the feeding response. This assay was validated using a starvation model in which starved hydra show an enhanced feeding response in comparison with daily fed hydra.

Here we describe a simple assay for the quantification of the feeding response in hydra induced by the reduced form of glutathione. This assay relies on measuring the distance between the apical end of the tentacle and mouth of hydra.

测量水润谷胱甘肽诱导饲养响应

Hydra is among the most primitive organisms possessing a nervous system and chemosensation for detecting reduced glutathione (GSH) for capturing the prey. ...

Cardiac Pressure-Volume Loop Analysis Using Conductance Catheters in Mice

Cardiac pressure-volume loop analysis is the &#8220;gold-standard&#8221; in the assessment of load-dependent and load-independent measures of ventricular systolic and diastolic function. Measures of ventricular contractility and compliance are obtained through examination of cardiac response to changes in afterload and preload. These techniques were originally developed nearly three decades ago to measure cardiac function in large mammals and humans. The application of these analyses to small mammals, such as mice, has been accomplished through the optimization of microsurgical techniques and creation of conductance catheters. Conductance catheters allow for estimation of the blood pool by exploiting the relationship between electrical conductance and volume. When properly performed, these techniques allow for testing of cardiac function in genetic mutant mouse models or in drug treatment studies. The accuracy and precision of these studies are dependent on careful attention to the calibration of instruments, systematic conduct of hemodynamic measurements and data analyses. We will review the methods of conducting pressure-volume loop experiments using a conductance catheter in mice.

Cardiac pressure-volume loop analysis is the most comprehensive way to measure cardiac function in the intact heart. We describe a technique to perform and analyze cardiac pressure volume loops, using conductance catheters.

心脏压力容量环分析小鼠使用电导导管

Cardiac pressure-volume loop analysis is the &#8220;gold-standard&#8221; in the assessment of load-dependent and load-independent measures of ventricular ...

Measuring the Stiffness of Ex Vivo Mouse Aortas Using Atomic Force Microscopy

Arterial stiffening is a significant risk factor and biomarker for cardiovascular disease and a hallmark of aging. Atomic force microscopy (AFM) is a versatile analytical tool for characterizing viscoelastic mechanical properties for a variety of materials ranging from hard (plastic, glass, metal, etc.) surfaces to cells on any substrate. It has been widely used to measure the stiffness of cells, but less frequently used to measure the stiffness of aortas. In this paper, we will describe the procedures for using AFM in contact mode to measure the ex vivo elastic modulus of unloaded mouse arteries. We describe our procedure for isolation of mouse aortas, and then provide detailed information for the AFM analysis. This includes step-by-step instructions for alignment of the laser beam, calibration of the spring constant and deflection sensitivity of the AFM probe, and acquisition of force curves. We also provide a detailed protocol for data analysis of the force curves.

Measuring the Stiffness of Ex Vivo Mouse Aortas Using Atomic Force Microscopy

We present detailed protocols for isolation of aortas from mouse and measurement of their elastic modulus using atomic force microscopy.

测量的刚度<em&gt;离体</em&gt;鼠标主动脉用原子力显微镜

Arterial stiffening is a significant risk factor and biomarker for cardiovascular disease and a hallmark of aging. Atomic force microscopy (AFM) is a ...

The Mouse Isolated Perfused Kidney Technique

The mouse isolated perfused kidney (MIPK) is a technique for keeping a mouse kidney under ex vivo conditions perfused and functional for 1 hr. This is a prerequisite for studying the physiology of the isolated organ and for many innovative applications that may be possible in the future, including perfusion decellularization for kidney bioengineering or the administration of anti-rejection or genome-editing drugs in high doses to prime the kidney for transplantation. During the time of the perfusion, the kidney can be manipulated, renal function can be assessed, and various pharmaceuticals administered. After the procedure, the kidney can be transplanted or processed for molecular biology, biochemical analysis, or microscopy.
This paper describes the perfusate and the surgical technique needed for the ex vivo perfusion of mouse kidneys. Details of the perfusion apparatus are given and data are presented showing the viability of the kidney&#39;s preparation: renal blood flow, vascular resistance, and urine data as functional, transmission electron micrographs of different nephron segments as morphological readouts, and western blots of transport proteins of different nephron segments as molecular readout.

The mouse isolated perfused kidney (MIPK) is a technique for keeping a mouse kidney under ex vivo conditions perfused and functional for 1 hr. The buffers and surgical technique are described in detail.

鼠标离体肾技术

The mouse isolated perfused kidney (MIPK) is a technique for keeping a mouse kidney under ex vivo conditions perfused and functional for 1 hr. This is a ...

测量压力容量环中的鼠标

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