Name: measuring fast calcium fluxes in cardiomyocytes
Uploaded: 2011-11-29T13:00:00
Description: Watch this Scientific Journal Video about Measuring Fast Calcium Fluxes in Cardiomyocytes at JoVE.com

这项工作是由美国国立卫生赠款2R01 GM53132和P50 GM071558研究院的支持。

1。载入细胞与钙指标<ol><li>中板35mm玻璃底的MatTek菜细胞（或任何其他的玻璃底部观看商会）。 </li><li>如果使用前用10μg/ml的层粘连蛋白大衣原发性心肌商会1天。板心肌细胞（成年犬，新生鼠，或其他生物隔离）35 KB缓冲区（K -逆转台氏缓冲mmol / L的HEPES，140 mmol / L的氯化钾，8 mmol / L的KHCO 3，2毫摩尔/ L的MgCl 2的 KH 2 PO 4，pH值7.5 0.4毫摩尔/ L）和逐步改变M199培养基?...

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我们已经开发出一种方法来查看和隔离生活心肌的快速钙信号。虽然这些测量的实验条件下，曾经被应用到其他细胞系统10以及心肌细胞 11，使用直方图和斌分析可以从很多的Ca 2 +事件被收购的数据。更快的事件，可以很容易地隔离较慢的适应，了解他们的底层机制的模型。培养心肌细胞，许多小学的细胞株，可就难了。虽然人们普遍认为，测量应尽快采取收购后...

<table border="1"><tbody><tr><td> 试剂名称 </td><td> 公司 </td><td> 目录编号 </td><td> 评论 </td></tr><tr><td> DMEM培养液</td><td> Invitrogen公司</td><td> ABCD1234 </td><td></td></tr></tbody></table>

measuring fast calcium fluxes in cardiomyocytes

心肌细胞有多个时间长短不一的Ca 2 +通量，共同努力，优化功能1,2。钙的变化外代理的活动主要是受这是刺激剂， 如乙酰胆碱3,4质膜磷脂酶Cβ-Gαq本地化的途径。最近，我们发现，质膜蛋白质称为小窝5,6域可以坑害激活GαQ 7。这包封的稳定Gαq激活状态，并在长期的Ca 2 +信号在心肌细胞和其它类型的细胞 8作用。我们发现这个令人吃惊的结果，通过规模快速测量动态生活心肌钙反应。简言之，装有一个荧光的Ca 2 +指示剂细胞。在我们的研究中，我们使用的Ca 2 +绿色（Invitrogen公司，在C.）展品钙离子结合后的荧光信号强度的增加。荧光强度，然后记录使用激光扫描共聚焦显微镜扫描模式。此方法允许沿选定的行的像素荧光强度的时间当然的快速采集，生产数百微秒的时间尺度上的时间痕迹。这些痕迹非常快转移到Excel，然后将进行分析Sigmaplot相比，电子噪音，免费染料和其他控制获得的痕迹。解剖的Ca 2 +不同的通量率的反应，我们进行了直方图分析，随着时间的推移，分级像素强度。分级让我们超过500扫描的痕迹和可视化编译的结果在一个单一的情节空间和时间。因此，缓慢的Ca 2 +波难以辨别时，扫描覆盖由于不同的峰的位置和噪声，可不难看出，在分级直方图。在时间尺度的测量速度非常快通量表明在很短的时间，而较长的Ca 2箱的强度分布窄 +波显示分级数据与广泛分布在较长的时间箱。这些不同的时间分布，让我们来剖析在细胞内的Ca 2 +通量的时机，并确定其影响各种细胞活动。

Watch this Scientific Journal Video about Measuring Fast Calcium Fluxes in Cardiomyocytes at JoVE.com

Measuring Fast Calcium Fluxes in Cardiomyocytes

我们提出了一个方法来隔离在生活细胞利用激光扫描共聚焦显微镜慢通量快速（微秒）钙事件。方法措施，由几百个像素单元格中的记录行扫描荧光强度的钙指标的波动。直方图分析，让我们来隔离不同的钙通量的时间尺度。

测量心肌细胞的快速钙助焊剂

Cardiomyocytes have multiple Ca2+ fluxes of varying duration that work together to optimize function 1,2. Changes in Ca2+ activity in response to ...

Research

JoVE Journal

Biology

Measuring Near Plasma Membrane and Global Intracellular Calcium Dynamics in Astrocytes

近质膜和全球星形胶质细胞内钙动力学测量

The brain contains glial cells.  Astrocytes, a type of glial cell, have long been known to provide a passive supportive role to neurons. However, ...

科研

生物学

Efficient Derivation of Human Cardiac Precursors and Cardiomyocytes from Pluripotent Human Embryonic Stem Cells with Small Molecule Induction

高效的推导人类心脏的前体和从多能干的小分子诱导人类胚胎干细胞的心肌细胞

To date, the lack of a suitable human cardiac cell source has been the major setback in regenerating the human myocardium, either by cell-based ...

Isolation and Kv Channel Recordings in Murine Atrial and Ventricular Cardiomyocytes

在小鼠的心房和心室肌细胞的分离及钾离子通道的录音

KCNE genes encode for a small family of Kv channel ancillary subunits that form heteromeric complexes with Kv channel alpha subunits to modify their ...

Isolation and Culture of Neonatal Mouse Cardiomyocytes

分离新生小鼠心肌文化和

Cultured neonatal cardiomyocytes have long been used to study  myofibrillogenesis and myofibrillar functions. Cultured cardiomyocytes allow  for easy ...

Measuring Glutathione-induced Feeding Response in 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. ...

Voltage and Calcium Dual Channel Optical Mapping of Cultured HL-1 Atrial Myocyte Monolayer

培养HL-1心房肌细胞单层的电压和钙双通道光映射

Optical mapping has proven to be a valuable technique to detect cardiac electrical activity on both intact ex vivo hearts and in cultured myocyte ...

Measuring RAN Peptide Toxicity in C. elegans

Measuring RAN Peptide Toxicity in C. elegans

测量在C.elegan的RAN肽毒性

C. elegans is commonly used to model age-related neurodegenerative diseases caused by repeat expansion mutations, such as Amyotrophic Lateral Sclerosis ...

Forward Genetic Screen Using Transgenic Calcium Reporter Aequorin to Identify Novel Targets in Calcium Signaling

使用转基因钙处理器 Aequorin 前向基因屏幕，以识别钙信号中的新目标

Forward genetic screens have been important tools in the unbiased identification of genetic components involved in several biological pathways. The basis ...

Light Sheet Microscopy of Fast Cardiac Dynamics in Zebrafish Embryos

斑马鱼胚胎快速心脏动力学的光片显微镜

Embryonic cardiac research has greatly benefited from advances in fast in vivo light sheet fluorescence microscopy (LSFM). Combined with the rapid ...

Measuring Mitochondrial Substrate Flux in Recombinant Perfringolysin O-Permeabilized Cells

测量重组透气溶性细胞中线粒体底物通量 O-透化细胞

Mitochondrial substrate flux is a distinguishing characteristic of each cell type, and changes in its components such as transporters, channels, or ...