Single-detector simultaneous optical mapping of V(m) and [Ca(2+)](i) in cardiac monolayers.
Single-detector simultaneous optical mapping of V(m) and [Ca(2+)](i) in cardiac monolayers.
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DOI：
10.1007/s10439-011-0478-z
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发表时间：
2012-05
影响因子：
3.8
通讯作者：
Bursac, Nenad
中科院分区：
文献类型：
--
作者：
Scull, James A.;McSpadden, Luke C.;Himel, Herman D.;Badie, Nima;Bursac, Nenad
同时绘制跨膜电压（Vm）和细胞内Ca2+浓度（Cai）已被用于心脏组织中正常和异常脉冲传播的研究。现有的双映射系统通常使用一个激发和两个发射带宽，需要两个具有精确像素配准的光电探测器。在这项研究中，我们描述了一种新颖的单检测器制图系统，该系统利用两个激发和一个发射带宽来同时记录新生大鼠心肌细胞单层中的动作电位和钙瞬态。用Ca2+敏感染料X-Rhod-1和电压敏感染料Di-4-ANEPPS染色的细胞被可编程的多色LED矩阵照亮。采用定制的双带通激发滤波器，以488.3Hz的速率反相闪烁蓝色和绿色LED脉冲，透射蓝光（482±6nm）和绿光（577±31nm）。使用长通发射滤波器（>605 nm）和504通道光电二极管阵列记录来自心肌细胞的组合信号。绿色激励产生Cai瞬态，没有明显的Vm串扰。用蓝色激励得到的Vm信号中存在的串扰通过减去适当缩放的Cai暂态来消除。应用该方法研究了各向异性心脏单分子膜动作电位与Cai瞬态之间的延迟。
Simultaneous mapping of transmembrane voltage (Vm) and intracellular Ca2+ concentration (Cai) has been used for studies of normal and abnormal impulse propagation in cardiac tissues. Existing dual mapping systems typically utilize one excitation and two emission bandwidths, requiring two photodetectors with precise pixel registration. In this study we describe a novel, single-detector mapping system that utilizes two excitation and one emission bandwidth for the simultaneous recording of action potentials and calcium transients in monolayers of neonatal rat cardiomyocytes. Cells stained with the Ca2+-sensitive dye X-Rhod-1 and the voltage-sensitive dye Di-4-ANEPPS were illuminated by a programmable, multicolor LED matrix. Blue and green LED pulses were flashed in opposite phase at a rate of 488.3Hz using a custom-built dual bandpass excitation filter that transmitted blue (482±6nm) and green (577±31nm) light. A long-pass emission filter (>605 nm) and a 504-channel photodiode array were used to record combined signals from cardiomyocytes. Green excitation yielded Cai transients without significant crosstalk from Vm. Crosstalk present in Vm signals obtained with blue excitation was removed by subtracting an appropriately scaled version of the Cai transient. This method was applied to study delay between onsets of action potentials and Cai transients in anisotropic cardiac monolayers.
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