Klauke Norbert, Smith Godfrey L, Cooper Jon
Department of Electronics, University of Glasgow, Glasgow G12 8LT, United Kingdom.
Biophys J. 2006 Oct 1;91(7):2543-51. doi: 10.1529/biophysj.106.085183. Epub 2006 Jul 14.
Open microfluidic channels were used to separate the extracellular space around a cardiomyocyte into three compartments: the cell ends and a central partition (insulating gap). The microchannels were filled with buffer solution and overlaid with paraffin oil, thus forming the cavities for the cell ends. The central part of the cardiomyocyte rested on the partition between two adjacent microchannels and was entirely surrounded by the paraffin oil. This arrangement increased the extracellular electrical resistance to > 20 MOmega and facilitated the recording of the time course of the change in extracellular voltage and current during subthreshold and suprathreshold stimuli. The waveform of the extracellular current and voltage in response to an extracellular depolarizing stimulus comprised an initial monophasic signal followed by a biphasic signal with a delay of 2-15 ms. The latter was associated with a transient contraction and therefore caused by an action potential. The biphasic signal became monophasic after the depolarization of one cell end by raised extracellular [K+]. This form of differential recording revealed the repolarization phase of the action potential. At rest, the sarcomere length within the gap was 12% +/- 4.8% longer than outside the gap, but intracellular Ca2+ transients occurred to the same extent as that observed in the outer pools. This data demonstrate the feasibility of the use of a microfluidic bath design to limit the extracellular resistance between two ends of an isolated cardiomyocyte.
细胞两端和一个中央分隔区(绝缘间隙)。微通道中充满缓冲溶液并覆盖石蜡油,从而形成细胞两端的腔室。心肌细胞的中央部分位于两个相邻微通道之间的分隔区上,并完全被石蜡油包围。这种布置将细胞外电阻增加到> 20 MΩ,并有助于记录阈下和阈上刺激期间细胞外电压和电流变化的时间过程。响应细胞外去极化刺激的细胞外电流和电压波形包括一个初始单相信号,随后是一个延迟2 - 15毫秒的双相信号。后者与短暂收缩相关,因此由动作电位引起。在细胞外[K +]升高使一个细胞端去极化后,双相信号变为单相。这种差分记录形式揭示了动作电位的复极化阶段。静息时,间隙内的肌节长度比间隙外长12%±4.8%,但细胞内Ca2 +瞬变的程度与在外部池观察到的相同。这些数据证明了使用微流体浴设计来限制分离心肌细胞两端之间细胞外电阻的可行性。