Imaging cellular signals in the heart in vivo:: Cardiac expression of the high-signal Ca2+ indicator GCaMP2
Imaging cellular signals in the heart in vivo:: Cardiac expression of the high-signal Ca2+ indicator GCaMP2
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DOI：
10.1073/pnas.0509378103
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发表时间：
2006-03-21
影响因子：
11.1
通讯作者：
Kotlikoff, MI
中科院分区：
文献类型：
--
作者：
Tallini, YN;Ohkura, M;Kotlikoff, MI
遗传编码的传感器蛋白提供了独特的机会，以促进了解复杂的细胞相互作用的生理相关的背景下，然而，以前描述的传感器已被证明是有限的使用报告细胞信号在哺乳动物体内。在这里，我们描述了一种改进的Ca2+传感器，GCaMP2，其在小鼠心脏中的诱导表达，以及其用于检查体内心脏细胞中的信号传导。GCaMP2的高亮度和稳定性使得能够测量跳动的小鼠心脏的所有区域中的肌细胞Ca2+瞬变，以及在离体灌注心脏中的长时间起搏和标测研究。在心肌细胞GCaMP2小鼠中，转基因表达在时间上被有效地调节，从而允许在转基因诱导后4周记录体内信号。在表达GCaMP2的胚胎中的Ca 2+波的高分辨率成像揭示了预分离心脏中电传导的关键方面。在胚胎日（e. d.）10.5，心房和心室传导发生迅速，符合早期形成专门的传导通路。然而，在e.d.，通过房室管的传导明显减慢。10.5在房室结发育之前，由于心室快速激动发生在远端房室管组织激动之后，因此，房室结在心脏中的作用形成了有效房室延迟的基础。与e.d.时房室管内肌肉层的消除一致。13.5，房室传导在此阶段被取消。这些研究表明，GCaMP2将在哺乳动物体内解剖许多复杂的细胞相互作用中具有广泛的用途。
Genetically encoded sensor proteins provide unique opportunities to advance the understanding of complex cellular interactions in physiologically relevant contexts; however, previously described sensors have proved to be of limited use to report cell signaling in vivo in mammals. Here, we describe an improved Ca2+ sensor, GCaMP2, its inducible expression in the mouse heart, and its use to examine signaling in heart cells in vivo. The high brightness and stability of GCaMP2 enable the measurement of myocyte Ca2+ transients in all regions of the beating mouse heart and prolonged pacing and mapping studies in isolated, perfused hearts. Transgene expression is efficiently temporally regulated in cardiomyocyte GCaMP2 mice, allowing recording of in vivo signals 4 weeks after transgene induction. High-resolution imaging of Ca2+ waves in GCaMP2-expressing embryos revealed key aspects of electrical conduction in the preseptated heart. At embryonic day (e.d.) 10.5, atrial and ventricular conduction occur rapidly, consistent with the early formation of specialized conduction pathways. However, conduction is markedly slowed through the atrioventricular canal in the e.d. 10.5 heart, forming the basis for an effective atrioventricular delay before development of the AV node, as rapid ventricular activation occurs after activation of the distal AV canal tissue. Consistent with the elimination of the inner AV canal muscle layer at e.d. 13.5, atrioventricular conduction through the canal was abolished at this stage. These studies demonstrate that GCaMP2 will have broad utility in the dissection of numerous complex cellular interactions in mammals, in vivo.