Intracellular calcium dynamics in cortical microglia responding to focal laser injury in the PC::G5-tdT reporter mouse
Intracellular calcium dynamics in cortical microglia responding to focal laser injury in the PC::G5-tdT reporter mouse
复制标题
PC::G5-tdT 报告小鼠皮质小胶质细胞对局灶性激光损伤的细胞内钙动力学反应
DOI：
10.3389/fnmo1.2015.00012
复制
发表时间：
2015-05-08
影响因子：
4.8
通讯作者：
Capecchi, Mario R.
中科院分区：
文献类型：
--
作者：
Pozner, Amir;Xu, Ben;Capecchi, Mario R.
小胶质细胞是脑实质的常驻免疫细胞，对组织损伤高度反应。在细胞损伤后，小胶质细胞过程将其运动性从随机搜索细胞外空间重新定向到特异性地到达受损组织。虽然这种防御机制的细胞形态方面的特点，这些反应的细胞内事件仍然在很大程度上是未知的。具体而言，由于技术上的困难，细胞内Ca2+动力学的作用尚未在急性激活的小胶质细胞中进行系统研究。在这里，我们使用活的双光子成像的小鼠皮质普遍表达的遗传编码的Ca2+指标GCaMP5G和荧光标记tdTomato在中枢神经系统小胶质细胞。我们发现小胶质细胞胞体和突起中自发的Ca 2+瞬变通常较低（所有小胶质细胞中只有4%在20分钟内表现出瞬变），但当动物在成像前12小时接受LPS治疗时，基线活性增加了约8倍。局部激光损伤时，胞体和突起中观察到额外的Ca2+活性激增。值得注意的是，在LPS处理的动物中偶尔检测到多个小胶质细胞中的连贯和同时的Ca2+升高。我们表明，Ca2+瞬变主要是通过嘌呤受体介导的。这项工作证明了有用的遗传编码的Ca2+指标的小胶质细胞生理学的调查。
Microglia, the resident immune cells of the brain parenchyma, are highly responsive to tissue injury. Following cell damage, microglial processes redirect their motility from randomly scouting the extracellular space to specifically reaching toward the compromised tissue. While the cell morphology aspects of this defense mechanism have been characterized, the intracellular events underlying these responses remain largely unknown. Specifically, the role of intracellular Ca2+ dynamics has not been systematically investigated in acutely activated microglia due to technical difficulty. Here we used live two-photon imaging of the mouse cortex ubiquitously expressing the genetically encoded Ca2+ indicator GCaMP5G and fluorescent marker tdTomato in central nervous system microglia. We found that spontaneous Ca2+ transients in microglial somas and processes were generally low (only 4% of all microglia showing transients within 20 min), but baseline activity increased about 8-fold when the animals were treated with LPS 12 h before imaging. When challenged with focal laser injury, an additional surge in Ca2+ activity was observed in the somas and protruding processes. Notably, coherent and simultaneous Ca2+ rises in multiple microglial cells were occasionally detected in LPS-treated animals. We show that Ca2+ transients were pre-dominantly mediated via purinergic receptors. This work demonstrates the usefulness of genetically encoded Ca2+ indicators for investigation of microglial physiology.