Imaging calcium microdomains within entire astrocyte territories and endfeet with GCaMPs expressed using adeno-associated viruses.
Imaging calcium microdomains within entire astrocyte territories and endfeet with GCaMPs expressed using adeno-associated viruses.
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DOI：
10.1085/jgp.201210949
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发表时间：
2013-05
期刊：
影响因子：
--
通讯作者：
Khakh BS
中科院分区：
文献类型：
--
作者：
Shigetomi E;Bushong EA;Haustein MD;Tong X;Jackson-Weaver O;Kracun S;Xu J;Sofroniew MV;Ellisman MH;Khakh BS
细胞内Ca2+瞬态被认为是星形胶质细胞与神经元和血管相互作用的主要信号。利用现有的常用方法，Ca2+仅在星形细胞体细胞和粗分支内进行了研究，而对最接近神经元突触和血管的远端细分枝和终足进行了研究。在这里，使用细胞质和膜系结形式的遗传编码Ca2+指标（GECIs；细胞- gcamp3和Lck-GCaMP3），我们报告了克服这些局限性的表征方法。我们在体内显微注射腺相关病毒，在星形胶质细胞中表达GECIs，并研究了感染后两周成年小鼠（年龄为80岁）体外急性海马切片中的Ca2+信号。我们的数据揭示了在急性海马切片内整个星形胶质细胞区域内意想不到的大量，频繁，同等比例和高度定位的Ca2+微域的波光全景，与使用电子显微镜描述的突触周围小枝的分布一致。来自尾足的信号被特别清晰地揭示出来。我们详细描述的工具和实验方法允许系统地研究整个星形胶质细胞内的Ca2+信号，包括精细的突触周围小枝和血管相关的终足，允许严格评估星形胶质细胞如何促进脑功能。
Intracellular Ca2+ transients are considered a primary signal by which astrocytes interact with neurons and blood vessels. With existing commonly used methods, Ca2+ has been studied only within astrocyte somata and thick branches, leaving the distal fine branchlets and endfeet that are most proximate to neuronal synapses and blood vessels largely unexplored. Here, using cytosolic and membrane-tethered forms of genetically encoded Ca2+ indicators (GECIs; cyto-GCaMP3 and Lck-GCaMP3), we report well-characterized approaches that overcome these limitations. We used in vivo microinjections of adeno-associated viruses to express GECIs in astrocytes and studied Ca2+ signals in acute hippocampal slices in vitro from adult mice (aged ∼P80) two weeks after infection. Our data reveal a sparkling panorama of unexpectedly numerous, frequent, equivalently scaled, and highly localized Ca2+ microdomains within entire astrocyte territories in situ within acute hippocampal slices, consistent with the distribution of perisynaptic branchlets described using electron microscopy. Signals from endfeet were revealed with particular clarity. The tools and experimental approaches we describe in detail allow for the systematic study of Ca2+ signals within entire astrocytes, including within fine perisynaptic branchlets and vessel-associated endfeet, permitting rigorous evaluation of how astrocytes contribute to brain function.
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DOI：
10.1523/jneurosci.0127-11.2011
发表时间：
2011-06-22
期刊：
The Journal of neuroscience : the official journal of the Society for Neuroscience
影响因子：
--
作者：
Reeves AM;Shigetomi E;Khakh BS
通讯作者：
Khakh BS
通过神经胶质调节突触连通性。
影响因子：
64.8
作者：
Eroglu, Cagla;Barres, Ben A.
通讯作者：
Barres, Ben A.
影响因子：
16.2
作者：
Agulhon, Cendra;Petravicz, Jeremy;McMullen, Allison B.;Sweger, Elizabeth J.;Minton, Suzanne K.;Taves, Sarah R.;Casper, Kristen B.;Fiacco, Todd A.;McCarthy, Ken D.
通讯作者：
McCarthy, Ken D.
影响因子：
18.2
作者：
Halassa MM;Haydon PG
通讯作者：
Haydon PG
影响因子：
5.3
作者：
Bushong, EA;Martone, ME;Ellisman, MH
通讯作者：
Ellisman, MH