Visualization of localized store-operated calcium entry in mouse astrocytes. Close proximity to the endoplasmic reticulum
Visualization of localized store-operated calcium entry in mouse astrocytes. Close proximity to the endoplasmic reticulum
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DOI：
10.1113/jphysiol.2005.085035
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发表时间：
2005-05-01
影响因子：
5.5
通讯作者：
Golovina, VA
中科院分区：
文献类型：
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作者：
Golovina, VA
内质网（ER） Ca2+储存的卸载通过大多数细胞（包括星形胶质细胞）的质膜（PM）中的“储存操作”Ca2+通道（soc）激活细胞外Ca2+的内流。关于SOC功能的一个关键尚未解决的问题是它们与ER Ca2+储存的空间关系。在这里，使用膜相关Ca2+指示剂FFP- 18的高分辨率成像显示，在原代培养的小鼠皮质星形胶质细胞中，储存操作的Ca2+进入（SOCE）发生在质膜-内质网连接处。在缺乏细胞外Ca2+的情况下，使用环吡唑酸（ER Ca2+ - atp酶抑制剂）和咖啡因消耗ER Ca2+储存，会在质膜和邻近ER之间的有限空间内短暂增加质膜下Ca2+浓度（[Ca2+](SPM)）。细胞外Ca2+的恢复引起局部Ca2+内流，首先在相同的受限区域增加[Ca2+](SPM)，然后在无er区域延迟增加。TRPC1基因的反义敲除被认为编码内源性SOCs，显著降低Fura-2测量的SOCE。抗trpc1抗体的高分辨率免疫细胞化学显示，这些trpc编码的soc局限于与潜在的“连接”ER相邻的PM微域。因此，通过trpc编码的soc的Ca2+进入不仅在功能上，而且在结构上与ER Ca2+储存密切相关。
Unloading of endoplasmic reticulum (ER) Ca2+ stores activates influx of extracellular Ca2+ through 'store-operated' Ca2+ channels (SOCs) in the plasma membrane (PM) of most cells, including astrocytes. A key unresolved issue concerning SOC function is their spatial relationship to ER Ca2+ stores. Here, using high resolution imaging with the membrane-associated Ca2+ indicator, FFP- 18, it is shown that store-operated Ca2+ entry (SOCE) in primary cultured mouse cortical astrocytes occurs at plasma membrane-ER junctions. In the absence of extracellular Ca2+, depletion of ER Ca2+ stores using cyclopiazonic acid, an ER Ca2+ -ATPase inhibitor, and caffeine transiently increases the sub-plasma-membrane Ca2+ concentration ([Ca2+](SPM)) within a restricted space between the plasma membrane and adjacent ER. Restoration of extracellular Ca2+ causes localized Ca2+ influx that first increases [Ca2+](SPM) in the same restricted regions and then, with a delay, in ER-free regions. Antisense knockdown of the TRPC1 gene, proposed to encode endogenous SOCs, markedly reduces SOCE measured with Fura-2. High resolution immunocytochemistry with anti-TRPC1 antibody reveals that these TRPC-encoded SOCs are confined to the PM microdomains adjacent to the underlying 'junctional' ER. Thus, Ca2+ entry through TRPC-encoded SOCs is closely linked, not only functionally, but also structurally, to the ER Ca2+ stores.