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First published online on February 24, 2005.
 Copyright © 2005 by The Physiological Society
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Received February 11, 2005
Revised February 23, 2005
Accepted after revision February 23, 2005

Visualization of Localized Store-Operated Calcium Entry in Mouse Astrocytes. Close Proximity to the Endoplasmic Reticulum

Vera A Golovina1*

1 University of Maryland School of Medicine

* To whom correspondence should be addressed. E-mail: vgolovin{at}umaryland.edu.

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 SOCs function is their spatial relationship to ER Ca2+ stores. Here, using high resolution imaging with the membrane-associated Ca2+ indicator, FFP-18, I show 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 TRPC1 protein, a candidate for 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.


Key words: Astrocyte • Ca2+ influx • Ca2+ stores




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