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Specialized distributions of mitochondria and endoplasmic reticulum proteins define Ca 2+ wave amplification sites in cultured astrocytes
Author(s) -
Simpson Peter B.,
Mehotra Surabhi,
Langley David,
Sheppard Carol A.
Publication year - 1998
Publication title -
journal of neuroscience research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.72
H-Index - 160
eISSN - 1097-4547
pISSN - 0360-4012
DOI - 10.1002/(sici)1097-4547(19980615)52:6<672::aid-jnr6>3.0.co;2-5
Subject(s) - calreticulin , ryanodine receptor , endoplasmic reticulum , mitochondrion , microbiology and biotechnology , astrocyte , biology , cytosol , immunocytochemistry , receptor , biochemistry , endocrinology , central nervous system , enzyme
This study was undertaken to examine the expression and role of the endoplasmic reticulum (ER) proteins calreticulin and ryanodine receptors, and mitochondria, in cultured astrocytes. Using several lines of investigation, we have identified a key role for mitochondria in astrocyte Ca 2+ signalling: (1) a significant correlation was found between sites of regenerative Ca 2+ wave amplification (possessing high amplitude ER Ca 2+ release) and the location of mitochondria in the cell; (2) norepinephrine (2 μM) caused a rapid‐onset increase in rhod 2 fluorescence in 34% of astrocyte mitochondria, indicating that cytosolic Ca 2+ responses result in mitochondrial Ca 2+ elevation; and (3) pretreatment with the protonophore carbonyl cyanide p ‐(trifluoromethoxy)phenylhydrazone to inhibit mitochondrial activity markedly reduced the amplitude of subsequent norepinephrine‐evoked cytosolic Ca 2+ responses. We then investigated the roles of several ER proteins in Ca 2+ signalling by immunocytochemistry. Ryanodine receptors and calreticulin were found to be expressed in heterogeneous patterns in astrocytes. The expression pattern of calreticulin corresponded closely with the distribution of mitochondria, whereas the expression of ryanodine receptors was not similar to that of either of these cellular factors. We measured Ca 2+ wave kinetics in a single astrocyte, then assessed protein distribution by immunocytochemistry in the same cell. Cross‐correlation between norepinephrine‐evoked Ca 2+ wave amplitude and calreticulin distribution indicated a close spatial relationship between this Ca 2+ ‐binding protein and sites of regenerative wave amplification. These results demonstrate that amplification sites for Ca 2+ waves in astrocytes are identifiable by accumulations of calreticulin (and type 2 InsP 3 Rs), and by the presence of mitochondria, which may regulate the ER Ca 2+ release process. J. Neurosci. Res. 52:672–683, 1998. Published 1998 Wiley‐Liss, Inc. This article is a US Government work and, as such, is in the public domain in the United States of America.