Intracellular sodium homeostasis in rat hippocampal astrocytes
1. We determined the intracellular Na+ concentration ([Na+]i) and mechanisms of its regulation in cultured rat hippocampal astrocytes using fluorescence ratio imaging of the Na+ indicator SBFI-AM (acetoxymethylester of sodium-binding benzofuran isophthalate, 10 microM). Dye signal calibration within...
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Published in | The Journal of physiology Vol. 491; no. Pt 2; pp. 291 - 305 |
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Main Authors | , |
Format | Journal Article |
Language | English |
Published |
England
The Physiological Society
01.03.1996
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Subjects | |
Online Access | Get full text |
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Summary: | 1. We determined the intracellular Na+ concentration ([Na+]i) and mechanisms of its regulation in cultured rat hippocampal
astrocytes using fluorescence ratio imaging of the Na+ indicator SBFI-AM (acetoxymethylester of sodium-binding benzofuran
isophthalate, 10 microM). Dye signal calibration within the astrocytes showed that the ratiometric dye signal changed monotonically
with changes in [Na+]i from 0 to 140 nM. The K+ sensitivity of the dye was negligible; intracellular pH changes, however,
slightly affected the 'Na+' signal. 2. Baseline [Na+]i was 14.6 +/- 4.9 mM (mean +/- S.D.) in CO2/HCO3(-)-containing saline
with 3 mM K+. Removal of extracellular Na+ decreased [Na+]i in two phases: a rapid phase of [Na+]i reduction (0.58 +/- 0.32
mM min-1) followed by a slower phase (0.15 +/- 0.09 mM min-1). 3. Changing from CO2/HCO3(-)-free to CO2/HCO3(-)-buffered saline
resulted in a transient increase in [Na+]i of approximately 5 mM, suggesting activation of inward Na(+)-HCO3- cotransport
by CO2/HCO3-. During furosemide (frusemide, 1 mM) or bumetanide (50 microM) application, a slow decrease in [Na+]i of approximately
2 mM was observed, indicating a steady inward transport of Na+ via Na(+)-K(+)-2Cl- cotransport under control conditions. Tetrodotoxin
(100 microM) did not influence [Na+]i in the majority of cells (85%), suggesting that influx of Na+ through voltage-gated
Na+ channels contributed to baseline [Na+]i in only a small subpopulation of hippocampal astrocytes. 4. Blocking Na+, K(+)-ATPase
activity with cardiac glycosides (ouabain or strophanthidin, 1 mM) or removal of extracellular K+ led to an increase in [Na+]i
of about 2 and 4 mM min-1, respectively. This indicated that Na+, K(+)-ATPase activity was critical in maintaining low [Na+]i
in the face of a steep electrochemical gradient, which would favour a much higher [Na+]i. 5. Elevation of extracellular K+
concentration ([K+]o) by as little as 1 mM (from 3 to 4 mM) resulted in a rapid and reversible decrease in [Na+]i. Both the
slope and the amplitude of the [K+]o-induced reductions in [Na+]i were sensitive to bumetanide. A reduction of [K+]o by 1
mM increased [Na+]i by 3.0 +/- 2.3 mM. In contrast, changing extracellular Na+ concentration by 20 mM resulted in changes
in [Na+]i of less than 3 mM. 6. These results implied that in hippocampal astrocytes low baseline [Na+]i is determined by
the action of Na(+)-HCO3- cotransport, Na(+)-K(+)-2Cl- cotransport and Na+, K(+)-ATPase, and that both Na+, K(+)-ATPase and
inward Na(+)-K(+)-2Cl cotransport are activated by small, physiologically relevant increases in [K+]o. These mechanisms are
well suited to help buffer increases in [K+]o associated with neural activity. |
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Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 |
ISSN: | 0022-3751 1469-7793 |
DOI: | 10.1113/jphysiol.1996.sp021216 |