Glutamate-induced swelling of single astroglial cells in primary culture.
Glutamate induced an increase in cell volume within one minute and evoked cytosolic Ca2+ transients in type 1 astroglial cells in primary culture obtained from the cerebral cortex of newborn rat. Even the metabotropic glutamate receptor agonists (1S,3R)-1-aminocyclopentane- 1,3-dicarboxylic acid (1S-3R-ACPD) and L(+)-2-amino-4 phosphonobutyric acid (L-AP4) induced a cell swelling with ACPD inducing a parallel Ca2+ transient while L-AP4 did not. A new method was used where rapid changes in relative cell volume could be followed at the single cell level. Relative volume changes in cultured single astroglial cells were examined by microspectrofluorimetry after loading the cells with the highly fluorescent intracellular probe fura-2/AM. At its isosbestic point, 358 nm, fura-2 is ion-insensitive and the fluorescent signals emitted are related only to the intracellular dye concentration. By varying the excitation wavelengths, changes in intracellular Ca2+ transients could be recorded simultaneously with the relative volume variations of the individual cells. Thus, as rapid changes in cell volume were followed, the results from this method could be of physiological significance. Glutamate-induced cell swelling was blocked by BaCl2 and by tetraethylammonium, suggesting that K+ channels are operative in glutamate-induced cell swelling. Furthermore, the glutamate-induced swelling was blocked by the Na+; K+, and 2Cl- co-transport inhibitor furosemide. The glutamate-induced swelling was partially blocked by pertussis toxin and partially blocked also by the glutamate carrier-blocker dihydroaspartate. When the ionotropic glutamate receptor alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid was blocked with the antagonist 2,3-dihydroxy-6-nitro-7- sulfamoyl-benzo(F)quinoxaline, glutamate still induced a swelling, suggesting that this receptor was not directly involved in the glutamate-induced volume increase. Even in situations of blocked or partially blocked swelling, intracellular Ca2+ transients could be obtained. Furthermore, the glutamate-induced swelling was evoked even in low extracellular Ca2+ concentrations. Our data suggest that glutamate-induced rapid swelling is a complex process at the molecular level. One hypothetical mechanism might be that glutamate interacts with metabotropic glutamate receptors and induces a release of Ca2+ from internal stores. Furthermore glutamate interacts with K+ channels, and probably at least one co-transporter and the sodium-dependent high-affinity uptake glutamate carrier, resulting in cell swelling.
Hansson
E
E
Institute of Neurobiology, University of Göteborg, Sweden.
Johansson
B B
BB
Westergren
I
I
Rönnbäck
L
L
D000613:Aminobutyrates / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn / Q000502:physiology; D001253:Astrocytes / Q000187:drug effects / Q000648:ultrastructure*; D002118:Calcium / Q000502:physiology; D048429:Cell Size / Q000187:drug effects; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000187:drug effects; D003515:Cycloleucine / Q000031:analogs & derivatives / Q000494:pharmacology; D005110:Extracellular Space / Q000187:drug effects / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism; D018698:Glutamic Acid / Q000494:pharmacology*; D007473:Ion Channels / Q000187:drug effects / Q000378:metabolism; D009498:Neurotoxins / Q000494:pharmacology; D037342:Pertussis Toxin; D015221:Potassium Channels / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D017470:Receptors, Glutamate / Q000187:drug effects / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D000613:Aminobutyrates; D007473:Ion Channels; D009498:Neurotoxins; D015221:Potassium Channels; D017470:Receptors, Glutamate; D010566:Virulence Factors, Bordetella; D003515:Cycloleucine; C036361:1-amino-1,3-dicarboxycyclopentane; D018698:Glutamic Acid; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins; C012729:2-amino-4-phosphonobutyric acid; D002118:Calcium