GLUTAMATE-INDUCED SWELLING OF SINGLE ASTROGLIAL CELLS IN PRIMARY CULTURE

Glutamate induced an increase in cell volume within one minute and evo ked cytosolic Ca2+ transients in type 1 astroglial cells in primary cu lture obtained from the cerebral cortex of newborn rat. Even the metab otropic glutamate receptor agonists (1S,3R)-1-aminocyclopentane 1,3-di carboxylic acid (1S-3R-ACPD) and L(+)-2-amino-4 phosphonobutyric acid (L-AP4) induced a cell swelling with ACPD inducing a parallel Ca2+ tra nsient 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. Re lative volume changes in cultured single astroglial cells were examine d by microspectrofluorimetry after loading the cells with the highly f luorescent 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 e xcitation 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 tetraethyl ammonium, suggesting that K+ channels art operative in glutamate-induc ed cell swelling. Furthermore, the glutamate-induced swelling was bloc ked by the Na+; K+, and 2Cl(-) co-transport inhibitor furosemide. The glutamate-induced swelling was partially blocked by pertussis toxin an d partially blocked also by the glutamate carrier-blocker dihydroaspar tate. When the ionotropic glutamate receptor alpha-amino-3-hydroxy-5-m ethyl-4-isoxazole proprionic acid was blocked with the antagonist -dih ydroxy-6-nitro7-sulfamoyl-benzo(F)quinoxaline, glutamate still induced a swelling, suggesting that this receptor was not directly involved i n 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 law extracellular Ca2+ concentrations. Our data suggest that gluta mate-induced rapid swelling is a complex process at the molecular leve l. One hypothetical mechanism might be that glutamate interacts with m etabotropic glutamate receptors and induces a release of Ca2+ from int ernal stores. Furthermore glutamate interacts with K+ channels, and pr obably at least one co-transporter and the sodium-dependent high-affin ity uptake glutamate carrier, resulting in cell swelling.