Characterization of the hypo-osmolarity-induced Ca2+ response in cultured rat astrocytes

The influence of astrocyte swelling on the cytosolic free calcium concentration [Ca2+]i was studied at the single cell level. Sudden exposure of normo-osmotically (305 mosmol/l) cultured astrocytes to hypo-osmotic medium induced a biphasic increase in cytosolic calcium with an initial peak followed by a sustained plateau. The response was osmolarity dependent and was maximal at 205 mosmol/l with respect to [Ca2+]i and the percentage of responding cells. Other modes of astrocyte swelling [gradual adjustment of hypo-osmolarity, normo-osmotic exposure of hyper-osmotic (405 mosmol/l) maintained cells] produced a much weaker [Ca2+]i response. Change from 405 to 205 mosmol/l, however, resulted in the entire peak and an increased plateau. Experiments with Ca(2+)-free medium and after pretreatment with BAPTA-AM, thapsigargin, phorbol myristate acetate, or nimodipine revealed that the peak mainly resulted from depletion of intracellular Ca2+ stores, whereas the plateau was probably due to capacitative Ca2+ entry and Ca2+ influx independent of store depletion including a nimodipin-sensitive component. Prior depletion of ryanodine-, bradykinin- or ATP-sensitive stores revealed that the initial hypo-osmolarity-induced Ca(2+)-release was from a Ca2+ pool also affected by ATP and bradykinin, but not by ryanodine. The recent finding, that the hypo-osmolarity-induced [Ca2+]i response was completely maintained if phospholipase C-mediated phosphatidylinositol hydrolysis was blocked, suggests that hypo-osmolarity may exert an inositol (1,4,5) triphosphate-independent access to these stores.