Analysis of intra- and intercellular calcium signaling in a mouse malignant glioma cell line.

Abstract

Intra- and intercellular calcium signaling in glioma cells was examined by mechanical stimulation of a monolayer cell line of methylcholanthrene-induced mouse ependymoblastoma, 203-glioma, with a fine round-tip glass needle. A fura-2 fluorescence image of the glioma revealed a four- to eightfold increase in the cytosolic calcium ion concentration in directly stimulated signal cells. The increased calcium spread to surrounding cells at a speed of 20 microns/sec for a distance of up to 200 microns. Calcium was transmitted between adjacent cells and even in cells up to 200 microns distant from the initially stimulated cell. Microinjection of Lucifer yellow dye showed no gap junctional communication between cells. Depletion of extracellular calcium ion inhibited both cytosolic calcium elevation and propagation to neighboring cells by mechanical stimulus. An intracellular calcium blocker, TMB-8, eliminated the cytosolic calcium mobilization in a mechanically stimulated cell, but had no effect on calcium diffusion to surrounding cells. Nifedipine and verapamil, antagonists of voltage-dependent calcium channels, did not act on the mechanically induced calcium response. This suggests that some stimulating factor may trigger transmission of calcium, which may be ejected directly from single stimulated cells and mediated via a membrane receptor but not through a gap junction. The calcium signaling in a mechanically stimulated cell may be related to both an influx and a redistribution of intracellular calcium from internal stores, while calcium propagation to neighboring cells may involve calcium influx alone.