Abstract: Astroglia are extensively coupled by gap junctions and form a functional syncytium. Astroglial gap junctions are thought to be involved in the spatial buffering of K+ in vivo and in the Ca2+ waves seen on glutamate receptor activation. The conductivity of gap junctions is regulated by several second messengers, with up-regulation by cyclic AMP and down-regulation through activation of protein kinase C, decreases in intracellular pH, or increases in the free cytosolic Ca2+ concentration. The results presented here indicate that dye coupling of astroglia is significantly up-regulated by membrane depolarization, both by increases in the extracellular K+ concentration and directly by ionophores. Furthermore, glutamate, kainate, and quisqualate, which depolarize astroglial cells through activation of ionotropic receptors, also increase dye coupling in astroglia. The effect of kainate and quisqualate was reversed by 6-cyano-7-nitroquinoxaline-2,3-dione, an inhibitor of the ionotropic glutamate receptor. A dose-dependent decrease in dye coupling was seen when the cells were injected with increasing concentrations of Ca2+. However, if the cells were simultaneously depolarized, the inhibitory effect of Ca2+ on gap junctional conductance was reversed. Significant increases over basal coupling was attained when the cells were injected with Ca2+ if they were treated with kainate or K+. These data suggest that ligands that depolarize astroglia enhance gap junction communication between astroglia and that this enhancement may be important in maintaining communication between astroglia in the face of elevated Ca2+ levels.