Depolarization of cultured astrocytes by KCl stimulated gamma-aminobutyric acid (GABA) release in a dose-dependent manner. At 60 mM KCl, the stimulatory effect was calcium- and sodium- independent, and was not altered by the presence of beta- alanine. The potassium-evoked GABA release was inhibited by furosemide and 4-acetamido-4'-isothiocyano- -2,2'-stilbene disulfonic acid (SITS), blockers of the chloride transporter across the plasma membrane, as well by chloride ion replacement with glucuronate. Other depolarizing agents, such as veratridine and ouabain, decreased basal GABA release; ouabain also inhibited the stimulatory effect of 60 mM KCl. The high K^+-induced GABA release may affect CNS excitability and may represent an important aspect of glial-neuronal interactions.