The endogenous benzodiazepine receptor ligand ODN increases cytosolic calcium in cultured rat astrocytes

We have investigated the production of diazepam-binding inhibitor (DBI)-related peptides by astrocytes in primary culture and we have determined the effect of the octadecaneuropeptide DBI[33-50] (ODN) on the intracellular calcium concentration ([Ca2+](i)) in astrocytes. Immunocytochemical labeling with antibodies against ODN showed that cultured astrocytes retain their ability to synthesize DBI in vitro. Cultured astrocytes were also found to release substantial amounts of ODN-immunoreactive material, and a brief exposure of astrocytes to a depolarizing potassium concentration resulted in a 5-fold increase in the rate of release of the ODN-like peptide. Microfluorimetric measurement of [Ca2+](i) with the fluorescent probe indo-1 showed that nanomolar concentrations of ODN induced a marked increase in [Ca2+](i). The stimulatory effect of ODN on [Ca2+](i) was not affected by calcium channel blockers or by incubation in Ca2+-free medium. in contrast, thapsigargin, an inhibitor of microsomal Ca2+-ATPase activity, totally abolished the ODN-induced increase in [Ca2+](i). Repeated pulses of ODN caused attenuation of the response, indicating the existence of a desensitization phenomenon. Preincubation of astrocytes with pertussis toxin totally blocked the effect of ODN on [Ca2+](i). The present study indicates that ODN-related peptides are synthesized and released by glial cells. Our results also show that synthetic ODN induces calcium mobilization from an intracellular store through stimulation of pertussis toxin-sensitive G protein. Taken together, these data suggest that endozepines act as paracrine and/or autocrine factors controlling the activity of astroglial cells.