Benzodiazepine binding and interactions with the GABA receptor complex in living cultures of rat cerebral cortex

Benzodiazepines bind to living cultures of dissociated rat cerebral cortex. This binding is saturable, and kinetic analyses indicate that the binding is to a single class on sites with kinetic constants very close to those obtained using neuronal membrane preparations. The efficacy of a number of benzodiazepines, xanthine derivatives and other drugs in competition experiments is similar to that seen in neuronal membrane preparations, and suggests that the benzodiazepine binding site studied in these investigations is the same as that found in neuronal membrane preparations and believed to be the pharmacologically active benzodiazepine binding site. GABA agonists increase the binding of benzodiazepines, and this increase has the same order of efficacy as their ability to hyperpolarize the neurons when applied at known concentrations with muscimol greater than GABA greater than THIP. At high concentrations THIP potentials benzodiazepine binding to the same level as GABA. Diazepam increases the ability of both GABA and THIP to hyperpolarize the neurons as well as the amplitude of spontaneous IPSPS which, in this system, are GABA-mediated. The competitive GABA antagonist bicuculline methiodide slightly decreased benzodiazepine binding and also antagonized the increase due to GABA. The non-competitive GABA antagonist picrotoxinin had no effect on benzodiazepine binding but did antagonize the GABA-induced increase in benzodiazepine binding. Replacement of Cl- in the incubation medium by acetate, which does not permeate the GABA-mediated Cl-- ionophore, increases benzodiazepine binding, and GABA no longer increases the binding. Picrotoxinin decreases the increase in benzodiazepine binding is Cl--free media, and this decrease is blocked by GABA. These results are discussed in terms of interactions at the GABA receptor complex consisting of a GABA recognition site, a benzodiazepine recognition site, a picrotoxinin recognition site, and a Cl- ionophore.