CHRONIC GABA EXPOSURE DOWN-REGULATES GABA-BENZODIAZEPINE RECEPTOR-IONOPHORE COMPLEX IN CULTURED CEREBRAL CORTICAL-NEURONS

Cerebral cortical cultured neurons were characterized for GABA-benzodiazepine (BZ) receptor complex, and the effect of chronic exposure of cortical neurons to GABA on GABA-BZ receptor system was investigated. In the intact cells, the [H-3]flunitrazepam binding was rapid and saturable, with an apparent K(d) of 4.2 +/- 1.5 nM and B(max) of 776 +/- 54 fmol/mg protein. Specifically bound [H-3]flunitrazepam was displaced in a concentration-dependent manner by various BZ receptor ligands such as Ro15-1788, DMCM, Ro15-4513, clonazepam, alprazolam, diazepam and zolpidem, and enhanced by GABA, muscimol and pentobarbital. GABA induced enhancement of Cl-36-influx in a concentration-dependent manner (EC50 = 9 +/- 2 muM). Chronic exposure of the cultured neurons to GABA resulted in a reduced [H-3]flunitrazepam, [H-3]GABA, [H-3]Ro15-1788, [H-3]Ro15-4513 and [S-35]TBPS binding, a reduced enhancement of [H-3]flunitrazepam binding by GABA, and a reduced GABA-induced Cl-36-influx susceptible to reversal by concomitant exposure of the cultures to R 5135, a GABA(A)-receptor antagonist. These findings indicate that cerebral cortical cultured neurons provide an ideal model to study GABA-BZ receptor complex using binding and Cl-36-influx assays, and chronic exposure of cortical cultures to GABA leads to a down-regulation of GABA-BZ receptor system. It is a GABA(A) receptor-mediated slow process.