Abstract

Benzodiazepine potentiation of γ-aminobutyric acid (GABA) neurotransmission is associated with the presence of agamma-2 subunit in the GABA_A receptor. A method was developed to modify the gamma-2 subunit expression in adult rat brain. Unilateral intracerebroventricular (i.c.v.) infusion of a 17-base phosphorothioate-modified antisense oligodeoxynucleotide (ASO) was performed every 12 hr for 3 days. Controls were treated with a sense oligodeoxynucleotide. Parasagittal brain sections were used for quantitative autoradiographic analysis of radioligand binding. ASO treatment caused a 15% to 25% decrease of specific [³H]flunitrazepam binding in most brain areas, with statistically significant decreases in frontal cortex, cerebellar molecular layer, zona reticulata of substantia nigra and CA3 of hippocampus. In contrast, [³H]muscimol binding was not changed. [³H]GABA binding was also unchanged, except for a 10% decrease in cerebellar granule cell layer. The effect on the chloride channel of the GABA_A receptor complex was examined by 4′-ethynyl-4-n-[2,3-³H₂]propylbicycloorthobenzoate binding; most brain areas showed small decreases in 4′-ethynyl-4-n-[2,3-³H₂]propylbicycloorthobenzoate binding. However, hippocampal regions showed much larger decreases. Binding of the adenosine A₁ receptor antagonist [³H]8-cyclopentyl-1,3-dipropylxanthine was used to examine possible secondary effects of the ASO. There was a decrease in [³H]8-cyclopentyl-1,3-dipropylxanthine binding, but this was much smaller than the change in [³H]flunitrazepam binding, and no area showed a significant effect. Quantitative immunoblotting with a monoclonal antibody that recognizes GABA_A receptor beta-2 andbeta-3 subunits showed no change in immunoreactivity in cerebellar tissue after ASO treatment. The results indicate a selective effect on benzodiazepine binding to GABA_A receptors and a possible change in receptor subunit composition.