Effects of treatment with GABA A receptor subunit antisense oligodeoxynucleotides on GABA-stimulated 36Cl − influx in the rat cerebral cortex

Abstract

GABA A receptor function was studied in cerebral cortical vesicles prepared from rats after intracerebroventricular microinjections of antisense oligodeoxynucleotides (aODNs) for α1, γ2, β1, β2 subunits. GABA A receptor α1 subunit aODNs decreased α1 subunit mRNA by 59±10%. Specific [ 3H]GABA binding was decreased by α1 or β2 subunit aODNs (to 63±3% and 64±9%, respectively) but not changed by γ2 subunit aODNs (94±5%). Specific [ 3H]flunitrazepam binding was increased by α1 or β2 subunit aODNs (122±8% and 126±11%, respectively) and decreased by γ2 subunit aODNs (50±13%). The “knockdown” of specific subunits of the GABA A receptor significantly influenced GABA-stimulated 36Cl − influx. Injection of α1 subunit aODNs decreased basal 36Cl − influx and the GABA E max; enhanced GABA modulation by diazepam; and decreased antagonism of GABA activity by bicuculline. Injection of γ2 subunit aODNs increased the GABA E max; reversed the modulatory efficacy of diazepam from enhancement to inhibition of GABA-stimulation; and reduced the antagonist effect of bicuculline. Injection of β2 subunit aODNs reduced the effect of diazepam whereas treatment with β1 subunit aODNs had no effect on the drugs studied. Conclusions from our studies are: (1) α1 subunits promote, β2 subunits maintain, and γ2 subunits suppress GABA stimulation of 36Cl − influx; (2) α1 subunits suppress, whereas β2, and γ2 subunits promote allosteric modulation by benzodiazepines; (3) diazepam can act as an agonist or inverse agonist depending on the relative composition of the receptor subunits; and (4) the mixed competitive/non-competitive effects of bicuculline result from activity at α1 and γ2 subunits and the lack of activity at β1 and β2 subunits.