Antidepressants act at the GABA(A) receptor to inhibit GABA-stimulated 36Cl(-) influx and GABA reduction of [35S]TBPS binding. This study examined how selective knock-down (via antisense oligodeoxynucleotides, aODNs) of GABA(A) receptor subunits modified antidepressant activity. The specific aODNs used were for the alpha1, beta1, beta2 or gamma2 subunits of the GABA(A) receptor. The aODN microinjections reduced corresponding GABA(A) receptor subunit mRNA levels by 30-40% as assessed by RT-PCR. The inhibitory effect of the antidepressants amitriptyline and mianserin on GABA-stimulated 36Cl(-) influx was decreased after microinjections of alpha1, beta1, or beta2 subunit aODNs but potentiated after microinjections of gamma2 subunit aODNs. This pattern of aODNs effect on amitriptyline and mianserin modulation of GABA-stimulated 36Cl(-) influx was the same for both antidepressants and similar to GABA but different than that of diazepam and bicuculline. We conclude that multiple subunits of the GABA(A) receptor regulate the effect of amitriptyline and mianserin on the GABA(A) receptor chloride ionophore complex. However, the exact identity of the subunit mediating the direct or allosteric modulation of the antidepressant effect on GABA-stimulated 36Cl(-) influx remains unclear.