A significant part of native γ-aminobutyric acidA receptors containing α4 subunits do not contain γ or δ subunits

Using a novel antibody directed against the alpha(4) subunit of gamma-aminobutyric acid(A) (GABA(A)) receptors, 5% of all [H-3]muscimol but only about 2% of all [H-3]Ro15-4513 binding sites present in brain membrane extracts could be precipitated. This indicated that part of the alpha(4) receptors containing [H-3]muscimol binding sites did not contain [H-3]Ro15-4513 binding sites. Immunoaffinity purification and Western blot analysis of alpha(4) receptors demonstrated that not only alpha(1), alpha(2), alpha(3), beta(1), beta(2), and beta(3) subunits but also gamma(1), gamma(2), gamma(3), and delta subunits can be colocalized with alpha(4) subunits in native GABA(A) receptors. Quantification experiments, however, indicated that only 7, 33, 4, or 7% of all alpha(4) receptors contained gamma(1), gamma(2), gamma(3), or delta subunits, respectively. These data not only explain the low percentage of [H-3]Ro15-4513 binding sites precipitated by the anti-alpha(4) antibody but also indicate that approximately 50% of the alpha(4) receptors did not contain gamma(1), gamma(2), gamma(3), or delta subunits. These receptors, thus, either are composed of alpha(4) and beta(1-3) subunits only, or additionally contain epsilon, pi, or so far unidentified GABA(A) receptor subunits.