ABSENCE OF ASSOCIATION BETWEEN DELTA-2 AND GAMMA-2 SUBUNITS IN NATIVEGABA(A) RECEPTORS FROM RAT-BRAIN

We investigated the possible association between delta and gamma 2 sub units in native GABA(A) receptors, from different rat brain regions, u sing subunit-specific anti-delta and anti-gamma 2 antibodies. Previous reports have provided somewhat controversial results, indicating both the presence and the absence of association between these two subunit s in native receptors. Our results indicate the absence of co-localiza tion between delta and gamma 2 subunits. In immunoprecipitation experi ments, anti-delta antibody consistently immunoprecipitated [H-3]muscim ol binding activity (GABA binding sites) from all brain areas tested ( 10-20% of the total binding). However, under the same conditions, no s ignificant [H-3]flumazenil or [H-3]ethyl -imidazol[1,5-a]-[1,4]benzodi azepine-3-carboxylate (Ro15-4513) binding (benzodiazepine binding site s) activity was detected in the immunopellets. These results indicate the absence of association between delta and gamma 2 subunits. This qu estion was directly addressed by immunopurification and Western blot e xperiments. As expected, no gamma 2 subunits were detected in anti-del ta immunoaffinity purified receptors. Conversely, no delta subunits we re identified in anti-gamma 2 immunopurified receptors. Thus, these re sults demonstrate the absence of association between delta and gamma 2 subunits in native GABA(A) receptors. Finally, our results also indic ate the relevance of the solubilization conditions on the apparent ass ociation between different subunits of the native GABA(A) receptor com plex. (C) 1998 Elsevier Science B.V.