gamma-Aminobutyric acid type A receptors in the rat brain can contain both gamma 2 and gamma 3 subunits, but gamma 1 does not exist in combination with another gamma subunit.
Antibodies specific for the gamma 1, gamma 2, and gamma 3 subunits of the gamma-aminobutyric acid (GABA)A receptor have been used to probe the composition of naturally occurring GABAA receptors in the rat brain. Most GABAA receptors contain at least one of these three subunits. The percentage of each, determined by immunoprecipitation of [3H]muscimol binding, was 11 +/- 1%, 59 +/- 3%, and 14 +/- 2% for gamma 1, gamma 2, and gamma 3 subunits, respectively. Receptors containing gamma 2 or gamma 3 subunits were labeled by benzodiazepine site ligands with high affinity, whereas gamma 1-containing receptors could be labeled only by [3H]muscimol. Receptors immunoprecipitated by anti-gamma 2 or anti-gamma 3 antibodies were labeled with [3H]Ro 15-1788 with similar affinities (Kd for anti-gamma 2-immunoprecipitated receptors, 1.9 nM; Kd for anti-gamma 3-immunoprecipitated receptors, 1.7 nM). Immunoprecipitation or Western blot analysis of GABAA receptors solubilized from rat cerebellar or whole-brain preparations indicated that gamma 1 was not present coassembled with any other gamma subunit. Western blot analysis of receptors purified on alpha-specific immunoaffinity resins showed that gamma 1 was predominantly assembled with the alpha 2 subunit. Some GABAA receptors may contain more than one type of gamma subunit. Quantitative immunoprecipitation and Western blot analysis both indicated that gamma 2 and gamma 3 subunits can exist in the same receptor complex. A large proportion of GABAA receptors immunopurified on a gamma 3 affinity resin also appeared to contain a gamma 2 subunit. In contrast, when receptors were purified on a gamma 2 affinity resin a small proportion also appeared to contain a gamma 3 subunit. We conclude that most gamma 1-containing receptors have no other gamma subunit in the same receptor complex but some GABAA receptors contain both gamma 2 and gamma 3 subunits.
Department of Biochemistry, Merck, Sharp, and Dohme Research Laboratories, Harlow, Essex, UK.
K
K
Quirk
N P
NP
Gillard
C I
CI
Ragan
P J
PJ
Whiting
R M
RM
McKernan
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism*; D002846:Chromatography, Affinity; D006801:Humans; D007106:Immune Sera; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism*; D011233:Precipitin Tests; D051381:Rats; D018079:Receptors, GABA / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism
D007106:Immune Sera; D010446:Peptide Fragments; D018079:Receptors, GABA; D011994:Recombinant Proteins