Analysis of the presence and abundance of GABAA receptors containing two different types of α subunits in murine brain using point-mutated α subunits

被引:60
作者
Benke, D
Fakitsas, P
Roggenmoser, C
Michel, C
Rudolph, U
Mohler, H
机构
[1] Univ Zurich, Inst Pharmacol & Toxicol, CH-8057 Zurich, Switzerland
[2] Fed Inst Technol, Dept Chem & Appl Biosci, CH-8057 Zurich, Switzerland
关键词
D O I
10.1074/jbc.M407154200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
gamma-Aminobutyric acid, type A (GABA(A)) receptors are pentameric proteins of which the majority is composed of two alpha subunits, two beta subunits and one gamma subunit. It is well documented that two different types of alpha subunits can exist in a singles GABA(A) receptor complex. However, information on the abundance of such GABA(A) receptors is rather limited. Here we tested whether mice containing the His to Arg point mutation in the alpha1, alpha2, or alpha3 subunit at positions 101, 101, and 126, respectively, which render the respective subunits insensitive to diazepam, would be suitable to analyze this issue. Immunodepletion studies indicated that the His to Arg point mutation solely rendered those GABA(A) receptors totally insensitive to diazepam binding that contain two mutated alpha subunits in the receptor complex, whereas receptors containing one mutated and one heterologous alpha subunit not carrying the mutation remained sensitive to diazepam binding. This feature permitted a quantitative analysis of native GABA(A) receptors containing heterologous alpha subunits by comparing the diazepam-insensitive binding sites in mutant mouse lines containing one mutated alpha subunit with those present in mouse lines containing two different mutated alpha subunits. The data indicate that the alpha1alpha1-containing receptors with 61% is the most abundant receptor subtype in brain, whereas the alpha1alpha2 (13%), alpha1alpha3 (15%), alpha2alpha2 (12%), alpha2alpha3 (2%), and alpha3alpha3 combinations (4%) are considerably less expressed. Only within the alpha1-containing receptor population does the combination of equal alpha subunits (84% alpha1alpha1, 7% alpha1alpha2,and 8% alpha1alpha3) prevail, whereas in the alpha2-containing receptor population (46% alpha2alpha2, 36% alpha2alpha1, and 19% alpha2alpha3) and particularly in the alpha3-containing receptor population (27% alpha3alpha3, 56% alpha3alpha1, and 19% alpha3alpha2), the receptors with two different types of alpha subunits predominate. This experimental approach provides the basis for a detailed analysis of the abundance of GABA(A) receptors containing heterologous alpha subunits on a brain regional level.
引用
收藏
页码:43654 / 43660
页数:7
相关论文
共 28 条
[1]   Molecular and pharmacological characterization of native cortical gamma-aminobutyric acid(A) receptors containing both alpha(1) and alpha(3) subunits [J].
Araujo, F ;
Tan, S ;
Ruano, D ;
Schoemaker, H ;
Benavides, J ;
Vitorica, J .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1996, 271 (44) :27902-27911
[2]  
Araujo F, 1999, J PHARMACOL EXP THER, V290, P989
[3]  
Barnard EA, 1998, PHARMACOL REV, V50, P291
[4]  
Baumann SW, 2003, J NEUROSCI, V23, P11158
[5]   Forced subunit assembly in α1β2γ2 GABAA receptors -: Insight into the absolute arrangement [J].
Baumann, SW ;
Baur, R ;
Sigel, E .
JOURNAL OF BIOLOGICAL CHEMISTRY, 2002, 277 (48) :46020-46025
[6]   IMMUNOCHEMICAL IDENTIFICATION OF THE ALPHA-1-SUBUNIT AND ALPHA-3-SUBUNIT OF THE GABA-A-RECEPTOR IN RAT-BRAIN [J].
BENKE, D ;
CICINSAIN, A ;
MERTENS, S ;
MOHLER, H .
JOURNAL OF RECEPTOR RESEARCH, 1991, 11 (1-4) :407-424
[7]  
Benke D, 1997, J NEUROCHEM, V69, P806
[8]   GABA(A) receptor subtypes differentiated by their gamma-subunit variants: Prevalence, pharmacology and subunit architecture [J].
Benke, D ;
Honer, M ;
Michel, C ;
Mohler, H .
NEUROPHARMACOLOGY, 1996, 35 (9-10) :1413-1423
[9]  
BENKE D, 1991, Molecular Neuropharmacology, V1, P103
[10]   Pharmacology of recombinant γ-aminobutyric acida receptors rendered diazepam-insensitive by point-mutated α-subunits [J].
Benson, JA ;
Löw, K ;
Keist, R ;
Mohler, H ;
Rudolph, U .
FEBS LETTERS, 1998, 431 (03) :400-404