Stoichiometry of a recombinant GABAA receptor deduced from mutation-induced rectification

Backus, Kurt H.; Arigoni, Michele; Drescher, Uwe; Scheurer, Louis; Malherbe, Pari; Möhler, Hanns; Benson, Jack A.

doi:10.1097/00001756-199312000-00026

Cited by 159 publications

(81 citation statements)

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“…This subunit composition can be reached by combining either three subunits of one type with one subunit each of the other two types or two subunits each from two types with one subunit of the third type. The former stoichiometry has been excluded for ␣ 3 ␤ 2 ␥ 2 and ␣ 1 ␤ 2 ␥ 2 receptors by electrophysiological characterization of mutant receptors (13,14). For FIG.…”

Section: Discussionmentioning

confidence: 99%

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Forced Subunit Assembly in α1β2γ2 GABAAReceptors

Baumann

Baur

Sigel

2002

Journal of Biological Chemistry

236

137

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The major isoform of the ␥-aminobutyric acid type A (GABA A ) receptor is thought to be composed of 2␣ 1 , 2␤ 2 , and 1␥ 2 subunit(s), which surround the ion pore. Definite evidence for the subunit arrangement is lacking. We show here that GABA A receptor subunits can be concatenated to a trimer that can be functionally expressed upon combination with a dimer. Many combinations did not result in the functional expression. In contrast, four different combinations of triple subunits with dual subunit constructs, all resulting in the identical pentameric receptor ␥ 2 ␤ 2 ␣ 1 ␤ 2 ␣ 1 , could be successfully expressed in Xenopus oocytes. We characterized the functional properties of these receptors in respect to agonist, competitive antagonist, and diazepam sensitivity. All properties were similar to those of wild type ␣ 1 ␤ 2 ␥ 2 GABA A receptors. Thus, together with information on the crystal structure of the homologous acetylcholine-binding protein (Brejc, K., van Dijk, W. J., Klaassen, R. V., Schuurmans, M., van Der Oost, J., Smit, A. B., and Sixma, T. K., (2001) Nature 411, 269 -276, we provide evidence for an arrangement ␥ 2 ␤ 2 ␣ 1 ␤ 2 ␣ 1 , counterclockwise when viewed from the synaptic cleft. Forced subunit assembly will also allow receptors containing different subunit isoforms or mutant subunits to be expressed, each in a desired position. The methods established here should be applicable to the entire ion channel family comprising nicotinic acetylcholine, glycine, and 5HT 3 receptors.

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Section: Discussionmentioning

confidence: 99%

“…The ␥ subunit has been shown to be required for functional modulation of the receptor channels by benzodiazepines (11,12). Different approaches have indicated a 2␣:2␤:1␥ subunit stoichiometry for this receptor (13)(14)(15)(16).…”

mentioning

confidence: 99%

Forced Subunit Assembly in α1β2γ2 GABAAReceptors

Baumann

Baur

Sigel

2002

Journal of Biological Chemistry

236

137

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“…The most common GABA A R subunit combination found in the brain contains two ␣ subunits, two ␤ subunits, and one ␥ subunit (Im et al, 1995;Chang et al, 1996;Li and De Blas, 1997;Jechlinger et al, 1998;Farrar et al, 1999), although combinations of two ␣, one ␤, and two ␥ subunits also occur in the brain (Backus et al, 1993;Khan et al, 1994aKhan et al, ,b, 1996.…”

Section: Individual Hippocampal Pyramidal Cells and Interneurons In Cmentioning

confidence: 99%

GABAergic Innervation Organizes Synaptic and Extrasynaptic GABA_AReceptor Clustering in Cultured Hippocampal Neurons

2002

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We have studied the effects of GABAergic innervation on the clustering of GABA A receptors (GABA A Rs) in cultured hippocampal neurons. In the absence of GABAergic innervation, pyramidal cells form small (0.36 Ϯ 0.01 m diameter) GABA A R clusters at their surface in the dendrites and soma. When receiving GABAergic innervation from glutamic acid decarboxylase-containing interneurons, pyramidal cells form large (1.62 Ϯ 0.08 m breadth) GABA A R clusters at GABAergic synapses. This is accompanied by a disappearance of the small GABA A R clusters in the local area surrounding each GABAergic synapse. Although the large synaptic GABA A R clusters of any neuron contained all GABA A R subunits and isoforms expressed by that neuron, the small clusters not localized at GABAergic synapses showed significant heterogeneity in subunit and isoform composition. Another difference between large GABAergic and small non-GABAergic GABA A R clusters was that a significant proportion of the latter was juxtaposed to postsynaptic markers of glutamatergic synapses such as PSD-95 and AMPA receptor GluR1 subunit. The densities of both the glutamate receptor-associated and non-associated small GABA A R clusters were decreased in areas surrounding GABAergic synapses. However, no effect on the density or distribution of glutamate receptor clusters was observed. The results suggest that there are local signals generated at GABAergic synapses that induce both assembly of large synaptic GABA A R clusters at the synapse and disappearance of the small GABA A R clusters in the surrounding area. In the absence of GABAergic innervation, weaker GABA A R-clustering signals, generated at glutamatergic synapses, induce the formation of small postsynaptic GABA A R clusters that remain juxtaposed to glutamate receptors at glutamatergic synapses.

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“…They are chloride ion channels composed of five subunits, which are classified into six subunit families [1][2][3]. a 1 b 2 c 2 is the major adult receptor isoform [1,2] and its subunit stoichiometry is 2a:2b:1c [4][5][6][7][8][9][10]. We concentrated on this major receptor isoform.…”

Section: Introductionmentioning

confidence: 99%

Two neighboring residues of loop A of the α₁ subunit point towards the benzodiazepine binding site of GABA_A receptors

et al. 2007

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Benzodiazepines are widely used drugs exerting sedative, anxiolytic, muscle relaxant, and anticonvulsant effects by acting through specific high affinity binding sites on some GA-BA A receptors. It is important to understand how these ligands are positioned in this binding site. We are especially interested here in the conformation of loop A of the a 1 b 2 c 2 GABA A receptor containing a key residue for the interaction of benzodiazepines: a 1 H101. We describe a direct interaction of a 1 N102 with a diazepam-and an imidazobenzodiazepine-derivative. Our observations help to better understand the conformation of this region of the benzodiazepine pocket in GABA A receptor.

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Stoichiometry of a recombinant GABAA receptor deduced from mutation-induced rectification

Cited by 159 publications

References 0 publications

Forced Subunit Assembly in α1β2γ2 GABAAReceptors

Forced Subunit Assembly in α1β2γ2 GABAAReceptors

GABAergic Innervation Organizes Synaptic and Extrasynaptic GABA_AReceptor Clustering in Cultured Hippocampal Neurons

Two neighboring residues of loop A of the α₁ subunit point towards the benzodiazepine binding site of GABA_A receptors

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Stoichiometry of a recombinant GABAA receptor deduced from mutation-induced rectification

Cited by 159 publications

References 0 publications

Forced Subunit Assembly in α1β2γ2 GABAAReceptors

Forced Subunit Assembly in α1β2γ2 GABAAReceptors

GABAergic Innervation Organizes Synaptic and Extrasynaptic GABAAReceptor Clustering in Cultured Hippocampal Neurons

Two neighboring residues of loop A of the α1 subunit point towards the benzodiazepine binding site of GABAA receptors

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GABAergic Innervation Organizes Synaptic and Extrasynaptic GABA_AReceptor Clustering in Cultured Hippocampal Neurons

Two neighboring residues of loop A of the α₁ subunit point towards the benzodiazepine binding site of GABA_A receptors