Molecular and Pharmacological Characterization of Native Cortical γ-Aminobutyric AcidA Receptors Containing Both α1 and α3 Subunits

Araujo, Francisco; Tan, Suan; Ruano, Diego; Schoemaker, Hans E.; Bénavidès, J.; Vitórica, Javier

doi:10.1074/jbc.271.44.27902

Cited by 53 publications

(62 citation statements)

References 44 publications

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“…The effect of the other tertiary subunits (δ, ε, π, and θ) on the properties of receptors containing different α subunit subtypes would also be of interest. Adding another layer of complexity is the relatively common occurrence of native receptors containing two different α subtypes within a single receptor (Araujo et al, 1996(Araujo et al, , 1999Jechlinger et al, 1998;Benke et al, 2004). Would the kinetic properties of these receptors be intermediate, or might they be dominated by the characteristics of one of the subtypes?…”

Section: Discussionmentioning

confidence: 99%

Effect of the α subunit subtype on the macroscopic kinetic properties of recombinant GABAA receptors

Picton

Fisher

2007

Brain Research

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The GABA A receptors (GABARs) are chloride-permeable ligand-gated ion channels responsible for fast inhibitory neurotransmission. These receptors are structurally heterogeneous, and in mammals can be formed from a combination of sixteen different subunit subtypes. Much of this variety comes from the six different α subunit subtypes. All neuronal GABARs contain an α subunit, and the identity of the α subtype affects the pharmacological properties of the receptors. The expression of each of the different α subtypes is regulated developmentally and regionally and changes with both normal physiological processes such development and synaptic plasticity, and pathological conditions such as epilepsy. In order to understand the functional significance of this structural heterogeneity, we examined the effect of the α subtype on the receptor's response to GABA. Each of the six α subtypes was transiently co-expressed with the β3 and γ2L subunits in mammalian cells. The sensitivity to GABA was measured with whole-cell recordings. We also determined the activation, deactivation, desensitization, and recovery kinetics for the six isoforms using rapid-application recordings from excised macropatches. We found unique characteristics associated with each α subunit subtype. These properties would be expected to influence the post-synaptic response to GABA, creating functional diversity among neurons expressing different α subunits.

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Section: Discussionmentioning

confidence: 99%

Effect of the α subunit subtype on the macroscopic kinetic properties of recombinant GABAA receptors

Picton

Fisher

2007

Brain Research

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“…Antibody Purification and Immunoprecipitations-When needed, the anti-␣1 and anti-␥2 antibodies (24,25) were purified through peptide affinity columns. The peptides were coupled to CNBr-activated Sepharose 4B (Amersham Pharmacia Biotech).…”

Section: Methodsmentioning

confidence: 99%

“…Thus, the ␥2 subunit from adult and aged hippocampus was first immunopurified through anti-␥2 affinity columns (our anti-␥2 antibody was made against a peptide of the N-terminal domain of the protein, and thus, it recognizes both short and long isoforms). The anti-␥2 immunoaffinity columns were synthesized as described (25). Fab fragments of the purified anti-␥2 antibodies were used in order to avoid any possible interference with the IgG heavy chain (55 kDa).…”

Section: Methodsmentioning

confidence: 99%

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GABAA and α-Amino-3-hydroxy-5-methylsoxazole-4-propionate Receptors Are Differentially Affected by Aging in the Rat Hippocampus

Ruano¹,

Araujo²,

Revilla³

et al. 2000

Journal of Biological Chemistry

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We have investigated the age-dependent modifications in the expression of eight different subunits of the ␥-aminobutyric acid, type A (GABA A ) receptor (␣1, ␣2, ␣3, ␣5, ␤2, ␤3, ␥2S, and ␥2L) and all four subunits of the ␣-amino-3-hydroxy-5-methylsoxazole-4-propionate (AMPA) receptor (GluR1-4) in the hippocampus of 24-month-old rats. All aged hippocampi displayed a remarkable increase (aged/adult ratio, 3.53 ؎ 0.54) in the mRNA levels of the short version of the ␥2 subunit in parallel with a similar increase in the ␥2 subunit protein (aged/adult ratio, 2.90 ؎ 0.62). However, this increase was not observed in the mature receptor. On the other hand, the expression of the different ␣ subunit mRNAs increased moderately with aging, displaying a heterogeneous pattern. The most frequent modification consisted in an increase in the expression of the ␣1 subunit mRNA (aged/adult ratio, 1.26 ؎ 0.18), in parallel with a similar increase on the ␣1 protein (aged/adult ratio, 1.27 ؎ 0.12) and in the ␣1 incorporated to the assembled GABA A receptor (tested by immunoprecipitation; aged/ adult ratio, ‫؍‬ 1.20 ؎ 0.10). However, in the same hippocampal samples, no major modifications were observed on the expression of the AMPA receptor subunits. As a whole, these results indicated the existence of an increased expression of the GABA A receptor subunits and a preservation of the AMPA receptor at the hippocampal formation. These modifications could reflect the existence of specific deficiencies (neuronal loss and/or deafferentiation) on the GABAergic system in the aged rats.Normal aging is associated with memory and/or learning impairments that could reflect modifications at the hippocampal formation (1). The GABA A 1 and AMPA receptors (major fast inhibitory and excitatory receptor complexes, respectively) could be implicated in these alterations (2, 3).Both neurotransmitter receptors are composed of a high number of subunits in a, probably, pentameric or tetrameric conformation. The GABA A receptors are formed by the combination of a total of 19 subunits grouped in eight families: ␣1-6, ␤1-3, ␥1-3, ␦, 1-3, ⑀, , and (Ref.5; for a review, see Ref. 4).The AMPA-preferring ionotropic glutamate receptor is composed by four subunits (GluR1-4) displaying different splicing isoforms (for a review, see Ref. 6). This high molecular heterogeneity can generate multiple receptor isotypes, displaying particular physiological and pharmacological properties. It is known that the sensitivity for benzodiazepines (anxiolytic/hypnotic drugs that interact to the GABA A receptor) increase during aging in humans and in rodents (7). Previous work from our group has demonstrated the existence of agingassociated modifications in both the pharmacological properties and the molecular composition of the GABA A receptors in rat hippocampus (8 -10). These changes could reflect a sensitization process of the GABA A receptor (see also Ref. 11). However, the age-dependent modifications on the expression of the different subunits of the GABA A receptor are currently ...

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“…This observation is consistent with the notion that the pyramidal cells express a population of GABA A Rs in which the two ␣ subunits present in the pentamer are of the same isoform. Nevertheless, these cells are also very likely to have pentamers that contain two different ␣ subunit isoforms (McKernan et al, 1991;Khan et al, 1994aKhan et al, , 1996Araujo et al, 1996Araujo et al, , 1999Jechlinger et al, 1998;Sigel and Baur, 2000). We do not know whether the small clusters that show colocalization of ␣ 1 and ␣ 2 or ␣ 1 and ␣ 3 have receptors that contain both isoforms or a mixture of receptors containing only one type of isoform.…”

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confidence: 98%

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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Molecular and Pharmacological Characterization of Native Cortical γ-Aminobutyric AcidA Receptors Containing Both α1 and α3 Subunits

Abstract: We have investigated the existence, molecular composition, and benzodiazepine binding properties of native cortical ␣ 1 -␣ 3 ␥-aminobutyric acid A (GABA A ) receptors using subunit-specific antibodies.The

Cited by 53 publications

References 44 publications

Effect of the α subunit subtype on the macroscopic kinetic properties of recombinant GABAA receptors

Effect of the α subunit subtype on the macroscopic kinetic properties of recombinant GABAA receptors

GABAA and α-Amino-3-hydroxy-5-methylsoxazole-4-propionate Receptors Are Differentially Affected by Aging in the Rat Hippocampus

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

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Molecular and Pharmacological Characterization of Native Cortical γ-Aminobutyric AcidA Receptors Containing Both α1 and α3 Subunits

Abstract: We have investigated the existence, molecular composition, and benzodiazepine binding properties of native cortical ␣ 1 -␣ 3 ␥-aminobutyric acid A (GABA A ) receptors using subunit-specific antibodies.The

Cited by 53 publications

References 44 publications

Effect of the α subunit subtype on the macroscopic kinetic properties of recombinant GABAA receptors

Effect of the α subunit subtype on the macroscopic kinetic properties of recombinant GABAA receptors

GABAA and α-Amino-3-hydroxy-5-methylsoxazole-4-propionate Receptors Are Differentially Affected by Aging in the Rat Hippocampus

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

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