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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A 1.45 kb DNA sequence encoding the rat alpha 6 GABAA receptor subunit (nucleotides 33-1483) was cloned from a Sprague-Dawley rat brain cDNA library by PCR amplification. Dideoxy sequencing of two individual clones revealed that the nucleotide sequence differed at only one basepair (T480-->G) from that published previously. This difference altered the deduced amino acid sequence, producing a conservative amino acid substitution (His121-->Gln). A Gln residue is present at the same location in the bovine alpha 6 subunit. Restriction endonuclease analysis of the total PCR product demonstrated that this variant of the rat alpha 6 subunit was the only allele found in this particular rat brain library, the original allele was not present. These results were further verified by RNAse protection assays performed with RNA isolated from individual rat cerebella. alpha 6, beta 1, and gamma 2S subunits were transiently expressed in L929 cells for electrophysiological analysis. Whole-cell recordings obtained from the cells demonstrated that GABAA receptor channels with the expected GABA and benzodiazepine pharmacology were produced. Excised outside out single channel recordings from the same cells revealed that GABA elicited brief duration openings to a 33 pS main conductance level and to at least one smaller (approximately 21 pS) subconductance level. Thus this allelic variant of rat alpha 6 subunit could assemble with other subunits to form a functional GABAA receptor channel with similar properties to the original allelic form.
Brain Res Mol Brain Res 1992 Nov
PMID:Molecular and electrophysiological characterization of a allelic variant of the rat alpha 6 GABAA receptor subunit. 128 Dec 55

We examined the expression of the beta subunit mRNA of the glycine receptor and the gamma subunit mRNA of the GABAA receptor in the rat dorsal root ganglion (DRG) using in situ hybridization histochemistry with oligonucleotide probes. About 44% and 37% of the all DRG neurons were labeled by the probes for glycine receptor beta subunit and GABAA receptor gamma subunit mRNAs. Labeled neurons were mostly large cells that simultaneously expressed both glycine receptor beta subunit and GABAA receptor gamma subunit mRNA as demonstrated using consecutive sections. Thus, we suggest the possibility that both GABA and glycine presynaptically regulate the activity of neurons involved in low-threshold mechanoreception at axo-axonic synapses in the spinal cord.
Brain Res Mol Brain Res 1992 Feb
PMID:Co-expression of glycine receptor beta subunit and GABAA receptor gamma subunit mRNA in the rat dorsal root ganglion cells. 131 5

The effects of protein kinase C (PKC) activators on gamma-aminobutyric acidA (GABAA) receptor function were studied by two-electrode voltage-clamp in Xenopus oocytes expressing brain mRNA or subunit cDNAs and in isolated mouse brain cerebellar membrane vesicles (microsacs), using 36Cl- uptake. Both oocytes and microsacs showed transient (desensitizing) and sustained (nondesensitizing) GABAA receptor responses. In oocytes expressing brain mRNA, the PKC activator phorbol myristoyl acetate (PMA), but not the inactive analog phorbol 12-monomyristate, inhibited both transient and sustained GABA-gated chloride currents. The inhibition by PMA was concentration dependent, with an EC50 of approximately 5 nM, and resulted in a decrease in the efficacy, but not the potency, of GABA. Additionally, PMA inhibited GABA-gated chloride currents in oocytes expressing alpha 1 beta 1 gamma 2L subunit cDNAs. The effect of PMA on recombinant receptors was significantly antagonized by PKC inhibitory peptide (PKCI). In the microsac preparation, the PKC activators (-)-7-octylindolactam V and PMA inhibited the sustained phase of 36Cl- flux without altering the transient phase. The action of PMA was blocked by kinase inhibitors and by depletion of Mg-ATP and was mimicked by protein phosphatase inhibitors. These results demonstrate that activation of PKC inhibits GABAA receptor function, and the results from the microsac experiments suggest that PKC-dependent phosphorylation preferentially inactivates a nondesensitized form or state of the receptor.
Mol Pharmacol 1992 Jun
PMID:Activation of protein kinase C selectively inhibits the gamma-aminobutyric acidA receptor: role of desensitization. 131 47

Many neurotransmitter receptors bind agonists with high affinity (Kd in the nanomolar range), whereas micromolar concentrations of the same agonists are required to elicit a functional effect. We have identified low affinity agonist binding sites for the gamma-amino-butyric acidA (GABAA) receptor-chloride channel under conditions normally used in 36Cl- uptake assays (a measure of receptor function). The GABAA agonist [3H]muscimol bound to a population of receptors with a Kd (2 microM) similar to its EC50 value for 36Cl- uptake. Binding was inhibited by the GABA agonist 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol and by the GABA antagonist bicuculline methiodide. A reduction in the number of [3H]muscimol binding sites (Bmax) by a thiol-modifying reagent produced a corresponding decrease in the Emax for muscimol. The benzodiazepine diazepam enhanced the potency of muscimol in ion flux experiments but did not alter the affinity of [3H]muscimol binding sites. We propose that benzodiazepines enhance GABAergic function by increasing receptor-ion channel coupling, rather than by increasing GABAA receptor affinity. These studies suggest that the study of physiologically relevant (low affinity) binding sites is necessary when examining regulation of receptors by cellular processes, drugs, and disease.
Mol Pharmacol 1992 Jun
PMID:Functionally relevant gamma-aminobutyric acidA receptors: equivalence between receptor affinity (Kd) and potency (EC50)? 131 48

We have studied the changes in the GABAergic system in the rat somatosensory cortex 1-14 d after sensory deprivation of the hind-limb representation area. Glutamate decarboxylase (GAD) activity was measured in the individual cortical layers using serial sections cut on a freezing microtome parallel to the cortical surface. Gamma-aminobutyric acid (GABA) high-affinity uptake was studied in cortical homogenates of the hind-limb representation area. There was a less than or equal to 13% decrease in GAD activity in layer II-IV in both cortical hemispheres 3 d after sciatic nerve injury. In contrast, we found that high-affinity uptake of GABA is not affected. The data mirror only small changes in GABAergic transmission, probably as a result of the methods employed. These changes correspond to electrophysiological studies suggesting that peripheral manipulation of the somatosensory system, e.g., nerve transection, is accompanied by changes in GABAergic transmission.
Mol Chem Neuropathol
PMID:High-affinity uptake of GABA and glutamate decarboxylase activity in rat primary somatosensory cortex after sciatic nerve injury. 132 1

1. The permeation of labeled Cl- across single nerve membranes microdissected from rabbit Deiters' neurons was studied in a microchamber system. The in----out permeation of the ions was evaluated under control conditions and in the presence of either 10(-6) M GABA or 10(-6) M GABA plus 10(-5) M bicuculline methiodide (BMI) on the membrane cytoplasmic side. 2. In 32 experiments, involving one animal each, at least two membranes served as controls and at least two others were studied with the addition of GABA. Within each experiment all the membranes were obtained from the same animal. 3. In an additional 10 experiments, involving one animal each, at least two membranes served as controls and at least two others were studied in the presence of GABA plus bicuculline methiodide on the membrane cytoplasmic side. 4. The data show that 10(-6) M GABA on the Deiters' membrane cytoplasmic side stimulates Cl- permeation in----out by 42% (P = 0.0000001). When 10(-5) M BMI was present together with GABA, no stimulation of Cl- in----out permeation occurred.
Cell Mol Neurobiol 1992 Aug
PMID:Further evidence for the presence of gamma-aminobutyric acidA (GABAA) receptors on the cytoplasmic side of Deiters' membrane. 132 32

Cerebral cortical cultured neurons were characterized for GABA-benzodiazepine (BZ) receptor complex, and the effect of chronic exposure of cortical neurons to GABA on GABA-BZ receptor system was investigated. In the intact cells, the [3H]flunitrazepam binding was rapid and saturable, with an apparent Kd of 4.2 +/- 1.5 nM and Bmax of 776 +/- 54 fmol/mg protein. Specifically bound [3H]flunitrazepam was displaced in a concentration-dependent manner by various BZ receptor ligands such as Ro15-1788, DMCM, Ro15-4513, clonazepam, alprazolam, diazepam and zolpidem, and enhanced by GABA, muscimol and pentobarbital. GABA induced enhancement of 36Cl-influx in a concentration-dependent manner (EC50 = 9 +/- 2 microM). Chronic exposure of the cultured neurons to GABA resulted in a reduced [3H]flunitrazepam, [3H]GABA, [3H]Ro15-1788, [3H]Ro15-4513 and [35S]TBPS binding, a reduced enhancement of [3H]flunitrazepam binding by GABA, and a reduced GABA-induced 36Cl-influx susceptible to reversal by concomitant exposure of the cultures to R 5135, a GABAA-receptor antagonist. These findings indicate that cerebral cortical cultured neurons provide an ideal model to study GABA-BZ receptor complex using binding and 36Cl-influx assays, and chronic exposure of cortical cultures to GABA leads to a down-regulation of GABA-BZ receptor system. It is a GABAA receptor-mediated slow process.
Brain Res Mol Brain Res 1992 Nov
PMID:Chronic GABA exposure down-regulates GABA-benzodiazepine receptor-ionophore complex in cultured cerebral cortical neurons. 133 97

Expression plasmids were constructed with cDNAs encoding the rat gamma-aminobutyric acid-A (GABAA) receptor alpha 1, beta 2, and gamma 2 subunits and were cotransfected into cultured human embryonic kidney 293 cells. A single cell line (WS-1) was established after G-418 treatment and clonal selection. This cell line contained saturable, high affinity binding sites for the benzodiazepines [3H] Ro 15-4513 and [3H]flunitrazepam that were modulated by GABA. Competition experiments with benzodiazepine receptor ligands suggest a profile characteristic of native "type I" benzodiazepine receptors, because strong correlations were observed between the Ki values of these ligands in WS-1 cells and in both cerebellar homogenates (r = 0.97, p < 0.0001) and 293 cells transiently transfected with the corresponding cDNAs (r = 0.96, p < 0.001). Fluorescence intensity in WS-1 cells loaded with the Cl(-)-specific probe 6-methoxy-N-(3-sulfopropyl)-quinolinium was reliably increased by GABA. This effect was blocked by bicuculline and augmented by midazolam, consistent with the presence of GABA-gated, benzodiazepine receptor-modulated, Cl- channels. Northern blot analysis revealed the presence of mRNAs encoding alpha 1 and gamma 2 receptor subunits. Southern blot analysis confirmed genomic integration of transfected alpha 1 and gamma 2 cDNAs. The beta 2 subunit was not detected in either Northern or Southern blot analysis, indicating that a functional type I GABAA/benzodiazepine receptor complex can be constituted without a beta subunit.
Mol Pharmacol 1992 Dec
PMID:Stable expression of type I gamma-aminobutyric acidA/benzodiazepine receptors in a transfected cell line. 133 19

Gamma-aminobutyric acid (GABA), the major inhibitory neurotransmitter in brain, opens chloride channels through actions on GABAA receptors. We now report base and amino acid sequences of the alpha 1, alpha 2, and alpha 3 subunits from GABAA receptors of audiogenic seizure-prone (DBA/2J) and -resistant (C57BL/6J) inbred strains of mice. Inbreeding had fixed different alleles of the alpha 1 subunit in the two strains, giving five base differences in the cDNAs. None of these affected amino acid sequence, but one did create a NsiI restriction site potentially useful in mapping genomic DNA. No base or amino acid sequence differences between the strains were detected for the other two subunits. Northern blots revealed no apparent strain differences in message levels for these three subunits in whole brains of the mice at 3 weeks of age, the peak of seizure susceptibility in DBA/2J, but did reveal distinct regional and developmental patterns of expression among the subunits in mouse brain.
J Mol Neurosci 1992
PMID:The alpha 1, alpha 2, and alpha 3 subunits of GABAA receptors: comparison in seizure-prone and -resistant mice and during development. 135 7

Incubation of rat brain synaptosomes and mitochondria with LPO inducers (Fe2+ and ascorbate) was accompanied by a decrease of deamination of serotonin (substrate of MAO-A) in mitochondria, but not in synaptosomes, with simultaneous stimulation of GABA and GLCA deamination, apparently owing to modification of catalytic properties of brain membrane-bound MAO. Oxidation of PEA (substrate of MAO-B) was insignificantly altered in both fractions. Reactions of deamination of serotonin, GABA, and GLCA (but not PEA), were highly sensitive to a selective inhibitor of MAO-A pyrazidol (pyrlindole). Isoniazid and hydrazides of quinoline carbonic acids (inhibitors of both modified MAO and copper-containing amine oxidases) strongly inhibited deamination of GABA and GLCA. During epileptiformic seizures in rats, genetically selected for high incidence of audiogenic epilepsia, stimulation in brain synaptosomes and mitochondria of LPO was observed. This was accompanied by a marked decrease in serotonin and PEA deamination, with a simultaneous increase in GABA and GLCA deamination in both fractions. The data obtained suggest that appearance of GABA-deaminating activity owing to modification of catalytic properties of MAO, might be an essential pathogenetic component in the development of epileptic seizures.
Mol Chem Neuropathol
PMID:The role of lipid peroxidation in the possible involvement of membrane-bound monoamine oxidases in gamma-aminobutyric acid and glucosamine deamination in rat brain. Focus on chemical pathogenesis of experimental audiogenic epilepsy. 152 Apr 3


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