Abstract
gamma-aminobutyric acid (GABA) is the principal inhibitory neurotransmitter in the mammalian brain. It acts via both ionotropic GABA-A and metabotropic GABA-B receptors. We evaluated the interaction of receptors with members of the inwardly rectifying potassium (Kir 3) channel family, which also play an important role in neuronal transmission and membrane excitability. These channels are functionally regulated by GABA-B receptors. Possible physical interactions between GABA-B receptor and Kir 3 channels expressed in HEK cells were evaluated using Bioluminescence Resonance Energy Transfer (BRET) experiments, co-immunoprecipitation and confocal microscopy. Our data indicate that Kir 3 channels and Gbetagamma subunits can interact with the GABA-B(1) subunits independently of the GABA-B(2) subunit or Kir 3.4 which are ultimately responsible for their targetting to the cell surface. Thus signalling complexes containing GABA-B receptors, G proteins and Kir channels are formed shortly after biosynthesis most likely in the endoplasmic reticulum.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Blotting, Western
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Cell Line
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Fluorescence Resonance Energy Transfer
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G Protein-Coupled Inwardly-Rectifying Potassium Channels / genetics
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G Protein-Coupled Inwardly-Rectifying Potassium Channels / metabolism*
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GTP-Binding Protein alpha Subunits, Gi-Go / metabolism
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GTP-Binding Protein beta Subunits / metabolism
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Green Fluorescent Proteins / genetics
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Green Fluorescent Proteins / metabolism
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Humans
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Immunoprecipitation
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Intracellular Fluid / metabolism
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Luciferases / genetics
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Luciferases / metabolism
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Microscopy, Confocal
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Plasmids / genetics
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Protein Binding
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Protein Transport / physiology
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Receptors, GABA-B / genetics
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Receptors, GABA-B / metabolism*
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Signal Transduction / physiology
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Transfection
Substances
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G Protein-Coupled Inwardly-Rectifying Potassium Channels
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GTP-Binding Protein beta Subunits
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Receptors, GABA-B
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Recombinant Fusion Proteins
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Green Fluorescent Proteins
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Luciferases
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GTP-Binding Protein alpha Subunits, Gi-Go