Evidence for oligomerization between GABAB receptors and GIRK channels containing the GIRK1 and GIRK3 subunits

Authors

  • Francisco Ciruela,

    1. Unitat de Farmacologia (4102), Departament Patologia i Terapèutica Experimental, Facultat de Medicina-Bellvitge, Universitat de IDIBELL-Universitat de Barcelona, L’Hospitalet de Llobregat, 08907 Barcelona, Spain
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  • Víctor Fernández-Dueñas,

    1. Unitat de Farmacologia (4102), Departament Patologia i Terapèutica Experimental, Facultat de Medicina-Bellvitge, Universitat de IDIBELL-Universitat de Barcelona, L’Hospitalet de Llobregat, 08907 Barcelona, Spain
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  • Kristoffer Sahlholm,

    1. Department of Neuroscience, Karolinska Institutet, Stockholm, Sweden
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  • Laura Fernández-Alacid,

    1. Departamento de Ciencias Médicas, Instituto de Investigación en Discapacidades Neurológicas (IDINE), Facultad de Medicina, Universidad de Castilla-La Mancha, Campus Biosanitario, C/Almansa 14, 02006 Albacete, Spain
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  • Joel C. Nicolau,

    1. Departamento de Ciencias Médicas, Instituto de Investigación en Discapacidades Neurológicas (IDINE), Facultad de Medicina, Universidad de Castilla-La Mancha, Campus Biosanitario, C/Almansa 14, 02006 Albacete, Spain
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  • Masahiko Watanabe,

    1. Department of Anatomy, Hokkaido University School of Medicine, Sapporo, Japan
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  • Rafael Luján

    1. Departamento de Ciencias Médicas, Instituto de Investigación en Discapacidades Neurológicas (IDINE), Facultad de Medicina, Universidad de Castilla-La Mancha, Campus Biosanitario, C/Almansa 14, 02006 Albacete, Spain
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Dr Francisco Ciruela, as above.
E-mail: fciruela@ub.edu
Dr Rafael Luján, as above.
E-mail: Rafael.Lujan@uclm.es

Abstract

The stimulation of inhibitory neurotransmitter receptors, such as γ-aminobutyric acid type B (GABAB) receptors, activates G protein-gated inwardly-rectifying K+ (GIRK) channels, which influence membrane excitability. There is now evidence suggesting that G protein-coupled receptors and G protein-gated inwardly-rectifying K+ [GIRK/family 3 of inwardly-rectifying K+ (Kir3)] channels do not diffuse freely within the plasma membrane, but instead there are direct protein–protein interactions between them. Here, we used bioluminescence resonance energy transfer, co-immunoprecipitation, confocal and electron microscopy techniques to investigate the oligomerization of GABAB receptors with GIRK channels containing the GIRK3 subunit, whose contribution to functional channels is still unresolved. Co-expression of GABAB receptors and GIRK channels in human embryonic kidney-293 cells in combination with co-immunoprecipitation experiments established that the metabotropic receptor forms stable complexes with GIRK channels. Using bioluminescence resonance energy transfer, we have shown that, in living cells under physiological conditions, GABAB receptors interact directly with GIRK1/GIRK3 heterotetramers. In addition, we have provided evidence that the receptor–effector complexes are also found in vivo and identified that the cerebellar granule cells are one neuron population where the interaction probably takes place. Altogether, our data show that signalling complexes containing GABAB receptors and GIRK channels are formed shortly after biosynthesis, probably in the endoplasmic reticulum and/or endoplasmic reticulum/Golgi apparatus complex, suggesting that this might be a general feature of receptor–effector ion channel signal transduction and supporting a channel-forming role for the GIRK3 subunit.

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