Endogenous RGS proteins enhance acute desensitization of GABAB receptor-activated GIRK currents in HEK-293T cells |
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Authors: | Manpreet Mutneja Frédérique Berton Ka-Fai Suen Christian Lüscher Paul A Slesinger |
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Institution: | (1) The Salk Institute for Biological Studies, La Jolla, CA 92037, USA;(2) Division of Biology, University of California–San Diego, La Jolla, CA 92093, USA;(3) Neurosciences Department, University of California–San Diego, La Jolla, CA 92093, USA;(4) Departments of Basic Neurosciences and Clinic of Neurology, University of Geneva, Geneva, Switzerland |
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Abstract: | The coupling of GABAB receptors to G-protein-gated inwardly rectifying potassium (GIRK) channels constitutes an important inhibitory pathway in the brain. Here, we examined the mechanism underlying desensitization of agonist-evoked currents carried by homomeric GIRK2 channels expressed in HEK-293T cells. The canonical GABAB receptor agonist baclofen produced GIRK2 currents that decayed by 57.3±1.4% after 60 s of stimulation, and then deactivated rapidly (time constant of 3.90±0.21 s) upon removal of agonist. Surface labeling studies revealed that GABAB receptors, in contrast to µ opioid receptors (MOR), did not internalize with a sustained stimulation for 10 min, excluding receptor redistribution as the primary mechanism for desensitization. Furthermore, heterologous desensitization was observed between GABAB receptors and MOR, implicating downstream proteins, such G-proteins or the GIRK channel. To investigate the G-protein turnover cycle, the non-hydrolyzable GTP analogue (GTPS) was included in the intracellular solution and found to attenuate desensitization to 38.3±2.0%. The extent of desensitization was also reduced (45.3±1.3%) by coexpressing a mutant form of the Gq G-protein subunit that has been designed to sequester endogenous RGS proteins. Finally, reconstitution of GABAB receptors with Go G-proteins rendered insensitive to RGS resulted in significantly less desensitization (28.5±3.2%). Taken together, our results demonstrate that endogenous levels of RGS proteins effectively enhance GABAB receptor-dependent desensitization of GIRK currents. |
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Keywords: | Desensitization GABA GIRK Kir3 Potassium channels RGS proteins |
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