title: Endogenous RGS proteins enhance acute desensitization of GABA(B) receptor-activated GIRK currents in HEK-293T cells.
abstract: The coupling of GABA(B) 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 GABA(B) 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 GABA(B) receptors, in contrast to micro 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 GABA(B) 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 (GTPgammaS) 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 Galphaq G-protein subunit that has been designed to sequester endogenous RGS proteins. Finally, reconstitution of GABA(B) receptors with Galphao 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 GABA(B) receptor-dependent desensitization of GIRK currents.
authors.affiliation: The Salk Institute for Biological Studies, La Jolla, CA, 92037, USA.
authors.forename: Manpreet
authors.initials: M
authors.lastname: Mutneja
authors.forename: Frédérique
authors.initials: F
authors.lastname: Berton
authors.forename: Ka-Fai
authors.initials: KF
authors.lastname: Suen
authors.forename: Christian
authors.initials: C
authors.lastname: Lüscher
authors.forename: Paul A
authors.initials: PA
authors.lastname: Slesinger
mesh_terms: D001418:Baclofen / Q000494:pharmacology; D002460:Cell Line; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D018755:GABA Agonists / Q000494:pharmacology; D006801:Humans; D024661:Potassium Channels, Inwardly Rectifying / Q000502:physiology*; D020710:RGS Proteins / Q000502:physiology*; D018080:Receptors, GABA-B / Q000502:physiology*
chemical_list: D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D018755:GABA Agonists; D024661:Potassium Channels, Inwardly Rectifying; D020710:RGS Proteins; D018080:Receptors, GABA-B; D001418:Baclofen