Abstract
It is well known that ethanol modulates the function of the Cys loop ligand-gated ion channels, which include the inhibitory glycine receptors (GlyRs). Previous studies have consistently shown that transmembrane and extracellular sites are essential for ethanol actions in GlyRs. In addition, recent evidence has shown that the ethanol modulation of GlyRs is also affected by G protein activation through Gβγ subunits. However, more specific roles of G protein α subunits on ethanol actions are unknown. Here, we show that the allosteric effect of ethanol on the human α1 GlyR is selectively enhanced by the expression of Gαs Q-L. For example, constitutively active Gαs, but not Gαq or Gαi, was able to displace the alcohol sensitivity of GlyRs toward low millimolar concentrations (17 ± 4 versus 48 ± 5% at 100 mM). Experiments under conditions that increased cAMP and protein kinase A (PKA)-mediated signaling, on the contrary, did not produce the same enhancement in sensitivity, suggesting that the Gαs Q-L effect was not dependent on cAMP/PKA-dependent signaling. On the other hand, the effect of Gαs Q-L was blocked by a Gβγ scavenger (9 ± 3% of control). Furthermore, two mutant receptors previously shown to have impaired interactions with Gβγ were not affected by Gαs Q-L, suggesting that Gβγ is needed for enhancing ethanol sensitivity. These results support the conclusion that activated Gαs can facilitate the Gβγ interaction with GlyRs in presence of ethanol, independent of increases in cAMP signaling. Thus, these data indicate that the activated form of Gαs is able to positively influence the effect of ethanol on a type of inhibitory receptor important for motor control, pain, and respiration.
Footnotes
This work was supported by the National Institutes of Health National Institute on Alcohol Abuse and Alcoholism [Grant AA15150]; and Comisión Nacional de Investigación Científica y Tecnológica [Grant AT-4040102].
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
doi:10.1124/jpet.111.184408.
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ABBREVIATIONS:
- LGIC
- ligand-gated ion channel
- GABAAR
- GABA receptor
- GlyR
- glycine receptor
- TM
- transmembrane
- GPCR
- G protein-coupled receptor
- BAPTA
- 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid
- PKA
- protein kinase A
- HEK
- human embryonic kidney
- GFP
- green fluorescent protein
- ANOVA
- analysis of variance
- ct-GRK
- carboxyl-terminal G protein-coupled receptor kinase.
- Received June 6, 2011.
- Accepted August 4, 2011.
- Copyright © 2011 by The American Society for Pharmacology and Experimental Therapeutics
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