Abstract
GABAA receptors (GABAARs) have long been a focus as targets for alcohol actions. Recent work suggests that tonic GABAergic inhibition mediated by extrasynaptic δ subunit-containing GABAARs is uniquely sensitive to ethanol and enhanced at concentrations relevant for human alcohol consumption. Ethanol enhancement of recombinant α4β3δ receptors is blocked by the behavioral alcohol antagonist Ro15-4513, suggesting that EtOH/Ro15-4513-sensitive receptors mediate important behavioral alcohol actions. Here we confirm alcohol/alcohol antagonist sensitivity of α4β3δ receptors using human clones expressed in a human cell line and test the hypothesis that discrepant findings concerning the high alcohol sensitivity of these receptors are due to difficulties incorporating δ subunits into functional receptors. To track δ subunit incorporation we use a functional tag, a single amino acid change (H68A) in a benzodiazepine binding residue, in which a histidine in the δ subunit is replaced by an alanine residue found at the homologous position in γ subunits. We demonstrate that the δH68A substitution confers diazepam sensitivity to otherwise diazepam-insensitive α4β3δ receptors. The extent of enhancement of α4β3δH68A receptors by 1 μM diazepam, 30 mM EtOH and 1 μM β-CCE (but not 1 μM Zn2+ block) is correlated in individual recordings, suggesting that δ subunit incorporation into recombinant GABAARs varies from cell to cell and that this variation accounts for the variable pharmacological profile. These data are consistent with the notion that δ subunit-incorporation is often incomplete in recombinant systems, yet necessary to confer high ethanol sensitivity, one of the features of native δ subunit-containing GABAARs.
- GABAA, GABAC
- Func. analysis receptor/ion channel mutants
- Structure/function/mechanism
- Alcohol
- Protein targets
- Alcohols
Footnotes
- The American Society for Pharmacology and Experimental Therapeutics