184 -191, 2008). The goal of our study was to determine the role of receptors containing this subunit in alcohol action. We designed an ␣2 subunit with serine 270 to histidine and leucine 277 to alanine mutations that was insensitive to potentiation by ethanol yet retained normal GABA sensitivity in a recombinant expression system. Knockin mice containing this mutant subunit were tested in a range of ethanol behavioral tests. These mutant mice did not develop the typical conditioned taste aversion in response to ethanol and showed complete loss of the motor stimulant effects of ethanol. Conversely, they also demonstrated changes in ethanol intake and preference in multiple tests. The knockin mice showed increased ethanolinduced hypnosis but no difference in anxiolytic effects or recovery from acute ethanol-induced motor incoordination.Overall, these studies demonstrate that the effects of ethanol at GABAergic synapses containing the ␣2 subunit are important for specific behavioral effects of ethanol that may be relevant to the genetic linkage of this subunit with human alcoholism.

The mechanism by which the inhaled anesthetic isoflurane produces amnesia and immobility is not understood. Isoflurane modulates GABA A receptors (GABA A -Rs) in a manner that makes them plausible targets. We asked whether GABA A -R ␣2 subunits contribute to a site of anesthetic action in vivo. Previous studies demonstrated that Ser270 in the second transmembrane domain is involved in the modulation of GABA A -Rs by volatile anesthetics and alcohol, either as a binding site or a critical allosteric residue. We engineered GABA A -Rs with two mutations in the ␣2 subunit, changing Ser270 to His and Leu277 to Ala. Recombinant receptors with these mutations demonstrated normal affinity for GABA, but substantially reduced responses to isoflurane. We then produced mutant (knockin) mice in which this mutated subunit replaced the wildtype ␣2 subunit. The adult mutant mice were overtly normal, although there was evidence of enhanced neonatal mortality and fear conditioning. Electrophysiological recordings from dentate granule neurons in brain slices confirmed the decreased actions of isoflurane on mutant receptors contributing to inhibitory synaptic currents. The loss of righting reflex EC 50 for isoflurane did not differ between genotypes, but time to regain the righting reflex was increased in N 2 generation knockins. This effect was not observed at the N 4 generation. Isoflurane produced immobility (as measured by tail clamp) and amnesia (as measured by fear conditioning) in both wild-type and mutant mice, and potencies (EC 50 ) did not differ between the strains for these actions of isoflurane. Thus, immobility or amnesia does not require isoflurane potentiation of the ␣2 subunit.

Background-Although many people consume alcohol (ethanol), it remains unknown why some become addicted. Elucidating the molecular mechanisms of tolerance and physical dependence (withdrawal) may provide insight into alcohol addiction. While the exact molecular mechanisms of ethanol action are unclear, γ-aminobutyric acid type A receptors (GABA A -Rs) have been extensively implicated in ethanol action. The α1 GABA A -R subunit is associated with tolerance and physical dependence, but its exact role remains unknown. In this report, we tested the hypothesis that α1-GABA A -Rs mediate in part these effects of ethanol.