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Logo of nihpaAbout Author manuscriptsSubmit a manuscriptHHS Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
Alcohol. Author manuscript; available in PMC 2007 October 22.
Published in final edited form as:
PMCID: PMC2040048

Ethanol acts directly on extrasynaptic subtypes of GABAA receptors to increase tonic inhibition


Based on the similarity of ethanol intoxication to the behavioral effects of drugs known to target GABAA receptors (GABARs) it has been suspected for decades that ethanol facilitates the activity of GABA. Even so, it has been surprisingly difficult to identify molecular targets of ethanol. Research conducted over the past several years suggests that a subclass of GABARs (those containing δ subunits) responds in a relevant concentration range to ethanol. Although δ subunit-containing GABARs are not ubiquitously expressed at inhibitory synapses like their γ subunit-containing, synaptic counterparts, they are found in many neurons in extrasynaptic locations. Here they give rise to a tonic form of inhibition that can potently suppress neuronal excitability. Studies have shown that both recombinant and native δ subunit-containing GABARs: 1) are modulated by behaviorally-relevant (i.e. low millimolar) concentrations of ethanol, 2) directly bind ethanol over the same concentration range, 3) show altered function upon single amino substitutions linked to changes in behavioral responsiveness to ethanol, and 4) are a site of action of Ro15-4513, a competitive antagonist of ethanol binding and a drug which prevents many of the behavioral aspects of ethanol intoxication. Despite such comprehensive evidence, however, the field is not free from controversy. This review evaluates published data for and against a central role of δ subunit-containing GABARs in ethanol actions and suggests future directions that might help settle points of controversy.

Keywords: alpha6 subunit, alpha4 subunit, delta subunit, alcohol, cerebellum, tonic inhibition


Determining the precise molecular target of a psychoactive drug is notoriously difficult and for few drugs has this been more challenging than for ethanol. Considering that ethanol is the most widely used drug in society and that ethanol abuse is by far the most common form of substance abuse, understanding where and how this compound acts in the brain is one of the key challenges of neuroscience.

It was first recognized decades ago that ethanol shares a pharmacological profile with drugs known to act on GABARs, however, experimental evidence that ethanol exerts effects on specific molecules involved in GABAergic signaling has been limited and controversial. Indeed, since the first published account suggesting that a specific GABAR subunit (γ2L) is required for ethanol sensitivity (Wafford et al., 1991), findings in this field have been difficult for other labs to replicate and there is currently no consensus regarding specific molecules at which ethanol might act in exerting its effects on behavior.

There are several reasons why progress in identifying targets has been slow and at times contentious. First, the actions of ethanol in the brain occur at unusually high concentrations. Behavioral signs of intoxication appear only when millimolar levels of ethanol are present in blood, implying a weak receptor-drug interaction. Such low affinity renders traditional biochemical approaches, so useful in defining molecular sites of action, impractical. Second, most candidate targets for ethanol are ion channels which are complex molecules. Native GABARs (whose role in ethanol effects is the subject of this review) exist as heteropentamers composed of combinations selected from19 homologous subunits. While stoichiometry and general rules for biosynthesis of the most common isoforms of native GABARs are becoming clearer (2 α subunits, 2 β subunits, and either a γ or δ subunit are thought to be required in the pentamer), many uncertainties remain. Third, even if the precise molecular makeup of a native GABAR were established, reconstitution of native GABARs with the desired subunit composition and spatial arrangement of subunits is problematic. Standard recombinant expression methods may not assure that multiple subunits will assemble with one another in their native spatial confirmations. Of course, issues such as accessory subunits, posttranslational modifications, and regulation by second messengers complicate matters further.

This review begins by summarizing positive evidence that δ subunit-containing GABARs are key targets for low (<30 mM) concentrations of ethanol. Much of this data has been obtained from complimentary studies of recombinant and native GABARs. Published objections to these findings are then discussed and, where possible, attempts are made to reconcile the negative findings with the supportive evidence. The review concludes with several suggestions for future experiments that might help clarify outstanding issues.

Evidence that ethanol acts on δ subunit-containing GABARs

In recombinant studies, low concentrations of ethanol enhance the function of extrasynaptic GABAR subtypes

A number of studies have shown that recombinant GABARs composed of specific combinations of subunits are special in that they respond to ethanol within a dosage range corresponding to blood ethanol concentrations achieved during moderate social consumption (Hanchar et al., 2005; Sundstrom-Poromaa et al., 2002; Wallner et al., 2003). The most common isoforms of GABARs contain γ subunits, and these receptors respond to ethanol at concentrations greater than 100 mM if at all (Wallner et al., 2006b). By contrast, GABARs composed from α1, α4, or α6 subunits, in combination with β3 subunits, and a δ subunit are uniquely ethanol-sensitive. GABA-activated currents mediated by recombinant receptors of this composition are enhanced by 30 to 50% by 10 mM ethanol and by more than 100% by 100 mM ethanol (Hanchar et al., 2005; Wallner et al., 2003). An example of current responses and ethanol dose response curves for GABARs composed of α6β3δ and α6β2δ is reproduced in Figure 1. Also displayed in this figure is the effect of a single amino acid variation in α6 found to occur naturally in rats. This polymorphism makes GABARs containing it even more sensitive to ethanol but only when composed with β3 and δ subunits (Hanchar et al., 2005). Cellular and behavioral ramifications of the α6 polymorphism make a strong case that such receptors function under physiological circumstances and will be discussed in more detail in sections 2.15 and 2.16 below.

Figure 1
GABARs composed of α6, β3- and δ subunits are sensitive to low millimolar ethanol concentrations

In neurons, low concentrations of ethanol enhance tonic but not synaptic inhibition

While the most abundant GABAR subunits of the α, β and γ subfamilies are found within symmetric postsynaptic densities in EM studies, δ subunits are confined to extrasynaptic membranes (Nusser et al., 1999; Nusser et al., 1998b; Peng et al., 2002; Wei et al., 2003). Here they are believed to be a defining component of the ‘extrasynaptic’ GABARs responsible for tonic inhibition (Farrant and Nusser, 2005). Numerous studies of knockout mice have suggested that GABARs containing α1/δ, α4/δ, or α6/δ are required for tonic inhibition (Brickley et al., 2001; Glykys et al., 2007; Nusser et al., 1999; Stell et al., 2003).

This physiological role, along with the high sensitivity of β3/δ subunit-containing GABARs, leads to the prediction that tonic GABA currents, rather than phasic (i.e., synaptic) GABA currents, should be enhanced by low concentrations of ethanol. Indeed, in three neuronal types known to express β3 and δ subunits, this is the case. Cerebellar granule cells (Hanchar et al., 2005), dentate gyrus granule cells (Fleming et al., 2007; Liang et al., 2006; Wei et al., 2004), and hippocampal interneurons (Glykys et al., 2007) exhibit tonic currents that are robustly enhanced by ethanol. These specific effects on tonic GABA currents have been observed for concentrations as low as 10 mM ethanol (Hanchar et al., 2005). To contrast, no enhancement of GABAR-mediated synaptic responses is observed at ethanol concentrations below 100 mM.

In certain brain areas, ethanol can also lead to increases in GABA release (Carta et al., 2004; Hanchar et al., 2005) which might increase tonic GABA current by an indirect, presynaptic rather than a direct action on extrasynaptic GABARs on the postsynaptic cell. However, even under conditions that prevent the presynaptic increases in GABA release, there is evidence that ethanol acts directly on native, extrasynaptic GABARs containing α6 subunits. In the presence of tetrodotoxin which inhibits presynaptic activity and under conditions in which GABA levels are stabilized (0.3 μM added GABA plus GABA uptake blockers), the effects of ethanol persist (see Figure 2). Moreover, ethanol effects on tonic GABA currents are significantly larger in animals homozygous for the ethanol hypersensitive polymorphism in the α6 subunit (Hanchar et al., 2005). Given that this gene is expressed only in the postsynaptic granule cell, such effects must be postsynaptic. Taken together such results strongly support the conclusion that ethanol directly affects extrasynaptic GABARs on cerebellar granule cells.

Figure 2
Low millimolar ethanol concentrations enhance tonic GABA current

Ethanol binds to recombinant and native δ subunit-containing GABARs

Arguably, the most straightforward evidence that a receptor - drug interaction occurs comes from biochemical experiments demonstrating binding of the drug to the receptor. Until recently it seemed unlikely that such evidence would be gathered for ethanol because of its relatively low apparent affinity for targets. However, Hanchar and colleagues unexpectedly discovered that a benzodiazepine derivative (Ro15-4513) binds with high affinity (Kd ~ 7 nM) to both recombinant and native, immunopurified β3/δ subunit-containing GABARs (Hanchar et al., 2006). How was this finding relevant to ethanol actions? First, it suggested that a non-classical, and therefore largely uncharacterized, benzodiazepine binding site was present on extrasynaptic GABARs. Second, although the underlying mechanisms were mysterious and somewhat controversial, Ro15-4513 had been reported to reverse many aspects of ethanol intoxication at a behavioral level (Paul, 2006; Suzdak et al., 1986). Given its high affinity binding to putative ethanol targets and the availability of Ro15-4513 in radiolabeled form, binding experiments made it possible to test whether ethanol was capable of displacing this compound when it was bound specifically to δ subunit-containing GABARs.

Remarkably, experiments showed that Ro15-4513 is displaced in a competitive and saturable manner by ethanol with a Kd of 8 mM (Hanchar et al., 2006; Wallner et al., 2006a). This was found to be the case for recombinant GABARs composed with α4, β3 and δ subunits, as well as immunopurified GABARs pulled down with an antibody to the δ subunit, but not for GABARs containing γ subunits (even though Ro15-4513 binds to them with roughly equal affinity). Intriguingly, flumazenil (a.k.a. Ro15-1788), the clinically important benzodiazepine site antagonist, was also found to bind to δ subunit-containing GABARs with high affinity and to displace Ro15-4513. However, flumazenil could remain bound in the presence of high ethanol concentrations. As illustrated in Figure 3, flumazenil and Ro15-4513 share a very similar chemical structure differing only in one position where a fluorine atom (flumazenil) takes the place of a larger three nitrogen atom azido group (Ro15-4513). Based on structure activity considerations, Hanchar et al. speculate that ethanol occupies the position where the azido group on Ro15-4513 interacts with the receptor. Thus, ethanol is able to bind to the GABAR at the same time as flumazenil but is competitively displaced by Ro15-4513. Coupled with the strong evidence that amino acid position 100 in α6 is a critical residue for benzodiazepine binding (Korpi et al., 1993; Mohler et al., 2001), and the effects on ethanol modulation of the ethanol hypersensitive polymorphism at this position (Hanchar et al., 2005), these results provide a specific hypothesis for where ethanol binds to this class of GABARs. They also strengthen the idea that ethanol acts directly on δ subunit-containing GABARs rather than through an unidentified, intermediate protein.

Figure 3
Ro15-4513 antagonizes ethanol effects on recombinant GABARs containing β3 and δ subunits and this antagonism is prevented by flumazenil or β-CCE

The ethanol antagonist prevents ethanol enhancement of recombinant and native GABARs

At a functional level, the binding data make several clear predictions: 1) Ro15-4513 should prevent ethanol enhancement of δ subunit-containing GABAR currents, 2) flumazenil should not interfere with this ethanol modulation, and 3) although inactive on its own, flumazenil should relieve the ability of Ro15-4513 to act as an ethanol antagonist. Figure 3 shows that all three predictions are found to be true when tested on α4/β3/δ subunit-containing GABARs expressed in oocytes (Wallner et al., 2006a). The β carboline β-CCE, another competitive, but structurally dissimilar ligand at classical benzodiazepine sites, was also found to antagonize the actions of Ro15-4513 in both binding (Hanchar et al., 2006) and electrophysiological experiments (Wallner et al., 2006a). Finally, a point mutation in a transmembrane region of the β3 subunit abolished the high dose (>100 mM) actions of ethanol, leaving a dose response relationship essentially identical to the binding curves reported in the accompanying study (Wallner et al., 2006a).

Is ethanol enhancement of native, extrasynaptic GABARs also prevented by Ro15-4513? We find that in cerebellar granule cells from α6100Q/Q rats the antagonist of ethanol binding does block ethanol modulation. Figure 4 summarizes previously unpublished results from 9 cells. In the presence of 300 nM added GABA, 30 mM ethanol causes a 31±7% increase in tonic GABA current. Coapplication of 100 nM Ro15-4513 and ethanol to the same cells results in no change in tonic current (1.9±9% increase) (p<0.01, ethanol alone vs. ethanol plus Ro15-4513). Application of 100 nM Ro15-4513 by itself to a separate group of 6 cerebellar granule cells from α6100Q/Q rats caused a significant increase in tonic current (29.2±8.8 %, n=6, p<0.004 as compared to no change), The finding that Ro15-4513 by itself increases tonic current argues that potentially confounding inverse agonist actions of this compound are not of concern. Thus, in line with previous work on acutely isolated and cultured neurons (Reynolds et al., 1992), we find that native α6/δ subunit-containing GABARs are also sensitive to the ethanol site antagonist.

Figure 4
The ethanol site antagonist Ro15-4513 blocks the ethanol-induced enhancement of tonic GABA current in cerebellar granule cells

These actions of Ro15-4513, flumazenil, and β-CCE in the binding and functional assays are striking. Not only do they provide specific information about a millimolar affinity ethanol binding site on a specific subtype of GABARs, but they shed light on the unexplained features of Ro15-4513 activity that have been reported in numerous behavioral studies. For example, the effects of low doses of ethanol on anxiety, memory, motor behavior, and self-administration have all been reported to be reduced by this drug (discussed in (Paul, 2006). Also explained is the puzzling finding that the benzodiazepine site antagonist flumazenil prevents the anti-intoxication actions of Ro15-4513 even though by itself it is without any effect on behavior (Suzdak et al., 1986).

A single amino acid change linked to ethanol hypersensitivity in vivo renders recombinant and native GABARs more ethanol sensitive in vitro

Some of the figures and text shown to this point include data on a genetic polymorphism in the α6 subunit of the GABAR. This single nucleotide difference in the gene encoding α6 gives rise to two alleles which differ in the amino acid residue at position 100 in the α6 protein (position 100 is either an arginine or a glutamine). Previous work had linked one of these alleles (α6100Q) to behavioral ethanol hypersensitivity (Korpi et al., 1993). Korpi and colleagues found that “alcohol non-tolerant” or ANT rats, a line selectively bred to be more sensitive to the motor impairing effects of ethanol, carried the α6100Q allele while their “alcohol tolerant” or AT counterparts carried the α6100R allele. Subsequent studies with other independently derived lines (e.g., “Sardinian non-preferring” rats) that had been selectively bred for enhanced sensitivity to ethanol, also uncovered an accumulation of the α6100Q allele (Saba et al., 2001). Interestingly, this amino acid change renders GABARs composed with α6 and γ2 much more sensitive to benzodiazepines as the arginine residue in this position prevents benzodiazepine sensitivity in α4 and α6 GABARs (Korpi et al., 1993; Korpi and Seeburg, 1993; Mohler et al., 2001; Santhakumar et al., 2006). However, at the time of the original discovery of the polymorphism, the connection between this single amino acid substitution and increased behavioral sensitivity to ethanol was unclear.

Using the Xenopus oocyte expression system, we found that when the α6100Q allele is expressed in combination with β3 and a δ subunit, the resulting GABAR is 2-3 fold more sensitive to ethanol (Hanchar et al., 2005). The increased sensitivity required that β3 and δ be expressed since the two alleles of α6 behave identically in other subunit combinations (Figure 1). Thus, the inability to link α6100Q to increased ethanol sensitivity in previous studies was likely due to the difficulty of expressing δ subunit-containing GABARs.

But does the polymorphism lead to alteration in the ethanol sensitivity of native GABARs? The only neurons in the nervous system that express both α6 and δ are cerebellar granule cells (CGCs) and tonic GABA currents in CGCs are known to rely on extrasynaptic GABARs composed of α6 and δ subunits (Brickley et al., 2001; Nusser et al., 1999; Nusser et al., 1995; Nusser et al., 1998b; Stell et al., 2003). Therefore, to examine the α6 alleles one would first need to determine whether tonic GABA currents from CGCs in brain slices were sensitive to ethanol, and then develop a strategy to test the effects of the single amino acid difference in intact neurons. With this logic we had originally intended to compare ethanol sensitivity of tonic GABA current in CGCs from ANT and AT rats. As we were developing our genotyping protocol we discovered to our surprise that the α6100Q allele has a high abundance in Sprague Dawley rats. To date we have genotyped 68 rats obtained from Charles River Laboratories and find that the α6100Q allele in this particular rat population has a frequency of 52%. Genotypic distributions are consistent with Mendelian inheritance. Of the 68 animals we have found 17 α6100R/R rats, 31 heterozygotes, and 20 α6100Q/Q rats. Indeed, the α6100Q allele is found in several lines of rats (Carr et al., 2003); in fact, the codon for this allele is in the Rat Genome Sequence Consortium database (Gibbs et al., 2004). Thus, rather than being a spontaneous mutation the α6100Q allele is an existing allele that has somehow been concentrated into the alcohol-hypersensitive groups in ANA,ANT, and Sardinian non-preferring lines by selective breeding (Carr et al., 2003; Korpi et al., 1993; Saba et al., 2001). In contrast to α6100Q, the alcohol-hyposensitive α6100R allele seems to be concentrated in the alcohol insensitive rats in all three studies. If these polymorphisms in α6 are unrelated to the behavioral alcohol phenotypes it is unclear why they would be segregated by the selective breeding.

The fortuitous discovery that the α6100Q allele is naturally abundant makes it possible to breed heterozygous animals and compare littermate offspring homozygous for the α6100R versus α6100Q alleles. Of course, such a strategy cannot completely rule out the remote possibility that unknown, cosegregating genetic factors influence ethanol sensitivity, but it is clearly superior to comparisons of populations isolated from one another by more than 40 generations of selective breeding. It seems likely that such groups harbor additional genetic differences.

Electrophysiological analysis of CGCs in brain slices from the two genotypes (α6100R/R and α6100Q/Q) showed that tonic GABA currents were enhanced by low millimolar concentrations of ethanol in both genotypes, and that significantly more enhancement was observed in slices from α6100Q/Q rats (Hanchar et al., 2005). As mentioned above, this was true even under conditions assuring that ethanol had no presynaptic effects to increase ethanol release (Figure 2). The genotypic difference in ethanol sensitivity of tonic currents indicates that at least some of the extrasynaptic GABARs in CGCs must be of the subunit composition α6β3δ.

In a subsequent study we also examined benzodiazepine sensitivity of synaptic currents in animals homozygous for these two alleles (Santhakumar et al., 2006). As predicted, analysis of spontaneous inhibitory postsynaptic currents (sIPSCs) in CGCs from α6100R/R and α6100Q/Q rats clearly revealed an enhanced benzodiazepine sensitivity of synaptic GABARs in α6100Q/Q slices. These findings confirm that some synaptic GABARs also contain α6 subunits. Furthermore, in analogy to our hypothesis regarding ethanol, the data offer a straightforward cellular mechanism for the enhanced behavioral sensitivity to benzodiazepines that has been thoroughly documented in ANT rats (Eriksson and Rusi, 1981; Hellevuo et al., 1989; Wong et al., 1996).

As mentioned earlier, ruling out presynaptic effects is particularly important because in the cerebellum ethanol causes a robust increase in GABA release if Golgi neurons are able to fire action potentials. Such ethanol-induced increases in GABA release can be observed as increases in the frequency of spontaneous but not miniature IPSCs recorded in CGCs (Carta et al., 2004; Hanchar et al., 2005). Intriguingly we found that the magnitude of increase in sIPSC frequency triggered by ethanol was significantly larger in the α6100Q/Q animals (Hanchar et al., 2005). This suggested to us that the mechanism by which ethanol increases GABA release involves α6 subunit-containing GABARs. In this way, the results also implicate a circuit mechanism whereby ethanol can increase GABA release by first acting on δ subunit-containing GABARs. We are actively pursuing this hypothesis.

Rats homozygous for the ethanol-hypersensitive allele in α6 are more severely impaired by low doses of ethanol in motor tasks

The much simpler genetic context provided by the α6100R/R and α6100Q/Q groups also made it possible to test whether differences in ethanol intoxication resulted from this amino acid substitution. Given that this polymorphism occurs in a gene expressed exclusively within a single class of cerebellar neurons we reasoned that cerebellum-dependent behaviour would be primarily affected. Furthermore, the dose dependence suggested from recombinant receptor recordings and receptor binding assays suggested that any differences would be most prominent at very low, i.e. mildly intoxicating, doses of ethanol. We found that very low doses of ethanol which leave α6100R/R rats unaffected significantly impaired the α6100Q/Q animals in the rotarod test, a classic assay for cerebellum-dependent motor coordination (Hanchar et al., 2005). Blood-alcohol concentration (BAC) assays performed on these animals showed that there was no difference in plasma concentration of ethanol between the genotypes; these measurements also showed that the dose of 1g/kg yielded a BAC of approximately 15 mM. Notably, the genotype-dependent differences in rotarod performance disappeared at higher doses (>1.5 g/kg) of ethanol. In addition to showing that the polymorphism correlates with a higher behavioral sensitivity to ethanol, these data also argue that tonic GABA currents in cerebellar granule neurons can have a profound effect on motor behavior.

Objections to the hypothesis that δ subunit-containing GABARs contribute to low dose ethanol actions in brain

Despite the comprehensive, cross-confirmatory evidence reviewed above, four papers have reported unsuccessful attempts to extend these results (Borghese et al., 2006; Casagrande et al., 2007; Valenzuela et al., 2005; Yamashita et al., 2006). Upon simple inspection, these failures seem impossible to reconcile with the extensive evidence implicating δ subunit-containing GABARs in ethanol actions. We remain puzzled by these reports as we have repeated all of the key experiments (many are replicated by different experimenters in subsequent publications) and find the results to be identical to those in our original reports. Of course, it is important to point out that failed experiments are difficult to interpret and even more difficult to troubleshoot when they have been done in another lab. That said, below we carefully review the methods and data in the prior studies identifying differences between those that yielded positive results from those that report negative results. In some instances the differences can clearly account for why experiments failed while in others we are left to speculate.

Recombinant experiments with δ subunit-containing GABARs

Two publications (Borghese et al., 2006; Yamashita et al., 2006) report that β3/δ subunit-containing GABARs are not sensitive to ethanol in the low millimolar range of concentrations. The first was a collaborative effort from several groups in which they examined transient expression of rat and human isoforms of α4/β3/δ GABARs in Xenopus oocytes and made use of L(tk-) cells constructed with the intent that α4, β3, and δ subunits be stably expressed (Borghese et al., 2006). We view their main conclusion as questionable in several respects. First, although they claim to have replicated our prior work, there are clear indications that the GABARs they studied are not the same GABARs as in the positive studies described above. Basic parameters of the expressed receptor pool - the average sizes of current, EC50 values, and relaxation dynamics upon GABA application - are notably different. For example, in the Borghese et al. study, the average GABA-induced current recorded 8 days after injection of α4/β3/δ into oocytes was 3800 nA, nearly as large a steady state current as they are able to record from α4/β3/γ2 subunit-containing receptors. In contrast, the papers reporting positive effects of ethanol observed approximately 400 nA of current 7-14 days after injection of the α4/β3/δ combination in oocytes (Hanchar et al., 2005; Wallner et al., 2003; Wallner et al., 2006a), much less current than can be recorded from α4/β3/γ2 GABARs (Hanchar et al., 2005; Wallner et al., 2003; Wallner et al., 2006a). EC50 values for GABA also differ for the α4β3 isoforms (0.6 μM for Borghese; 22 μM for Wallner). Lastly, the extent of current relaxation to less than 40% of the peak current (see Figure 5) is much greater than is observed in the studies that report low dose ethanol effects (Fig. 1, this paper; Fig. 3, Wallner et al., 2003; Figs. Figs.11 and and2,2, Wallner et al., 2006). How might such discrepancies explain why GABARs in the Borghese study are less sensitive? Although it cannot be established with certainty, the differences raise serious doubt about whether Borghese and colleagues actually replicated the conditions in the previous work. The strong enhancement that certain modulators exert on δ subunit-containing GABARs is clearly related to the low efficacy (i.e. maximal open probability) of these GABAR subtypes (Bianchi and Macdonald, 2003; Wallner et al., 2003). It remains possible that the extremely large currents observed in the Borghese study reflect receptors in a higher efficacy state. Alternatively, despite the claims of the paper, the subunit composition they have expressed may not be the same as in those studies reporting ethanol sensitive GABARs.

Figure 5
The example trace showing “ethanol insensitivity” in Borghese et al., 2006

Curiously, the rat and human isoforms of GABARs within the Borghese study are also quite different from one another with regard to some of these basic parameters. Rat and human α4β3δ GABARs are expected to show identical functional properties because the sequences are approximately 95% identical in protein sequence, with almost all differences found in the poorly conserved M3-M4 intracellular loops. Nonetheless, the human channels studied by Borghese et al. give rise to forty times smaller currents (Storustovu and Ebert, 2006). The basis for this large difference is unclear but it calls into question whether the rat and human GABAR isoforms, which have been studied in the same expression system, have the same subunit composition.

Even ignoring such discrepancies, it is strange that the authors reject the hypothesis that β3/δ subunit-containing GABARs are not responsive to low doses of ethanol when their own figure shows otherwise. Figure 1A in Borghese et al. (reproduced as Figure 5 here) illustrates clear increases in the GABA evoked current in response to 10 and 30 mM ethanol. Such effects apparently were not observed in every recording and are not reflected in the average, but their presence raises the possibility that a fraction of the GABARs in their study were ethanol-sensitive.

The recombinant experiments in the Yamashita et al. paper focused on GABARs composed with α6, β3, and δ subunits. In our view this study has an obvious technical shortcoming: they used a 2:2:1 ratio of α/β/δ cDNAs in the transfections. In order to ensure adequate expression of δ subunit-containing GABARs, we and others transfect/inject with a five- to ten-fold abundance of δ cDNA or cRNA (Borghese et al., 2006; Hanchar et al., 2006; Hanchar et al., 2005; Storustovu and Ebert, 2006; Sundstrom-Poromaa et al., 2002; Wallner et al., 2003; Wallner et al., 2006a). Indeed, even for γ2 subunit-containing GABARs, transfections must be conducted with an excess of γ2 cRNA in order to ensure that all GABARs incorporate this subunit (Boileau et al., 2002; Minier and Sigel, 2004). Thus, the lack of ethanol-sensitivity in the recombinant experiments in the Yamashita paper could simply be due to a lack of δ subunits in the receptors.

Native, extrasynaptic GABARs are not directly responsive to ethanol

Four reports have argued that native, δ subunit-containing GABARs are not directly sensitive to ethanol (Borghese et al., 2006; Carta et al., 2004; Casagrande et al., 2007; Yamashita et al., 2006). Borghese et al. show data that ostensibly conflict with previously and subsequently published reports demonstrating a sensitivity to 30 mM ethanol of tonic GABA current in dentate gyrus granule cells (Fleming et al., 2007; Glykys et al., 2007 Suzdak, 1986 #23; Liang et al., 2006; Wei et al., 2004). Comparison of these studies shows minor methodological differences, but no indication of why Borghese failed to observe sensitivity in the native GABARs. In the studies reporting sensitivity, ethanol modulation of tonic current was observed in slices from adult mice under conditions in which tonic GABA concentrations were purposefully elevated in the slices (Glykys et al., 2007; Wei et al., 2004) and in adult rats with no added GABA (Fleming et al., 2007; Liang et al., 2006). In the negative report, slices from younger mice were used, and extracellular GABA concentrations were not manipulated (Borghese et al., 2006).The recent report by Fleming et al., (2007) finds a large effect (~100% increase) of 30 mM ethanol on tonic current in adolescent dentate gyrus granule cells.

It is apparent that basic aspects of the mIPSCs are unusual in the Borghese study. The average quantal size (mIPSC amplitude) is approximately 50% larger than those recorded in a wide range of studies under similar conditions in rats (Harney et al., 2003; Naylor et al., 2005; Nusser et al., 1998a; Otis et al., 1994; Otis and Mody, 1992a; Otis and Mody, 1992b; Overstreet et al., 2000; Shao and Dudek, 2005) and mice (Glykys et al., 2007; Payne et al., 2006; Stell et al., 2003; Wei et al., 2004). This cannot be explained by elevated GABA concentrations leading to desensitization of synaptic GABARs because GABA concentrations were not manipulated in most of these other studies. Decay rates of mIPSCs are also considerably slower than in previous studies. While these features do not point to an obvious explanation for the different results, they do indicate significant, perhaps critical differences between the preparations.

In two studies, authors examined tonic GABA currents on cultured granule cells and found variable but weak responsiveness to ethanol (Casagrande et al., 2007; Yamashita et al., 2006), one of these studies suggested that this seemed to depend on whether recordings were made in whole-cell or perforated-patch mode(Yamashita et al., 2006). In several respects it is difficult to reconcile these results with those obtained in slices (Hanchar et al., 2005; Santhakumar et al., 2006) where it is clear that tonic current is mediated by GABARs composed with α6 and δ (Brickley et al., 2001; Santhakumar et al., 2006; Stell et al., 2003). Certainly it cannot be assumed that in the culture dish a mature complement of GABARs develops normally during the 7-14 day time period these cells spend in vitro. In fact, in vivo, the expression of α6 subunits has not reached adult levels until after postnatal day (PND) 35 (Brickley et al., 2001). Reflecting this, tonic GABA currents increase at least five-fold between PND7 and PND35 (Brickley et al., 1996; Brickley et al., 2001; Wall and Usowicz, 1997). Thus, even assuming development in the culture dish proceeds as in the brain, the tonic current has only reached one-third of its adult level in the oldest cultures studied.

Yet another problem is that culture conditions dramatically alter the expression levels of both α6 and δ subunits (Martikainen et al., 2004; Salonen et al., 2006). Even the ability to detect δ subunit protein on Western blots does not indicate that it is functional because in culture much of the protein is sequestered intracellularly (Follesa et al., 2005). Despite these concerns, the authors in the studies performed no direct assays for the presence of δ or α6 subunits (Casagrande et al., 2007; Yamashita et al., 2006). While Yamashita et al., did perform experiments with the α6 selective drug furosemide, this compound is not selective for δ subunit-containing GABARs (Tia et al., 1996) and so the sensitivity could simply indicate the presence of α6/γ2 subunits.

A third paper recorded from cerebellar granule cells in acute slices and described robust increases in tonic current in response to doses of 20 mM ethanol and higher (Carta et al., 2004). However, contrary to the interpretation of Hanchar et al., they attribute this enhancement of tonic current entirely to an increase in GABA release. The key finding leading them to this interpretation is that they do not detect ethanol-induced increases in tonic current in the presence of tetrodotoxin which blocks the stimulatory effect of ethanol on the presynaptic GABA-releasing neuron.

Several of our own results conflict with this picture. First, we find that increases in tonic current can be detected in tetrodotoxin (Hanchar et al., 2005). Second, rats homozygous for the ethanol-hypersensitive allele of α6, in addition to exhibiting increased benzodiazepine sensitivity for synaptic receptors (Santhakumar et al., 2006), also show larger increases in tonic current in response to ethanol (Hanchar et al., 2005). Since α6 is expressed only in granule cells and the mutation leads to an increased ethanol sensitivity in recombinant receptors, it seems most plausible that ethanol is exerting a direct postsynaptic effect. Third, we found that the magnitude of increase in GABA release (measured by changes in sIPSC frequency) was larger in animals homozygous for the ethanol hypersensitive allele (Hanchar et al., 2005), suggesting that even the presynaptic effect may arise indirectly from ethanol action on an α6/δ subunit-containing GABAR. Finally, Ro15-4513, the competitive antagonist at the ethanol binding site, blocks the ability of ethanol to increase the tonic current (Figure 4).

Why were Carta et al. unable to see this increase in TTX? Here we can only speculate, but, once again, differences in the experimental conditions may be critical. For the experiments in tetrodotoxin we added 300 nM GABA and inhibited GABA uptake in order to increases the size of the baseline tonic current and prevents fluctuations in GABA concentration. We reasoned that these manipulations would ensure a sizable tonic current and be more likely to reveal the effects on GABARs if the receptors were intrinsically sensitive to ethanol. In part we feel this is justified because tonic GABA current in brain slices is almost certainly smaller than tonic currents in vivo (Chadderton et al., 2004; Santhakumar et al., 2006). In any case, this positive result, coupled with the new data presented in Figure 4, has a more straightforward interpretation than the negative one.

The ethanol hypersensitive mutation has no effect on native GABARs

In response to our report comparing α6100R/R and α6100Q/Q rats, the same group went on to test ethanol effects in two lines of rats selectively bred to be differentially sensitive to ethanol (Valenzuela et al., 2005). These lines were chosen because they accumulate the two α6 alleles such that ANT rats are typically homozygous for the ethanol hypersensitive allele, α6100Q, while AT rats are homozygous for α6100R (Korpi et al., 1993). They report that in these two groups of rats ethanol increases tonic current in cerebellar granule cells to the same extent. They also saw no difference in the extent to which ethanol increases sIPSC frequency (i.e. GABA release).

Previously, we responded in some detail to these findings (Otis et al., 2005), and so here it is appropriate to summarize our main objection: given the likelihood that many other ethanol-related genes segregate during the 40 generations of selective breeding, this is a suspect test of whether the α6 polymorphism affects ethanol sensitivity of a native GABAR. By comparison our strategy allowed us simply to distinguish rats by genotype, breed heterozygotes, and test littermates homozygous for either allele. This strikes us as a more easily interpreted experiment. Indeed, this approach has shown us that the genotypes differ at a cellular level in their responsiveness to benzodiazepines (Santhakumar et al., 2006) as well as to ethanol (Hanchar et al., 2005) and we conclude that these cellular mechanisms play important roles in the different behavioral responsiveness to benzodiazepines (Wong et al., 1996) and ethanol (Hanchar et al., 2005). It should also be noted that, unlike the Valenzuela study, our experiments provides an explanation of why the two α6 alleles co-segregate so reliably during the selective breeding in several lines of rats (Carr et al., 2003; Korpi et al., 1993; Saba et al., 2001). If α6100Q has nothing to do with ethanol sensitivity it is puzzling why it accumulates in ethanol hypersensitive and is excluded from ethanol hyposensitive rats in various breeding paradigms.

Suggestions for future studies

While a review is rarely able to settle all outstanding issues, especially with regard to a complicated and controversial topic of this sort, such an effort does hold the possibility of clarifying key issues and suggesting critical future experiments. On the bright side, there is detailed, substantial, and accumulating evidence that extrasynaptic GABARs are responsive to low doses of ethanol. In addition, the negative findings reviewed here, although confusing, have come from experiments that have not replicated conditions in the original experiments. Indeed, in many cases there are specific concerns that provide straightforward explanations for why the experiments were not successful. But the important question remains: what can be done to settle the issue of whether δ subunit-containing GABARs play a key role in ethanol actions?

Here we have a few suggestions. First, care must be taken in recombinant studies to ensure the molecular composition of GABARs is as intended. Ideally this would involve a tag accessible only when the receptor is present on the surface membrane. Second, analysis of the ethanol sensitivity of native GABARs, along with determination of their molecular makeup, should be a priority. Third, in vivo manipulations of key components of hypothesized ethanol-sensitive GABARs subtypes should be attempted and thoroughly studied. Special attention should be paid to mildly intoxicating concentrations of ethanol in behavioral analysis of such animals. Considering the success that knock-in genetic strategies have shown in delineating the molecular mechanisms of benzodiazepine and anesthetic sensitivity (Mohler et al., 2001), there is reason to be hopeful that similar approaches will prove fruitful in understanding ethanol action.


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  • Bianchi MT, Macdonald RL. Neurosteroids shift partial agonist activation of GABAA receptor channels from low- to high-efficacy gating patterns. J. Neurosci. 2003;23:10934–10943. [PubMed]
  • Boileau AJ, Baur R, Sharkey LM, Sigel E, Czajkowski C. The relative amount of cRNA coding for γ2 subunits affects stimulation by benzodiazepines in GABAA receptors expressed in Xenopus oocytes. Neuropharmacology. 2002;43:695–700. [PubMed]
  • Borghese CM, Storustovu S, Ebert B, Herd MB, Belelli D, Lambert JJ, Marshall G, Wafford KA, Harris RA. The δ subunit of γ-aminobutyric acid type A receptors does not confer sensitivity to low concentrations of ethanol. J. Pharmacol. Exp. Ther. 2006;316:1360–1368. [PubMed]
  • Brickley SG, Cull-Candy SG, Farrant M. Development of a tonic form of synaptic inhibition in rat cerebellar granule cells resulting from persistent activation of GABAA receptors. J. Physiol. 1996;497(Pt 3):753–759. [PubMed]
  • Brickley SG, Revilla V, Cull-Candy SG, Wisden W, Farrant M. Adaptive regulation of neuronal excitability by a voltage-independent potassium conductance. Nature. 2001;409:88–92. [PubMed]
  • Carr LG, Spence JP, Peter Eriksson CJ, Lumeng L, Li TK. AA and ANA rats exhibit the R100Q mutation in the GABAA receptor alpha 6 subunit. Alcohol. 2003;31:93–97. [PubMed]
  • Carta M, Mameli M, Valenzuela CF. Alcohol enhances GABAergic transmission to cerebellar granule cells via an increase in Golgi cell excitability. J. Neurosci. 2004;24:3746–3751. [PubMed]
  • Casagrande S, Cupello A, Pellistri F, Robello M. Only high concentrations of ethanol affect GABAA receptors of rat cerebellum granule cells in culture. Neurosci. Lett. 2007;414:273–276. [PubMed]
  • Chadderton P, Margrie TW, Hausser M. Integration of quanta in cerebellar granule cells during sensory processing. Nature. 2004;428:856–860. [PubMed]
  • Eriksson K, Rusi M. Finnish selection studies on alcohol-related behaviors: general outline. In: McClearn GE, Deitrich RA, Erwin VG, editors. Development of animal models as pharmacogenetic tools. NIAAA Research Monograph; Rockville, MD: 1981. pp. 87–117.
  • Farrant M, Nusser Z. Variations on an inhibitory theme: phasic and tonic activation of GABAA receptors. Nat. Rev. Neurosci. 2005;6:215–229. [PubMed]
  • Fleming RL, Wilson WA, Swartzwelder HS. The magnitude and ethanol sensitivity of tonic GABAA receptor mediated inhibition in dentate gyrus changes from adolescence to adulthood. J. Neurophysiol. 2007 doi: 10.1152/jn.00101.2007. [PubMed] [Cross Ref]
  • Follesa P, Mostallino MC, Biggio F, Gorini G, Caria S, Busonero F, Murru L, Mura ML, Sanna E, Biggio G. Distinct patterns of expression and regulation of GABA receptors containing the δ subunit in cerebellar granule and hippocampal neurons. J. Neurochem. 2005;94:659–671. [PubMed]
  • Gibbs RA, Weinstock GM, Metzker ML, Muzny DM, Sodergren EJ, Scherer S, Scott G, Steffen D, Worley KC, Burch PE, et al. Genome sequence of the Brown Norway rat yields insights into mammalian evolution. Nature. 2004;428:493–521. [PubMed]
  • Glykys J, Peng Z, Chandra D, Homanics GE, Houser CR, Mody I. A new naturally occurring GABAA receptor subunit partnership with high sensitivity to ethanol. Nat. Neurosci. 2007;10:40–48. [PubMed]
  • Hanchar HJ, Chutsrinopkun P, Meera P, Supavilai P, Sieghart W, Wallner M, Olsen RW. Ethanol potently and competitively inhibits binding of the alcohol antagonist Ro15-4513 to α4/6β3δ GABAA receptors. Proc. Natl. Acad. Sci. USA. 2006;103:8546–8551. [PubMed]
  • Hanchar HJ, Dodson PD, Olsen RW, Otis TS, Wallner M. Alcohol-induced motor impairment caused by increased extrasynaptic GABAA receptor activity. Nat. Neurosci. 2005;8:339–345. [PMC free article] [PubMed]
  • Harney SC, Frenguelli BG, Lambert JJ. Phosphorylation influences neurosteroid modulation of synaptic GABAA receptors in rat CA1 and dentate gyrus neurones. Neuropharmacology. 2003;45:873–883. [PubMed]
  • Hellevuo K, Kiianmaa K, Korpi ER. Effect of GABAergic drugs on motor impairment from ethanol, barbital and lorazepam in rat lines selected for differential sensitivity to ethanol. Pharmacol. Biochem. Behav. 1989;34:399–404. [PubMed]
  • Korpi ER, Kleingoor C, Kettenmann H, Seeburg PH. Benzodiazepine-induced motor impairment linked to point mutation in cerebellar GABAA receptor. Nature. 1993;361:356–359. [PubMed]
  • Korpi ER, Seeburg PH. Natural mutation of GABAA receptor α6 subunit alters benzodiazepine affinity but not allosteric GABA effects. Eur. J. Pharmacol. 1993;247:23–27. [PubMed]
  • Liang J, Zhang N, Cagetti E, Houser CR, Olsen RW, Spigelman I. Chronic intermittent ethanol-induced switch of ethanol actions from extrasynaptic to synaptic hippocampal GABAA receptors. J. Neurosci. 2006;26:1749–1758. [PubMed]
  • Martikainen IK, Lauk K, Moykkynen T, Holopainen IE, Korpi ER, Uusi-Oukari M. Kainate down-regulates a subset of GABAA receptor subunits expressed in cultured mouse cerebellar granule cells. Cerebellum. 2004;3:27–38. [PubMed]
  • Minier F, Sigel E. Positioning of the alpha-subunit isoforms confers a functional signature to gamma-aminobutyric acid type A receptors. Proc. Natl. Acad. Sci. USA. 2004;101:7769–7774. [PubMed]
  • Mohler H, Crestani F, Rudolph U. GABAA-receptor subtypes: a new pharmacology. Curr. Opin. Pharmacol. 2001;1:22–25. [PubMed]
  • Naylor DE, Liu H, Wasterlain CG. Trafficking of GABAA receptors, loss of inhibition, and a mechanism for pharmacoresistance in status epilepticus. J. Neurosci. 2005;25:7724–7733. [PubMed]
  • Nusser Z, Ahmad Z, Tretter V, Fuchs K, Wisden W, Sieghart W, Somogyi P. Alterations in the expression of GABAA receptor subunits in cerebellar granule cells after the disruption of the α6 subunit gene. Eur. J. Neurosci. 1999;11:1685–1697. [PubMed]
  • Nusser Z, Hajos N, Somogyi P, Mody I. Increased number of synaptic GABAA receptors underlies potentiation at hippocampal inhibitory synapses. Nature. 1998a;395:172–177. [PubMed]
  • Nusser Z, Roberts JD, Baude A, Richards JG, Somogyi P. Relative densities of synaptic and extrasynaptic GABAA receptors on cerebellar granule cells as determined by a quantitative immunogold method. J. Neurosci. 1995;15:2948–2960. [PubMed]
  • Nusser Z, Sieghart W, Somogyi P. Segregation of different GABAA receptors to synaptic and extrasynaptic membranes of cerebellar granule cells. J. Neurosci. 1998b;18:1693–1703. [PubMed]
  • Otis TS, De Koninck Y, Mody I. Lasting potentiation of inhibition is associated with an increased number of γ-aminobutyric acid type A receptors activated during miniature inhibitory postsynaptic currents. Proc. Natl. Acad. Sci. USA. 1994;91:7698–7702. [PubMed]
  • Otis TS, Hanchar HJ, Dodson PD, Olsen RW, Wallner M. Letters to the editor. Alcohol. Clin. Exp. Res. 2005;29:1358.
  • Otis TS, Mody I. Differential activation of GABAA and GABAB receptors by spontaneously released transmitter. J. Neurophysiol. 1992a;67:227–235. [PubMed]
  • Otis TS, Mody I. Modulation of decay kinetics and frequency of GABAA receptor-mediated spontaneous inhibitory postsynaptic currents in hippocampal neurons. Neuroscience. 1992b;49:13–32. [PubMed]
  • Overstreet LS, Jones MV, Westbrook GL. Slow desensitization regulates the availability of synaptic GABA(A) receptors. J. Neurosci. 2000;20:7914–7921. [PubMed]
  • Paul SM. Alcohol-sensitive GABA receptors and alcohol antagonists. Proc. Natl. Acad. Sci. USA. 2006;103:8307–8308. [PubMed]
  • Payne HL, Donoghue PS, Connelly WM, Hinterreiter S, Tiwari P, Ives JH, Hann V, Sieghart W, Lees G, Thompson CL. Aberrant GABAA receptor expression in the dentate gyrus of the epileptic mutant mouse stargazer. J. Neurosci. 2006;26:8600–8608. [PMC free article] [PubMed]
  • Peng Z, Hauer B, Mihalek RM, Homanics GE, Sieghart W, Olsen RW, Houser CR. GABAA receptor changes in δ subunit-deficient mice: altered expression of α4 and γ2 subunits in the forebrain. J. Comp. Neurol. 2002;446:179–197. [PubMed]
  • Reynolds JN, Prasad A, MacDonald JF. Ethanol modulation of GABA receptor-activated Cl-currents in neurons of the chick, rat and mouse central nervous system. Eur. J. Pharmacol. 1992;224:173–181. [PubMed]
  • Saba L, Porcella A, Congeddu E, Colombo G, Peis M, Pistis M, Gessa GL, Pani L. The R100Q mutation of the GABAA α6 receptor subunit may contribute to voluntary aversion to ethanol in the sNP rat line. Brain Res. Mol. Brain Res. 2001;87:263–270. [PubMed]
  • Salonen V, Kallinen S, Lopez-Picon FR, Korpi ER, Holopainen IE, Uusi-Oukari M. AMPA/kainate receptor-mediated up-regulation of GABAA receptor δ subunit mRNA expression in cultured rat cerebellar granule cells is dependent on NMDA receptor activation. Brain Res. 2006;1087:33–40. [PubMed]
  • Santhakumar V, Hanchar HJ, Wallner M, Olsen RW, Otis TS. Contributions of the GABAA receptor α6 subunit to phasic and tonic inhibition revealed by a naturally occurring polymorphism in the α6 gene. J. Neurosci. 2006;26:3357–3364. [PMC free article] [PubMed]
  • Shao LR, Dudek FE. Changes in mIPSCs and sIPSCs after kainate treatment: evidence for loss of inhibitory input to dentate granule cells and possible compensatory responses. J. Neurophysiol. 2005;94:952–960. [PubMed]
  • Stell BM, Brickley SG, Tang CY, Farrant M, Mody I. Neuroactive steroids reduce neuronal excitability by selectively enhancing tonic inhibition mediated by δ subunit-containing GABAA receptors. Proc. Natl. Acad. Sci. USA. 2003;100:14439–14444. [PubMed]
  • Storustovu SI, Ebert B. Pharmacological characterization of agonists at δ-containing GABAA receptors: Functional selectivity for extrasynaptic receptors is dependent on the absence of γ2. J. Pharmacol. Exp. Ther. 2006;316:1351–1359. [PubMed]
  • Sundstrom-Poromaa I, Smith DH, Gong QH, Sabado TN, Li X, Light A, Wiedmann M, Williams K, Smith SS. Hormonally regulated α4β2δ GABAA receptors are a target for alcohol. Nat. Neurosci. 2002;5:721–722. [PMC free article] [PubMed]
  • Suzdak PD, Glowa JR, Crawley JN, Schwartz RD, Skolnick P, Paul SM. A selective imidazobenzodiazepine antagonist of ethanol in the rat. Science. 1986;234:1243–1247. [PubMed]
  • Tia S, Wang JF, Kotchabhakdi N, Vicini S. Developmental changes of inhibitory synaptic currents in cerebellar granule neurons: role of GABAA receptor α6 subunit. J. Neurosci. 1996;16:3630–3640. [PubMed]
  • Valenzuela CF, Mameli M, Carta M. Single-amino-acid difference in the sequence of α6 subunit dramatically increases the ethanol sensitivity of recombinant GABAA receptors. Alcohol. Clin. Exp. Res. 2005;29:1356–1357. author reply 1358. [PubMed]
  • Wafford KA, Burnett DM, Leidenheimer NJ, Burt DR, Wang JB, Kofuji P, Dunwiddie TV, Harris RA, Sikela JM. Ethanol sensitivity of the GABAA receptor expressed in Xenopus oocytes requires 8 amino acids contained in the γ2L subunit. Neuron. 1991;7:27–33. [PubMed]
  • Wall MJ, Usowicz MM. Development of action potential-dependent and independent spontaneous GABAA receptor-mediated currents in granule cells of postnatal rat cerebellum. Eur. J. Neurosci. 1997;9:533–548. [PubMed]
  • Wallner M, Hanchar HJ, Olsen RW. Ethanol enhances α4b3δ and α6β3δ γ-aminobutyric acid type A receptors at low concentrations known to affect humans. Proc. Natl. Acad. Sci. USA. 2003;100:15218–15223. [PubMed]
  • Wallner M, Hanchar HJ, Olsen RW. Low-dose alcohol actions on α4β3δ GABAA receptors are reversed by the behavioral alcohol antagonist Ro15-4513. Proc. Natl. Acad. Sci. USA. 2006a;103:8540–8545. [PubMed]
  • Wallner M, Hanchar HJ, Olsen RW. Low dose acute alcohol effects on GABAA receptor subtypes. Pharmacol. Ther. 2006b;112:513–528. [PMC free article] [PubMed]
  • Wei W, Faria LC, Mody I. Low ethanol concentrations selectively augment the tonic inhibition mediated by δ subunit-containing GABAA receptors in hippocampal neurons. J. Neurosci. 2004;24:8379–8382. [PubMed]
  • Wei W, Zhang N, Peng Z, Houser CR, Mody I. Perisynaptic localization of delta subunit-containing GABAA receptors and their activation by GABA spillover in the mouse dentate gyrus. J. Neurosci. 2003;23:10650–10661. [PubMed]
  • Wong G, Sarviharju M, Toropainen M, Matecka D, Korpi ER. Pharmacologic actions of subtype-selective and novel GABAergic ligands in rat lines with differential sensitivity to ethanol. Pharmacol. Biochem. Behav. 1996;53:723–730. [PubMed]
  • Yamashita M, Marszalec W, Yeh JZ, Narahashi T. Effects of ethanol on tonic GABA currents in cerebellar granule cells and mammalian cells recombinantly expressing GABAA receptors. J. Pharmacol. Exp. Ther. 2006;319:431–438. [PubMed]