Associate Editor: H. Bönisch
α2-containing GABAA receptors: A target for the development of novel treatment strategies for CNS disorders

https://doi.org/10.1016/j.pharmthera.2012.08.006Get rights and content

Abstract

GABAA receptors have important physiological functions, as revealed by pharmacological studies and experiments involving gene-targeted mouse models, and are the target of widely used drugs such as the benzodiazepines. In this review, we are summarizing current knowledge about the function of α2-containing GABAA receptors, a receptor subtype representing approximately 15–20% of all GABAA receptors. This receptor subtype mediates anxiolytic-like, reward-enhancing, and antihyperalgesic actions of diazepam, and has antidepressant-like properties. Secondary insufficiency of α2-containing GABAA receptors has been postulated to play a role in the pathogenesis of schizophrenia, and may be involved in cognitive impairment in other disorders. Moreover, polymorphisms in the GABRA2 gene encoding the GABAA receptor α2 subunit have been found to be linked to chronic alcohol dependence and to polydrug abuse. Thus, α2-containing GABAA receptors are involved in the regulation and/or modulation of emotional behaviors and of chronic pain, and appear to be a valid target for novel therapeutic approaches for the treatment of anxiety, depression, schizophrenia and chronic pain.

Introduction

γ-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the central nervous system (CNS). GABAergic inhibition is essential for temporal modulation and maintenance of the excitation/inhibition balance and for the entrainment of oscillatory activities (Klausberger & Somogyi, 2008). GABA acts at ionotropic GABAA receptors and at metabotropic GABAB receptors.

GABAA receptors are formed as pentameric combinations of homologous subunits. There are 19 genes known encoding GABAA receptor subunits (α1-6, β1-3, γ1-3, δ, ε, θ, π, ρ1-3) (Olsen & Sieghart, 2008), with receptors formed by α, β and γ subunits being the most common. The α subunits provide the largest diversity, and for practical purposes GABAA receptors are most frequently classified by their α subunits.

This diverse set of receptors are the targets of a variety of therapeutically useful CNS-active compounds, e.g. benzodiazepines, zolpidem, zopiclone, barbiturates, etomidate, propofol, and neurosteroids (Rudolph and Antkowiak, 2004, Rudolph and Knoflach, 2011). Out of these compounds, benzodiazepines have been the most extensively studied, largely due to their widespread clinical use in the treatment of anxiety disorders. Benzodizepines are positive allosteric modulators (PAM) of the GABAA receptors, with a variety of actions, including sedation, hypnosis, muscle relaxation, and anticonvulsant and amnestic effects, in addition to anxiolysis. This wide array of behavioral effects reduces the therapeutic utility of clinically used benzodiazepines (e.g., diazepam, chlordiazepoxide, midazolam), as even when they are effective in reaching the desired therapeutic targets (e.g., anxiolysis in patients with anxiety disorders), they are associated with a number of unwanted side effects (e.g., sedation and hypnosis). Thus, understanding the molecular substrates for each of these behavioral endpoints has been a major research interest with the hope of producing more therapeutically-specific GABAergic compounds in the future. Differential quantitative and anatomical distribution of GABAA receptor subunits in the CNS, and differential modulation by benzodiazepine site ligands of different GABAA receptors based on subunit composition provided the first clues in terms of an understanding of benzodiazepine behavioral pharmacology.

Whereas the GABA binding sites on GABAA receptors are formed by residues on the α and β subunits, the binding sites for benzodiazepines are formed by residues on α and γ subunits (Sigel & Luscher, 2011). Benzodiazepines bind to GABAA receptors containing the α1, α2, α3, and α5 subunits and a γ subunit (in addition to the obligatory β subunit) (Mohler et al., 2002). Receptors containing the α4 or α6 subunit do not bind these compounds. Whereas the three β subunits β1-3 appear to confer identical properties to the receptor complex with respect to modulation by benzodiazepines (as well as by barbiturates and neurosteroids) (Hadingham et al., 1993), the γ subunits contribute somewhat differentially to this process (Benke et al., 1996). Approximately 85% of all γ subunits are γ2 subunits (Benke et al., 1996), and it has been shown that the γ2 subunit is required for behavioral actions of benzodiazepines (Gunther et al., 1995). Thus, GABAA receptors containing a γ2 subunit, in combination with α1, α2, α3, or α5 subunits likely mediate all of the above-mentioned behavioral effects of benzodiazepines.

As seen in Fig. 1, approximately 60% of all GABAA receptors contain the α1 subunit, 15–20% the α2 subunit, 10–15% the α3 subunit, and 5% the α5 subunit (H. Mohler et al., 2002). In addition to this quantitative difference in their expression levels, the α subunits also show anatomically differential patterns of expression in the CNS. The finding that functionally different neuronal circuits express GABAA receptors of specific α subunit compositions preferentially, as well as the differential subcellular, sometimes synapse-specific expression of α subunits within individual neurons, suggested that the α subunits may confer different behavioral functions and thus may constitute targets for specific therapeutic endpoints.

While a few compounds exhibiting relative subtype-selectivity have been developed, no truly subtype-specific compounds have been generated so far. Thus, the traditional pharmacological approach of defining the functions of GABAA receptor subtypes by synthesizing novel compounds, while extremely valuable for medication development, was met with limited success for clear assignment of functions to receptor subtypes. This limitation stimulated interest in the development of genetic mouse models that would allow unequivocal elucidation of subunit function. To date, several mouse models involving knockouts or point mutations of specific α subunits have been generated and the studies employing these models have provided invaluable information about the functions of different GABAA receptor subtypes. For instance, mice carrying histidine to arginine point mutations in one of the four diazepam-sensitive α subunits [α1(H101R), α2(H101R), α3(H126R) and α5(H105R)] that transform the diazepam-sensitive α subunits into diazepam-insensitive α subunits have been employed extensively to understand the GABAA receptor subtypes that mediate different behavioral actions of benzodiazepines (Rudolph et al., 1999, Low et al., 2000, McKernan et al., 2000, Crestani et al., 2002, Smith et al., 2012). In these mice, the “missing” diazepam actions in mice carrying the mutated α subunit can then be ascribed to the mutant α subunit. The first studies using these point-mutated mice revealed that while wild type mice are sedated by diazepam, α1(H10R)mice are not sedated by diazepam, but diazepam still has an anxiolytic-like action in these mice (Rudolph et al., 1999), suggesting that the sedative effects of diazepam are mediated by the α1-containing GABAA receptors.

Since these early studies, many other studies employing genetic mouse models revealed specific roles for different α subunits, in addition to refining and re-affirming the earlier findings. In a different line of work, human genetics studies revealed associations between different disorders and personality traits and different α subunits. One subunit, the α2 subunit, has stood out in both lines of research as being linked to many diverse physiological processes and several disorders of the nervous system, suggesting that this subunit might constitute an extremely fruitful drug target. Here, we summarize findings regarding this subunit in relation to different CNS disorders and their treatment.

In each of the sections below, we first summarize findings from human genetics and/or postmortem human studies, followed by findings from animal research, most commonly from gene-targeted mouse models. In many cases the suggestive evidence from human genetics studies can be corroborated by genetically modified mouse models with mechanistic demonstrations of loss or gain of function in animals following genetic manipulations. A review of the two literatures together also provides an easy way to spot gaps in data in either research approach as a guide for future investigations in this area. We start with the well-replicated studies investigating the role of GABRA2 in anxiety, followed by findings relating GABRA2 to alcohol dependence and drug abuse. Emerging evidence linking GABRA2 to depression is discussed, followed by a section suggesting the role of GABRA2 in the brain reward system as the potential common ground for the role of α2-containing GABAA receptors in substance abuse and depression. We finally discuss the role of GABRA2 in other CNS disorders, such as schizophrenia and Alzheimer's disease, in the sleep EEG and in pain.

One caveat to note before we begin our discussion is that in many cases other GABAA receptor subtypes have also been implicated in a single disorder, as these disorders represent complex neuropathology that involve several brain structures and neurotransmitter systems. The main goal of this review is to demonstrate the potential of α2-containing GABAA receptors as valuable drug targets for diverse neuropathology rather than a full discussion of GABAergic mechanisms involved in a certain disorder. While we do discuss other GABAA receptor subtypes briefly at most points, there are also instances where we leave out the discussion of other subunits in order focus on α2-containing GABAA receptors. More extensive discussion of other GABAA receptor subtypes are available in other recent reviews (Rudolph and Mohler, 2006, Rudolph and Knoflach, 2011).

Section snippets

GABRA2 and anxiety

The evidence linking the GABA system to anxiety and anxiety disorders is extensive [see (Kalueff and Nutt, 2007, Mohler, 2012) for reviews]. GABAergic drugs, such as benzodiazepines have been used to treat anxiety disorders for more than half a century. Surprisingly, human genetics studies linking GABAA receptor subunits and anxiety phenotypes are limited in number and have revealed relatively weak results. A few studies suggested a possible link between the GABRA2 gene and anxiety. For

GABRA2 and alcohol dependence

While it is known that ethanol can potentiate GABAA receptors in vitro, the contribution of this potentiation to the effects of ethanol continues to be controversial. Most synaptic GABAA receptors show sensitivity to ethanol only at concentrations above the drunk driving limit in many states. One reported exception to this, although not universally accepted (Borghese et al., 2006, Yamashita et al., 2006) are the extrasynaptic α4β3δ GABAA receptors, which are modulated by “social” concentrations

GABRA2 and substance abuse

In addition to alcohol dependence, GABRA2 polymorphisms are also associated with other drug dependence, both in adolescence and adulthood (Enoch et al., 2010). A study on African-American men with lifetime DSM-IV single and comorbid diagnoses of alcohol, cocaine, and heroin dependence found one unique haplotype predicting heroin addiction, and another one which was more common in controls and appeared to confer resilience to addiction after exposure to severe childhood trauma. An unlinked SNP

GABRA2 and depression

The neurotransmitter GABA has been implicated to play a role in depression. For instance, GABA levels are reduced in the brains of patients with major depression (MDD) and are normalized following chronic treatment with antidepressants (Sanacora et al., 2002, Sanacora et al., 2004). Genetic associations have been reported between MDD and the genes encoding several GABAA receptor subtypes, such as GABRA1 and GABRA5 (Oruc et al., 1997, Horiuchi et al., 2004), but none of these studies have

GABRA2 and reward

In the last three sections we have reviewed evidence suggesting a role for α2-containing GABAA receptors in alcohol dependence and other substance abuse, and in depression. One common characteristic of addictive disorders and depression is the dysfunction of the brain reward system. Thus, it is likely that the α2-containing GABAA receptors play an important role in the functioning of the brain reward system. Moreover, benzodiazepine abuse and misuse has been long documented among both patients

GABRA2 and sleep EEG

Sleep EEG is altered in several psychiatric and neurodegenerative disorders, suggesting that an altered sleep EEG could be an important biomarker for several disorders, as well as a possible factor that exacerbates the neuropathology (see Petit et al., 2004, Steiger and Kimura, 2010 for reviews). The patients in one of the studies linking polymorphisms in the GABRA2 gene to alcohol dependence had the quantitative endophenotype of increased power in the β frequency band (13–28 Hz) of the EEG (

GABRA2 and schizophrenia

GABA abnormalities in the cortex of schizophrenic patients have been a consistent finding of postmortem studies (see Stan & Lewis, 2012 for a review). Human genetics studies have associated several GABAA receptor subunit genes, such as GABRA1, GABRA6 and GABRB2, with schizophrenia (Petryshen et al., 2005). While no study has linked GABRA2 polymorphisms to schizophrenia so far, there is evidence from postmortem studies of schizophrenia brains, from pharmacological studies and from gene-targeted

GABRA2 and pain

While diazepam is not usually considered to be an analgesic agent, intrathecal administration of diazepam in rodents which does not cause motor sedation has an antihyperalgesic effect, e.g. in paradigms for neuropathic and inflammatory pain, where it increases the paw withdrawal latencies. Studies with mice carrying point-mutated diazepam-insensitive α subunits indicated that diazepam-induced analgesia is mediated by α2- and α3- containing GABAA receptors in an animal model of inflammatory pain

Conclusion

While the role of α2-containing GABAA receptors in benzodiazepine-induced anxiolysis has been known for a long time (Low et al., 2000), more recent studies reveal the involvement of these receptors in processes as diverse as reward and alcohol/drug abuse, schizophrenic symptomatology, depression and chronic pain. Since α2-containing GABAA receptors do not mediate sedation (Rudolph et al., 1999, McKernan et al., 2000), it is predicted that α2-specific agonists would not be compounded by

Conflict of interest statement

In the last three years, UR has provided professional services for Sunovion and for Concert Pharmaceuticals. JL and EE declare that they have no conflict of interest.

Acknowledgements

Research by the authors on GABAA receptors was or is supported by grants from the National Institutes of Health to UR (award numbers GM086448, MH080006, MH085149, DA027571, DA026578, MH094834 and MH095905). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institute of General Medical Sciences, the National Institute of Mental Health and the National Institute of Drug Abuse, or the National Institutes of Health. EE was

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