Associate editor: A.L. Morrow
Neurosteroid modulation of synaptic and extrasynaptic GABAA receptors

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Abstract

Certain naturally occurring pregnane steroids act in a nongenomic manner to potently and selectively enhance the interaction of the inhibitory neurotransmitter GABA with the GABAA receptor. Consequently such steroids exhibit anxiolytic, anticonvulsant, analgesic, sedative, hypnotic, and anesthetic properties. In both physiological and pathophysiological scenarios, the pregnane steroids may function as endocrine messengers (e.g., produced in the periphery and cross the blood–brain barrier) to influence behaviour. However, additionally “neurosteroids” can be synthesised in the brain and spinal cord to act in a paracrine or autocrine manner and thereby locally influence neuronal activity. Given the ubiquitous expression of the GABAA receptor throughout the mammalian central nervous system (CNS), physiological, pathophysiological, or drug-induced pertubations of neurosteroid levels may be expected to produce widespread changes in brain excitability. However, the neurosteroid/GABAA receptor interaction is brain region and indeed neuron specific. The molecular basis of this specificity will be reviewed here, including (1) the importance of the subunit composition of the GABAA receptor; (2) how protein phosphorylation may dynamically influence the sensitivity of GABAA receptors to neurosteroids; (3) the impact of local steroid metabolism; and (4) the emergence of extrasynaptic GABAA receptors as a neurosteroid target.

Introduction

It is now over 20 years since Harrison and Simmonds (1984) first demonstrated that the synthetic steroid anesthetic alphaxalone (5α-pregnan-3α-ol-11,20-dione) potently enhances the function of the major inhibitory receptor in the mammalian central nervous system (CNS), the GABAA receptor. Subsequently, a number of naturally occurring metabolites of progesterone and deoxycorticosterone (e.g., 5α-pregnan-3α-ol-20-one [3α,5α-THP] and 5α-pregnan-3α,21-diol-20-one [3α,5α-THDOC], respectively) were discovered to share this property (reviewed in Lambert et al., 2003, Belelli and Lambert, 2005). The enhancement of inhibitory neurotransmission by such steroids is consistent with their behavioural properties (anxiolytic, anticonvulsant, analgesic, sedative/hypnotic, and anesthetic; Lambert et al., 1995, Gasior et al., 1999, Rupprecht, 2003).

The GABAA receptor is the target for a variety of clinically important therapeutic drugs, including benzodiazepines, certain general anesthetics and anticonvulsants (Frolund et al., 2002, Whiting, 2003). The receptor is composed of 5 subunits arranged to form a central anion selective channel. To date, 19 subunits have been identified and divided into subfamilies according to amino acid sequence homology (α1-6, β1-3, γ1-3, δ, ε, θ, π, ρ1-3; Barnard et al., 1998). This subunit repertoire permits the expression of ∼ 20 to 30 distinct GABAA receptor isoforms in the CNS, and they have a distinct expression pattern (Sieghart and Sperk, 2002, Fritschy and Brunig, 2003). The subunit composition confers distinct biophysical and pharmacological properties upon GABAA receptors. Key amino acid residues that contribute to binding domains on the receptor for benzodiazepines (Wieland et al., 1992, Sigel and Buhr, 1997) and particular general anesthetics have been identified (Belelli et al., 1997, Mihic et al., 1997, Li et al., 2006). These findings have led to the generation of mice engineered to express GABAA receptor isoforms, which by virtue of mutating critical amino acids, are rendered insensitive to such compounds (Reynolds et al., 2003, Rudolph and Mohler, 2004, Mohler et al., 2004). Such mice are proving invaluable in delineating which GABAA receptor subtypes mediate the various behavioural effects of benzodiazepines and general anesthetics. The recent identification of critical residues for neurosteroid enhancement of the GABA response (located on the α subunit) and for direct activation of the GABAA receptor by the steroid (located within an α/β subunit interface) should now permit the putative physiological and pathophysiological role of neurosteroids to be explored in similar mouse models (Hosie et al., 2006).

GABAA receptor subunit composition can additionally influence the distribution of the receptor within a neuron. For example, receptors incorporating the γ2 subunit in combination with an α and a β subunits are usually, although not exclusively, clustered within postsynaptic aggregates, where they mediate the transient “phasic” response to quantally released GABA (Fritschy and Brunig, 2003, Farrant and Nusser, 2005). By contrast, receptors incorporating a δ (rather than a γ subunit), in combination with an α4, or an α6, subunit and a β subunit, are located almost exclusively out with the synapse, where they mediate a persistent “tonic” form of inhibition in response to a maintained exposure to relatively low concentrations of ambient GABA (Farrant and Nusser, 2005, Mody, 2005, Chandra et al., 2006).

Endogenous steroids derived either from peripheral or central sources (“neurosteroids”) are present within the brain at concentrations sufficient to modulate GABAA receptor function, suggesting a physiological role for these agents (Paul and Purdy, 1992, Mellon and Vaudry, 2001, Mellon and Griffin, 2002). Furthermore, the expression of steroidogenic enzymes in certain neurons implies that these neurosteroids, in addition to serving as endocrine messengers, may function in a paracrine, or indeed an autocrine, fashion as local neuromodulators (Agis-Balboa et al., 2006; see Fig. 1).

The endogenous levels of neurosteroids are not static but are subject to dynamic change in response to a variety of conditions including: development, puberty, stress, pregnancy, ovarian cycle, and treatment with certain psychoactive drugs (e.g., ethanol, γ-hydroxybutyrate, fluoxetine; Purdy et al., 1991, Paul and Purdy, 1992, Uzunova et al., 1998, Barbaccia, 2004, Kumar et al., 2004, Morrow et al., 2004). Pertubations in neurosteroid levels may contribute to neurological disorders, such as catamenial epilepsy, and have been implicated in a variety of psychological conditions, including panic attacks, major depression, postpartum depression, premenstrual tension, and schizophrenia (Purdy et al., 1991, Smith, 2004, Sundstrom-Poromaa, 2004, Eser et al., 2006, Finn et al., 2006, Marx et al., 2006).

GABAA receptors are expressed throughout the CNS, therefore changes in neurosteroid levels would be expected to globally influence neuronal activity, a scenario incongruent with their proposed physiological role. However, steroidogenic enzyme expression and consequently local neurosteroid production are brain region specific (Mellon and Vaudry, 2001, Mellon and Griffin, 2002, Mellon, 2004, Agis-Balboa et al., 2006). Furthermore, the neurosteroid–GABAA receptor interaction is highly selective, being neuron specific and indeed within certain neurons, neurosteroids may differentially interact with distinct pools of GABAA receptors (Stell et al., 2003, Belelli and Lambert, 2005). Clearly such selectivity is fundamental to the effects of the neurosteroids on neuronal circuitry and consequent behaviour. Here we will review the current understanding of the molecular basis of this specificity and consider the relative importance of synaptic and extrasynaptic GABAA receptors as targets for neurosteroid action.

Section snippets

Neurosteroid modulation of synaptic GABAA receptors

Electrophysiological approaches have been used to unveil the molecular mechanisms underpinning neurosteroid depressant actions in the CNS. Such studies exploit the ability of presynaptic terminal boutons of neurons maintained in culture or within brain slices to spontaneously release vesicles of GABA, even in the absence of action potential drive. This approach permits an evaluation of the effects of neurosteroids at both pre- and post-synaptic GABAA receptors.

GABAA receptor subunit composition

As outlined above, the mammalian CNS expresses ∼20 to 30 distinct GABAA receptor isoforms which are differentially distributed and exhibit distinct physiological and pharmacological properties. Given that the GABA-enhancing actions of benzodiazepines and certain general anesthetics are highly dependent upon GABAA receptor subunit composition (Sigel and Buhr, 1997, Belelli et al., 1997), the neuronal selectivity of neurosteroids may reflect the differential expression of GABAA receptor isoforms.

Neurosteroid modulation of extrasynaptic GABAA receptors

Although facilitation of inhibitory synaptic (“phasic”) transmission has been considered the primary mechanism whereby neurosteroids influence neuronal excitability, recent studies have highlighted the importance of a “tonic” form of inhibition to neuronal information processing (Semyanov et al., 2004, Farrant and Nusser, 2005). Importantly, in some brain regions, the perisynaptic and extrasynaptic GABAA receptors, which mediate tonic inhibition, appear more sensitive to neurosteroids than

Plasticity of neurosteroid action at synaptic and extrasynaptic GABAA receptors

The functional properties of mammalian GABAA receptors are subject to dynamic change in response to a variety of physiological and pathophysiological stimuli including development (Hollrigel and Soltesz, 1997, Vicini et al., 2001, Keller et al., 2004), hormonal status (e.g., menstrual/estrous cycle, pregnancy, puberty; Brussaard et al., 1997, Smith et al., 1998a, Smith et al., 1998b, Maguire et al., 2005, Maguire and Mody, 2007, Shen et al., 2007), drug withdrawal (Devaud et al., 1996, Devaud

Summary and future directions

Although steroids have traditionally been ascribed to act through classical intracellular receptors, extensive evidence over the past 2 decades has established that neurosteroids such as 3α,5α-THP exert rapid, highly potent actions at the cell membrane via an allosteric enhancement of GABAA receptor function. The realisation that neurons and glia possess the enzymatic machinery required for de novo synthesis of such steroids raises the intriguing possibility that locally synthesised

Acknowledgements

Some of the work reported here was supported by the BBSRC, Tenovus Tayside, and the Anonymous Trust. The authors thank Professor J.A. Peters for helpful discussions.

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