The cellular, molecular and ionic basis of GABAA receptor signalling

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Abstract

GABAA receptors mediate fast synaptic inhibition in the CNS. Whilst this is undoubtedly true, it is a gross oversimplification of their actions. The receptors themselves are diverse, being formed from a variety of subunits, each with a different temporal and spatial pattern of expression. This diversity is reflected in differences in subcellular targetting and in the subtleties of their response to GABA. While activation of the receptors leads to an inevitable increase in membrane conductance, the voltage response is dictated by the distribution of the permeant Cl and HCO3 ions, which is established by anion transporters. Similar to GABAA receptors, the expression of these transporters is not only developmentally regulated but shows cell-specific and subcellular variation. Untangling all these complexities allows us to appreciate the variety of GABA-mediated signalling, a diverse set of phenomena encompassing both synaptic and non-synaptic functions that can be overtly excitatory as well as inhibitory.

Introduction

The spectacular progress of biosciences during recent decades has led to a vast accumulation of knowledge concerning mechanisms that govern the development and functions of living beings. It has also shown that much of the variation among species and organ systems, as well as among distinct brain regions and nerve cells, is largely due to what can be thought of as “combinatorics” of basic molecular elements and signalling pathways. In fact, given the large number of possible conformational states and allosteric interactions that proteins and complexes thereof can assume, any potential signalling molecule can bind to an enormous variety of biologically relevant binding sites or receptors in diverse target cells. In the case of a neurotransmitter substance such as GABA, these ligand-binding site interactions take place at ionotropic and metabotropic receptors, at vesicular and plasmalemmal transporters and with metabolic enzymes. In the nervous system, such interactions of GABA culminate in its most important action — inhibition of neuronal firing (Kuffler and Edwards, 1958; Krnjevic, 1974).

The role of GABA as the main inhibitory transmitter in the mammalian brain is undoubtedly fundamental, but it should be noted that GABA also has excitatory actions (and “dual” effects — both inhibitory and excitatory) in a number of cell types (Cherubini et al., 1991; Voipio and Kaila, 2000; Ben-Ari, 2001; Marty and Llano, 2005). This is, only to be expected, because the properties of the ligand-binding site of a transmitter receptor channel do not dictate or even influence the biophysical properties of the channel's ionic filter or the transmembrane ion gradients that set the polarity and driving force of the channel-mediated current. As inhibition and excitation are central issues in the present review, we want to note here that postsynaptic inhibition is best defined as a transient decrease in the probability of firing in the target cell, while postsynaptic excitation has the opposite effect. Very often, hyperpolarization and depolarization are taken as synonyms of inhibition and excitation, which is an error, as is evident from the above definition (and from what is described below). Importantly, the actions of GABA can be manifest as conventional synaptic transmission, involving the transient or “phasic” activation of receptors, or they can reflect the persistent or “tonic” activation of receptors in a manner temporally divorced form identifiable presynaptic events (Mody, 2001; Semyanov et al., 2004; Farrant and Nusser, 2005). Such tonic receptor activation can also have varied functional consequences.

GABA's role as a neurotransmitter goes far beyond its immediate effects on the excitability of an individual target cell. Extensive work on cortical structures has shown that the activity of GABAergic neurons is crucially involved in the assembly of neurons into functional networks, and in shaping oscillations and transient population events at the network level (McBain and Fisahn, 2001; Buzsaki, 2002; Freund, 2003; Whittington and Traub, 2003; Buzsaki and Draguhn, 2004; Jonas et al., 2004; Vida et al., 2006). Many aspects of these network-level phenomena are based on GABA-mediated changes in the integrative properties of principal neurons that depend critically on synapse location and the timing of activity. Thus, numerous examples illustrate the importance of spatially distinct GABAergic inputs (axonic, perisomatic, proximal or distal dendritic) in enabling or regulating important neuronal behaviours in the cortex (Spruston et al., 1995; Miles et al., 1996; Pouille and Scanziani, 2001; Somogyi and Klausberger, 2005; Szabadics et al., 2006) and basal ganglia (Plenz, 2003; Tepper and Bolam, 2004; see also Chapters 6–11 in this volume). While a great deal of work has been carried out on GABAergic transmission in the basal ganglia, it is quite obvious that the amount of information available is not comparable to that for GABA actions in cortical neurons and networks. The aim of the present review is to provide a general overview of GABAA receptor-mediated actions in an attempt to facilitate future work on GABAA receptor-mediated transmission at the cellular and network level in the basal ganglia. Our main focus is on the electrophysiological, biophysical and ion-regulatory mechanisms that shape GABA-mediated postsynaptic potentials and tonic signalling.

Section snippets

Molecular basis of GABAA receptor function

Ionotropic GABA receptors are members of the “Cys-loop” superfamily of ligand-gated ion channels (cl-LGIC), so named for a conserved motif in the amino-terminal domain in which a pair of cysteines forms a disulphide bridge (Simon et al., 2004). Other family members include the nicotinic acetylcholine receptors, glycine receptors and 5-HT3 receptors. In each case, the receptors are pentameric assemblies of subunits (each with four transmembrane domains; M1–4) that form a central ion channel,

Ionic permeability of GABAA receptors

Irrespective of their subunit composition, all GABAA receptors are permeable to the same ions. Based on measurements of the permeation of a series of large, weakly hydrated probe anions, the diameter of the narrowest part of the GABAA receptor channel has been estimated at about 0.55 nm (Inomata et al., 1986; Bormann et al., 1987; Akaike et al., 1989; Fatima-Shad and Barry, 1993; Kaila, 1994; Wotring et al., 1999). However, the GABAA channel is not a simple water-filled pore (Takeuchi and

GABAA receptor channel conductance

Although the channels formed by GABAA receptors are similarly ion selective, they do display differences in single-channel conductance, i.e., the rate at which they allow ions to flow. Most receptors display a prominent main conductance state and one or more subconductance states. The absolute conductance depends on the concentrations of permeant ions present, but with near symmetrical [Cl] (∼130–160 mM; room temperature in outside-out patches) the main conductance state of homomeric ρ1

GABA concentration changes at the synapse

GABA is synthesized in the cytosol and accumulated into synaptic vesicles by a vesicular transporter (VGAT/VIAAT (McIntire et al., 1997; Chaudhry et al., 1998; Wojcik et al., 2006)) that is able to generate an intra-lumenal concentration thought to be in the range of several hundred millimolar (Axmacher et al., 2004). Thus, the fusion of a single vesicle liberates many thousands of GABA molecules into the synaptic cleft (∼20 nm wide and 0.05–0.2 μm2 in area), generating a GABA concentration

GABAA receptor gating and the IPSC

GABAA receptors undergo considerable spontaneous motion while in the closed state (Bera and Akabas, 2005), suggesting a level of flexibility appropriate for rapid gating (Maconochie et al., 1994; Jones and Westbrook, 1995; McClellan and Twyman, 1999; Burkat et al., 2001; Chakrapani and Auerbach, 2005). In theory, the channel can open in the absence of agonist, albeit with an extremely low probability (Chang and Weiss, 1999a; Campo-Soria et al., 2006). Of note, some recombinant (Sigel et al.,

Tonic activity of GABAA receptors

GABAA receptors are not activated only during “phasic”, i.e., synaptic signalling. Persistent or “tonic” activation of receptors, independent of any identified release event, occurs prior to synapse formation in embryonic (Valeyev et al., 1993; LoTurco et al., 1995; Owens et al., 1999; Demarque et al., 2002) and immature (Sipila et al., 2007) as well as newly derived postnatal neurons (Nguyen et al., 2003; Liu et al., 2005; Ge et al., 2006), as well as in a variety of mature (synaptically

Neuronal ion regulation and the driving force for GABAA receptor-mediated currents

As noted above, GABAA receptors are permeable to two physiologically relevant anions, Cl and HCO3, with a HCO3/Cl permeability ratio (PHCO3-/PCl-) of around 0.2–0.4. A frequent and erroneous assumption in the GABA literature is that this permeability ratio translates directly into identical quantitative relations of the respective anion currents during synaptic and tonic responses. With an intracellular pH (pHi) of about 7.1–7.2 and the above PHCO3-/PCl- ratio, the intraneuronal HCO3

Neuronal chloride regulation

Neuronal Cl homeostasis is mainly controlled by the SLC12A family of cation-chloride co-transporters (CCCs; Fig. 3). The CCCs are composed of glycoproteins (MW of monomers in the range of 120–200 kDa) with 12 putative transmembrane segments flanked on one side by a small intracellular amino-terminus and by a large intracellular carboxy-terminus on the other (Payne et al., 2003). CCCs are secondary active transporters that do not directly consume ATP, but derive the energy for Cl translocation

Neuronal pH regulation and EGABA

In virtually all kinds of cells, including neurons, acid extrusion mechanisms (extrusion of H+, uptake of HCO3) maintain pHi at a significantly more alkaline level than what would be predicted on the basis of a passive distribution of H+ and HCO3 ions (Roos and Boron, 1981). Therefore, the equilibrium potential for HCO3 (which equals the equilibrium potential of H+) is much more positive than the resting membrane potential (typically about −10 to −15 mV). Unlike Cl that can mediate either

The “developmental switch” from depolarization to hyperpolarization

In a pioneering study, Obata et al. (1978) showed that GABA has an excitatory action in co-cultures of muscle and spinal neurons taken from 6- to 8–day-old chick embryos, while an inhibitory effect was observed when culturing was started on day 10. Immature neurons are known to have very high input impedance that makes recordings with conventional sharp microelectrodes difficult and unreliable. However, the excitatory effect of GABA (and glycine) demonstrated by Obata and co-workers was clearly

Ionic plasticity of GABAA receptor-mediated signalling

Ionotropic glutamatergic transmission does not appear to be heavily modulated by activity-dependent changes in the driving force of the postsynaptic currents. In contrast to this, GABAergic signalling has the unique property of “ionic plasticity” which is based on both short-term and long-term shifts in the concentrations of Cl and HCO3 in postsynaptic neurons. This type of plasticity phenomena fall into two categories: short-term modulatory changes that are solely attributable to

SN pars compacta

There is little information about the ionic mechanisms that shape GABAergic actions in basal ganglia. However, a rather exceptional finding was made in recent work (Gulacsi et al., 2003) on dopaminergic (DA) neurons in the substantia nigra (SN) pars compacta. In contrast to most of the neurons in the CNS, these DA neurons do not express KCC2, as shown by several kinds of immunohistochemical techniques. In contrast to this, GABAergic neurons in the SN were immunopositive for KCC2, with high

Conclusion

If there is one straightforward conclusion to be drawn from the preceding sections, it might be that extrapolation of observations regarding GABAA receptor-mediated signalling in one neuron or network to another cell type or brain region is not easy. Of course, this does not mean that it should not be attempted, but rather the opposite. A comparative approach — at the level of different species or brain regions — is a fundamental strategy of neurobiological research that has generated lots of

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