Psychopharmacology Unit, University of Bristol
Correspondence: D. J. Nutt, Psychopharmacology Unit, School of Medical Sciences, University of Bristol, University Walk, Bristol BS8 ITD, UK. Tel: 0117 925 3066; fax: 0117 927 7057; e-mail: David.J.Nutt{at}bristol.ac.uk
Declaration of interest D.J.N. has received grants from various pharmaceutical companies with an interest in drugs acting at the GABAbenzodiazepine receptor.
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ABSTRACT |
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Aims To present current knowledge about the role of the GABAAbenzodiazepine receptor in anxiety disorders, new insights into the molecular biology of the receptor complex and neuroimaging studies suggesting involvement of these receptors in disease states.
Method An overview of published literature, including some recent data.
Results The molecular biology of this receptor is detailed. Molecular genetic studies suggesting involvement of the GABAAbenzodiazepine receptor in animal behaviour and learning are outlined; possible parallels with human psychopathology are discussed.
Conclusions Current insights into the role of the GABAAbenzodiazepine receptor in the action of benzodiazepines and as a factor in disease states, in both animals and humans, may lead to new, more sophisticated interventions at this receptor complex and potentially significant therapeutic gains.
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INTRODUCTION |
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THE GABAABENZODIAZEPINE RECEPTOR |
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GABA: REINING IN THE NERVOUS SYSTEM |
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AGONISTS, ANTAGONISTS, INVERSE AGONISTS |
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Further insights into the sophistication of the benzodiazepine site came 15 years ago with the discovery of drugs which bind to it, yet have the opposite effects to the classical benzodiazepine receptor agonists. These drugs decrease the probability of the chloride channel opening in response to GABA and have stimulant, anxiogenic and proconvulsant properties. They are now termed inverse agonists. Subsequently, benzodiazepine receptor antagonists, notably flumazenil, have been discovered which block the activities of both agonists and inverse agonists. Moreover, the recent discovery of both partial agonists and partial inverse agonists shows that the benzodiazepine receptor mediates a spectrum of different actions (see Fig. 2).
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WHY DOES THE BENZODIAZEPINE SITE EXIST? |
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It is also possible that benzodiazepine receptors exist for exactly the opposite purpose to mediate the activities of endogenous inverse agonists. Such compounds could keep brain arousal optimal and if levels fell, sleep could result. In the search for an endogenous benzodiazepine receptor ligand, several compounds with inverse agonist activity have emerged. One early candidate, ethyl-ß-carboline-3-carboxylate (ß-CCE), was the first compound shown to promote anxiety by a direct action at a receptor in the brain (Braestrup et al, 1980). But it was later shown that ß-CCE was not endogenous, being formed in the extraction process. Another, called tribulin (Hucklebridge et al, 1998), is found in urine and its levels are elevated in many conditions where there is increased anxiety (post-traumatic stress disorder and alcohol withdrawal, for example; reviewed by Glover, 1998). However, its structure has not yet been determined and its status remains uncertain.
A third theory suggests that there is no endogenous benzodiazepine receptor ligand and that the site may simply be a particular protein conformation that fine tunes GABA function, possibly altering maximal efficacy, or the rate of desensitisation. Recent evidence suggests that the benzodiazepine receptor spectrum is not fixed and that the set-point where drugs bind, but have no effect can be moved, perhaps as a result of differential sub-unit expression (see below). Among the effects of a set-point shift could be tolerance of, and/or dependence on, benzodiazepine receptor agonists, differential sensitivity to alcohol, anxiety predisposition, panic disorder and stress responsiveness (see Fig. 3). Genetically determined factors may also lead to different positions of the set-point (see also benzodiazepine dependence).
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ANXIETY: A BENZODIAZEPINE RECEPTOR ABNORMALITY? |
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When patients with panic disorder were given an intravenous 2 mg dose of flumazenil, enough to occupy more than half of the receptors in the brain, it provoked panic in most of the patients but was quite innocuous in the control subjects (Nutt et al, 1990). This finding, which has been replicated in a number of studies (though not in all) (Woods et al, 1991; Maddock, 1998; Strohle et al, 1999) clearly demonstrates that panic disorder is not due to the actions of an inverse agonist, as in this case the antagonisit would reduce anxiety. The anxiogenic effect of flumazenil could reflect displacement of an endogenous agonist, but this would only be present in patients (since controls did not experience an increase in anxiety). Another, and more likely, hypothesis is that the set-point of the benzodiazepine receptor has moved in the inverse agonist direction, making flumazenil a weak inverse agonist, thus generating anxiety. Further, we could expect the effects of full agonists to be reduced in patients with panic disorder and this is indeed the case. It has been shown that patients with panic disorder are subsensitive to the central effects of diazepam (Roy-Byrne et al, 1990) and we also know, from clinical experience, that treatment of these patients requires either a high-dose or a high-potency benzodiazepine.
Abnormality at the GABAbenzodiazepine receptor may be specific to, or more pronounced in, severe episodic anxiety as patients with generalised anxiety disorder, post-traumatic stress disorder and depression do not panic when given flumazenil. Since there is a significant hereditary factor in panic disorder, the receptor abnormality could be due to the transmission of a defective receptor gene (see Identifying the role of receptor sub-units, below).
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IMAGING STUDIES IN ANXIETY |
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These findings are consistent with the idea that some anxiety disorders may be due to defective neuroinhibitory processes. The greatest decreases observed in benzodiazepine binding occurred in areas thought to be involved in the experience of anxiety in man, such as the orbitofrontal and temporal cortex and insula. Interestingly, in a recent PET study of anxiety generation, we have shown that the anxiolytic effects of the benzodiazepine most closely relate to modulation in brain metabolism in insula and orbitofrontal cortex, among others (Malizia, 2000) (see Fig. 5). Care should be taken in drawing too many conclusions from these small studies, but they do strengthen the case that abnormalities in basal or adaptive inhibitory neuromodulation are of pathological significance in anxiety disorders.
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ROLE OF GABA IN MEMORY AND ANXIETY |
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There may be many clinical sequelae from this relationship. Traumatic memories, such as those which occur in post-traumatic stress disorder, are extremely deeply encoded, probably because of the high level of emotion and arousal at the time of the original insult as well as the negative reinforcement on re-experience of the events. Anxiety leads to rapid and profound learning of escape and avoidance behaviours, as in agoraphobia following a panic attack, or anxious avoidance after being bitten by a dog. Apart from animal studies, clinical evidence of reduced GABA in the genesis of anxiety comes from the use of pentylenetetrazol as a convulsant agent before electroconvulsive therapy was introduced in clinical practice. Pentylenetetrazol acts by blocking GABAA receptor function and has been reported to produce extreme anxiety, traumatic memories and extreme avoidance behaviour when used clinically (reviewed in Kalueff & Nutt, 1997). Recently, Goddard et al (2001) have reported decreased cortical GABA levels in patients with panic disorder using magnetic resonance spectroscopy.
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INSIGHTS FROM MOLECULAR BIOLOGY |
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At the last count, 19 subtypes coded by different genes had been found in
mammals. Six different isoforms of the sub-unit have so far been
identified and different subtypes have different sensitivities to
benzodiazepine receptor ligands. Receptors with the
6
sub-unit, for example, are essentially insensitive to all hypnotic/anxiolytic
benzodiazepine agonists. The type of
sub-unit is also critical as only
the
2 sub-unit gives GABAAbenzodiazepine
receptor which is responsive to benzodiazepines. The genes that encode for
these various subtypes seem to occur in clusters, suggesting that there is
some coherence in the expression of receptor complexes with different forms of
the sub-units. The most important and most prevalent
GABAAbenzodiazepine receptor in the brain is made up from
1, ß2 and
2 sub-units, encoded by the same cluster of genes
on chromosome 5.
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IDENTIFYING THE ROLE OF RECEPTOR SUB-UNITS |
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Knock-out mice with GABAAbenzodiazepine receptors deficient in other types of sub-unit have also been produced. When the gene for the expression of the ß3 subunit is knocked out, for instance, it produces mice that are hyperactive, have poor motor coordination and have spontaneous seizures. They have an attenuated response to GABA and it would seem that the ß3 sub-unit has been replaced by a ß2 sub-unit. Interestingly, there is also reduced sensitivity to anaesthetic agents, including halothane, enflurane, etomidate and the benzodiazepine, midazolam, the first direct evidence that GABAbenzodiazepine receptors may be important in mediating the anaesthetic effects of these agents.
More recently, knock-in technology has been used to produce mice in which
the 1 subtype gene has been mutated to become insensitive to
benzodiazepines but still responsive to GABA (knock-in mutations). In these
mice the sedative actions of benzodiazepines are abolished but the anxiolytic,
anticonvulsant and hypnotic ones remain
(Rudolph et al, 1999;
Tobler et al, 2001).
Subsequent mutation of two remaining subtypes has shown that the anxiolytic
effect is lost if the
2 but not the
3
subtype is mutated (Low et al,
2000). The localisation of the
2 subtype in the
limbic system supports the role of this circuit in anxiety. This growing
evidence for the specificity of benzodiazepine actions being mediated via
receptor subtypes is leading to the search for subtype-selective drugs such as
2 or
3 agonists as non-sedating
anxiolytics and
5 inverse agonists (which act mainly in the
hippocampus) as memory enhancers
(Lingford-Hughes et al,
2001).
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CURRENT CLINICAL ISSUES |
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WHY SHOULD RECEPTOR SENSITIVITY CHANGE? |
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NEW THERAPEUTIC HORIZONS |
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Clinical Implications and Limitations |
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LIMITATIONS
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Received for publication August 7, 2000. Revision received January 26, 2001. Accepted for publication January 31, 2001.