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GABAB receptors make it to the top – as dimers
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GABAB receptors make it to the top – as dimers Hanns Möhler and Jean-Marc Fritschy The GABAB receptor, the metabotropic receptor for the neurotransmitter g-aminobutyric acid, was once termed the ‘last wallflower on the receptors’ dance’1, but lost this status when the GABABR1 receptor was cloned2,3. However, it was at the recent 4th International GABAB Symposium* that the GABAB receptor rose to stardom by revealing its partner, the GABABR2 receptor. In reporting that the GABAB receptor operates as a heterodimer, a new mechanism in signal transduction for G protein-coupled receptors was celebrated.
GABAB receptors
GABAB receptors are present in most regions of the mammalian brain on presynaptic terminals and postsynaptic neurones. Activation of presynaptic GABAB receptors located on GABA-containing nerve terminals (autoreceptors) or terminals of various other neurones (heteroreceptors) suppresses the release of neurotransmitters, whereas the stimulation of postsynaptic receptors produces a prolonged neuronal hyperpolarization. The former mechanism appears to be mediated by inhibition of an inward Ca21 conductance whereas the latter is produced by an increase in K1 conductance. In both cases the GABAB receptor is coupled via G proteins to its conductance mechanism. The receptor was structurally identified by expression cloning of the GABABR1 receptor, which occurs in two splice variants termed GABABR1a and GABABR1b (130 and 100 kDa, respectively). They display an affinity for antagonists and an overall distribution corresponding to native GABAB receptors. However, the more GABABR1 was investigated the more it became apparent that it was ‘handicapped’. Its agonist affinity was lower than
that of native receptors by a factor of 100–150. Furthermore, it coupled only weakly to adenylate cyclase and its coupling to other effector systems, such as K1 and Ca21 channels, remained elusive. In addition, when expressed in mammalian cells GABABR1 remained largely trapped in the endoplasmic reticulum and did not reach the cell membrane.
GABAB receptors as heterodimers The solution to these problems came with the identification of GABABR2 receptor protein, a second component of GABAB receptors, as reported independently by three groups working in industry and one in academia. The groups from Novartis (K. Kaupmann et al., Basle, Switzerland), GlaxoWellcome (F. H. Marshal et al., Stevenage, UK), Synaptic Pharmaceutical (K. A. Jones et al., Paramus, NJ, USA) and the National Institutes of Health (T. I. Bonner et al., Bethesda, MD, USA) succeeded in isolating the GABABR2 cDNA by homology screening based on expressed sequence tags. In addition, the GlaxoWellcome group identified GABABR2 as a binding partner by way of the C terminus of GABABR1 in the yeast two hybrid system. The GABABR2 protein is a seven transmembrane domain (7TM) protein that corresponds in size to the GABABR1 receptor and displays a sequence homology of about 35%. Its distribution corresponds largely to that of GABABR1 as demonstrated by in situ hybridization. GABABR2 appears to fulfill several functions: (1) It acts as a translocator: it permits GABABR1 to reach the cell membrane, where both proteins are present as dimers as shown by co-immunoprecipitation not only in heterologous expression systems but also in situ. (2) It influences ligand affinity: the heterodimer displays
0165-6147/99/$ – see front matter © 1999 Elsevier Science. All rights reserved.
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an agonist affinity that is identical (GlaxoWellcome group) or lower only by a factor of ten (Novartis group) than that of native receptors. (3) The heterodimer shows robust coupling to effector systems and displays strong inhibition of forskolinstimulated adenylate cyclase activity. In addition, its coupling to a K1 channel (Kir 3.1) could be demonstrated for the first time. Results on the coupling of the heterodimer to Ca21 channels are eagerly awaited. Thus, it is now apparent that functional GABAB receptors are heterodimers made up of two closely related transmembrane proteins that interact at the C terminus in a stoichiometry of 1:1.
A new tune for 7TM receptors: heterodimerization In contrast to classical 7TM G protein-coupled receptors such as adrenoceptors or opioid receptors, which are fully functional as monomers, problems with processing and ligand selectivity of certain receptors has been accumulating in recent years (Fig. 1). Cloning of the receptor for the calcitonin gene-related peptide (CGRP) is a case in point: the initially cloned CRLR receptor (calcitonin receptor-like receptor) lacked proper ligand selectivity and was retained in the endoplasmic reticulum. Only when co-expressed with RAMP1, a single transmembranedomain protein, were cell-surface receptors generated that had a ligand specificity characteristic of CGRP receptors. In combination with RAMP2 the CRLR receptor formed a heterodimer with a ligand specificity of adrenomedullin receptors4. The help of a translocator protein odorant-4 (ODR4) was also required for a subset of olfactory receptors to proceed from the endoplasmic reticulum to the cilia in olfactory neurones of Caenorhabditis elegans5. ODR4 encodes a membrane protein that is expressed on intracellular membranes acting to facilitate receptor folding or localization, or both. However, the GABAB receptor heterodimer represents a totally new
PII: S0165-6147(99)01323-1
TiPS – March 1999 (Vol. 20)
H. Möhler Director, and J-M. Fritschy, Research Scientist, Institute of Pharmacology, Swiss Federal Institute of Technology (ETH) and University of Zurich, Winterthurerstrasse 190, CH-8057 Zurich, Switzerland. *4th International GABAB Symposium, 5–7 November 1998, Los Angeles, USA.
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endoplasmatic reticulum
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cell surface
classical receptors e.g. adrenoceptor, opioid
7TM receptor
RAMP1
CGRP receptor
CRLR RAMP2
Adrenomedullin receptor
Olfactory receptor ODR4
Olfactory receptor (C. elegans)
GABABR1
R1
GABABR2
R2 ?
R2
GABAB receptor
?
Fig. 1. G protein-coupled receptors: transport and heterodimerization. Seven transmembrane domain G protein-coupled receptors (7TM receptors) occur either as monomers, e.g. adrenoceptors or opioid receptors, or as heterodimers, e.g. calcitonin gene-related peptide (CGRP) receptor [calcitonin receptor-like receptor (CRLR) and RAMP1]4, adrenomedullin receptor (CRLR and RAMP2)4, GABAB receptor (GABABR1 and GABABR2, this meeting). The double bar in the endoplasmic reticulum indicates the retention of the respective receptor component unless it is associated with its dimerization partner. In the case of the olfactory receptor in Caenorhabditis elegans the heterodimerization with odorant 4 (ODR4) is transient5.
principle of receptor processing and signal transduction in that two 7TM receptor proteins associate. This opens the intriguing possibility that GABABR2 is not only involved in receptor translocation and the modulation of agonist affinity, but might also take part in signal transduction. Thus, in the heterodimer not only R1 but also R2 could display binding sites for GABA and both GABABR1 and GABABR2 could potentially interact with G proteins.
Where is the receptor heterogeneity? The therapeutic potential of GABAB receptor ligands can best be realized if receptor subtypes can be structurally identified. At present, the archetypal receptor agonist 88
TiPS – March 1999 (Vol. 20)
baclofen is used in the treatment of spasticity, but agonist therapeutics could potentially extend to the treatment of certain types of pain, such as neuropathic pain, and cocaine withdrawal symptoms. By contrast, the effects of GABAB receptor antagonists in humans have yet to be established, but many exciting possibilities have arisen including the treatment of cognitive impairment and absence epilepsy. Subtyping of GABAB receptors might therefore provide the template for future drug targeting. Although the heterodimer appears to be the basic molecular architecture of GABAB receptors, variations on this theme could arise. These are given below: (1) GABABR1 splice variants: the 1a and 1b isoforms differ in their
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N-terminal domain, with the implication that the heterodimer might therefore display different ligandbinding characteristics depending on the splice variant present. Furthermore, the splice variants might interact with different extracellular anchoring proteins providing distinct subcellular localizations. (2) GABABR2 splice variants: there are indications that C-terminal splice variants exist for GABABR2. This is of major interest because this region is crucial for the interaction of GABABR1 and GABABR2 and it might also modulate G protein coupling. (3) GABABR2 as a potential monomer: although the heterodimer R1–R2 appears to be the prevalent structure of GABAB receptors, the R2 protein can reach the cell membrane by itself when expressed in heterologous systems in the absence of R1. It is presently not clear whether GABABR2 is a functional receptor in its own right. Although the GlaxoWellcome group suggested that R2 alone was not functional, the Novartis group reported that it couples to a K1 channel. Nevertheless, the possibility cannot be ruled out that R2 – in addition to its role in the heterodimer – might act as a receptor in its own right, thereby adding to GABAB receptor heterogeneity. (4) Components yet to be discovered: although GABABR2 appears to be a rather ubiquitous constituent of GABAB receptors in the brain, the striatum showed a surprisingly low level of GABABR2 mRNA relative to GABABR1, raising the possibility that there might be other receptor components. Similarly, in peripheral organs, the co-distribution of GABABR1 and GABABR2 mRNA has been found only in lung and thymus, leaving room for the discovery of further peripheral GABAB receptor components. (5) Heterogeneity in targeting: the location of GABAB receptors at particular subcellular sites might ensure their interactions with selective effector systems and thereby display particular pharmacological actions. Because the functional heterogeneity
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might arise from the subcellular distribution of GABAB receptors, an extensive structural heterogeneity might not be required.
Cellular and subcellular location of GABAB receptors
The GABAB receptor dimer appears to be the basic building block for most, if not all, GABAB receptors in the brain. In situ hybridization and light microscropic immunohistochemistry revealed their co-localization in agreement with the distribution of GABAB receptor binding sites previously identified by autoradiography. However, difficulties were encountered in visualizing, using electron microscopy, the GABAB receptors on the subcellular level using either pre- or postembedding immunohistochemistry. It has not yet been possible to visualize the receptors on GABAcontaining terminals or terminals of other neurones. This failure contrasts with the extensive visualization of GABAB receptors in the vicinity (extrasynaptic) of glutamatergic synapses (F-M. Fritschy, Zurich, Switzerland; R. Shigemoto, Kyoto, Japan). For example, GABAB receptors are highly expressed in cerebellar Purkinje cells where they were found to be largely located on dendritic spines that receive glutamatergic input from the parallel fibres. The GABAB receptors are not located within the postsynaptic density but rather found at extrasynaptic sites, particularly at the shaft of the spine.
Functional roles of GABAB receptors A distinction between pre- and postsynaptic GABAB receptors has been outlined in several electrophysiological studies in the neocortex, hippocampus, amygdala and hypothalamus. Presynaptic receptors are generally activated by lower concentrations of baclofen than postsynaptic receptors. This distinction was extended to antagonists (J. P. Gallagher, Galveston, USA) potentially opening the way for the development of subtype-specific drugs. A
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receptor with an unexpectedly high baclofen affinity was noted in a cell line (S. J. Morris, Kansas City, USA). The coupling of postsynaptic GABAB receptors to inwardly rectifying K1 channels (Kir3.1 and Kir3.2) is now well established, in particular from studies using the weaver mouse (U. Misgeld, Heidelberg, Germany). Postsynaptic GABAB receptors appear to require strong stimuli for activation. In paired recordings of neocortical and hippocampal neurones, GABAB receptors were detected in a small fraction of neurones only upon application of a train of stimuli (A. Thompson, London, UK). This non-linearity in stimulus–response properties of GABAB receptors was shown by computational modelling to underlie the generation of spike-and-wave discharges (SWDs) in absence seizures and sleep spindles in the thalamo-cortical system (A. Destexhe, Québec, Canada). However, the contribution of GABAB receptors for SWDs was questioned by in vivo recordings in genetic absence epilepsy rats (V. Crunelli and N. Leresche, Cardiff, UK and Paris, France). Unlike previous data from in vitro studies, these results suggest that SWDs in this model of absence epilepsy do not depend on GABAB receptor activation in vivo. A number of presentations focused on the role of GABAB receptors as potential therapeutic targets. These included the control of epilepsy (Y. Ito, Funabashi, Japan; L. S. Leung, London, Canada; G. D. Ritchie, Wright-Patterson Base, USA), the regulation of the hypothalamo– pituitary axis (M. J. Kelly, Portland, USA; I. Shibuya, Kitakyushu, Japan), the enhancement of anti-depressant drug action (F. Artigas, Barcelona, Spain), the suppression of cocainewithdrawal symptoms (J. P. Gallager, Galveston, USA) and the alleviation of pain (D. H. Hammond, Chicago, USA). With regard to pain, clear-cut evidence suggests that baclofen can reduce pain perception in animal models of central sensitization but fails to do so in humans. A
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possible explanation might be the ‘uncoupling’ of GABAB receptors induced by chronic pain, as suggested by autoradiographic data in a model of monoarthritic pain (N. G. Bowery, Birmingham, UK).
Concluding remarks Most, if not all, GABAB receptors are heterodimers formed from GABABR1 and GABABR2 subunits. The interaction of these two 7TM receptor proteins represents a novel principle of receptor assembly and signal transduction which will be a major focus of future GABAB receptor research. In addition, gene-targeting experiments are expected to determine whether GABABR2, apart from being a partner in the heterodimer, functions as a receptor in its own right. Furthermore, knockout experiments, performed in a regionor cell-specific manner, will help identify the role of GABAB receptors in complex CNS functions and pathologies. Recognizing the regional and cellular relevance of GABAB receptors will be essential in identifying novel drug targets. Thus, based on the new biology of GABAB receptors, a new pharmacology could be on the horizon at the time of the next GABAB receptor meeting.
Note added in proof Recently, five groups have had papers published or are in press on the cloning of GABABR2 (Refs 6–10).
References 1 Bowery, N. G. and Brown, D. A. (1997) Nature 386, 223–224 2 Kaupmann, K. et al. (1997) Nature 386, 239–246 3 Bettler, B., Kaupmann, K. and Bowery, N. G. (1998) Curr. Opin. Neurobiol. 8, 345–350 4 McLatchie, L. M. et al. (1998) Nature 393, 333–339 5 Dwyer, N. D. et al. (1998) Cell 93, 455–466 6 Kaupmann, K. et al. (1998) Nature 396, 683–687 7 White, J. H. et al. (1998) Nature 386, 679–682 8 Jones, K. A. et al. (1998) Nature 386, 674–679 9 Kuner, R. et al. (1999) Science 283, 74–77 10 Ng, G. Y. K. J. Biol. Chem. (in press)
TiPS – March 1999 (Vol. 20)
Acknowledgement The 4th International GABAB Symposium was made possible by the support of the American Society for Pharmacology and Experimental Therapeutics and Novartis Pharma AG.
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GABAB receptors make it to the top – as dimers Hanns Möhler and Jean-Marc Fritschy The GABAB receptor, the metabotropic receptor for the neurotransmitter g-aminobutyric acid, was once termed the ‘last wallflower on the receptors’ dance’1, but lost this status when the GABABR1 receptor was cloned2,3. However, it was at the recent 4th International GABAB Symposium* that the GABAB receptor rose to stardom by revealing its partner, the GABABR2 receptor. In reporting that the GABAB receptor operates as a heterodimer, a new mechanism in signal transduction for G protein-coupled receptors was celebrated.
GABAB receptors
GABAB receptors are present in most regions of the mammalian brain on presynaptic terminals and postsynaptic neurones. Activation of presynaptic GABAB receptors located on GABA-containing nerve terminals (autoreceptors) or terminals of various other neurones (heteroreceptors) suppresses the release of neurotransmitters, whereas the stimulation of postsynaptic receptors produces a prolonged neuronal hyperpolarization. The former mechanism appears to be mediated by inhibition of an inward Ca21 conductance whereas the latter is produced by an increase in K1 conductance. In both cases the GABAB receptor is coupled via G proteins to its conductance mechanism. The receptor was structurally identified by expression cloning of the GABABR1 receptor, which occurs in two splice variants termed GABABR1a and GABABR1b (130 and 100 kDa, respectively). They display an affinity for antagonists and an overall distribution corresponding to native GABAB receptors. However, the more GABABR1 was investigated the more it became apparent that it was ‘handicapped’. Its agonist affinity was lower than
that of native receptors by a factor of 100–150. Furthermore, it coupled only weakly to adenylate cyclase and its coupling to other effector systems, such as K1 and Ca21 channels, remained elusive. In addition, when expressed in mammalian cells GABABR1 remained largely trapped in the endoplasmic reticulum and did not reach the cell membrane.
GABAB receptors as heterodimers The solution to these problems came with the identification of GABABR2 receptor protein, a second component of GABAB receptors, as reported independently by three groups working in industry and one in academia. The groups from Novartis (K. Kaupmann et al., Basle, Switzerland), GlaxoWellcome (F. H. Marshal et al., Stevenage, UK), Synaptic Pharmaceutical (K. A. Jones et al., Paramus, NJ, USA) and the National Institutes of Health (T. I. Bonner et al., Bethesda, MD, USA) succeeded in isolating the GABABR2 cDNA by homology screening based on expressed sequence tags. In addition, the GlaxoWellcome group identified GABABR2 as a binding partner by way of the C terminus of GABABR1 in the yeast two hybrid system. The GABABR2 protein is a seven transmembrane domain (7TM) protein that corresponds in size to the GABABR1 receptor and displays a sequence homology of about 35%. Its distribution corresponds largely to that of GABABR1 as demonstrated by in situ hybridization. GABABR2 appears to fulfill several functions: (1) It acts as a translocator: it permits GABABR1 to reach the cell membrane, where both proteins are present as dimers as shown by co-immunoprecipitation not only in heterologous expression systems but also in situ. (2) It influences ligand affinity: the heterodimer displays
0165-6147/99/$ – see front matter © 1999 Elsevier Science. All rights reserved.
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an agonist affinity that is identical (GlaxoWellcome group) or lower only by a factor of ten (Novartis group) than that of native receptors. (3) The heterodimer shows robust coupling to effector systems and displays strong inhibition of forskolinstimulated adenylate cyclase activity. In addition, its coupling to a K1 channel (Kir 3.1) could be demonstrated for the first time. Results on the coupling of the heterodimer to Ca21 channels are eagerly awaited. Thus, it is now apparent that functional GABAB receptors are heterodimers made up of two closely related transmembrane proteins that interact at the C terminus in a stoichiometry of 1:1.
A new tune for 7TM receptors: heterodimerization In contrast to classical 7TM G protein-coupled receptors such as adrenoceptors or opioid receptors, which are fully functional as monomers, problems with processing and ligand selectivity of certain receptors has been accumulating in recent years (Fig. 1). Cloning of the receptor for the calcitonin gene-related peptide (CGRP) is a case in point: the initially cloned CRLR receptor (calcitonin receptor-like receptor) lacked proper ligand selectivity and was retained in the endoplasmic reticulum. Only when co-expressed with RAMP1, a single transmembranedomain protein, were cell-surface receptors generated that had a ligand specificity characteristic of CGRP receptors. In combination with RAMP2 the CRLR receptor formed a heterodimer with a ligand specificity of adrenomedullin receptors4. The help of a translocator protein odorant-4 (ODR4) was also required for a subset of olfactory receptors to proceed from the endoplasmic reticulum to the cilia in olfactory neurones of Caenorhabditis elegans5. ODR4 encodes a membrane protein that is expressed on intracellular membranes acting to facilitate receptor folding or localization, or both. However, the GABAB receptor heterodimer represents a totally new
PII: S0165-6147(99)01323-1
TiPS – March 1999 (Vol. 20)
H. Möhler Director, and J-M. Fritschy, Research Scientist, Institute of Pharmacology, Swiss Federal Institute of Technology (ETH) and University of Zurich, Winterthurerstrasse 190, CH-8057 Zurich, Switzerland. *4th International GABAB Symposium, 5–7 November 1998, Los Angeles, USA.
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endoplasmatic reticulum
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cell surface
classical receptors e.g. adrenoceptor, opioid
7TM receptor
RAMP1
CGRP receptor
CRLR RAMP2
Adrenomedullin receptor
Olfactory receptor ODR4
Olfactory receptor (C. elegans)
GABABR1
R1
GABABR2
R2 ?
R2
GABAB receptor
?
Fig. 1. G protein-coupled receptors: transport and heterodimerization. Seven transmembrane domain G protein-coupled receptors (7TM receptors) occur either as monomers, e.g. adrenoceptors or opioid receptors, or as heterodimers, e.g. calcitonin gene-related peptide (CGRP) receptor [calcitonin receptor-like receptor (CRLR) and RAMP1]4, adrenomedullin receptor (CRLR and RAMP2)4, GABAB receptor (GABABR1 and GABABR2, this meeting). The double bar in the endoplasmic reticulum indicates the retention of the respective receptor component unless it is associated with its dimerization partner. In the case of the olfactory receptor in Caenorhabditis elegans the heterodimerization with odorant 4 (ODR4) is transient5.
principle of receptor processing and signal transduction in that two 7TM receptor proteins associate. This opens the intriguing possibility that GABABR2 is not only involved in receptor translocation and the modulation of agonist affinity, but might also take part in signal transduction. Thus, in the heterodimer not only R1 but also R2 could display binding sites for GABA and both GABABR1 and GABABR2 could potentially interact with G proteins.
Where is the receptor heterogeneity? The therapeutic potential of GABAB receptor ligands can best be realized if receptor subtypes can be structurally identified. At present, the archetypal receptor agonist 88
TiPS – March 1999 (Vol. 20)
baclofen is used in the treatment of spasticity, but agonist therapeutics could potentially extend to the treatment of certain types of pain, such as neuropathic pain, and cocaine withdrawal symptoms. By contrast, the effects of GABAB receptor antagonists in humans have yet to be established, but many exciting possibilities have arisen including the treatment of cognitive impairment and absence epilepsy. Subtyping of GABAB receptors might therefore provide the template for future drug targeting. Although the heterodimer appears to be the basic molecular architecture of GABAB receptors, variations on this theme could arise. These are given below: (1) GABABR1 splice variants: the 1a and 1b isoforms differ in their
R
T
N-terminal domain, with the implication that the heterodimer might therefore display different ligandbinding characteristics depending on the splice variant present. Furthermore, the splice variants might interact with different extracellular anchoring proteins providing distinct subcellular localizations. (2) GABABR2 splice variants: there are indications that C-terminal splice variants exist for GABABR2. This is of major interest because this region is crucial for the interaction of GABABR1 and GABABR2 and it might also modulate G protein coupling. (3) GABABR2 as a potential monomer: although the heterodimer R1–R2 appears to be the prevalent structure of GABAB receptors, the R2 protein can reach the cell membrane by itself when expressed in heterologous systems in the absence of R1. It is presently not clear whether GABABR2 is a functional receptor in its own right. Although the GlaxoWellcome group suggested that R2 alone was not functional, the Novartis group reported that it couples to a K1 channel. Nevertheless, the possibility cannot be ruled out that R2 – in addition to its role in the heterodimer – might act as a receptor in its own right, thereby adding to GABAB receptor heterogeneity. (4) Components yet to be discovered: although GABABR2 appears to be a rather ubiquitous constituent of GABAB receptors in the brain, the striatum showed a surprisingly low level of GABABR2 mRNA relative to GABABR1, raising the possibility that there might be other receptor components. Similarly, in peripheral organs, the co-distribution of GABABR1 and GABABR2 mRNA has been found only in lung and thymus, leaving room for the discovery of further peripheral GABAB receptor components. (5) Heterogeneity in targeting: the location of GABAB receptors at particular subcellular sites might ensure their interactions with selective effector systems and thereby display particular pharmacological actions. Because the functional heterogeneity
M
E
might arise from the subcellular distribution of GABAB receptors, an extensive structural heterogeneity might not be required.
Cellular and subcellular location of GABAB receptors
The GABAB receptor dimer appears to be the basic building block for most, if not all, GABAB receptors in the brain. In situ hybridization and light microscropic immunohistochemistry revealed their co-localization in agreement with the distribution of GABAB receptor binding sites previously identified by autoradiography. However, difficulties were encountered in visualizing, using electron microscopy, the GABAB receptors on the subcellular level using either pre- or postembedding immunohistochemistry. It has not yet been possible to visualize the receptors on GABAcontaining terminals or terminals of other neurones. This failure contrasts with the extensive visualization of GABAB receptors in the vicinity (extrasynaptic) of glutamatergic synapses (F-M. Fritschy, Zurich, Switzerland; R. Shigemoto, Kyoto, Japan). For example, GABAB receptors are highly expressed in cerebellar Purkinje cells where they were found to be largely located on dendritic spines that receive glutamatergic input from the parallel fibres. The GABAB receptors are not located within the postsynaptic density but rather found at extrasynaptic sites, particularly at the shaft of the spine.
Functional roles of GABAB receptors A distinction between pre- and postsynaptic GABAB receptors has been outlined in several electrophysiological studies in the neocortex, hippocampus, amygdala and hypothalamus. Presynaptic receptors are generally activated by lower concentrations of baclofen than postsynaptic receptors. This distinction was extended to antagonists (J. P. Gallagher, Galveston, USA) potentially opening the way for the development of subtype-specific drugs. A
E
T
I
N
G
receptor with an unexpectedly high baclofen affinity was noted in a cell line (S. J. Morris, Kansas City, USA). The coupling of postsynaptic GABAB receptors to inwardly rectifying K1 channels (Kir3.1 and Kir3.2) is now well established, in particular from studies using the weaver mouse (U. Misgeld, Heidelberg, Germany). Postsynaptic GABAB receptors appear to require strong stimuli for activation. In paired recordings of neocortical and hippocampal neurones, GABAB receptors were detected in a small fraction of neurones only upon application of a train of stimuli (A. Thompson, London, UK). This non-linearity in stimulus–response properties of GABAB receptors was shown by computational modelling to underlie the generation of spike-and-wave discharges (SWDs) in absence seizures and sleep spindles in the thalamo-cortical system (A. Destexhe, Québec, Canada). However, the contribution of GABAB receptors for SWDs was questioned by in vivo recordings in genetic absence epilepsy rats (V. Crunelli and N. Leresche, Cardiff, UK and Paris, France). Unlike previous data from in vitro studies, these results suggest that SWDs in this model of absence epilepsy do not depend on GABAB receptor activation in vivo. A number of presentations focused on the role of GABAB receptors as potential therapeutic targets. These included the control of epilepsy (Y. Ito, Funabashi, Japan; L. S. Leung, London, Canada; G. D. Ritchie, Wright-Patterson Base, USA), the regulation of the hypothalamo– pituitary axis (M. J. Kelly, Portland, USA; I. Shibuya, Kitakyushu, Japan), the enhancement of anti-depressant drug action (F. Artigas, Barcelona, Spain), the suppression of cocainewithdrawal symptoms (J. P. Gallager, Galveston, USA) and the alleviation of pain (D. H. Hammond, Chicago, USA). With regard to pain, clear-cut evidence suggests that baclofen can reduce pain perception in animal models of central sensitization but fails to do so in humans. A
R
E
P
O
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T
possible explanation might be the ‘uncoupling’ of GABAB receptors induced by chronic pain, as suggested by autoradiographic data in a model of monoarthritic pain (N. G. Bowery, Birmingham, UK).
Concluding remarks Most, if not all, GABAB receptors are heterodimers formed from GABABR1 and GABABR2 subunits. The interaction of these two 7TM receptor proteins represents a novel principle of receptor assembly and signal transduction which will be a major focus of future GABAB receptor research. In addition, gene-targeting experiments are expected to determine whether GABABR2, apart from being a partner in the heterodimer, functions as a receptor in its own right. Furthermore, knockout experiments, performed in a regionor cell-specific manner, will help identify the role of GABAB receptors in complex CNS functions and pathologies. Recognizing the regional and cellular relevance of GABAB receptors will be essential in identifying novel drug targets. Thus, based on the new biology of GABAB receptors, a new pharmacology could be on the horizon at the time of the next GABAB receptor meeting.
Note added in proof Recently, five groups have had papers published or are in press on the cloning of GABABR2 (Refs 6–10).
References 1 Bowery, N. G. and Brown, D. A. (1997) Nature 386, 223–224 2 Kaupmann, K. et al. (1997) Nature 386, 239–246 3 Bettler, B., Kaupmann, K. and Bowery, N. G. (1998) Curr. Opin. Neurobiol. 8, 345–350 4 McLatchie, L. M. et al. (1998) Nature 393, 333–339 5 Dwyer, N. D. et al. (1998) Cell 93, 455–466 6 Kaupmann, K. et al. (1998) Nature 396, 683–687 7 White, J. H. et al. (1998) Nature 386, 679–682 8 Jones, K. A. et al. (1998) Nature 386, 674–679 9 Kuner, R. et al. (1999) Science 283, 74–77 10 Ng, G. Y. K. J. Biol. Chem. (in press)
TiPS – March 1999 (Vol. 20)
Acknowledgement The 4th International GABAB Symposium was made possible by the support of the American Society for Pharmacology and Experimental Therapeutics and Novartis Pharma AG.
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