Serotonin Receptors and their Function

Serotonin Receptors and their Function

G.A. Kennett, Assistant Director
Neurobehavioural Research
SmithKline Beecham Pharmaceuticals
New Frontiers Science Park, Third Avenue
Harlow, Essex, CM19 5AW, UK

Introduction

5-HT has been implicated in the aetiology of many disease states and may be particularly important in mental illness, such as depression, anxiety, schizophrenia, eating disorders, obsessive compulsive disorder (OCD), migraine and panic disorder. Indeed, many currently used treatments of these disorders are thought to act by modulating serotoninergic tone. During the last decade, multiple 5-HT receptor subtypes have been characterised. This has led to the realisation that many treatments acting via the serotonergic system, such as selective serotonin reuptake inhibitor (SSRI) antidepressants which increase presynaptic 5HT function, or migraine prophylactics like cyproheptadine which are 5-HT receptor antagonists, have non-selective effects on postsynaptic 5-HT receptor subtypes. The development of more selective ligands may therefore lead to treatments with increased efficacy and reduced side effects. Alternatively, selective ligands may form completely novel therapies.

Initially, receptor subtypes were characterised using pharmacological tools only. On the basis of the receptor binding profiles, common secondary messenger coupling and the functional activity of ligands, four main subgroups of 5-HT receptors, termed 5-HT1, 5-HT2, 5HT3 and 5-HT4, were identified. More recently, molecular biological techniques have both confirmed this classification, in that each subgroup has been found to have relatively dissimilar protein structures, and led to the identification of novel 5-HT receptors (5-HT1F, 5-HT5, 5-HT6 and 5HT7) enabling them to be cloned, expressed in cultured cell lines and pharmacologically and functionally characterised. Knowledge of 5-HT receptor cDNA sequences has also allowed antibody and antisense techniques to be employed.

The 5-HT1 receptor family

At least five 5-HT1 receptor subtypes have been recognised, 5-HT1A, 5-HT1B (formerly also termed 5-HT1Db), 5-HT1D (formerly 5HT1Da), 5-HT1E and 5-HT1F. All are seven transmembrane, G-protein coupled receptors (via Gi or Go), encoded by intronless genes, of between 365 and 422 amino acids with an overall sequence homology of 40%. These receptors are all thought to be negatively linked to adenylyl cyclase.1

5-HT1A

This receptor subtype which is located on human chromosome 5cen-q11 is widely distributed in the CNS, particularly the hippocampus, septum and amygdala, areas thought to be associated with the control of mood. The receptor is negatively coupled to adenylyl cyclase, and principally causes hyperpolarisation. Interestingly, 5-HT1A receptors in the raphe nuclei, act as somatodendritic autoreceptors which inhibit neuronal cell firing and 5-HT release onto postsynaptic sites. Blockade of 5-HT1A somatodendritic autoreceptors in rodents has little effect on extraneuronal 5-HT levels alone, but potentiates the increase seen after SSRIs. It has therefore been proposed that desensitisation of these 5HT1A autoreceptors may underlie the ability of chronic, but not acute SSRI administration to raise synaptic cleft 5-HT.1 Coadministration of the 5HT1A receptor antagonist, pindolol, is also reported to enhance the therapeutic efficacy and shorten the onset of action of SSRIs in depressive patients.2 If true, this may argue against preclinical electrophysiological evidence that the postsynaptic 5-HT1A receptors mediate the antidepressant effects of SSRIs2 as this should also be antagonised by pindolol.

Activation of the postsynaptic 5HT1A receptor in rats results in a characteristic 5-HT syndrome consisting of flat body posture, forepaw treading and headweaving, hypothermia and ACTH release.3 Stimulation of postsynaptic 5-HT1A receptors may also cause anxiogenic-like responses.3 In contrast, activation of presynaptic 5-HT1A receptors induces both hyperphagia and anxiolytic-like effects in rats and hence may account for the clinical anxiolytic efficacy of the 5HT1A receptor agonists, buspirone and gepirone.3, 4 5-HT1A receptor agonists are also active in animal models of depression such as the forced swimming test1 which is deemed consistent with evidence that SSRIs are antidepressant through 5-HT1A receptor activation,2 and with clinical evidence that 5-HT1A agonists may have antidepressant efficacy.3, 4

Several agonists show selectivity for the 5-HT1A receptor, particularly 8-hydroxy-di-n-propylamino tetralin (8-OH-DPAT, table 1), which acts as a full agonist in most systems, while the non benzodiazepine anxiolytics, buspirone and gepirone, and other ligands such as MDL 72832 are partial agonists.1 The synthesis of selective and silent antagonists at this receptor has proved more difficult. Several apparent antagonists have been characterised, such as NAN 190, BMY 7378, MDL 73005EF, WAY 100,135, UH 301, spiroxatrine and SDZ 216525. However, all have demonstrated partial agonist properties in studies of somatodendritic autoreceptor function,4 perhaps due to the much larger receptor reserve associated with these as opposed to postsynaptic receptors.1 To date, the only selective high affinity silent antagonist at this receptor is WAY 100,6355 (table 1).

5-HT1B

The 5-HT1B receptor is located on human chromosome 6q13 and is concentrated in the basal ganglia, striatum and frontal cortex. The receptor is negatively coupled to adenylyl cyclase. The receptor was originally defined by its pharmacology and, due to species differences in the binding affinity of key ligands such as the b-adrenoceptor antagonist, cyanopindolol (which has a higher affinity for the rat and mouse homologue, table 1), was thought to exist only in rodents. More recently, the amino acid sequence of the receptor has been characterised and found to be 93% identical overall and 96% identical within the transmembrane domains with that of the 5-HT1Db receptor, a close homologue found in higher species, with similar distribution and function.1 Indeed, the differences in the pharmacology of these two homologues are now attributed to the mutation of a single amino acid in the transmembrane spanning region.1 Thus, it has recently been agreed to classify the receptors as species homologues of the same receptor termed h5-HT1B (formerly 5-HT1Db) and r5-HT1B with the h and r prefix referring to the human and rat species respectively.6

Studies have established that the terminal autoreceptor controlling 5-HT release in both the rat, guinea pig and man is of the 5-HT1B type.1 It has therefore been argued that selective antagonists might prevent 5-HT negative feedback via this site, increasing extraneuronal 5-HT as do SSRI antidepressants. However, the first potent (pKD 9.9) and selective 5-HT1B/1D antagonist7 to be reported, GR 127935, decreased extraneuronal 5-HT in the brain when administered systemically to guinea pigs. This may be due for to its partial agonist activity at the 5-HT1B receptor8 or due to the presence of 5HT1D receptors on the raphe acting as somatodendritic autoreceptors9 at which GR 127935 also has agonist properties.10 This interpretation is supported by the failure of the recently characterised selective silent 5-HT1B antagonist, SB-224289 (pKD 8.0 >60 fold selective over 5-HT1D and other receptors, table 1), to increase 5-HT release in the frontal cortex.11 Preliminary evidence suggests that a small population of 5HT1B receptors may also act as somatodendritic autoreceptors in the raphe.12 In addition to its actions as an autoreceptor, the 5-HT1B receptor may act as a terminal heteroreceptor controlling the release of other neurotransmitters, such as acetylcholine, glutamate1 and dopamine.13 With the exception of GR 127935 and SB-224289 and GR 5556214 (table 1), there are few selective antagonists for the 5-HT1B receptor. The most commonly used, pindolol, cyanopindolol (table 1) and SDZ 21009 are equipotent at the 5-HT1A receptor, where they have antagonist or partial agonist properties, and are more potent as b-adrenoceptor antagonists1 (table 1) while isamoltane, has approximately 30 fold selectivity for the 5HT1B over the 5HT1A receptor, but also has greater affinity for b-adrenergic receptors.15

Stimulation of central postsynaptic 5-HT1B receptors in mice,16 but not rats,17 causes hyperlocomotion while penile erection and hypophagia are also reportedly 5-HT1B receptor mediated on the basis of the pharmacological studies carried out to date.3 Postsynaptic 5-HT1B receptor activation in another species, the guinea-pig is reported to induce hypothermia,18 while the hypothermic response to 5HT1B agonists in the rat remains to be fully characterised.19 The putative 5HT1B receptor agonist, anpirtoline, has analgesic and antidepressant-like properties in rodents20 and it is of interest that mutant mice lacking the 5-HT1B receptor are reported to be both highly aggressive21 and have an increased preference for alcohol.22

Interest in 5-HT1B receptor agonists has been generated by the antimigraine properties of sumatriptan, a non selective 5-HT1D and 5-HT1B receptor agonist with low selectivity against other receptors in functional studies1, 23 (table 1). This compound may act either via constriction-mediating 5HT1B receptors on cerebral arteries or by blocking neurogenic inflamation and nociceptive activity within trigeminovascular afferents. This latter action has been argued to be 5HT1B receptor-mediated as protein extravasation induced by trigeminal ganglion stimulation is blocked by sumatriptan, the selective 5-HT1B receptor agonist, CP-93,129 (>100 fold selectivity over 5-HT1A, h5-HT1B and 5-HT2 receptors,24 table 1) and the close structural analogue of sumatriptan, CP-122,288,25 in wild type, but not in mutant mice lacking the 5-HT1B receptor.26 However, while 5-HT1B receptor mRNA has been detected in rat, only 5-HT1D mRNA was detected in the guinea pig and human trigeminal ganglia.25 Thus, in man the antimigraine properties of sumatriptan may be either 5-HT1B or 5HT1D receptor-mediated (although see section on 5-HT1F).

RU 24969 was the first reported full agonist at the 5-HT1B receptor, but, in binding studies, it is only 5 fold selective over the 5-HT1A and 5-HT1D1 receptor. CGS 12066B, a compound of a different structural class, also possesses over 10 fold selectivity over the 5-HT1A receptor in functional studies and is a full agonist,27 while anpirtoline has only five fold selectivity in binding assays.20 Trifluoromethylphenylpiperazine (TFMPP) and m-chlorophenylpiperazine (mCPP, table 1), are both poorly selective partial agonists.27 Other selective 5-HT1B agonists characterised include MK 464,28 and the brain penetrant agonists, BW 311C90,29 SKF 99101H,30 and GR 46611.18 The most potent agonist reported is L-694,247 (pKD 10.0),31 while 5(nonyloxy)tryptamine has the greatest selectivity over the 5-HT1A receptor.32 None of these, however, differentiates between the 5-HT1B and the 5-HT1D receptor subtypes and only MK 464 and 5-(nonyloxy)tryptamine have pronounced selectivity over the 5HT1A receptor.

5-HT1D

The 5-HT1D receptor (formerly termed 5HT1Da) has 63% overall structural homology with the 5-HT1B receptor (formerly 5-HT1Db) and a 77% amino acid sequence homology in the seven transmembrane domains. The receptor is located on human gene 1p36.3-p34.3 and is negatively linked to adenylyl cyclase. Low levels of the 5-HT1D receptor mRNA are found in the rat brain, predominantly in the caudate putamen, nucleus accumbens, hippocampus and cortex, but also in the dorsal raphe and locus coeruleus.1

It has been proposed that neurogenic inflamation and nociceptive activity within trigeminovascular afferents may be 5-HT1D receptor mediated due to the presence of 5-HT1D, but not 5-HT1B receptor mRNA in the guinea pig and human trigeminal ganglia24 (see 5-HT1B and 5-HT1F sections). Thus, antagonism of plasma extravasation induced by electrical stimulation of the trigeminal nerves by the antimigraine 5HT1B/1D receptor agonist, sumatriptan (table 1), may be 5-HT1D receptor mediated. There are, however, no selective 5-HT1D agonists at the present time, although a number of potent and relatively selective 5-HT1B/1D receptor agonists have been characterised (sumatriptan, CP-122,288,25 MK 464,28 BW 311C90,29 and the more brain penetrant agonists, SKF 99101H,30 GR 4661118 and L-694,247 (pKD 10.0),31 as described in the section on the 5-HT1B receptor above.

The location of 5-HT1D receptor mRNA in the raphe,1 suggests that it may function as an 5-HT autoreceptor. Indeed, there is electrophysiological, release33 and voltammetric evidence13 to this effect. This data could be further substantiated by the use of ketanserin and ritanserin, which in addition their high affinity for the 5-HT2A site also have high affinity for the 5-HT1D, but not the 5HT1B site (table 1).34

5-HT1E

The 5-HT1E receptor was first characterised in man as a [3H]-5-HT binding site in the presence of 5-carboxyamidotryptamine (5-CT) to block binding to the 5HT1A and 5-HT1D receptors. Human brain binding studies have reported that 5-HT1E receptors (representing up to 60% of 5HT1 receptor binding) are concentrated in the caudate putamen with lower levels in the amygdala, frontal cortex and globus pallidus.1 This is consistent with the observed distribution of 5HT1E mRNA.1 The receptor has been mapped to human chromosome 6q14-q15, is negatively linked to adenylyl cyclase and consists of a 365 amino acid protein with seven transmembrane domains.1 There are no reported selective or high affinity ligands for this receptor (except for 5HT itself) and its function is currently unknown.

5-HT1F

This receptor subtype is most closely related to the 5-HT1E receptor with 70% sequence homology across the 7 transmembrane domains. mRNA coding for the receptor is concentrated in the dorsal raphe, hippocampus and cortex of the rat and also in the striatum, thalamus and hypothalamus of the mouse.1 5-HT1F receptor mRNA has been detected in human brain and is also present in the mesentery and uterus.1 The receptor is negatively linked to adenylyl cyclase. The antimigraine 5-HT1B/1D agonist, sumatriptan, has almost equal affinity for the 5HT1F (pKi 7.6)35 as the 5-HT1B/1D receptors (pKI 8.4, 8.1 respectively, table 1). Thus, it has been hypothesised that the 5-HT1F receptor might be be a target for drugs with antimigraine properties. 5-HT1F mRNA has been detected in the trigeminal ganglia, whose stimulation leads to plasma extravasation in the dura, a component of neurogenic inflammation which is thought to be a possible cause of migraine.36 The first 5-HT1F receptor selective agonist, LY 334370 (pKD 9.4, see table 1)35 with >100 fold separation over the 5-HT1B/1D receptors has been claimed to block the effects of trigeminal nerve stimulation37 as does sumatriptan (see 5-HT1B section). LY 334370 has also been used as a radioligand and demonstrated a reasonable correlation between the receptor protein and mRNA distribution with highest binding in the cortical areas, striatum, hippocampus and olfactory bulb.38

The 5-HT2 receptor family

The 5-HT2 receptor family consists of three subtypes termed 5-HT2A, 5-HT2B and 5-HT2C. The latter site was was previously termed 5-HT1C before its structural similarity to the 5-HT2 family members was recognised1. All three are single protein molecules of 458-471 amino acids with a overall homology of approximately 50% rising to between 70-80% in the seven transmembrane domains. All three are thought to be linked to the phosphoinositol hydrolysis signal transduction system via the a subunit of the Gq GTP binding protein, although it is yet to be proven that the 5-HT2B receptor is so coupled in native tissue.1 Indeed, in human pulmonary artery endothelial cells, 5-HT2B receptor stimulation causes intracellular calcium release via a mechanism independent of phosphatidylinositol hydrolysis.39 A similar mechanism has been detected in the rat stomach fundus.40 In neither tissue was the 5-HT2B receptor coupled to phosphatidylinositol hydrolysis.

5-HT2A

This receptor (previously termed 5-HT2) is located on human chromosome 13q14-q21 and is widely distributed in peripheral tissues where it mediates contractile responses of many vascular, urinary, gastrointestinal and uterine smooth muscle preparations, platelet aggregation and increased capillary permeability in both rodent and human tissue.1 Centrally, it is principally located in the cortex, claustrum and basal ganglia.1 Stimulation of central 5-HT2A receptors in rodents causes head shaking and may mediate the effects of hallucinogens such as lysergic acid diethylamide (LSD) in man, the release of glutamate from the rat cerebellum and the release of b-endorphin, corticosterone, luteinising hormone and prolactin as well as adrenaline from the rat adrenal medulla.1

5-HT2A receptor antagonism may account for the enhanced efficacy against negative symptoms and low propensity for inducing extrapyramidal side effects of the atypical antipsychotic, clozapine, perhaps by increasing striatal dopamine release.41 Clinical trials of mixed dopamine D2/5HT2A receptor antagonists such as olanzapine42 and risperidone and of selective 5-HT2A receptor antagonists such as amperozide and MDL 100,907 are currently underway.43 5HT2A receptor antagonists such as ritanserin are also reported to improve sleep quality.44 As the vasoconstrictor effects of 5-HT2A receptor stimulation are markedly potentiated in hypertension and atherosclerosis, while low levels of 5HT potentiate the thrombogenic and vasoconstrictor effects of other neurotransmitters, selective 5-HT2A receptor antagonists, notably ketanserin, are in clinical use for hypertension.43

Despite the large number of compounds which have high affinity for the site, few are truly selective. Thus, ketanserin (70 fold selective over 5-HT2B and 5HT2C receptors, table 1) is a potent adrenergic a1 receptor antagonist while ritanserin has high affinity for 5-HT2C, 5-HT2B and H1 receptors. The recently developed ligand MDL 100,907 (reportedly 300 fold selective over the rat 5-HT2C receptor) may offer advantages over ketanserin, while spiperone (100 fold selective over 5HT2B and 5-HT2C, table 1), risperidone and pirenperone have good separation over the 5-HT2B and 5-HT2C receptors but are potent dopamine D2 or histamine H1 receptor antagonists. ICI 170,809 is relatively selective for the 5-HT2 receptor subtypes, lacking significant affinity for adrenergic or histamine receptors, but LY 53857 has marked affinity for 5-HT3 and only 30 fold selectivity over 5HT1A sites. Amperozide, whilst almost 100 fold selective over the 5-HT2C receptor is only 10 fold selective over adrenergic a1 and 24 fold selective over dopamine D2 sites. Most 5-HT2A agonists, such as LSD, 2,5-dimethoxy-4-iodophenylisopropylamine (DOI) and (-) 2,5-dimethoxy-4-iodoamphetamine (DOM) have no selectivity over other 5HT2 receptor subtypes and are not well characterised, while a-methyl 5-HT (table 1) is 10 fold more potent at 5-HT2C and 100 fold more potent at 5-HT2B sites.44, 45

5-HT2B

This receptor located on chromosome 2q36.3-2q37.1 mediates contraction of the rat stomach fundus and endothelium-dependent relaxation of the rat and cat jugular veins and possibly of the pig pulmonary artery, via nitric oxide release.45 5-HT2B receptor mRNA has been detected throughout the mouse, rat and guinea pig colon and small intestine.46 Interestingly, the 5-HT2B receptor mRNA was much more abundant in foetal small and large intestine at E13-16, after which levels declined suggesting a role in development47 and a splice variant has been reported in guinea pig, but not rat, apparently lacking the third and part of the fourth transmembrane domains.48 In man, 5-HT2B receptor mRNA is expressed in low levels in the brain, and at much higher levels in the placenta, lung, liver, kidney, heart, intestine and stomach.44 Recent receptor specific antibody studies in the rat have reported the presence of receptor protein in the amygdala, septum, hypothalamus and cerebellum.49 In rodent studies, stimulation of 5-HT2B receptors has been reported to cause modest anxiolysis50 hyperphagia and reduced grooming.51 They may also be involved in the precipitation of migraine and the action of the 5-HT2 receptor antagonist migraine prophylactics, cyproheptadine, pizotifen and mianserin.52 The 5-HT2B receptor has also been postulated to mediate mesenteric artery contraction in hypertensive, but not normotensive rats.53 Finally, it has been reported that activation of 5-HT2B receptors stably expressed in a mouse fibroblast cell line caused mitogenesis via MAP kinase activation which was linked to tumor transforming activity.54

The receptor has been relatively easy to characterise in in vitro models as non-selective 5-HT2 receptor antagonists such as ICI 170,809, ritanserin, metergoline and LY 53857 antagonise 5-HT2B-mediated effects, unlike 5-HT2A receptor selective antagonists such as ketanserin (table 1). Also the adrenergic a2 receptor antagonists yohimbine and rauwolscine are potent antagonists and have low affinity for the 5-HT2C and 5HT2A sites.45 Recently, SB 200646A and SB 206553 have been reported as selective 5-HT2C/2B receptor antagonists with 50-100 fold lower affinity for the 5-HT2A and other sites55 (table 1), while SB 204741 has been reported as the first selective 5-HT2B receptor antagonist with approximately 100 fold selectivity over the 5-HT2C and 5-HT2A sites45 (table 1). A series of tetrahydro-b-carbolines have been characterised with improved potency for the 5-HT2B receptor.56 Of these, LY 266097 has a pKI of 9.7 on the human cloned 5-HT2B receptor with 100 fold selectivity over the human 5-HT2C and 5-HT2A sites. Unfortunately, selectivity over other sites is unknown at present. Agonists with some selectivity are also available. a-methyl 5-HT is a full agonist with high affinity for the 5-HT2B site (pEC50 8.4) and lower affinity for both 5-HT2C (pEC50 7.3) and 5-HT2A (pEC50 6.1, table 1). 5-Methoxytryptamine is also 25 and 400 fold selective over the 5HT2A and 5HT2C sites respectively. Finally, BW 723C86 has recently been reported to have 10 fold selectivity over the 5HT2C and 100 fold selectivity over the 5HT2A receptors in the rat, although less against cloned human receptors45, 50 (table 1). These tools should allow the functional role of the 5-HT2B receptor to be explored.

5-HT2C

5-HT2C specific antibodies have recently been used to show the presence of the receptor protein in the choroid plexus (highest density) and at a lower level in the cerebral cortex, hippocampus, striatum, and substantia nigra of rat and a similar distribution in man. This is consistent with previous autoradiographic studies and with mRNA distribution. There is, at present, no evidence of the existence of this receptor or its mRNA in peripheral tissues.1,57 The receptor has been mapped to human chromosome Xq24. No splice variants have been reported but the receptor is capable of posttranscriptional modification whereby adenosine residues can be represented as guanosine in the second loop to yield 4 variants. To date no pharmacological differences between the variants has been detected, but when expressed in cell lines they respond to 5-HT, as assessed by phosphatidylinositol hydrolysis, with different EC50 values (between 4 and 95 nM).58

The presence of very high levels of the 5-HT2C receptor in the choroid plexus has led to the suggestion that it may regulate cerebral spinal fluid production. It has also been hypothesised to mediate the migraine prophylactic effects of the non-selective 5-HT2 receptor antagonists, methysergide, pizotifen, cyproheptadine and mianserin as well as the anxiogenic and panic precipitating properties of the 5-HT2C agonist, mCPP (table 1), in man and rats59 and this is consistent with the anxiolytic-like actions of selective 5-HT2C receptor antagonists in animal models.60 This latter effect may relate to the mode of action of SSRIs which desensitise 5-HT2C receptor responsivity.44 mCPP also induces hypophagia, penile erections, oral dyskinesias and hyperthermia in rats, all of which are thought to be 5-HT2C receptor mediated.44, 61 In addition, mCPP can disrupt sleep in man while the non-selective 5-HT2 receptor antagonist, ritanserin, improves sleep quality, possibly via an action at the 5HT2C receptor.44 Finally, mutant mice lacking the 5-HT2C receptor are reported to suffer spontaneous convulsions, exhibit proconvulsant properties, obesity through increased food intake and cognitive impairment. They are also insensitive to the hypolocomotor and anxiogenic effects of mCPP.62, 63 Proconvulsant and hyperphagic properties were not observed in adult rats treated with selective 5HT2C receptor antagonists, suggesting that developmental or neuroadaptive changes may be responsible for these latter effects.60

Much of the above evidence relies on the clinical and preclinical studies that have used mCPP as a pharmacological tool. However, this compound is a partial agonist at the 5-HT2C receptor and has, at best, approximately 10 fold selectivity as for the 5-HT2C site in rat and man.44 A more selective alternative to mCPP is MK 212 which has 25 fold selectivity for the 5-HT2C over the 5-HT2A site and is an agonist at both receptors, while mCPP is an antagonist at the 5HT2A receptor (table 1). Recently, a new and highly selective 5-HT2C receptor full agonist, Ro 60-0175, has been characterised with a reportedly high, but as yet unpublished affinity for the human cloned 5-HT2C receptor and a claimed separation of at least 2.5 log units over 40 other receptors.64 Most 5HT2 receptor antagonists such as LY 53857, ICI 170,809, ritanserin and mianserin do not discriminate between the subtypes.44 Fortunately, novel ligands have been developed that should help unravel the functional role of the 5HT2C site. SB 200646A59 and SB 20655355 have respectively 50 and 100 fold selectivity for the 5-HT2C and 5HT2B receptor over the 5-HT2A site and higher selectivity over all other sites at which they have been tested. These compounds are now complemented by the characterisation of two truly selective 5-HT2C receptor antagonists, SB 242084 with an affinity of 9.0 for the 5-HT2C receptor and over 100 fold selectivity over other sites60 (table 1) and RS-102221 with an affinity of 8.4 for the human 5-HT2C receptor and at least 100 fold selectivity over all other receptor binding sites assesssed65 (table 1). It is of interest that these two compounds have rather different profiles in vivo which may indicate different degrees of brain penetration.

The 5-HT3 receptor

The 5-HT3 receptor binding site is widely distributed both centrally and peripherally and has been detected in a number of neuronally derived cells. The highest densities are found in the area postrema, nucleus tractus solitarius, substantia gelatinosa at all levels and nuclei of the lower brainstem such as the trigeminal nucleus and the dorsal vagal complex. It is also found in higher brain areas such as the cortex, hippocampus amygdala and medial habenula, but at lower densities.1 Peripherally, it is principally found on the neurones of the sensory and enteric nervous systems and pre-and postganglionic autonomic neurones. Unlike other 5-HT receptors, 5-HT3 receptor subunits form a pentameric cation channel that is selectively permeable to Na+, K+ and Ca++ ions causing depolarisation.1 The receptor displays marked species variation, but there is little evidence for receptor subtypes within a species, although 5-HT3 ligands have different affinities in mouse cortex and ileum.1 This may be accounted for by multiple receptor binding sites.66

In vivo, administration of 5-HT3 receptor ligands can either stimulate or inhibit cardiac function, induce vasodilation, affect lung and intestinal function, cause pain and sensitisation of nociceptive neurons and induce nausea and vomiting. This latter action is thought to underlie the emetic side effects of cancer chemotherapy and radiotherapy and has led to the use of selective 5-HT3 receptor antagonists as antiemetic agents.1 Central 5-HT3 receptor antagonists are reported to have anxiolytic actions in some, but not all rodent and primate models of anxiety and clinical studies are underway. 5-HT3 receptor antagonists have also been reported to induce cognitive enhancing effects and to reduce dopamine function in rats suggesting utility as procognitive and antipsychotic agents.1

Highly potent and selective 5-HT3 receptor antagonists are available including ondansetron, granisetron, BRL 46470A67 (table 1), GR 65630 and MDL 72222. Another ligand is tropisetron. However, unlike the other ligands mentioned, this has mM affinity for the 5-HT4 site.1 The number of agonists available is rather more limited. 2-methyl 5-HT and phenylbiguanide (PBG) have been widely used, while chlorophenylbiguanide (CPBG) is the agonist with highest affinity (1.4 nM compared with PBG 141 nM and 2-Methyl 5-HT 207 nM vs [3H]-GR 67330) (table 1) for the site. Both PBG and CPBG have greater selectivity than 2-methyl 5-HT for the 5-HT3 receptor, but both potently block dopamine reuptake.1, 68 All three are partial agonists at the receptor which may explain why CPBG is considerably less potent functionally than predicted by its high affinity.

The 5-HT4 receptor

Receptor binding studies have established that the 5-HT4 receptor is highly concentrated in areas of the rat brain associated with dopamine function such as the striatum, basal ganglia and nucleus accumbens where they may be located on GABAergic or cholinergic interneurons and/or on GABAergic projections to the substantia nigra.69 Recently, two C terminal splice variants of 387 (short) and 407 (long) amino acids have been identified with different distributions.70 The receptor is functionally coupled via the G G-protein.

Peripherally, stimulation of 5-HT4 receptors on the myenteric plexus of the guinea pig and rat oesophagus, and guinea pig and human colon cause contractions.71 Thus, 5-HT4 receptor antagonists may be of use in iritable bowel syndrome. 5-HT4 receptors in the mucosa of the rat colon and elsewhere are involved in secretory processes.71 They may also mediate vomiting in ferrets, possibly by activating vagal afferents.71 5-HT4 receptors in the heart induce tachycardia and positive inotropic effects when stimulated and may be of importance in cardiovascular pathology. 5-HT4 receptor activation may also cause cortisol secretion and bladder contraction in man.71 Centrally, 5-HT4 agonists can increase striatal dopamine release, an effect blocked by selective antagonists.72 If this interaction with dopamine were also observed in the nucleus accumbens, 5-HT4 receptor antagonists might be of use as antipsychotic agents. However, the selective 5-HT4 antagonist, SB-207266A had no effect on several models of in vivo dopamine function such as amphetamine, morphine and nicotine-induced hyperactivity.73 In the rat frontal cortex, 5-HT4 receptor stimulation facilitates acetylcholine release74 and may thus have cognition enhancing effects while in the hippocampus, it may result in increased 5-HT release,75 suggesting a possible role in the mediation of anxiety. This latter area has evoked some controversy as the selective 5-HT4 receptor antagonists, GR 113808, SB 204070A and SB 207266A, have modest anxiolytic-like actions in some rodent models of anxiety, yet GR 113808 is reported to antagonise the anxiolytic-like effects of the benzodiazepine anxiolytic, diazepam, in others.76

There are several highly potent and selective 5-HT4 receptor antagonists such as GR 11380877 (3000 fold selective over all other receptors, table 1), RS 23597-190 (125 or more selectivity over dopamine and 5-HT3 receptors, LY 297582 (250 fold selective over 5-HT3 sites,71 SB 207266 (3000 fold selective over all other sites,78 (table 1), SB 204070 (5000 fold selective over all other sites78) and SB 203186 (pKB 8.7 in human atrium79). Another compound, SDZ 205-557, has only 30 fold selectivity over the 5-HT3 site and is less well characterised.71 Recently, a novel ketone compound, RS 39604, has been reported which is perhaps the most potent (pKI 9.5), orally active and selective 5-HT4 antagonist with 1000 fold selectivity over 5HT3, 5-HT1A, 5-HT2C and other receptors80 (table 1).

The first 5-HT4 agonists reported were benzamides such as renzapride (pKI guinea pig striatum, 7.0, IA 1.3), cisapride (pKI 7.5, IA 1.3) and zacopride (pKI 6.8, IA 1.4).71 Later, benzimidazolones, such as BIMU8 (pKI 7.9, IA 1.2) and BIMU1 (pKI 7.8, IA 0.7), were characterised,71 however, all have substantial, or in the case of BIMU1 and BIMU8, higher affinity for the 5-HT3 receptor.7

Page Last Updated: 06/29/2005