Neurotransmitters Agonists And Antagonists




10.1.1 Introduction 388 Neurotransmitters 388 Receptors 390 Neuronal signal effectors 391 FURTHER READING 391




10.2.1 Introduction 391

10.2.2 Subpopulations of 5-HT Receptors 392

10.2.3 Receptor populations and ligands 394

10.2.4 Possible applications of 5-HT agonists and antagonists 407

10.2.5 Epilogue 408 FURTHER READING 408


10.3.1 Introduction 410

10.3.2 H1-Receptors 411

10.3.3 H2-Receptors 415

10.3.4 H3-Receptors 417 FURTHER READING 421


10.4.1 Introduction 422

10.4.2 Dopamine agonists 423 Aminotetralins and related compounds 423 Aporphine alkaloids 425 Ergot alkaloids 425 Benzazepines 427 Miscellaneous structures 427 Selective agonists 428

10.4.3 Dopamine antagonists 429 FURTHER READING 432

10.1 OVERVIEW Robert D.E.Sewell

10.1.1 Introduction

Over recent years, our concepts about neurotransmitters have been transformed firstly, because the number of putative transmitters has increased to upwards of 40 candidates and secondly, because our understanding of neurotransmitter function has widened to cover a more diverse range of effects. The aim of this chapter, is not to embrace the medicinal chemistry of all the neurotransmitters and their agonists and antagonists, but more to concentrate on three selected candidates, namely serotonin, histamine and dopamine. Amongst these three examples, there are receptor types which typify both of the major functional classes of neuronal receptor (i.e. ionotropic and metabotropic receptors). Neurotransmitters

Neurotransmitters are a varied assortment of substances implicated in the transfer of signals across chemical synapses. These neuronal elements consist of narrow clefts which separate presynaptic nerve terminals from receptors located on postsynaptic membranes found on subsequent neurones, muscles or glands. In addition to postsynaptic receptors, there are receptors sited presynaptically on the nerve terminals themselves. These are sometimes designated as autoreceptors; capable of responding to released neurotransmitter, providing a negative feedback function concerned with regulation of transmitter release. Other neuronal synapses exist which rely on electrical rather than chemical transmission and this represents a very fast mode of communication between cells.

The chemical neurotransmitters (Figure 10.1) may be classed into three major categories:

(1) Simple amino acids like glutamate, y-amino butyric acid (GABA) and glycine account for transmission at a high proportion of CNS synapses. They are relatively fast communicators and occur at the highest levels of all the neurotransmitter groups in brain tissue (micromoles per gram of tissue).

(2) The amine neurotransmitters are composed of 'classical' transmitters such as acetylcholine, serotonin (5-hydroxytryptamine—5-HT) histamine and the catecholamines (dopamine and noradrenaline). Also included in this group are purine neurotransmitters like adenosine and adenosine triphosphate (ATP). All the neurotransmitters in the group are found in moderate concentrations in neuronal tissue (nanomoles per gram of whole brain tissue) and tend to exert a slower modulatory type of action.

(3) The neuropeptides occur at the lowest neuronal levels of all three groups (<nanomoles per gram of whole brain tissue). In addition to release at synapses, they may also be released at non-synaptic locations to diffuse more freely onto receptor sites to produce slow long-lasting effects and, in such circumstances, might be more appropriately considered as local neuromodulators. Glutamics might be more appropriately considered as local neuromodulators. Glutamics

Neuromodulator Drug Examples
Figure 10.1 Examples of neurotransmitters.

It is not uncommon for neurones to contain more than one neurotransmitter, a typical example being neuropeptides occurring in combination with amines in one and the same neurone. Receptors

Neurotransmitter effects are mediated via receptors located in neuronal membranes and these are thought to be of two functional classes:

Metabotropic receptors

Metabotropic receptors are linked to intracellular proteins which transduce signals across the cell membrane. These proteins are known as G-proteins (so-called because they are coupled to the guanosine nucleotides GDP or GTP). They are comprised of three subunits (a, p, and y) of which the a subunit possesses GTP-ase activity. The DNA sequences coding for the majority of G-protein coupled metabotropic receptors do have some sequence homology so they are viewed as a superfamily and over 100 constituent members have been cloned.

Binding of a neurotransmitter or an agonist with a metabotropic receptor often stimulates the formation of an intracellular second messenger by means of an effector enzyme. The second messengers include adenosine 3',5'-cyclic phosphate (c-AMP), inositol phosphates, diacylglycerol and arachidonate. The overall mechanism invariably involves an amplification process since a single neurotransmitter/agonist-receptor complex may activate several G-protein molecules in turn to generate many secondary messenger molecules intracellularly. There appear to be subtypes of virtually all metabotropic receptors which differ either in location and/or second messenger coupling, but they all possess seven trans-membrane hydrophobic spanning regions in their peptide sequences, with an extracellular amino terminus and an intracellular C-terminus. The membrane spanning portions of receptor proteins form a-helices and possess similarities within each receptor group so it is probable that the agonist binding site resides at least partly in the membrane spanning region.

Ionotropic receptors

Ionotropic receptors are linked directly to ion channels in the neuronal membrane. They are responsible for transient (submillisecond) increases in the conductance of specific ions in each instance and this gives rise to rapid synaptic transmission. There are several examples which include the nicotinic acetylcholine receptor, the 5-HT3 receptor and receptors for the amino acids such as GABAA, glycine and N-methyl-D-aspartate (NMDA). Ionotropic receptors are comprised of 4-5 protein subunits in a complex linked to an ion channel. The basic structure of each subunit consists of a protein which loops in and out of the neuronal membrane. In the case of the nicotinic receptor, which has been comprehensively studied, there are 5 subunits (a2, p, y, 5) each traversing the membrane a total of five times. Four of the spanning segments are hydrophobic in nature and are considered as truly integral transmembrane domains. The fifth segment has only one face of its a-helix structure which is hydrophobic while the other face is hydrophilic and, along with counterparts in the other 4 subunits, constitutes the lining of the ion channel interfacing with an aqueous environment. When an agonist binds to this pentameric receptor, a conformational change occurs in the complex which allows the passage of ions through the channel. Neuronal signal effectors

Both metabotropic and ionotropic receptors are coupled to effectors involving enzymes or ion channels and the modulation of cytoplasmic Ca2+ concentration features in several of these systems. Thus, ionotropic receptors can regulate cytoplasmic Ca2+ concentration either directly by conducting Ca2+ itself, or via permeability to monovalent cations which depolarise the membrane as Na+ influxes and this subsequently activates Ca2+ channels in the proximity of the receptor.

Metabotropic receptors may activate slower biochemical effector processes which modulate Ca2+ concentration intracellularly through the following G-proteins:

(1) Gs which catalyses the conversion of adenosine triphosphate (ATP) to cAMP by stimulating adenylate cyclase whilst concomitantly activating Ca2+ channels.

(2) Gi/Go which inhibits both adenylate cyclase and Ca2+ channels whilst simultaneously activating K+ channels.

(3) G11/Gq which activates phospholipase C to catalyse the hydrolysis of phosphatidylinositol biphosphate (PIP2) to 1,4,5-inositol triphosphate (IP3) and diacylglycerol (DAG). IP3 may cause Ca2+ release from such intracellular storage sites as the endoplasmic reticulum. DAG on the other hand, stimulates the enzyme protein kinase C (PKC) which is capable of phosphorylating other cellular or membrane proteins, enzymes and ion channels. If ion channel proteins are phosphorylated, this may alter ion fluxes albeit in a slower manner than that generated by ionotropic receptors.


Cascieri, M.A., Fong, T.M. and Strader, C.D. (1995) Molecular characterization of a common binding site for small molecules within the transmembrane domain of G-protein coupled receptors. Journal of Pharmacological and Toxicological Methods 33, 170-185.

Gilman, A.G. (1987) G proteins: transducers of receptor-generated signals. Annual Review of Biochemistry 56, 615-649.

Nicholls, D.G. (1994) Proteins, transmitters and synapses. Oxford: Blackwell Scientific Publications.

Strader, C.A., Fong, T.M., Tota, M.R. and Underwood, D. (1994) Structure and function of G-protein-coupled receptors. Annual Review of Biochemistry 63, 101132.

Strader, C.A., Fong, T.M., Graziano, M.P. and Tota, M.R. (1995) The family of G-protein-coupled receptors. FASEB 9, 745-754.

Unwin, N. (1993) Nicotinic acetylcholine receptor at 9A resolution. Journal of Molecular Biology 229, 1101-1124.

Watson, S. and Girdlestone, D. (1995) Receptor and ion channel nomenclature. Supplement (1995) Trends in Pharmacological Sciences.

10.2 SEROTONIN RECEPTORS AND LIGANDS Richard A.Glennon and Malgorzata Dukat

10.2.1 Introduction

Serotonin (5-hydroxytryptamine, 5-HT; 10.1) was discovered about 50 years ago and was the subject of extensive investigation during the 1950s and 1960s. Although it was

(10,1); 5-HT

implicated in numerous physiological functions and pathological and psychopathological conditions, research was hampered by a lack of suitable pharmacological tools with which to study serotonergic function, and by problems associated with interpretation of pharmacological results which seemed to vary depending upon the particular pharmacological model being used or physiological response being monitored. During the late 1960s and into the 1970s, the pace of 5-HT research slowed to a crawl. With the advent of radioligand binding techniques, particularly the rapid filtration method, and with the development of new psychopharmacological assay techniques and methodologies, there was renewed interest in centrally acting agents and neurotransmitters, in general, and in 5-HT in particular. Interest in 5-HT was further heightened with the discovery in late 1979 of two different populations (i.e., 5-HT1 and 5-HT2) of central 5-HT binding/receptor sites. For the first time, it was thought possible to explain some of the confounding results reported in the years before; that is, different populations of 5-HT receptors might mediate different responses and, further, might behave differently toward various serotonergic agents.

Studies during the 1980s and into the 1990s focussed broadly on, (i) discovery and characterization of additional populations of 5-HT receptors, (ii) identification of the functional significance of these receptors, and (iii) development of novel subtype-selective agonists and antagonists (Herndon and Glennon 1993). Radioligand binding techniques accounted for the early discovery of several new populations of 5-HT receptors; indeed, the number of proposed populations of sites began to increase at an alarming pace. Questions were raised concerning whether so many different populations of 5-HT receptors could actually exist, or whether they simply represented tissue or species homologues, minor variants of known 5-HT receptors, or artifacts resulting from the selectivity (or, more accurately, the non-selectivity) characteristics of the serotonergic agents being used to investigate the receptors. Subsequently, molecular biology confirmed that many of these populations represented different receptors and that certain others were species homologs. Such studies also resulted in the discovery of additional populations of sites.

10.2.2 Subpopulations of 5-HT receptors

At least 15 populations of 5-HT receptors have now been cloned (Table 10.1). Transmembrane homology amongst these populations ranges from about 30% to

>90%. As might be expected, low homology is associated with distantly related populations of 5-HT receptors whereas high homology is typically found between 5-HT receptors that display similar pharmacological profiles and second messenger systems. Likewise, species

Table 10.1 5-HT Receptor populations, second messengers and useful agents.

Selective antagonist

































DOXd (10.19)



DOXd (10.19)



DOXd (10.19)

GR 127935c (10.17) GR 127935c (10.17)



GR 127935c (10.17) GR 127935c (10.17)

5-HT3 Ion channel fflCPBGf (10.28) Zacopride 2-Me 5-HT (10.32)




SB 204070

(10.39) SB 207710




a 5-HT1 receptors are negatively coupled to adenylate cyclase [(-)cAMP] whereas certain other receptors are positively coupled [(+)cAMP]. The 5-HT2 family of receptors is coupled to phosphoinositide hydrolysis (IP3).

See text for discussion and additional examples. These agents are not necessarily specific for a given population of receptors but are merely useful agents that display some selectivity. The (-) symbol indicates that no selective agents have yet been identified. c No high-affinity agents have yet been found to discriminate between the two subpopulations.

d DOX refers to 1-(2,5-dimethoxy-4-X-phenyl)-2-aminopropane where X=-CH3 (DOM), -Br (DOB), or -I (DOI). e Ketanserin binds with lower affinity at 5-HT2B receptors than at 5-HT2A/2C receptors.

f fflCPBG=ffletachlorophenylbiguanide.

homologues usually display >90% homology. Populations of 5-HT receptors belong both to the G-protein and ion channel superfamilies of neurotransmitter receptors; the former category can be further categorized according to their second messenger coupling. Table 10.1 shows some of the best studied and more recent populations of 5-HT receptors and their second messenger systems.

5-HT receptor nomenclature can be rather bewildering; fortunately, this nomenclature has been recently revised (Martin and Humphrey 1994; Hoyer et al. 1994). Nevertheless, older literature must be read very carefully. For example, 5-HT1B receptors were once thought to exist in various animal species, including human; the term "5-HT1B" now refers only to rodent 5-HT1B receptors, whereas humans possess 5-HT1D receptors in corresponding anatomical locations. In fact, two different populations of 5-HT1D receptors have now been identified: 5-HT1Da and 5-HT1Dp; 5-HT1Dp receptors appear to be the human counterpart (i.e., species homolog) of rodent 5-HT1B receptors. An even more complex situation exists with the 5-HT2 family of receptors. Because 5-HT1C receptors were found to display greater sequence homology, similar second messenger coupling, and related pharmacology to what were once termed 5-HT2 receptors, they have been renamed 5-HT2C receptors. Thus, the term "5-HT1C" is no longer used. Correspondingly, and for purpose of distinction, the original 5-HT2 receptors have been renamed 5-HT2A receptors. Today, the term "5-HT2" is used only to refer to the 5-HT2 family of receptors. For several years, there was no 5-HT2B receptor population; 5-HT2F receptors were renamed 5-HT2B receptors shortly after their initial characterization to fill the obvious gap. To make matters even more confusing, 5-HT2 (now 5-HT2A) receptors, have been demonstrated to exist both in a high-affinity (5-HT2H) and low-affinity (5-HT2L) state. During a period of controversy, when it was thought that these two affinity states might represent distinct populations of sites, the terms 5-HT2A and 5-HT2B were introduced to describe 5-HT2H and 5-HT2L binding behavior. Thus, certain terms, such as 5-HT2A receptors for example, have different meanings or refer to different populations of receptors depending upon their chronological occurrence in the literature.

As a further note of caution, care must be given to what constitutes a site-selective agent. Certain agents were once considered quite selective for a given population of receptors. However, with the proliferation of 5-HT (and other neurotransmitter) receptor populations, and with more extensive investigation of older agents, it is now recognized that many of these agents are considerably less selective than once supposed. Table 10.1 lists some of the agents typically used to investigate the various populations of 5-HT receptors; although these agents are listed as being "selective" agonists or antagonists, no claims are made, or intended, as to their absolute selectivity. Rather, these are merely relatively selective or semi-selective agents that display some selectivity for that particular population of receptors versus most (but not necessarily all) other populations of 5-HT receptors. The interested reader is referred to the primary literature to obtain more information about the selectivity of a specific agent; for comparative binding data on ligands at various populations of 5-HT receptors, see Hoyer et al. (1994) and Ziffa and Fillion (1992).

The present chapter will focus primarily on some of the more widely used or standard serotonergic agents, and on some of the more recent agents. For the most part, the earlier work, including the medicinal chemistry and structure-activity relationships of serotonergic agents (Glennon et al. 1991; Glennon and Dukat 1993; King 1994), has been already reviewed and further information about uncited material can be found in these review articles. Serotonergic signal transduction pathways have also been reviewed.

10.2.3 Receptor populations and ligands

Not so long ago, it was thought that the chemical class to which a serotonergic agent belonged might be readily related to selectivity. For example, indolealkylamines, like 5-HT itself, were considered to be nonselective agents, aminotetralins were 5-HT1A ligands (and, more specifically, 5-HT1A agonists), arylpiperazines were 5-HT1B agonists, and benzoate esters and benzamides were 5-HT3 antagonists. None of these generalizations has held up over time. Most serotonergic agents can be conveniently categorized into one of several different large chemical families (Glennon and Dukat 1993); however, few (if any) of these families contain only agents that are exclusively agonists or antagonists for a specific population of 5-HT receptors. Selectivity, and functional activity, seem more controlled by pendent substituent groups rather than by the chemical class to which the agent belongs. Subtle structural changes can result in significantly different pharmacological consequences. Some of the general structure-activity and structure-affinity relationships (SAR and SAFIR) of various chemical classes will be discussed. Because space does not allow a detailed discussion of these relationships, only selected examples will be provided. More extensive reviews of this topic can be found in the literature (e.g. Glennon etal. 1991; Glennon and Dukat 1993).

Because 5-HT1A receptors were one of the first to be identified, there is considerable information available on 5-HT1A ligands. 5-Carboxamidotryptamine (5-CAT; 10.2) has seen application as a 5-HT1A agonist; although this agent is non-selective, it was once used to classify a receptor population as belonging to the 5-HT1 family. Interestingly, this generality is no longer valid in that although 5-CAT binds at most populations of 5-HT1 receptors, it displays low affinity for 5-HT1E receptors; 5-CAT also binds at certain other populations of non-5-HT1, 5-HT receptors. Nevertheless, it is still a relatively widely used agent. Two of the most important structure types include the aminotetralins and the arylpiperazines. The standard 5-HT1A agonist, 8-OH DPAT or 8-hydroxy-2-(dipropylamino)tetralin (10.3), remains one of the more selective serotonergic agents available; its selectivity is likely related to the observation that the intact indole nucleus of 5-HT is not a requirement for activation of 5-HT1A receptors. Although racemic 8-OH DPAT is the most commonly used form of the agent, it has been demonstrated that R(+)8-OH DPAT is a full agonist whereas its S(-)-enantiomer is a partial agonist. Incorporation of a 5-fluoro group further shifts the functional activity of this compound; S(-)5-F 8-OH DPAT or S(-)UH-301 is a potent 5-HT1A antagonist. Tritiated 8-OH DPAT is the radioligand of choice for binding studies.

No single chemical class of agents has been as extensively investigated for serotonergic activity as the arylpiperazines (reviewed Glennon and Dukat 1993). N1-(Aryl)piperazines can be divided into two broad classes: the N4-unsubstituted or short-chain arylpiperazines, and the long-chain arylpiperazines (LCAPs). Short-chain, and especially N4-unsubstituted arylpiperazines are notoriously non-selective serotonergic agents with different binding profiles and functional activities; in fact, certain agents may be agonists or partial agonists at one receptor population and antagonists at another. The long-chain arylpiperazines seem much more selective for 5-HT1A versus other populations of receptors, and selected examples bind at 5-HT1A receptors with Ki values as high as 0.1 nM. It might be noted, however, depending upon the particular agent, that certain of these agents also bind with high affinity at dopamine, a-adrenergic, and/or at members of the 5-HT2 family of receptors. The general structure of the arylpiperazines is given by: Ar-PIPERAZINE-(CH2)n-Terminus. The aryl (Ar) group can vary widely amongst phenyl, substituted phenyl, naphthyl, and heteroaryl. The terminus is usually an amide (or imide) or an aromatic group, and there appears to be considerable bulk tolerance. Thus, different combinations of functionalities can result in a vast array of structures that retain affinity for 5-HT1A receptors. With regard to n, two to four methylene groups appear optimal. However, the length of this chain can influence selectivity and, furthermore, the nature of the amide substituent can influence optimal chain length. For example, n=4 is optimal when the



terminus is a heteroarylamide, but when the terminus is an alkylamide, n=2 is optimal. Electronic distribution in the amide region also plays a role in 5-HT1A affinity. Typical examples of this class of agents include the agonists or partial agonists buspirone (10.4), gepirone (10.5), and ipsapirone (10.6); several very low efficacy partial agonists, including BMY 53857 (10.7) and NAN-190 (10.8), have also been reported and have been widely used as 5-HT1A antagonists, but only WAY-100,135 (10.9) is considered a silent antagonists.

A final, although less well investigated, class of agents that has been explored is the aryloxyalkylamines, such as ^-adrenergic antagonists propranolol (10.10) and pindolol. Though these agents bind at 5-HT1B and/or ^-adrenergic receptors with greater affinity than they display for 5-HT1A receptors, they were among the first compounds shown to behave as 5-HT1A anatagonists. Interest with these agents continues (Langlois et al. 1993), and mechanistic studies that have implicated f-adrenergic involvement for a particular action solely on the basis of antagonism by propranolol or pindolol may be in need of re-investigation.

Arylpiperazines were once the mainstay of 5-HT1B receptor research because they were considered selective for this population. With the realization that these agents are far less

(10.4); bLiipirtsne

selective than originally reported, their use has diminshed. Another agent that played a pivotal role in 5-HT1B research was RU-24969; however, this agent displays only several-fold selectivity for 5-HT1B versus 5-HT1A receptors. A newer series of agents developed by Pfizer, e.g. CP-93,129 (10.11), is related to RU-24969 but seem to be somewhat more selective. Interest in 5-HT1B receptors dampened once it was shown that rodent 5-HT1B receptors are structurally distinct from their species homolog, the human 5-HT1DiS receptors. These two receptor populations display a high (>90%) degree of homology and most agents that bind at 5-HT1B receptors also bind at 5-HT1DiS receptors. As a consequence, attention was refocused on the latter population. Interestingly, aryloxyalkylamines such as propranolol are amongst the few agents that can differentiate between these two populations of receptors in that they bind with high affinity at 5-HT1B receptors, but with > 100-fold lower affinity at 5-HT1D^ receptors.

Sumatriptan (10.12) is considered a prototypical 5-HT1D agonist. Structure-affinity relationships for the binding of 5-HT1D ligands at bovine receptors were described prior to cloning of human 5-HT1Da and 5-HT1DiS receptors. Much less is known about the SAFIR for binding at human 5-HT1D receptors. But, several new agents have been recently introduced including an oxadiazole (10.13), carbazole (10.14), and the 5-(pyridylamino)indole (10.15). One of the problems facing sumatriptan and many other 5-HT1D agonists is their high affinity for 5-HT1A receptors; 10.12-10.15 typically display <50-fold 5-HT1D selectivity. ALX-1323 (NOT; 10.16) is a new agonist with high affinity and >300-fold selectivity for 5-HT1D/S receptors relative to 5-HT1A receptors. Recently, a novel series of 5-HT1D antagonists has been reported; typical examples include piperazines (10.17), where R=5-methylisoxazol-3-yl and 2-methyl-4-(N,N-dimethylcarboxamido)phenyl (Clitherow et al. 1994).

Human 5-HT1E receptors were first decribed in brain homogenates six years ago using radioligand binding techniques. The receptors were not fully characterized at that time but it was demonstrated that even minor molecular modification of the 5-HT structure resulted in reduced affinity. For example, O-methyl 5-HT and 5-CAT (10.2), agents typically binding with high affinity at most populations of 5-HT! receptors, displayed low affinity for these receptors. Several years later, several groups independently cloned 5-HT1E receptors and demonstrated binding profiles similar to those which had been reported earlier. Methiothepin serves as a nonselective 5-HT1E antagonist. At this time, no 5-HT1E selective agents have been reported.

This is the most recent population of human 5-HT! receptors to be identified. No selective agonists or antagonists are currently available.

The first two populations of 5-HT receptors to be identified were the 5-HT! and 5-HT2 receptors. It is now recognized that these populations actually consist of subpopulations. However, although this was realized early on for 5-HT! receptors, recognition of subpopulations of 5-HT2 receptors did not occur until the late 1980s and was not fully appreciated until into the 1990s. Consequently, much of the early work on "5-HT2" receptors may in fact reflect results that can now be dissociated into 5-HT2A, 5-HT2B, and 5-HT2C. Nevertheless, it is only very recently that work has been initiated on attempting to develop agents with selectivity for 5-HT2 receptor subpopulations.

Much of the early work on 5-HT2 agonists and antagonists was previously reviewed (Herndon and Glennon 1993; Glennon et al. 1991). Ketanserin (10.18) was one of the first, and is still one of the most widely used, 5-HT2 antagonists. Tritiated ketanserin has been the radioligand of choice for investigating 5-HT2 receptors, but it appears that ketanserin binds with lower affinity at 5-HT2B receptors than it displays for 5-HT2A or 5-HT2C receptors. Additionally, ketanserin has been variously reported to bind with as little as 2-fold to as much as 140-fold selectively for 5-HT2A versus 5-HT2C receptors. Various other 5-HT2 antagonists have been described. Additional SAFIR and binding hypotheses have been suggested to account for the binding of various antagonists at 5-HT2A receptors. a-Methylserotonin, although not particularly selective, has been employed as a 5-HT2 agonist. The DOX series of compounds represent another widely used group of 5-HT2 agonists; these are typified by DOB and DOI (10.19, where X=Br and I, respectively). [125I] DOI is available for use as a radioligand. In certain functional o

(10.18); ketanserin

(10.21): spiperone

AMI 193

studies, 1-(3-chlorophenyl)piperazine (mCPP; 10.20) has been reported to be a 5-HT2C (or 5-HT2B/2C) agonist but a 5-HT2A antagonist; however, it also binds at other populations of 5-HT receptors.

With the recent reclassification of 5-HT2 receptors has come attempts to develop new agents with subtype selectivity. Spiperone (10.21) and AMI-193 (10.22) display >1,000-fold selectivity for 5-HT2A versus 5-HT2C receptors. MDL 100,907 (10.23) has also been reported to bind with 200-fold higher affinity at 5-HT2A versus 5-HT2C receptors. However, these results were reported prior to the discovery of 5-HT2B receptors. SB 200646A (10.24) was the first reported 5-HT2C-selective antagonist, but was later

(10.26); SDZSER-082

shown to also be a 5-HT2B antagonist; structural modification subsequently led to the development of the 5-HT2B-selective antagonist SB 204741 (10.25). SDZ SER-082 (10.26) and SB 206553 (10.27) are other examples of a 5-HT2B/2C versus 5-HT2A-selective antagonists. Thus, even though there are a great number of agents that have been termed "5-HT2 antagonists", the identification of subtypes of 5-HT2 receptors has initiated a search for more selective agents. Even while this search progresses, molecular biological and functional studies have identified new species homologs of 5-HT2 receptors and have also raised the possibility of additional members of the 5-HT2 family of receptors.

The 5-HT3 population of receptors was first studied using isolated peripheral tissue preparations and it was several years before a suitable radioligand was identified and 5-HT3 receptors were characterized in the brain. Two of the most commonly used 5-HT3 agonists are (3-chlorophenyl)biguanide (mCPBG; 10.28) and 2-methyl 5-HT (10.29). The N,N,N-trimethyl quaternary salt of 5-HT (5-HTQ; 10.30) also seems to be a selective 5-HT3 agonist. The SAR of 5-HT3 agonists has not been well investigated. In contrast, numerous 5-HT3 antagonists have been reported and a detailed discussion of their SAR, although beyond the scope of this chapter, has recently appeared (King 1994). The first useful 5-HT3 antagonist, MDL 72222 (10.31), resulted from the observations that metoclopramide and cocaine are weak 5-HT3 antagonists. MDL 72222 was subsequently shown to possess those basic features important for 5-HT3 antagonist activity i.e. arylcarbonyl linker-basic side chain. Many of the early agents were aryl-substituted benzoate esters and benzamides, but structurally related agents were also developed. Some of the older and more widely used 5-HT3 antagonists include: zacopride (10.32), renzapride (10.33), zatosetron (LY 277359; 10.34), tropisetron (ICS 205-930; 10.35), granisetron (BRL 43694; 10.36), and ondansetron (10.37). A significant amount of structural latitude is permitted, particularly in the basic side chain. This has resulted in the development of hundreds of 5-HT3 antagonists. Tritiated tropisetron, zacopride, granisetron, and related compounds have been used in radioligand binding studies.

A population of receptors originally identified in primary cell cultures of mouse embryo colliculi, and subsequently investigated in detail using peripheral functional assays, has been shown to exist in the brain and has been termed 5-HT4. 5-HT4 receptors were very recently cloned and display <50% homology in their transmembrane domains with other

Para Tres Lavocal

5-HT receptors. Two isoforms or splice variants, 5-HT4L and 5-HT4S, that vary only in the length and sequence of their C-terminal chain, have been identified. Many 5-HT3 ligands had been shown to be structurally unique relative to other serotonergic agents;

this was probably not unusual given that 5-HT3 receptors are the only population of ion channel receptors within the 5-HT family. Interestingly, however, the first agents used to investigate 5-HT4 receptors were some of these same agents, even though 5-HT4

Agonist Drug

receptors represent G-protein receptors. Certain 5-HT3 agonists were found to behave as 5-HT4 antagonists whereas others even acted as (partial) agonists. More selective agents have now been identified including the agonist SC 53116 (10.38), and the antagonists SB 204 070 (10.39), SB 207 710 (10.40), RS-23597-190 (10.41), GR 113808 (10.42), and SDZ 205,557 (10.43). [3H]GR 113808 and [125I]SB 207 710 have been used as radioligands. A 5-HT4 agonist pharmacophore model has been proposed and new agonists, derivatives of carbazimid-amide (10.44) have been synthesized on the basis of this model.

(10,38J: SC 53H(i

(10LJ9); X = CI, (SB 204070) (10.40); X = I, (SB 207710)

These three populations represent the newest families of 5-HT receptors to be identified; (see Glennon and Dukat (1995) and Lucas and Hen (1995) and references therein for a recent overview). Both a rat 5-HT5A and 5-HT5B receptor, but only the human 5-HT5A receptor, have been cloned. Rat and human 5-HT6 as well as human, rat, mouse, and hamster 5-HT7 receptors have been cloned. To date, no selective ligands have been identified. Interestingly, the 5-HT1 ligand 5-CAT (10.2) binds at 5-HT7 receptors with high affinity and the 5-HT1A agonist 8-OH DPAT (10.3) also binds at 5-HT7 receptors. Certain typical and atypical neuroleptic agents and tricyclic antidepressants bind at 5-HT6 and 5-HT7 receptors suggesting that these receptor populations may play a role in the mechanism of action of psychotherapeutic agents.

10.2.4 Possible applications of 5-HT agonists and antagonists

Experimental reports suggest that 5-HT receptors in the CNS may be implicated in the control of several basic physiological functions and behaviours which include food intake, thermoregulation, sexual behaviour, aggression, panic attacks as well as sleep, circadian rhythm, cognitive processes, cardiovascular disorders and schizophrenia (see Glennon and Dukat 1995). Agonists and antagonists for 5-HT receptor sub-types may therefore represent an array of possible therapeutic targets for the medicinal chemist (see Leonard 1994). Thus, it has been proposed that agonist stimulation of 5-HT1A receptors may suppress central 5-HT activity to increase the feeding response and this might have potential in the drug treatment of anorexia nervosa. 5-HT2 agonists, in contrast, have the opposite effect in that they suppress appetite in a 5-HT2 antagonist reversible manner. Moreover, 5-HT1A agonists tend to induce hypothermia whereas 5-HT1B and 5-HT2 stimulation produces hyperthermia in animal studies. Even though investigations in humans suggest that 5-HT2 receptors are perturbed during depression and this malfunction tends to be normalized during effective antidepressant therapy in some patients, the most frequent current treatment relating to these effects, involves indirect receptor stimualtion via inhibition of neuronal reuptake of 5-HT itself.

Both experimental and clinical data support the view that 5-HT1A agonists or partial agonists (e.g. buspirone), in addition to 5-HT2 and 5-HT3 antagonists, may have potential as drug treatments for anxiety. Though there are few reports that 5-HT3 antagonism is effective in the treatment of schizophrenia, it has been proposed that antagonists at 5-HT3 receptors may be valuable in the management or treatment of drug abuse in humans.

Migraine is largely thought to be due to vasodilatation in brain tissue probably in association with localised inflammation. In this context, 5-HT1D receptors are located on both cerebral and extracerebral blood vessels and are stimulated by the agonist sumatriptan to include vascular constriction and reduced release of inflammatory mediators. This mechanism is thought to be responsible not only for migraine prophylaxis but also in the alleviation of the headache and nausea associated with a migraine attack once it has been established.

Other clinical applications for 5-HT receptor ligands include: antagonists at 5-HT3 (eg. ondansetron) which are employed to treat chemotherapy-induced emesis and certain newer antidepressants (eg. trazodone and mianserin) which may share a common 5-HT2 mechanism of action. Moreover, 5-HT2A agonists (eg. DOM and DOB) possess hallucinogenic properties.

10.2.5 Epilogue

At least 15 different types of 5-HT receptors (Table 10.1), and numerous species homologs, have now been cloned. Several other, less well-investigated populations of 5-HT receptors have also been described in the literature. This has become an enormous challenge for medicinal chemists who have attempted to formulate and understand the SAR and SAFIR of various agents in order to develop newer compounds with subtype selectivity. This chapter, though not comprehensive, has described some of the most useful agents having evolved to date. In the past, serendipity accounted for the discovery of some of the most useful compounds in the armamentarium of serotonergic agents (including, for example, 8-OH DPAT and ketanserin). Futhermore, the design of selective agents was often hampered by confounding pharmacological results stemming from interference by populations of 5-HT receptors that had yet to be discovered. With advances in molecular biology, the identification and characterization of novel receptor populations, and an increased awareness of SAR and SAFIR, newer agents are being designed with greater rationale. The availability of the amino acid sequences of 5-HT receptors populations from molecular biological studies is spurring molecular modeling investigations of the receptors themselves. Increasingly, these graphics models, coupled with site directed mutagenesis and the investigation of chimeric receptors, are being employed to explain the binding of ligands to 5-HT receptors, by identifying or implicating specific amino acid residues that may be contributing to binding interactions, and to aid the design of newer agents. The ever increasing knowledge of receptor populations and the results of studies described above, bode well for things to come and should eventually allow the design of high-affinity site-selective agents.


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10.3 HISTAMINE RECEPTORS Holger Stark and Walter Schunack

10.3.1 Introduction

The story of histamine (10.45, specific histamine numbering is shown on this structure) began in the early 1900s with classical pharmacological investigations concerning its physiological and pathophysiological effects. Histamine has been recognized as an important chemical messenger communicating information from one cell to another. A large variety of cell types including smooth muscles, endocrine and exocrine glands, blood cells and cells of the immune system of mainly vertebrates respond to histamine stimuli with large differences from species to species. New aspects were brought into this

(Ift.45); hiSLamine (10.46)

field of research by the discovery that histamine does not only act as a local hormone but also acts as a neurotransmitter. Although histamine is widely distributed within mast cells in almost all mammalian peripheral tissues it plays an important role in the mammalian brain displaying powerful neuromodulatory, immunomodulatory, and neurotransmitter effects. Histamine itself does not cross the blood-brain barrier. Physiologically, it is produced by decarboxylation of L-histidine, mainly by specific histidine decarboxylase. This enzyme could be selectively inactivated by (S)-a-fluoromethylhistidine (10.46) being a "suicide" substrate.

Histamine catabolism occurs along two alternative pathways. One metabolic route is via methylation by the specific histamine N-methyltransferase to NT-methylhistamine. The other one is the oxidative deamination by diamine oxidase to imidazole acetaldehyde and further oxidative and coupling products. The first pathway seemed to be the only one to operate in the mammalian brain. Therefore, the methylated metabolite is the most important catabolite for investigations on central histamine levels for neurotransmitter function.

According to their chronological order of discovery three subtypes of histamine receptors are pharmacologically accepted at present: histamine H1-, H2-, and H3-receptors. The effects following direct stimulation of these G-protein coupled receptors are manifold and depend on species and tissue. H1-receptors are coupled to the phosphatidylinositol cycle, H2-receptors to adenylate cyclase; the signaling system for H3-receptors is unknown so far. Whereas H1- and H2-receptors are characterized by classical pharmacology as well as by molecular biology in different species the cloning of the H3-receptor is at present still under investigation. H1- and H2-receptors are postsynaptically located. The H3-receptor was identified at first as a presynaptically located autoreceptor inhibiting the synthesis and release of histamine in histaminergic neurons. Later on, the newly found function of H3-heteroreceptors modulating the release of a number of different neurotransmitter (noradrenaline, acetylcholine, dopamine, serotonin, neuropeptides) gave further hints for therapeutic indications of H3-receptor ligands.

Histamine possesses two basic moieties: the primary nitrogen on the side-chain (Na) protonated under physiological conditions (pKa1=9.73) and the protonable aromatic imidazole nucleus (pKa2=5.91). The first H1-receptor agonists developed followed the minimal structural requirements having an aromatic ring and an ethylamine side-chain. This approach was more or less successful with N-methyl-2-(2-pyridyl)ethanamine (betahistine, 10.47) and 2-(2-thiazolyl)ethanamine (10.48). Although they show less than 30% activity compared to the endogenous ligand (histamine 100%) they were used as pharmacological tools for a long time. Recent developments in the class of 2-substituted

10.3.2 H1-receptors

(10,47); bitahistine
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