Progress in Histo- and Cytochemistry, Vol. 26 W. Graumann / J. Drukker (Eds.), Histochemistry of Receptors © Fischer Verlag . Stuttgart· Jena . New York . 1992
6.2 Role and localization of serotonin-5HT2 receptors
J. E. LEYSEN, A. SCHOTIE Department of Biochemical Pharmacology, Janssen Research Foundation, Beerse (Belgium)
Introduction The classification and identification of serotonin receptor subtypes has been largely based on radioligand binding studies. The earliest identified subtype was the SHT2 receptor, originally labeled in the rat frontal cortex with 3H-spiperone, which is a neuroleptic with mixed dopamine and serotonin antagonistic properties (LEYSEN et al. 1978). Early studies already identified pharmacological and possible pathophysiological roles for SHT2 receptors, but to this day their role under normal physiological conditions remains enigmatic (LEYSEN et al. 1982; LEYSEN 1990). In this review we will attempt to integrate information about the serotonergic neuroanatomy and -activity, the biochemical and pharmacological characteristics of the SHT2 receptors and their distribution, the pharmacological roles of the SHT2 receptors, the regulation of the SHT2 receptors and the pathophysiological role of SHT2 receptors; finally, hypotheses on the physiological role of these receptors will be reviewed.
Serotonergic neuroanatomy and activity of serotonergic neurons Serotonin is phylogenetically one of the most ancient neurotransmitters. Serotonergic neurons mature early during embryonic development and show a most widespread distribution in the central nervous and peripheral system. Two types of central serotonergic neurons originate in the raphe and project into the entire forebrain and the spinal cord: a thin varicose axon system (D fibers) with probable synaptic contacts originates in the dorsal raphe, and a basket axon system (M fibers) which form synapses originates in the median raphe (TORK 1990). DES CARRIES et al. (1990) showed that SHT innervation in three anatomical regions of the adult rat telencephalon (the cerebral cortex, the neostriatum and the hippocampus - which receive the majority of ascending SHT neurons from the dorsal and median raphe) is predominantly non-junctional. A quantitative analysis of the synaptic contacts of SHT neurons revealed a frequency of junctional varicosities of less than 20% in the neostriatum, of 30 to 40% in the cerebral cortex and a «very low one» in the hippocampus. This is in sharp contrast with the high junctional frequency (90%) of nonserotonergic neurons in these areas. It has been suggested that the nonjunctional anatomy of the serotonergic neurones may be related to the phylogenic antiquity of the SHT system.
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Indeed, a similar lack of synaptic differentiation was demonstrated for SHT innervation in the spinal cord of the lampreyeel. In such a system SHT supposedly acts by «drip irrigation». JACOBS et al. (1990) reported another conspicuous peculiarity of the serotonergic system. In a study where serotonergic neuronal activity was recorded in normally behaving cats, dorsal raphe nucleus serotonergic neurons were found to show a characteristic slow and rhythmic activity in a quiet waking state (3 spikes/sec), with this phasically increasing to 6 spikes/sec in response to an arousing stimulus, gradually declining on drowsiness and entry into slow wave sleep, and falling silent on entering REM sleep. The activity pattern in cats apparently is determined primarily by the behavioural state of the animal and is not influenced by the light-dark cycle. Also, strong environmental stressors (placement of the cat in a canvas bag, loud white noise, confrontation with a dog) or physiological challenges (effects on body temperature, cardiovascular system, glucoregulation) failed to activate brain serotonergic neurons above the level seen during an undisturbed active waking state. This was despite the fact that the manipulations produced behavioral arousal plus a significant activation of the sympathetic nervous system and the noradrenergic neurons, in the locus coeruleus. The above considerations regarding the peculiar neuroanatomy and neuronal activity of the serotonergic system in brain may be of key importance for the interpretation of observations regarding the physiological and pathophysiological role of serotonin receptor subtypes and their regulation.
5HT2 receptors: biochemical and pharmacological characteristics SHT2 receptors exhibit high nanomolar affinities when binding a large number and great variety of drugs endowed with SHT2 antagonistic properties. In a series of 150 miscellaneous neurotransmitter antagonists or uptake blockers, over 50 compounds showed nanomolar binding affinity for SHT2 receptors. The compounds further revealed widely different receptor binding profiles and belonged to more than 20 different chemical classes. In spite of the plethora of high affinity SHT2 antagonists, only 2 selective compounds were found: pipamperone and cinanserin; yet, surprisingly, these two compounds showed no structural match. SHT binds to SHT2 receptors with an affinity constant of 0.5 I-tM - which is of the same order as the SHT affinity constant for SHT3 and SHT4 receptor subtypes, but which is two orders of magnitude higher than its affinity constant for SHT 1 receptor subtypes. No selective agonists for SHT2 receptors are known; methoxyphenetylamine derivatives have the highest affinity and are the least non-selective (SHT2 /SHT 1C agonists). Details on structures, SHT2 receptor binding affinity and selectivity of drugs are reported in LEYSEN (1989, 1990). As mentioned above, SHT2 receptors were first labeled using the neuroleptic [3H] spiperone, though later eH] ketanserin was proposed as a more selective ligand; it allowed an extensive characterization of the SHT2 receptor binding properties and investigation of its role and localization (LEYSEN et al. 1982). Nevertheless, it appeared that the compound could also label nonserotonergic binding sites (LEYSEN et al. 1988). Several other eH] and 1251 labeled antagonists and agonists (in particular methoxyphenethylamines) have been used for SHT2 receptor labeling, however, none of the ligands appeared to be absolutely selective and, depending on the tissue and the ligand, occluding agents are required to prevent labeling of various other receptors (LEYSEN 1989; RAnJA et al. 1991).
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Activation of 5HT2 receptors stimulates, via a coupled G-protein, phospholipase C to hydrolyse phosphatidylinositol 4,5-biphosphate with the attendant formation of inositol trisphosphate and diacylglycerol; also stimulated in a subsequent rise in intracellular Ca++ and activation of protein kinase C. An extensive characterization of this 5HT2 receptor coupled signal transducing system was first carried out in human platelets (DE CHAFFOY DE COURCELLES et al. 1985). 5HT2 receptor coupled stimulation of phosphatidylinositol turnover was detected in various central and peripheral tissues (see below) (SANDERS-BUSH et al. 1990). Recently it has been demonstrated that 5HT2 receptor stimulation, in an independent way, also activates phospholipase A2 with the release of arachidonic acid (FELDER et al. 1990). Electrophysiological studies on brain slices revealed that 5HT2 receptor stimulation caused neuronal depolarization in association with an increase in input resistance, followed by a slow depolarization and a decrease in spike frequency (ARANEDA and ANDRADE 1991). The genes of rat and human 5HT2 receptors have been cloned, but only the rat sequence was published. The receptor protein shows the general monomeric structure, with 7 transmembrane domains, of the G-protein coupled receptor family (see WEINSHANK et al. this workshop). While 5HT2 receptors in various mammalian species and tissues show a very high similarity, the alleged existence of 5HT2A and 5HT2B receptor subtypes suggested by apparent differences in agonist binding in tissues from different species, (PIERCE and PEROUTKA 1989) is probably an experimental artefact (LEYSEN 1990).
Location of 5HT2 receptors Using brain tissue homogenates, a particularly high concentration of 5HT2 receptors in the prefrontal cortex was demonstrated, which was similarly found in all mammalian species including humans (LEYSEN et al. 1982, 1983; SCHOTTE 1983). Autoradiographic techniques revealed details on the 5HT2 receptor distribution in the brain. 5HT2 receptors are highly enriched in the claustrum, the neocortex (mainly in layer IV but also in layer I and V; in human cortex layer III and V were more heavily labeled than layer IV), the tuberculum olfactorium, the nucleus accumbens and the postero-lateral extension of the nucleus caudatus. 5HT2 receptors have further been detected in the brain stem and spinal cord (for review see RADJA et al. 1991). Electrophysiological (ARANEDA and ANDRADE 1991), immunohistological and lesion studies (LEYSEN et al. 1983) as well as studies of brains of Alzheimer patients (CROSS 1990) have indicated that 5HT2 receptors are located on pyramidal cells in the neocortex, and may be localized on GABA-ergic and somatostatin containing neurones. In the cortex of Alzheimer patients markers of these neurons were disappeared concomitantly with 5HT2 receptors (CROSS 1990). By studying 5HT2 receptor mediated functions, such as platelet aggregation or shape change, smooth muscle contraction and phosphatidylinositol hydrolysis, 5HT2 receptors were demonstrated on platelets, smooth muscle tissue (vascular, tracheal, uterine), various cells in primary culture (astroglial cells, calf aorta smooth muscle cells) or cell lines (C 6 glioma cells, WRKI rat mammary tumor cells, A 7rs rat aorta smooth muscle cells, and Pll pituitary tumor cells) (reviewed in SANDERS-BuSH et al. 1990).
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Pharmacological roles of 5HT2 receptors Agonist-triggered effects: Various exogenous agonists (methoxyphenetylamines, LSD, quipazine) - or else very high dosages of tryptamine or 1-5 hydroxy tryptophane or 1-5 hydroxytryptophane combined with a monoamine oxidase inhibitor or a 5HT reuptake blocker - induce in rodents behavioral excitation (head twitches, tremor, myoclonus) and can induce discriminative stimulus effects. Exogenous agonists induce in humans hallucinations and vivid dreams, in susceptible pigs malignant hyperthermia, and in pithed rats hypertension. In vitro, the exogenous agonists and 5HT cause smooth muscle contraction in various tissues (blood vessels, trachea, uterus) and activate platelets. Moreover, in these tissues 5HTz receptor stimulation amplifies the effect of various triggers such as norepinephrine and thromboxane. All the effects triggered in vivo and vitro are antagonized by 5HTz antagonists with potencies highly significantly correlated with their 5HTz receptor binding affinity. 5HTz antagonists also prevent the avoidance learning deficit caused by p-chloroamphetamine-induced 5HT release (for review see LEYSEN 1989). Serotonin is further thought to play a role in primary drives such as feeding and sexual behavior. There might be a role for 5HT2 receptors in these behaviour patterns, but experimental data in laboratory animals are conflicting. Effects of 5HTz antagonists acting on their own 5HT2 antagonists were shown to prolong slow-wave sleep in laboratory animals, in healthy human volunteers and in dysthymic patients. 5HTz antagonists abolish rat behavioral inhibition induced by natural aversive stimuli such as bright light and open space. In contrast to other neurotransmitter antagonists (e.g. dopamine, noradrenaline, acetylcholine), 5HTz antagonists appear not to affect motoric, intellectual or autonomic functions (see references in LEYSEN and PAUWELS 1985). Hence in normal mammalian species, SHT2 antagonists acting on their own have only very mild effects; moreover, the effects are only apparent under certain conditions or during short periods of the sleep cycle. Neurochemical effects Blockade of central 5HT2 receptors by 5HTz antagonists does not affect 5HT turnover (i.e. there is no effect on 5HT or 5HIAA levels) measured in various brain areas (LEYSEN et a1. 1985). Hence 5HTz receptor blockade appeared not to induce a feedback activation of 5HT neurons; this is in marked contrast with the dopaminergic system, where feedback activation of dopamine turnover in the brain upon dopamine-Dz receptor blocking is a well known phenomenon. Functional relationship between 5HT2 and 5HT1A receptors Various behavioral studies with 5HT agonists and antagonists have provided evidence for a mutual inhibitory action between 5HT 1A and 5HTz receptors stimulation (DARMANI et a1. 1990;
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BACKUS et al. 1990). In electrophysiological studies 5HT lA and 5HTz receptors were demonstrated to co-exist on pyramidal cells in the neocortex, where they mediate opposing effects: stimulation of the former induced membrane hyperpolarization and of the latter membrane depolarization. 5HTz receptors desensitize rapidly, whereas 5HT 1A receptors do not easily desensitize. It seems puzzling that there are two distinct receptors for serotonin on the same cell mediating opposing electrophysiological responses (ARANEDA and ANDRADE 1991). Since the 5HT 1A has nM affinity for serotonin and the 5HTz receptor 11M affinity, it seems likely that the former receptor would be the first one to become stimulated. The latter receptor would only be activated when the local serotonin concentration becomes high. ARANEDA and ANDRADE discuss the various possible functional consequences of the coexistence of the two 5HT receptor subtypes (ARANEDA and ANDRADE 1991).
Role of 5HT2 receptors in the dopaminergic system
5HTz antagonists reduce neuroleptic-induced catalepsy in rats. In patients they alleviate neuroleptic-induced extrapyramidal symptoms and Parkinson tremor (BERSANI et al. 1990). In a recent study, SALLER et al. (1990) showed that 5HTz antagonists can enhance the neurolepticinduced inrease of dopamine turnover in the basal ganglia, though not in certain mesolimbic brain areas. These observations have provided a possible explanation for the reduction by 5HTz antagonists of the neuroleptic-induced impairment of motoric functions: the enhanced dopamine turnover in the basal ganglia would partially counteract the dopamine receptor blockade in these areas.
5HT2 antagonists and drugs of abuse Ritanserin, a very potent, long acting and relatively selective 5HTz antagonist, has recently been shown to reduce forced preference of rats for various addictive drugs such as alcohol, cocaine and fentanyl (MEERT et al. 1991). A role for 5HTz receptors in these conditions seems worthy of further exploration.
Regulation of 5HT2 receptors 5HTz receptors become very rapidly desensitized and downregulated. This was apparent from in vivo studies on rats, where the behavioral response to 5HTz agonists was already significantly reduced by the second injection of a methoxyphenetylamine derivative and was completely disappeared after 4 injections. Concomitantly 5HTz receptor numbers, in the frontal cortex, measured ex vivo using radio ligand binding techniques, were markedly reduced (LEYSEN and PAUWELS 1990). Also, the early observations in humans of rapid and long-lasting tolerance to LSD-induced psychotic and behavioral effects (CHOLDEN et al. 1955) indicate that tolerance is probably to be ascribed to 5HTz receptor down regulation. In cells in culture, 5HTz receptor induced phospholipid turnover became 80% reduced upon 15 min exposure of the cells to serotonin (PAUWELS et al. 1990); moreover, a similar rapid desen-
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sitization of 5HTz receptor-mediated neuronal depolarization of cortical slices was observed in electrophysiological studies (ARANEDA and ANDRADE 1991). Rapid desensitization following agonist stimulation was also reported for the 5HT3 and the 5HT4 receptors, but in contrast the 5HT1A receptor appeared to be hard to desensitize. Hence, a clear distinction in the regulatory response of various 5HT receptor subtypes appears to exist. Serotonergic denervation by lesioning serotonergic neurons, and chronic blockading of 5HTz receptors by antagonists failed to induce 5HTz receptor up regulation (LEYSEN et al. 1983, 1986). By contrast, chronic treatment with 5HTz antagonists caused receptor desensitization and down regulation, an anomalous response not in agreement with the generally accepted receptor regulation theory (LEYSEN et al. 1986). The reported 5HTz receptor down regulation by tricyclic antidepressants, which has been related to the therapeutic action of the compounds (PEROUTKA and SNYDER 1980), is probably a consequence of the 5HTz receptor blocking properties of the compounds and not of their 5HT reuptake inhibitory effects, as was first suggested. Indeed selective 5HT uptake blockers, such as citalopram failed to induce 5HTz receptor down regulation (for review see CONN and SANDERS-BuSH 1987). Besides 5HTz antagonists, 5HT 1A mixed agonists can also cause 5HTz receptor down regulation when chronically administered. The precise mechanism of the anomalous 5HTz receptor down regulation is not known, but it seems likely that heterologous phenomena are involved. It remains possible that 5HTz receptor down regulation induced by the various drugs plays a role in the therapeutic action of the drugs in depression and anxiety (LEYSEN and PAUWELS 1990).
Pathophysiological role of 5HT2 receptors In double-blind controlled clinical studies with the 5HTz antagonist, ritanserin, therapeutic effects have been obtained in dysthymia (lack of drive, depressed mood, neurotic depression), generalized anxiety and negative symptoms of schizophrenia (REYNTJENS et al. 1986; BERSANI et al. 1991). The drug also reduced neuroleptic induced extrapyramidal symptoms and Parkinson tremor (BERSANI et al. 1990). 5HTz antagonists increased periods of slow-wave sleep in dysthymic patients and regularized disturbed sleep in psychotic patients (IDZIKOWSKI et al. 1991). It has been suggested that improvement of the quality of sleep may be an important factor in the therapeutic action of the compounds OANSSEN 1987). These findings with ritanserin corroborate the early hypothesis of a possible role for 5HTz antagonism in sleep regulation, which was proposed when the particular clinical observations made with pipamperone were related to its specificity for 5HTz receptors (LEYSEN et al. 1978). Recent studies with risperidone, a compound exhibiting combined predominant 5HTz and potent dopamine-Dz antagonism, revealed therapeutic effects on both positive and negative symptoms of schizophrenia with a very low attendant extrapyramidal side effect. In view of the findings with ritanserin, the two latter properties have been ascribed to the predominant 5HTz antagonistic action of the compound. Possible therapeutic use of 5HTz antagonists was suggested for the treatment of non-bulimic anorexia nervosa and for the treatment of alcohol and cocaine withdrawal.
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Physiological role of 5HT2 receptors As appears from the above, serotonergic neuron anatomy and aCtIVIty, response to 5HT2 receptor stimulation and blockading and 5HT2 receptor regulation show marked differences with known classical neurotransmitter systems. Poor synapsis formation and the low, barely influencable activity of serotonergic neurons; the presence of two 5HT receptor subtypes on the same cells with opposing electrophysiological responses; detrimental effects by 5HT2 agonists and the very rapid and profound desensitization induced by these effects; the lack of effect by 5HT2 antagonists acting on their own on behavior, motoric, intellectual and autonomic functions; the failure of 5HT2 receptors to up regulate following denervation or chronic blockading - all these observations point to a very low, if indeed there is any activation by 5HT2 receptors under normal physiological conditions. We have therefore proposed the following hypothesis: In mammals 5HT2 receptor stimulation is scarce under normal physiological conditions, with activation probably only occurring during short periods of the day-night cycle (e.g. to interrupt slow wave sleep), in emergency (e.g. in the periphery upon cardiovascular accident to induce vasoconstriction and platelet aggregation) or under pathological conditions (e.g. dysthymia, anxiety, social withdrawal impaired blood circulation). Hence under normal conditions acute blockading does not disrupt the normal situation, which explains the lack of directly observable effects by 5HT2 antagonists. Because of the low stimulation, the receptors may exist in a supersensitive state and chronic blockading or denervation cannot produce further supersensitivity or upregulation. The rapid and profound homologous desensitization and receptor down regulation is an effective defense mechanism against the detrimental effects of enhanced agonist stimulation. In addition, 5HT2 receptors are sensitive to heterologous desensitization, such as by 5HT2A receptor stimulation. It seems not unlikely that under pathological conditions which are responsive to 5HT2 antagonists, 5HT2 receptor stimulation is enhanced and homologous receptor downregulation impaired. Under such conditions, 5HT2 antagonists will exert therapeutic effects, both acutely by blocking the receptors and chronically by downregulating them via heterologous mechanisms.
References ARANEDA, R., ANDRADE, R.: 5-Hydroxytryptaminez and 5-hydroxytryptaminelA receptors mediate opposing responses on membrane excitability in rat association cortex. - Neurosci. 40, 399-412 (1991). BACKUS, L. 1., SHARP, T., GRAHAME-SMITH, D. G.: Behavioural evidence for a functional interaction between central 5-HT z and 5-HT 1A receptors. - Brit.]. Pharmacol. 100, 793-799 (1990). BERSANI, G., GRISPINI, A., MARINI, S., PASINI, A., VALDUCCI, M., CIANI, N.: 5-HTz Antagonist ritanserin in neuroleptic-induced Parkinsonism: A double-blind comparison with orphenadrine and placebo. - Clin. Neuropharmacol. 13, 500-506 (1990). BERSANI, G., POZZI, F., MARINI, S., GRISPINI, A., PASINI, A., CIANI, N.: 5-HTz receptor antagonism in dysthymic disorder: a double-blind placebo-controlled study with ritanserin. - Acta psychiat. scand. 83, 244-248 (1991). . CHOLDEN, L. S., KURLAND, A., SAVAGE, c.: Clinical reactions and tolerance to LSD in chronic schizophrenia. - J. nerv. ment. Dis. 122, 211-221 (1955). CONN, P. J., SANDERS-BuSH, E.: Central serotonin receptors: Effector systems, physiological roles and regulation. - Psychopharmacol. 92, 267-277 (1987).
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CROSS, A. J.: Serotonin in Alzheimer-type dementia and other dementing illnesses. - In: The Neuropharmacology of Serotonin (WHITAKER-AzMITIA, P. M., PEROUTKA, S. J., eds.), Vol. 600, pp. 405-417. - The New York Academy of Sciences, New York 1990. DARMANI, N. A., MARTIN, B. R., PANDEY, U., GLENNON, R. A.: Do functional relationships exist between 5HTIA and 5-HT2 receptors? - Pharmacol. Biochem. Behav. 36,901-906 (1990). DE CHAFFOY DE COURCELLES, D., LEYSEN, J. E., DE CLERCK, F., VAN BELLE, H., JANSSEN, P. A. J.: Evidence that phospholipid turnover is the signal transducing system coupled to serotonin-S2 receptor sites. - J. bioI. Chern. 260, 7603-7608 (1985). DEs CARRIES, L., AUDET, M.-A., DOUCET, G., GARCIA, S., OLESKEVICH, S., SEGUE LA, P. H., SOGHOMORIAN, J.-J., WATKINS, K. c.: Morphology of central serotonin neurons. Brief review of quantified aspects of their distribution and ultrastructural relationships. - In: The Neuropharmacology of Serotonin (WHITAKERAZMITIA, P. M., PEROUTKA, S. J., eds.), Vol. 600, pp. 81-92. - The New York Academy of Sciences, New York 1990. FELDER, C. c., KANTERMAN, R., Y., MA, A. L., AxELROD, J.: Serotonin stimulates phospholipase A2 and the release of arachidonic acid in hippocampal neurons by a type 2 serotonin receptor that is independent of inositolphospholipid hydrolysis. - Proc. nat!. Acad. Sci. USA 87, 2187-2191 (1990). IDZIKOWSKI, C., MILLS, F. J., JAMES, R. J.: A dose-response study examing the effects of ritanserin on human slow wave sleep. - Brit. J. clin. Pharmacol. 31,193-196 (1991). JACOBS, B. L., WILKENSON, L. 0., FORNAL, C. A.: The role of brain serotonin. A neurophysiological perspective. - Neuropsychopharmacol. 3,473-479 (1990). JANSSEN, P. A. J.: Does ritanserin, a potent serotonin-S2 antagonist restore energetic functions during the night? - J. roy. Soc. Med. 80, 409-413 (1987). LEYSEN, J. E.: Gaps and peculiarities in 5-HT2 receptor studies. - Neuropsychopharmacol. 3,361-369 (1990). -: Use of 5-HT receptor agonists and antagonists for the characterization of their respective receptor sites. In: Neuromethods, Drugs as Tools in Neurotransmitter Research (BOULTON, A. B., BAKER, G. B., AUGuSTO, V. J., eds.), Vol. 12, pp. 299-350. - Humana Press., Clifton, NJ 1989). LEYSEN, J. E., EENS, A., GOMMEREN, W., VAN GOMPEL, P., WYNANTS, J., JANSSEN, P. A. J.: Identification of nonserotonergic eH]ketanserin binding sites associated with nerve terminals in rat brain and with platelets; relation with release of biogenic amine metabolites induced by ketanserin- and tetrabenazine-like drugs. J. Pharmacol. expo Ther. 244, 310-321 (1988). LEYSEN, J. E., GOMMEREN, W., VAN GOMPEL, P., WYNANTS, J., JANSSEN, P. F. M., LADURON, P. M.: Receptor-binding properties in vitro and in vivo of ritanserin. A very potent and long acting serotonin-S2 antagonist. - Mol. Pharmacol. 27, 600-611 (1985). LEYSEN, J. E., NIEMEGEERS, C. J. E., TOLLENAERE, J. P., LADURON, P. M.: Serotonergic component of neuroleptic receptors. - Nature (Lond.) 272, 168-171 (1978). LEYSEN, J. E., NIEMEGEERS, C. J. E., VAN NUETEN, J. M., LADURON, P. M.: eH]Ketanserin (R 41468), a selective 3H-ligand for serotonin2 receptor binding sites. - Mol. Pharmacol. 21, 301-314 (1982). LEYSEN, J. E., PAUWELS, P.: 5-HT2 Receptors, roles and regulation. - In: The Neuropharmacology of Serotonin (WHlTAKER-AzMITIA, P. M., PEROUTKA, S. J., eds.), Vol. 600, pp. 183-193. - The New York Academy of Sciences, New York 1990. LEYSEN, J. E., VAN GOMPEL, P., GOMMEREN, W., WOESTENBORGHS, R., JANSSEN, P. A. J.: Down regulation of serotonin-S2 receptor sites in rat brain by chronic treatment with the serotonin-S2 antagonists: ritanserin and setoperone. - Psychopharmacol. 88, 434-444 (1986). LEYSEN, J. E., VAN GOMPEL, P., VERWIMP, M., NIEMEGEERS, C. J. E.: Role and localization of serotonin (S2)receptor-binding sites: Effects of neuronal lesions. - In: CNS-Receptors-From Molecular Pharmacology to Behavior (MANDEL, P., DEFEUDlS, F. V., eds.), pp. 373-383. - Raven Press, New York 1983. MEERT, T. F., AWOUTERS, F., NIEMEGEERS, C. J. E., SCHELLEKENS, K. H. L., JANSSEN, P. A. J.: Ritanserin reduces abuse of alcohol, cocaine and fentanyl in rats. - Pharmacopsychiatry, in press (1991). PAUWELS, P. J., VAN GOMPEL, P., LEYSEN, J. E.: Rapid desensitization and resensitization of 5-HT2 receptor mediated phosphatidyl inositol hydrolysis by serotonin agonists in quiescent calf aortic smooth muscle cells. - Life Sci. 47, 2009-2019 (1990). PEROUTKA, S. J., SNYDER, S. H.: Regulation of serotonin2 (5-HTz) receptors labeled with eH]spiroperidol by chronic treatment with the antidepressant amitriptyline. - J. Pharmacol. expo Ther. 215, 582-587 (1980). PIERCE, P. A., PEROUTKA, S. J.: Evidence for distinct 5-hydroxytryptamine2 binding site subtypes in cortical membrane preparations. - J. Neurochem. 52, 656-658 (1989).
Serotonin-5HT2 receptors . 249 RAD]A, F., LAPORTE, A.-M., DAVAL, G., VERGE, D., GOZLAN, H., HAMON, M.: Autoradiography of serotonin receptor subtypes in the central nervous system. - Neurochem. Int. 18, 1-15 (1991). REYNT]ENS, A., GELDERS, Y. G., HOPPENBROUWERS, M.-L. J. A., VANDEN BUSSCHE, G.: Thymostenic effects of ritanserin (R 55 667), a centrally acting serotonin-S2 receptor blocker. - Drug Dev. Res. 8, 205-211 (1986). SALLER, C. F., CZUPRYNA, M. J., SALAMA, A. I.: 5-HT2 Receptor blockade by ICI 169,369 and other 5-HT2 antagonists modulates the effects of D-2 dopamine receptor blockade. - J. Pharmacol. expo Ther. 253, 1162-1170 (1990). SANDERS-BuSH, E., TSUTSUMI, M., BURRIS, K. D.: Serotonin receptors and phosphatidylinositol turnover. In: The Neuropharmacology of Serotonin (WHITAKER-AzMITIA, P. M., PEROUTKA, S. J., eds.), Vol. 600, pp. 224-236. - The New York Academy of Sciences, New York 1990. SCHOTTE, A., MALOTEAUX, J. M., LADURON, P. M.: Characterization and regional distribution of serotonin Srreceptors in human brain. - Brain Res. 276, 231-235 (1983). TORK, I.: Anatomy of the serotonergic system. - In: The Neuropharmacology of Serotonin (WHITAKERAZMITIA, P. M., PEROUTKA, S. J., eds.), Vol. 600, pp. 9-35. - The New York Academy of Sciences, New York 1990.
6.2 Role and localization of serotonin-5HT2 receptors
J. E. LEYSEN, A. SCHOTIE Department of Biochemical Pharmacology, Janssen Research Foundation, Beerse (Belgium)
Introduction The classification and identification of serotonin receptor subtypes has been largely based on radioligand binding studies. The earliest identified subtype was the SHT2 receptor, originally labeled in the rat frontal cortex with 3H-spiperone, which is a neuroleptic with mixed dopamine and serotonin antagonistic properties (LEYSEN et al. 1978). Early studies already identified pharmacological and possible pathophysiological roles for SHT2 receptors, but to this day their role under normal physiological conditions remains enigmatic (LEYSEN et al. 1982; LEYSEN 1990). In this review we will attempt to integrate information about the serotonergic neuroanatomy and -activity, the biochemical and pharmacological characteristics of the SHT2 receptors and their distribution, the pharmacological roles of the SHT2 receptors, the regulation of the SHT2 receptors and the pathophysiological role of SHT2 receptors; finally, hypotheses on the physiological role of these receptors will be reviewed.
Serotonergic neuroanatomy and activity of serotonergic neurons Serotonin is phylogenetically one of the most ancient neurotransmitters. Serotonergic neurons mature early during embryonic development and show a most widespread distribution in the central nervous and peripheral system. Two types of central serotonergic neurons originate in the raphe and project into the entire forebrain and the spinal cord: a thin varicose axon system (D fibers) with probable synaptic contacts originates in the dorsal raphe, and a basket axon system (M fibers) which form synapses originates in the median raphe (TORK 1990). DES CARRIES et al. (1990) showed that SHT innervation in three anatomical regions of the adult rat telencephalon (the cerebral cortex, the neostriatum and the hippocampus - which receive the majority of ascending SHT neurons from the dorsal and median raphe) is predominantly non-junctional. A quantitative analysis of the synaptic contacts of SHT neurons revealed a frequency of junctional varicosities of less than 20% in the neostriatum, of 30 to 40% in the cerebral cortex and a «very low one» in the hippocampus. This is in sharp contrast with the high junctional frequency (90%) of nonserotonergic neurons in these areas. It has been suggested that the nonjunctional anatomy of the serotonergic neurones may be related to the phylogenic antiquity of the SHT system.
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Indeed, a similar lack of synaptic differentiation was demonstrated for SHT innervation in the spinal cord of the lampreyeel. In such a system SHT supposedly acts by «drip irrigation». JACOBS et al. (1990) reported another conspicuous peculiarity of the serotonergic system. In a study where serotonergic neuronal activity was recorded in normally behaving cats, dorsal raphe nucleus serotonergic neurons were found to show a characteristic slow and rhythmic activity in a quiet waking state (3 spikes/sec), with this phasically increasing to 6 spikes/sec in response to an arousing stimulus, gradually declining on drowsiness and entry into slow wave sleep, and falling silent on entering REM sleep. The activity pattern in cats apparently is determined primarily by the behavioural state of the animal and is not influenced by the light-dark cycle. Also, strong environmental stressors (placement of the cat in a canvas bag, loud white noise, confrontation with a dog) or physiological challenges (effects on body temperature, cardiovascular system, glucoregulation) failed to activate brain serotonergic neurons above the level seen during an undisturbed active waking state. This was despite the fact that the manipulations produced behavioral arousal plus a significant activation of the sympathetic nervous system and the noradrenergic neurons, in the locus coeruleus. The above considerations regarding the peculiar neuroanatomy and neuronal activity of the serotonergic system in brain may be of key importance for the interpretation of observations regarding the physiological and pathophysiological role of serotonin receptor subtypes and their regulation.
5HT2 receptors: biochemical and pharmacological characteristics SHT2 receptors exhibit high nanomolar affinities when binding a large number and great variety of drugs endowed with SHT2 antagonistic properties. In a series of 150 miscellaneous neurotransmitter antagonists or uptake blockers, over 50 compounds showed nanomolar binding affinity for SHT2 receptors. The compounds further revealed widely different receptor binding profiles and belonged to more than 20 different chemical classes. In spite of the plethora of high affinity SHT2 antagonists, only 2 selective compounds were found: pipamperone and cinanserin; yet, surprisingly, these two compounds showed no structural match. SHT binds to SHT2 receptors with an affinity constant of 0.5 I-tM - which is of the same order as the SHT affinity constant for SHT3 and SHT4 receptor subtypes, but which is two orders of magnitude higher than its affinity constant for SHT 1 receptor subtypes. No selective agonists for SHT2 receptors are known; methoxyphenetylamine derivatives have the highest affinity and are the least non-selective (SHT2 /SHT 1C agonists). Details on structures, SHT2 receptor binding affinity and selectivity of drugs are reported in LEYSEN (1989, 1990). As mentioned above, SHT2 receptors were first labeled using the neuroleptic [3H] spiperone, though later eH] ketanserin was proposed as a more selective ligand; it allowed an extensive characterization of the SHT2 receptor binding properties and investigation of its role and localization (LEYSEN et al. 1982). Nevertheless, it appeared that the compound could also label nonserotonergic binding sites (LEYSEN et al. 1988). Several other eH] and 1251 labeled antagonists and agonists (in particular methoxyphenethylamines) have been used for SHT2 receptor labeling, however, none of the ligands appeared to be absolutely selective and, depending on the tissue and the ligand, occluding agents are required to prevent labeling of various other receptors (LEYSEN 1989; RAnJA et al. 1991).
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Activation of 5HT2 receptors stimulates, via a coupled G-protein, phospholipase C to hydrolyse phosphatidylinositol 4,5-biphosphate with the attendant formation of inositol trisphosphate and diacylglycerol; also stimulated in a subsequent rise in intracellular Ca++ and activation of protein kinase C. An extensive characterization of this 5HT2 receptor coupled signal transducing system was first carried out in human platelets (DE CHAFFOY DE COURCELLES et al. 1985). 5HT2 receptor coupled stimulation of phosphatidylinositol turnover was detected in various central and peripheral tissues (see below) (SANDERS-BUSH et al. 1990). Recently it has been demonstrated that 5HT2 receptor stimulation, in an independent way, also activates phospholipase A2 with the release of arachidonic acid (FELDER et al. 1990). Electrophysiological studies on brain slices revealed that 5HT2 receptor stimulation caused neuronal depolarization in association with an increase in input resistance, followed by a slow depolarization and a decrease in spike frequency (ARANEDA and ANDRADE 1991). The genes of rat and human 5HT2 receptors have been cloned, but only the rat sequence was published. The receptor protein shows the general monomeric structure, with 7 transmembrane domains, of the G-protein coupled receptor family (see WEINSHANK et al. this workshop). While 5HT2 receptors in various mammalian species and tissues show a very high similarity, the alleged existence of 5HT2A and 5HT2B receptor subtypes suggested by apparent differences in agonist binding in tissues from different species, (PIERCE and PEROUTKA 1989) is probably an experimental artefact (LEYSEN 1990).
Location of 5HT2 receptors Using brain tissue homogenates, a particularly high concentration of 5HT2 receptors in the prefrontal cortex was demonstrated, which was similarly found in all mammalian species including humans (LEYSEN et al. 1982, 1983; SCHOTTE 1983). Autoradiographic techniques revealed details on the 5HT2 receptor distribution in the brain. 5HT2 receptors are highly enriched in the claustrum, the neocortex (mainly in layer IV but also in layer I and V; in human cortex layer III and V were more heavily labeled than layer IV), the tuberculum olfactorium, the nucleus accumbens and the postero-lateral extension of the nucleus caudatus. 5HT2 receptors have further been detected in the brain stem and spinal cord (for review see RADJA et al. 1991). Electrophysiological (ARANEDA and ANDRADE 1991), immunohistological and lesion studies (LEYSEN et al. 1983) as well as studies of brains of Alzheimer patients (CROSS 1990) have indicated that 5HT2 receptors are located on pyramidal cells in the neocortex, and may be localized on GABA-ergic and somatostatin containing neurones. In the cortex of Alzheimer patients markers of these neurons were disappeared concomitantly with 5HT2 receptors (CROSS 1990). By studying 5HT2 receptor mediated functions, such as platelet aggregation or shape change, smooth muscle contraction and phosphatidylinositol hydrolysis, 5HT2 receptors were demonstrated on platelets, smooth muscle tissue (vascular, tracheal, uterine), various cells in primary culture (astroglial cells, calf aorta smooth muscle cells) or cell lines (C 6 glioma cells, WRKI rat mammary tumor cells, A 7rs rat aorta smooth muscle cells, and Pll pituitary tumor cells) (reviewed in SANDERS-BuSH et al. 1990).
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Pharmacological roles of 5HT2 receptors Agonist-triggered effects: Various exogenous agonists (methoxyphenetylamines, LSD, quipazine) - or else very high dosages of tryptamine or 1-5 hydroxy tryptophane or 1-5 hydroxytryptophane combined with a monoamine oxidase inhibitor or a 5HT reuptake blocker - induce in rodents behavioral excitation (head twitches, tremor, myoclonus) and can induce discriminative stimulus effects. Exogenous agonists induce in humans hallucinations and vivid dreams, in susceptible pigs malignant hyperthermia, and in pithed rats hypertension. In vitro, the exogenous agonists and 5HT cause smooth muscle contraction in various tissues (blood vessels, trachea, uterus) and activate platelets. Moreover, in these tissues 5HTz receptor stimulation amplifies the effect of various triggers such as norepinephrine and thromboxane. All the effects triggered in vivo and vitro are antagonized by 5HTz antagonists with potencies highly significantly correlated with their 5HTz receptor binding affinity. 5HTz antagonists also prevent the avoidance learning deficit caused by p-chloroamphetamine-induced 5HT release (for review see LEYSEN 1989). Serotonin is further thought to play a role in primary drives such as feeding and sexual behavior. There might be a role for 5HT2 receptors in these behaviour patterns, but experimental data in laboratory animals are conflicting. Effects of 5HTz antagonists acting on their own 5HT2 antagonists were shown to prolong slow-wave sleep in laboratory animals, in healthy human volunteers and in dysthymic patients. 5HTz antagonists abolish rat behavioral inhibition induced by natural aversive stimuli such as bright light and open space. In contrast to other neurotransmitter antagonists (e.g. dopamine, noradrenaline, acetylcholine), 5HTz antagonists appear not to affect motoric, intellectual or autonomic functions (see references in LEYSEN and PAUWELS 1985). Hence in normal mammalian species, SHT2 antagonists acting on their own have only very mild effects; moreover, the effects are only apparent under certain conditions or during short periods of the sleep cycle. Neurochemical effects Blockade of central 5HT2 receptors by 5HTz antagonists does not affect 5HT turnover (i.e. there is no effect on 5HT or 5HIAA levels) measured in various brain areas (LEYSEN et a1. 1985). Hence 5HTz receptor blockade appeared not to induce a feedback activation of 5HT neurons; this is in marked contrast with the dopaminergic system, where feedback activation of dopamine turnover in the brain upon dopamine-Dz receptor blocking is a well known phenomenon. Functional relationship between 5HT2 and 5HT1A receptors Various behavioral studies with 5HT agonists and antagonists have provided evidence for a mutual inhibitory action between 5HT 1A and 5HTz receptors stimulation (DARMANI et a1. 1990;
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BACKUS et al. 1990). In electrophysiological studies 5HT lA and 5HTz receptors were demonstrated to co-exist on pyramidal cells in the neocortex, where they mediate opposing effects: stimulation of the former induced membrane hyperpolarization and of the latter membrane depolarization. 5HTz receptors desensitize rapidly, whereas 5HT 1A receptors do not easily desensitize. It seems puzzling that there are two distinct receptors for serotonin on the same cell mediating opposing electrophysiological responses (ARANEDA and ANDRADE 1991). Since the 5HT 1A has nM affinity for serotonin and the 5HTz receptor 11M affinity, it seems likely that the former receptor would be the first one to become stimulated. The latter receptor would only be activated when the local serotonin concentration becomes high. ARANEDA and ANDRADE discuss the various possible functional consequences of the coexistence of the two 5HT receptor subtypes (ARANEDA and ANDRADE 1991).
Role of 5HT2 receptors in the dopaminergic system
5HTz antagonists reduce neuroleptic-induced catalepsy in rats. In patients they alleviate neuroleptic-induced extrapyramidal symptoms and Parkinson tremor (BERSANI et al. 1990). In a recent study, SALLER et al. (1990) showed that 5HTz antagonists can enhance the neurolepticinduced inrease of dopamine turnover in the basal ganglia, though not in certain mesolimbic brain areas. These observations have provided a possible explanation for the reduction by 5HTz antagonists of the neuroleptic-induced impairment of motoric functions: the enhanced dopamine turnover in the basal ganglia would partially counteract the dopamine receptor blockade in these areas.
5HT2 antagonists and drugs of abuse Ritanserin, a very potent, long acting and relatively selective 5HTz antagonist, has recently been shown to reduce forced preference of rats for various addictive drugs such as alcohol, cocaine and fentanyl (MEERT et al. 1991). A role for 5HTz receptors in these conditions seems worthy of further exploration.
Regulation of 5HT2 receptors 5HTz receptors become very rapidly desensitized and downregulated. This was apparent from in vivo studies on rats, where the behavioral response to 5HTz agonists was already significantly reduced by the second injection of a methoxyphenetylamine derivative and was completely disappeared after 4 injections. Concomitantly 5HTz receptor numbers, in the frontal cortex, measured ex vivo using radio ligand binding techniques, were markedly reduced (LEYSEN and PAUWELS 1990). Also, the early observations in humans of rapid and long-lasting tolerance to LSD-induced psychotic and behavioral effects (CHOLDEN et al. 1955) indicate that tolerance is probably to be ascribed to 5HTz receptor down regulation. In cells in culture, 5HTz receptor induced phospholipid turnover became 80% reduced upon 15 min exposure of the cells to serotonin (PAUWELS et al. 1990); moreover, a similar rapid desen-
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sitization of 5HTz receptor-mediated neuronal depolarization of cortical slices was observed in electrophysiological studies (ARANEDA and ANDRADE 1991). Rapid desensitization following agonist stimulation was also reported for the 5HT3 and the 5HT4 receptors, but in contrast the 5HT1A receptor appeared to be hard to desensitize. Hence, a clear distinction in the regulatory response of various 5HT receptor subtypes appears to exist. Serotonergic denervation by lesioning serotonergic neurons, and chronic blockading of 5HTz receptors by antagonists failed to induce 5HTz receptor up regulation (LEYSEN et al. 1983, 1986). By contrast, chronic treatment with 5HTz antagonists caused receptor desensitization and down regulation, an anomalous response not in agreement with the generally accepted receptor regulation theory (LEYSEN et al. 1986). The reported 5HTz receptor down regulation by tricyclic antidepressants, which has been related to the therapeutic action of the compounds (PEROUTKA and SNYDER 1980), is probably a consequence of the 5HTz receptor blocking properties of the compounds and not of their 5HT reuptake inhibitory effects, as was first suggested. Indeed selective 5HT uptake blockers, such as citalopram failed to induce 5HTz receptor down regulation (for review see CONN and SANDERS-BuSH 1987). Besides 5HTz antagonists, 5HT 1A mixed agonists can also cause 5HTz receptor down regulation when chronically administered. The precise mechanism of the anomalous 5HTz receptor down regulation is not known, but it seems likely that heterologous phenomena are involved. It remains possible that 5HTz receptor down regulation induced by the various drugs plays a role in the therapeutic action of the drugs in depression and anxiety (LEYSEN and PAUWELS 1990).
Pathophysiological role of 5HT2 receptors In double-blind controlled clinical studies with the 5HTz antagonist, ritanserin, therapeutic effects have been obtained in dysthymia (lack of drive, depressed mood, neurotic depression), generalized anxiety and negative symptoms of schizophrenia (REYNTJENS et al. 1986; BERSANI et al. 1991). The drug also reduced neuroleptic induced extrapyramidal symptoms and Parkinson tremor (BERSANI et al. 1990). 5HTz antagonists increased periods of slow-wave sleep in dysthymic patients and regularized disturbed sleep in psychotic patients (IDZIKOWSKI et al. 1991). It has been suggested that improvement of the quality of sleep may be an important factor in the therapeutic action of the compounds OANSSEN 1987). These findings with ritanserin corroborate the early hypothesis of a possible role for 5HTz antagonism in sleep regulation, which was proposed when the particular clinical observations made with pipamperone were related to its specificity for 5HTz receptors (LEYSEN et al. 1978). Recent studies with risperidone, a compound exhibiting combined predominant 5HTz and potent dopamine-Dz antagonism, revealed therapeutic effects on both positive and negative symptoms of schizophrenia with a very low attendant extrapyramidal side effect. In view of the findings with ritanserin, the two latter properties have been ascribed to the predominant 5HTz antagonistic action of the compound. Possible therapeutic use of 5HTz antagonists was suggested for the treatment of non-bulimic anorexia nervosa and for the treatment of alcohol and cocaine withdrawal.
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Physiological role of 5HT2 receptors As appears from the above, serotonergic neuron anatomy and aCtIVIty, response to 5HT2 receptor stimulation and blockading and 5HT2 receptor regulation show marked differences with known classical neurotransmitter systems. Poor synapsis formation and the low, barely influencable activity of serotonergic neurons; the presence of two 5HT receptor subtypes on the same cells with opposing electrophysiological responses; detrimental effects by 5HT2 agonists and the very rapid and profound desensitization induced by these effects; the lack of effect by 5HT2 antagonists acting on their own on behavior, motoric, intellectual and autonomic functions; the failure of 5HT2 receptors to up regulate following denervation or chronic blockading - all these observations point to a very low, if indeed there is any activation by 5HT2 receptors under normal physiological conditions. We have therefore proposed the following hypothesis: In mammals 5HT2 receptor stimulation is scarce under normal physiological conditions, with activation probably only occurring during short periods of the day-night cycle (e.g. to interrupt slow wave sleep), in emergency (e.g. in the periphery upon cardiovascular accident to induce vasoconstriction and platelet aggregation) or under pathological conditions (e.g. dysthymia, anxiety, social withdrawal impaired blood circulation). Hence under normal conditions acute blockading does not disrupt the normal situation, which explains the lack of directly observable effects by 5HT2 antagonists. Because of the low stimulation, the receptors may exist in a supersensitive state and chronic blockading or denervation cannot produce further supersensitivity or upregulation. The rapid and profound homologous desensitization and receptor down regulation is an effective defense mechanism against the detrimental effects of enhanced agonist stimulation. In addition, 5HT2 receptors are sensitive to heterologous desensitization, such as by 5HT2A receptor stimulation. It seems not unlikely that under pathological conditions which are responsive to 5HT2 antagonists, 5HT2 receptor stimulation is enhanced and homologous receptor downregulation impaired. Under such conditions, 5HT2 antagonists will exert therapeutic effects, both acutely by blocking the receptors and chronically by downregulating them via heterologous mechanisms.
References ARANEDA, R., ANDRADE, R.: 5-Hydroxytryptaminez and 5-hydroxytryptaminelA receptors mediate opposing responses on membrane excitability in rat association cortex. - Neurosci. 40, 399-412 (1991). BACKUS, L. 1., SHARP, T., GRAHAME-SMITH, D. G.: Behavioural evidence for a functional interaction between central 5-HT z and 5-HT 1A receptors. - Brit.]. Pharmacol. 100, 793-799 (1990). BERSANI, G., GRISPINI, A., MARINI, S., PASINI, A., VALDUCCI, M., CIANI, N.: 5-HTz Antagonist ritanserin in neuroleptic-induced Parkinsonism: A double-blind comparison with orphenadrine and placebo. - Clin. Neuropharmacol. 13, 500-506 (1990). BERSANI, G., POZZI, F., MARINI, S., GRISPINI, A., PASINI, A., CIANI, N.: 5-HTz receptor antagonism in dysthymic disorder: a double-blind placebo-controlled study with ritanserin. - Acta psychiat. scand. 83, 244-248 (1991). . CHOLDEN, L. S., KURLAND, A., SAVAGE, c.: Clinical reactions and tolerance to LSD in chronic schizophrenia. - J. nerv. ment. Dis. 122, 211-221 (1955). CONN, P. J., SANDERS-BuSH, E.: Central serotonin receptors: Effector systems, physiological roles and regulation. - Psychopharmacol. 92, 267-277 (1987).
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CROSS, A. J.: Serotonin in Alzheimer-type dementia and other dementing illnesses. - In: The Neuropharmacology of Serotonin (WHITAKER-AzMITIA, P. M., PEROUTKA, S. J., eds.), Vol. 600, pp. 405-417. - The New York Academy of Sciences, New York 1990. DARMANI, N. A., MARTIN, B. R., PANDEY, U., GLENNON, R. A.: Do functional relationships exist between 5HTIA and 5-HT2 receptors? - Pharmacol. Biochem. Behav. 36,901-906 (1990). DE CHAFFOY DE COURCELLES, D., LEYSEN, J. E., DE CLERCK, F., VAN BELLE, H., JANSSEN, P. A. J.: Evidence that phospholipid turnover is the signal transducing system coupled to serotonin-S2 receptor sites. - J. bioI. Chern. 260, 7603-7608 (1985). DEs CARRIES, L., AUDET, M.-A., DOUCET, G., GARCIA, S., OLESKEVICH, S., SEGUE LA, P. H., SOGHOMORIAN, J.-J., WATKINS, K. c.: Morphology of central serotonin neurons. Brief review of quantified aspects of their distribution and ultrastructural relationships. - In: The Neuropharmacology of Serotonin (WHITAKERAZMITIA, P. M., PEROUTKA, S. J., eds.), Vol. 600, pp. 81-92. - The New York Academy of Sciences, New York 1990. FELDER, C. c., KANTERMAN, R., Y., MA, A. L., AxELROD, J.: Serotonin stimulates phospholipase A2 and the release of arachidonic acid in hippocampal neurons by a type 2 serotonin receptor that is independent of inositolphospholipid hydrolysis. - Proc. nat!. Acad. Sci. USA 87, 2187-2191 (1990). IDZIKOWSKI, C., MILLS, F. J., JAMES, R. J.: A dose-response study examing the effects of ritanserin on human slow wave sleep. - Brit. J. clin. Pharmacol. 31,193-196 (1991). JACOBS, B. L., WILKENSON, L. 0., FORNAL, C. A.: The role of brain serotonin. A neurophysiological perspective. - Neuropsychopharmacol. 3,473-479 (1990). JANSSEN, P. A. J.: Does ritanserin, a potent serotonin-S2 antagonist restore energetic functions during the night? - J. roy. Soc. Med. 80, 409-413 (1987). LEYSEN, J. E.: Gaps and peculiarities in 5-HT2 receptor studies. - Neuropsychopharmacol. 3,361-369 (1990). -: Use of 5-HT receptor agonists and antagonists for the characterization of their respective receptor sites. In: Neuromethods, Drugs as Tools in Neurotransmitter Research (BOULTON, A. B., BAKER, G. B., AUGuSTO, V. J., eds.), Vol. 12, pp. 299-350. - Humana Press., Clifton, NJ 1989). LEYSEN, J. E., EENS, A., GOMMEREN, W., VAN GOMPEL, P., WYNANTS, J., JANSSEN, P. A. J.: Identification of nonserotonergic eH]ketanserin binding sites associated with nerve terminals in rat brain and with platelets; relation with release of biogenic amine metabolites induced by ketanserin- and tetrabenazine-like drugs. J. Pharmacol. expo Ther. 244, 310-321 (1988). LEYSEN, J. E., GOMMEREN, W., VAN GOMPEL, P., WYNANTS, J., JANSSEN, P. F. M., LADURON, P. M.: Receptor-binding properties in vitro and in vivo of ritanserin. A very potent and long acting serotonin-S2 antagonist. - Mol. Pharmacol. 27, 600-611 (1985). LEYSEN, J. E., NIEMEGEERS, C. J. E., TOLLENAERE, J. P., LADURON, P. M.: Serotonergic component of neuroleptic receptors. - Nature (Lond.) 272, 168-171 (1978). LEYSEN, J. E., NIEMEGEERS, C. J. E., VAN NUETEN, J. M., LADURON, P. M.: eH]Ketanserin (R 41468), a selective 3H-ligand for serotonin2 receptor binding sites. - Mol. Pharmacol. 21, 301-314 (1982). LEYSEN, J. E., PAUWELS, P.: 5-HT2 Receptors, roles and regulation. - In: The Neuropharmacology of Serotonin (WHlTAKER-AzMITIA, P. M., PEROUTKA, S. J., eds.), Vol. 600, pp. 183-193. - The New York Academy of Sciences, New York 1990. LEYSEN, J. E., VAN GOMPEL, P., GOMMEREN, W., WOESTENBORGHS, R., JANSSEN, P. A. J.: Down regulation of serotonin-S2 receptor sites in rat brain by chronic treatment with the serotonin-S2 antagonists: ritanserin and setoperone. - Psychopharmacol. 88, 434-444 (1986). LEYSEN, J. E., VAN GOMPEL, P., VERWIMP, M., NIEMEGEERS, C. J. E.: Role and localization of serotonin (S2)receptor-binding sites: Effects of neuronal lesions. - In: CNS-Receptors-From Molecular Pharmacology to Behavior (MANDEL, P., DEFEUDlS, F. V., eds.), pp. 373-383. - Raven Press, New York 1983. MEERT, T. F., AWOUTERS, F., NIEMEGEERS, C. J. E., SCHELLEKENS, K. H. L., JANSSEN, P. A. J.: Ritanserin reduces abuse of alcohol, cocaine and fentanyl in rats. - Pharmacopsychiatry, in press (1991). PAUWELS, P. J., VAN GOMPEL, P., LEYSEN, J. E.: Rapid desensitization and resensitization of 5-HT2 receptor mediated phosphatidyl inositol hydrolysis by serotonin agonists in quiescent calf aortic smooth muscle cells. - Life Sci. 47, 2009-2019 (1990). PEROUTKA, S. J., SNYDER, S. H.: Regulation of serotonin2 (5-HTz) receptors labeled with eH]spiroperidol by chronic treatment with the antidepressant amitriptyline. - J. Pharmacol. expo Ther. 215, 582-587 (1980). PIERCE, P. A., PEROUTKA, S. J.: Evidence for distinct 5-hydroxytryptamine2 binding site subtypes in cortical membrane preparations. - J. Neurochem. 52, 656-658 (1989).
Serotonin-5HT2 receptors . 249 RAD]A, F., LAPORTE, A.-M., DAVAL, G., VERGE, D., GOZLAN, H., HAMON, M.: Autoradiography of serotonin receptor subtypes in the central nervous system. - Neurochem. Int. 18, 1-15 (1991). REYNT]ENS, A., GELDERS, Y. G., HOPPENBROUWERS, M.-L. J. A., VANDEN BUSSCHE, G.: Thymostenic effects of ritanserin (R 55 667), a centrally acting serotonin-S2 receptor blocker. - Drug Dev. Res. 8, 205-211 (1986). SALLER, C. F., CZUPRYNA, M. J., SALAMA, A. I.: 5-HT2 Receptor blockade by ICI 169,369 and other 5-HT2 antagonists modulates the effects of D-2 dopamine receptor blockade. - J. Pharmacol. expo Ther. 253, 1162-1170 (1990). SANDERS-BuSH, E., TSUTSUMI, M., BURRIS, K. D.: Serotonin receptors and phosphatidylinositol turnover. In: The Neuropharmacology of Serotonin (WHITAKER-AzMITIA, P. M., PEROUTKA, S. J., eds.), Vol. 600, pp. 224-236. - The New York Academy of Sciences, New York 1990. SCHOTTE, A., MALOTEAUX, J. M., LADURON, P. M.: Characterization and regional distribution of serotonin Srreceptors in human brain. - Brain Res. 276, 231-235 (1983). TORK, I.: Anatomy of the serotonergic system. - In: The Neuropharmacology of Serotonin (WHITAKERAZMITIA, P. M., PEROUTKA, S. J., eds.), Vol. 600, pp. 9-35. - The New York Academy of Sciences, New York 1990.
