Ligands and tracers for PET studies of the 5-HT system—current status

The status of the radiochemical development and biological evaluation of radioligands and tracers for PET studies of the serotonergic system is reviewed, indicating those agents with present value and those with future potential. Practical recommendations are given for the preparation of two useful radioligands for PET studies of central 5-HT₂ receptors, namely [¹⁸F]setoperone and [¹⁸F]altanserin. Though, it has not proved possible to recommend tracers or radioligands for the study of other aspects of the serotonergic system, prospects for future radiochemical development are indicated, especially for developing radioligands for the 5-HT re-uptake site, and for the 5-HT₁ and 5-HT₃ receptors.     

Do imipramine and dihydroergosine possess two components--one stimulating 5-HT1 and the other inhibiting 5-HT2 receptors?

The mechanisms by which imipramine and dihydroergosine stimulate the 5-HT syndrome in rats and inhibit the head-twitch response in rats and mice were studied. Imipramine- and dihydroergosine-induced stimulation of the 5-HT syndrome was inhibited stereoselectively by propranolol, a high affinity ligand for 5-HT1 receptor sites, but not by ritanserin, a specific 5-HT2 receptor antagonist. (-)-Propranolol potentiated the inhibitory effect of imipramine, but not of dihydroergosine on the head-twitch response, while ritanserin was without effect. Neither imipramine nor dihydroergosine were able to stimulate the 5-HT syndrome in the animals pretreated with p-chlorophenylalanine. As expected, 8-OH-DPAT, a selective 5-HT1A receptor agonist, stimulated, and 5-HT1B agonists CGS 12066B and 1-(trifluoromethylphenyl)piperazine (TFMPP) failed to stimulate the 5-HT syndrome induced in rats by pargyline and 5-HTP administration. A higher dose of ritanserin inhibited the syndrome. While 8-OH-DPAT alone produced all behavioral components of the 5-HT syndrome, dihydroergosine or imipramine alone even at very high doses never produced tremor or a more intensive forepaw padding as seen when these drugs were given in combination with pargyline and 5-HTP. A single administration of (-)-propranolol also inhibited the head-twitch response. This effect lasted in mice longer than after ritanserin administration. In in vitro experiments dihydroergosine expressed approximately twenty-fold higher affinity for 3H-ketanserin binding sites than imipramine. The results suggest that imipramine and dihydroergosine possess two components--one stimulating the 5-HT syndrome in rats by a presynaptic, presumably 5-HT1A-mediated mechanism, and the other inhibiting 5-HT2 binding sites.     

Molecular and Pharmacological Characterization of Serotonin 5-HT2α and 5-HT7 Receptors in the Salivary Glands of the Blowfly Calliphora vicina

Secretion in blowfly (Calliphora vicina) salivary glands is stimulated by the biogenic amine serotonin (5-hydroxytryptamine, 5-HT), which activates both inositol 1,4,5-trisphosphate (InsP₃)/Ca²⁺ and cyclic adenosine 3′,5′-monophosphate (cAMP) signalling pathways in the secretory cells. In order to characterize the signal-inducing 5-HT receptors, we cloned two cDNAs (Cv5-ht2α, Cv5-ht7) that share high similarity with mammalian 5-HT₂ and 5-HT₇ receptor genes, respectively. RT-PCR demonstrated that both receptors are expressed in the salivary glands and brain. Stimulation of Cv5-ht2α-transfected mammalian cells with 5-HT elevates cytosolic [Ca²⁺] in a dose-dependent manner (EC₅₀ = 24 nM). In Cv5-ht7-transfected cells, 5-HT produces a dose-dependent increase in [cAMP]_i (EC₅₀ = 4 nM). We studied the pharmacological profile for both receptors. Substances that appear to act as specific ligands of either Cv5-HT_2α or Cv5-HT₇ in the heterologous expression system were also tested in intact blowfly salivary gland preparations. We observed that 5-methoxytryptamine (100 nM) activates only the Cv5-HT_2α receptor, 5-carboxamidotryptamine (300 nM) activates only the Cv5-HT₇ receptor, and clozapine (1 µM) antagonizes the effects of 5-HT via Cv5-HT₇ in blowfly salivary glands, providing means for the selective activation of each of the two 5-HT receptor subtypes. This study represents the first comprehensive molecular and pharmacological characterization of two 5-HT receptors in the blowfly and permits the analysis of the physiological role of these receptors, even when co-expressed in cells, and of the modes of interaction between the Ca²⁺- and cAMP-signalling cascades.     

4-Aminoethylpiperazinyl aryl ketones with 5-HT₁A/5-HT₇ selectivity

The well-known 5-HT(1A)/5-HT(7) selectivity issue was tackled by a new series of 4-aminoethylpiperazinyl aryl ketones (1a-1l) specifically designed to distinguish the two hydrophobic sites centered at the anchoring salt bridge. The 4-aminoethylpiperazinyl aryl ketones showed a wide spectrum of activity and selectivity for the 5-HT receptors depending on the type of the hydrophobic groups attached at the aryl piperazinyl ketone scaffold. Docking study of the most active compounds against 5-HT(7)R and 5-HT(1A)R revealed that both receptors have two hydrophobic pockets around the anchoring salt bridge. These two binding sites are perpendicular to each other in 5-HT(7)R but parallel in 5-HT(1A)R, and this observation is well matched with the previous report which claimed that 5-HT(7)R affinity arises from bent conformation of the bound ligand whereas an extended one is best suited for 5-HT(1A)R selectivity. Also, as these pockets have different size and shape, inhibitory activity as well as selectivity of the 4-aminoethylpiperazinyl aryl ketones against 5-HT(7)R and 5-HT(1A)R seemed to be determined by combination of two hydrophobic substituents attached at both ends of the title compounds.     

中枢5-HT与酒精依赖有关

动物及临床实验表明,一些药物能选择性地与5-HT系统相互作用,从而降低机体对酒精的摄取。由酒精滥用与依赖所导致的社会及自身健康与经济上的后果给社会带来严重的问题,例如,美国1990年用于酒精滥用与依赖的经济费用就高达1363亿美元。在过去40年,对酒精滥用及酒精依赖患者主要给予抗抑郁、抗焦虑、镇静催眠等治疗精神病的药物,但效果甚微。     

The pharmacology of the isolated foregut of the locust Schistocerca gregaria—II. characterization of a 5-HT2-like receptor

1. 5-HT (10⁻⁸−5 × 10⁻⁶ M) relaxed isolated locust foreguts.2. The effects of 5-HT were mimicked by 5,6-DHT, 5-MeOT, tryptamine, 5-MeT, MK212 and methysergide while 5-hydroxyindole, 5-hydroxyindole acetic acid, 5-hydroxytryptophol, NN-DMT, 8-OH DPAT and RU 24969 were without effect.3. Ketanserin (pA₂ = 5.65) was a competitive antagonist of the effects of 5-HT, MK212 and methysergide.4. Mianserin (pA₂ = 6.3) was also a competititve antagonist of 5-HT but ICS 205-930 had no antagonistic effect on 5-HT-induced relaxation.5. It is concluded that 5-HT relaxes the locust foregut by interacting with 5-HT₂-like receptors.     

Sulfonamides with the N-alkyl-N′-dialkylguanidine moiety as 5-HT7 receptor ligands

A series of arylsulfonamides containing guanidine incorporated in the structure of secondary amines (piperidine, piperazine) was synthesized on SynPhase Lanterns and evaluated for 5-HT_1A, 5-HT_2A, and 5-HT₇ receptors. The results demonstrated that N-alkyl-N′-dialkylguanidines displayed good 5-HT₇/5-HT_1A selectivity and may be regarded as promising structural core for development of 5-HT₇ ligands.     

5-HT受体亚型在大鼠颌下腺的免疫组织化学定位及5-HT的功能研究

目的 研究探讨了大鼠颌下腺中5-羟钩胺受体亚型的分布以及5-HT功能。方法 免疫组织化学法和免疫酶联检测法,结果 大鼠颌下腺的浆液性腺泡上皮细胞,闰管,颗粒曲管,纹状管和排泄管的管壁上皮细胞均呈5-HT1AR离体培养的颌下腺分泌神经生长因子（NGF）,但是,当外源5-HT浓度大于10^-7时却抑制NGF的分泌。结论 提示5-HT对颌下腺NGF的分泌可能起双向调节的作用。     

Computational exploration of polymorphisms in 5-hydoxytryptamine 5-HT₁A and 5-HT₂A receptors associated with psychiatric disease

The huge polymorphic data have been prioritized towards a specific disease based on sequence and structure homology tools to a large extent. In this study, we have explored the potential non-synonymous Single Nucleotide Polymorphism (nsSNP) in serotonin (5-HT) receptors involved in psychotic syndromes and their response pathway. The most damaging point mutations were screened from 12 classes of serotonin receptors comprising 7743 variants. In 5HT(1A) receptor, two alleles were found to be highly deleterious located at ligand binding extracellular-2 and one at intracellular loop-3 domains. Similarly, we found two alleles predicted to be highly damaging in 5HT(2A) residing at N and C-Terminal domains. The above alleles were further confirmed based on their flexibility and stability difference using the molecular dynamic simulation analysis. Integrating these results appeared promising for being able to filter out potential non-synonymous Single Nucleotide Polymorphisms for neuropsychiatric disorders.     

荷叶中的生物碱及其对5-HT 2A和5-HT 2C受体的激动作用研究CSCD

对荷叶中的生物碱进行了分离、鉴定和调脂减肥活性研究。本研究结合传统酸提碱沉法与现代高效液相色谱制备技术,从荷叶中分离、纯化到11个生物碱,分别被鉴定为N-氧基原荷叶碱(1)、原荷叶碱(2)、莲碱(3)、降氧化北美黄连次碱(4)、荜茇宁(5)、巴婆碱(6)、O-去甲基荷叶碱(7)、N-去甲基荷叶碱(8)、荷叶碱(9)、衡州乌药碱(10)和亚美罂粟碱(11),其中,化合物1、4和5为首次从荷叶中分得。测试所得化合物对5-HT_(2A)和5-HT_(2C)受体的激动作用,结果表明11个生物碱对5-HT_(2A)受体均具有一定的激动作用,进一步揭示了荷叶调脂减肥的可能药效基础和作用机理。     

Biochemical and pharmacological study of N-linked glycosylation of the human serotonin 5-HT₇a receptor

The 5-hydroxytryptamine (5-HT)(7(a)) receptor is a G-protein-coupled receptor critically involved in human psychiatric and neurological disorders. In the present study, we evaluate the presence and the functional role of N-glycosylation of the human 5-HT(7) receptor. Western blot analysis of HEK293T cells transiently expressing the 5-HT(7(a)) receptor in the presence of tunicamycin gave rise to a band shift, indicating the existence of an N-glycosylated form of the 5-HT(7(a)) receptor. To further investigate this, we mutated the two predicted N-glycosylation sites (N5Q and N66Q) and compared the molecular mass of the immunoreactive bands with those of the wild-type receptor, indicating that both asparagines were N-glycosylated. The mutant receptors had the same binding affinity for [(3) H]5-CT and the same potency and efficacy with regard to 5-HT-induced activation of adenylyl cyclase. However, there was a reduction in maximal ligand binding for the single and double mutants compared to the wild-type receptor. Next, membrane labelling and immunocytochemical studies demonstrated that the N-glycosylation mutants were expressed at the cell surface. We conclude that N-glycosylation is not important for cell surface expression of the 5-HT(7) receptor.     

Are 5-HT receptors involved in the sprouting of serotoninergic terminals following neonatal 5,7-dihydroxytryptamine treatment in the rat?

The neonatal administration of 5,7-dihydroxytryptamine to rats (100 mg kg^?1 s.c. on the 1st and 2nd day after birth) resulted in marked reductions in serotoninergic presynaptic markers ([³H]-5-HT synaptosomal uptake, tryptophan hydroxylase activity and endogenous 5-HT content) in various forebrain areas, particularly the cerebral cortex and the hippocampus. In contrast, this treatment produced an increased outgrowth of serotoninergic terminals in the brain stem as judged by the significant increments of these presynaptic markers in this region. Both in the hippocampus and the brain stem, these 5,7-dihydroxytryptamine-induced changes in serotoninergic innervation were associated with a transient increase in 5-HT-sensitive adenylate cyclase activity. No significant alteration of the specific high affinity binding of [³H]-5-HT to synaptosomal membranes from various brain regions was detected in 5,7-dihydroxytryptamine-treated rats for at least the first postnatal month.The chronic blockade of 5-HT receptors by metergoline (5 mg kg^?1 day^?1 from day 3 to day 22 after birth) altered neither the changes in presynaptic markers nor the evolution of [³H]-5-HT high affinity binding in 5,7-dihydroxytryptamine-treated rats.These findings further illustrate that the high affinity binding sites for [³H]-5-HT do not correspond to postsynaptic 5-HT receptors coupled to adenylate cyclase in the rat brain. Apparently, 5-HT receptors play no role in the increased outgrowth of serotoninergic systems in the brain stem following neonatal 5,7-dihydroxy-tryptamine treatment.     

5-HT2和5-HT3受体在外周初级感觉神经末梢痛反应和痛调制中的相互作用

目的：探讨5-HT2和5-HT3受体亚型在5-HT引起外周痛反应和痛调制中的相互作用及其机制;方法：在大鼠三又神经节神经元标本上应用全细胞膜片钳技术记录5-羟色胺激活电流（15_HT）,并结合痛行为实验进行观察。结果：在大多数受检细胞（54／88,61．4％）特别是中、小型细胞外加5-HT可引起一快去敏感的内向电流,此内向电流能被5-HT,受体特异性激动剂2-甲基-5-羟色胺所模拟,被5-HT3受体拮抗剂ICS250-930可逆性阻断,而5-HT2受体激动剂α-甲基-5-羟色胺则有明显增强15-HT的作用,5-HT1受体激动剂R-（＋）-UH301无明显反应。在进一步的整体清醒动物的行为学试验中我们观察到,大鼠后肢掌底皮下注射5-HT（10-5,10-4和10-3mol／L）引起浓度依赖性的痛行为反应,而用5-HT2和5-HT3受体特异性拮抗剂Cyproheptadine和ICS250-930分别阻断相应受体亚型后,5-HT引起的痛行为反应的强度序列为：5-HT〉5-HT＋ICS〉5-HT＋Cyp。结论：本文结果提示：5-HT所引起的痛反应中,在初级感觉神经元水平5-HT3受体可能仅起着启始作用,而5-HT,受体则在伤害性信息的维持和调制过程中发挥更大的作用。     

Novel brain penetrant benzofuropiperidine 5-HT₆ receptor antagonists

7-Arylsulfonyl substituted benzofuropiperidine was discovered as a novel scaffold for 5HT(6) receptor antagonists. Optimization by substitution at C-1 position led to identification of selective, orally bioavailable, brain penetrant antagonists with reduced hERG liability. An advanced analog tested in rat social recognition model showed significant activity suggesting potential utility in the enhancement of short-term memory.     

Synthesis and structure–affinity relationships of novel small molecule natural product derivatives capable of discriminating between serotonin 5-HT1A, 5-HT2A, 5-HT2C receptor subtypes

Efforts to develop ligands that distinguish between clinically relevant 5-HT2A and 5-HT2C serotonin receptor subtypes have been challenging, because their sequences have high homology. Previous studies reported that a novel aplysinopsin belonging to a chemical class of natural products isolated from a marine sponge was selective for the 5-HT2C over the 5-HT2A receptor subtype. Our goal was to explore the 5-HT2A/2C receptor structure–affinity relationships of derivatives based on the aplysinopsin natural product pharmacophore. Twenty aplysinopsin derivatives were synthesized, purified and tested for their affinities for cloned human serotonin 5-HT1A, 5-HT2A, and 5-HT2C receptor subtypes. Four compounds in this series had >30-fold selectivity for 5-HT2A or 5-HT2C receptors. The compound (E)-5-((5,6-dichloro-1H-indol-3-yl)methylene)-2-imino-1,3-dimethylimidazolidin-4-one (UNT-TWU-22, 16) had approximately 2100-fold selectivity for the serotonin 5-HT2C receptor subtype: an affinity for 5-HT2C equal to 46 nM and no detectable affinity for the 5-HT1A or 5-HT2A receptor subtypes. The two most important factors controlling 5-HT2A or 5-HT2C receptor subtype selectivity were the combined R1,R3-alkylation of the imidazolidinone ring and the type and number of halogens on the indole ring of the aplysinopsin pharmacophore.     

Is nitric oxide involved in 5-HT2B receptor-mediated contraction in the rat stomach fundus?

Although contraction of the rat stomach fundus by 5-HT is known to be mediated by the 5-HT2B receptor, the second messenger pathways involved in this response remain unclear. Since nitric oxide (NO) has been associated with contraction of certain gastrointestinal smooth muscle, the purpose of this study was to determine if NO is involved in 5-HT-induced contraction in the rat stomach fundus. The arginine analogs L-NAME and L-NMMA, at a concentration (100 μM) established to inhibit NO synthase in the rat stomach fundus by inhibiting depolarizationinduced relaxation in this tissue, had no effect on 5-HT contraction. Furthermore, the NO donors sodium nitroprusside and SNAP did not contract rat stomach fundus under basal tone, whereas 5-HT was a potent contractile agonist. These data do not support a role for NO in 5-HT_2B receptormediated contraction in the rat stomach fundus.     

The discovery and SAR of indoline-3-carboxamides—A new series of 5-HT6 antagonists

Antagonists of the 5-HT₆ receptor have been shown to improve cognitive function in a wide range of animal models and as such may prove to be attractive agents for the symptomatic treatment of cognitive disorders such as Alzheimer’s disease (AD) and schizophrenia. We report herein the identification and SAR around N-(2-aminoalkyl)-1-(arylsulfonyl)indoline-3-carboxamides—a novel chemotype of 5-HT₆ antagonists.