N-(4-Bromo-2,5-Dimethoxyphenethyl)-6-(4-Phenylbutoxy)Hexan-1-Amine (XOB): A Novel Phenylalkylamine Antagonist of Serotonin 2A Receptors and Voltage-Gated Sodium Channels.

Bipolar disorder impacts millions of patients in the United States but the mechanistic understanding of its pathophysiology and therapeutics is incomplete. Atypical antipsychotic serotonin2A (5-HT2A) receptor antagonists, such as quetiapine and olanzapine, and mood-stabilizing voltage-gated sodium channel (VGSC) blockers, such as lamotrigine, carbamazepine, and valproate, show therapeutic synergy and are often prescribed in combination for the treatment of bipolar disorder. Combination therapy is a complex task for clinicians and patients, often resulting in unexpected difficulties with dosing, drug tolerances, and decreased patient compliance. Thus, an unmet need for bipolar disorder treatment is to develop a therapeutic agent that targets both 5-HT2A receptors and VGSCs. Toward this goal, we developed a novel small molecule that simultaneously antagonizes 5-HT2A receptors and blocks sodium current. The new compound, N-(4-bromo-2,5-dimethoxyphenethyl)-6-(4-phenylbutoxy)hexan-1-amine (XOB) antagonizes 5-HT-stimulated, Gq-mediated, calcium flux at 5-HT2A receptors at low micromolar concentrations while displaying negligible affinity and activity at 5-HT1A, 5-HT2B, and 5-HT2C receptors. At similar concentrations, XOB administration inhibits sodium current in heterologous cells and results in reduced action potential (AP) firing and VGSC-related AP properties in mouse prefrontal cortex layer V pyramidal neurons. Thus, XOB represents a new, proof-of-principle tool that can be used for future preclinical investigations and therapeutic development. This polypharmacology approach of developing a single molecule to act upon two targets, which are currently independently targeted by combination therapies, may lead to safer alternatives for the treatment of psychiatric disorders that are increasingly being found to benefit from the simultaneous targeting of multiple receptors. SIGNIFICANCE STATEMENT: The authors synthesized a novel small molecule (XOB) that simultaneously antagonizes two key therapeutic targets of bipolar disorder, 5-HT2A receptors and voltage-gated sodium channels, in heterologous cells, and inhibits the intrinsic excitability of mouse prefrontal cortex layer V pyramidal neurons in brain slices. XOB represents a valuable new proof-of-principle tool for future preclinical investigations and provides a novel molecular approach to the pharmacological treatment of complex neuropsychiatric disease, which often requires a combination of therapeutics for sufficient patient benefit.

Serotonin 2C receptor in a rat model of temporal lobe epilepsy: From brainstem expression to pharmacological blockade in relation to ventilatory function.

Because of its involvement in breathing control and neuronal excitability, dysregulation of the serotonin (5-HT) 2C receptor (5-HT2C) might play a key role in sudden unexpected death in epilepsy. Seizure-induced respiratory arrest is thus prevented by a 5-HT2B/C agonist in different seizure model. However, the specific contribution of 5-HT2C in chronic epilepsy-related respiratory dysfunction remains unknown. In a rat model of temporal lobe epilepsy (EPI rats), in which we previously reported interictal respiratory dysfunctions and a reduction of brainstem 5-HT tone, quantitative reverse transcriptase polymerase chain reaction showed overexpression of TPH2 (5-HT synthesis enzyme), SERT (5-HT reuptake transporter), and 5-HT2C transcript levels in the brainstem of EPI rats, and of RNA-specific adenosine deaminase (ADAR1, ADAR2) involved in the production of 5-HT2C isoforms. Interictal ventilation was assessed with whole-body plethysmography before and 2 h after administration of SB242084 (2 mg/kg), a specific antagonist of 5-HT2C. As expected, SB242084 administration induced a progressive decrease in ventilatory parameters and an alteration of breathing stability in both control and EPI rats. However, the size of the SB242084 effect was lower in EPI rats than in controls. Increased 5-HT2C gene expression in the brainstem of EPI rats could be part of a compensatory mechanism against epilepsy-related low 5-HT tone and expression of 5-HT2C isoforms for which 5-HT affinity might be lower.

[18F]RS-127445 radiosynthesis and evaluation as a 5-HT2B receptor PET radiotracer in rat brain.

Serotonin (5-hydroxytryptamine, 5-HT) is a neurotransmitter involved in many physiological and pathological mechanisms through its numerous receptors. Among these, the 5-HT2B receptor is known to play a key role in multiple brain disorders but remains poorly understood. Positron emission tomography (PET) can contribute to a better understanding of pathophysiological mechanisms regulated by the 5-HT2B receptor. To develop the first PET radiotracer for the 5-HT2B receptor, RS-127445, a well-known 5-HT2B receptor antagonist, was labeled with fluorine-18. [18F]RS-127445 was synthesized in a high radiochemical purity and with a good molar activity and radiochemical yield. Preliminary PET scans in rats showed good brain penetration of [18F]RS-127445. However, competition experiments and in vitro autoradiography showed high non-specific binding, especially to brain white matter.

Desloratadine alleviates ALS-like pathology in hSOD1G93A mice via targeting 5HTR2A on activated spinal astrocytes.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with progressive loss of motor neurons in the spinal cord, cerebral cortex and brain stem. ALS is characterized by gradual muscle atrophy and dyskinesia. The limited knowledge on the pathology of ALS has impeded the development of therapeutics for the disease. Previous studies have shown that autophagy and astrocyte-mediated neuroinflammation are involved in the pathogenesis of ALS, while 5HTR2A participates in the early stage of astrocyte activation, and 5HTR2A antagonism may suppress astrocyte activation. In this study, we evaluated the therapeutic effects of desloratadine (DLT), a selective 5HTR2A antagonist, in human SOD1G93A (hSOD1G93A) ALS model mice, and elucidated the underlying mechanisms. HSOD1G93A mice were administered DLT (20 mg·kg-1·d-1, i.g.) from the age of 8 weeks for 10 weeks or until death. ALS onset time and lifespan were determined using rotarod and righting reflex tests, respectively. We found that astrocyte activation accompanying with serotonin receptor 2 A (5HTR2A) upregulation in the spinal cord was tightly associated with ALS-like pathology, which was effectively attenuated by DLT administration. We showed that DLT administration significantly delayed ALS symptom onset time, prolonged lifespan and ameliorated movement disorders, gastrocnemius injury and spinal motor neuronal loss in hSOD1G93A mice. Spinal cord-specific knockdown of 5HTR2A by intrathecal injection of adeno-associated virus9 (AAV9)-si-5Htr2a also ameliorated ALS pathology in hSOD1G93A mice, and occluded the therapeutic effects of DLT administration. Furthermore, we demonstrated that DLT administration promoted autophagy to reduce mutant hSOD1 levels through 5HTR2A/cAMP/AMPK pathway, suppressed oxidative stress through 5HTR2A/cAMP/AMPK/Nrf2-HO-1/NQO-1 pathway, and inhibited astrocyte neuroinflammation through 5HTR2A/cAMP/AMPK/NF-κB/NLRP3 pathway in the spinal cord of hSOD1G93A mice. In summary, 5HTR2A antagonism shows promise as a therapeutic strategy for ALS, highlighting the potential of DLT in the treatment of the disease. DLT as a 5HTR2A antagonist effectively promoted autophagy to reduce mutant hSOD1 level through 5HTR2A/cAMP/AMPK pathway, suppressed oxidative stress through 5HTR2A/cAMP/AMPK/Nrf2-HO-1/NQO-1 pathway, and inhibited astrocytic neuroinflammation through 5HTR2A/cAMP/AMPK/NF-κB/NLRP3 pathway in the spinal cord of hSOD1G93A mice.

Metabolism, Disposition, Excretion, and Potential Transporter Inhibition of 7-16, an Improving 5-HT2A Receptor Antagonist and Inverse Agonist for Parkinson's Disease.

Compound 7-16 was designed and synthesized in our previous study and was identified as a more potential selective 5-HT2A receptor antagonist and inverse agonist for treating Parkinson's disease psychosis (PDP). Then, the metabolism, disposition, and excretion properties of 7-16 and its potential inhibition on transporters were investigated in this study to highlight advancements in the understanding of its therapeutic mechanisms. The results indicate that a total of 10 metabolites of 7-16/[14C]7-16 were identified and determined in five species of liver microsomes and in rats using UPLC-Q Exactive high-resolution mass spectrometry combined with radioanalysis. Metabolites formed in human liver microsomes could be covered by animal species. 7-16 is mainly metabolized through mono-oxidation (M470-2) and N-demethylation (M440), and the CYP3A4 isozyme was responsible for both metabolic reactions. Based on the excretion data in bile and urine, the absorption rate of 7-16 was at least 74.7%. 7-16 had weak inhibition on P-glycoprotein and no effect on the transport activity of OATP1B1, OATP1B3, OAT1, OAT3, and OCT2 transporters. The comprehensive pharmacokinetic properties indicate that 7-16 deserves further development as a new treatment drug for PDP.

Flexible versus Fixed Spatial Self-Ordered Response Sequencing: Effects of Inactivation and Neurochemical Modulation of Ventrolateral Prefrontal Cortex.

Previously, studies using human neuroimaging and excitotoxic lesions in non-human primate have demonstrated an important role of ventrolateral prefrontal cortex (vlPFC) in higher order cognitive functions such as cognitive flexibility and the planning of behavioral sequences. In the present experiments, we tested effects on performance of temporary inactivation (using GABA receptor agonists) and dopamine (DA) D2 and 5-HT2A-receptor (R) blockade of vlPFC via local intracerebral infusions in the marmoset. We trained common marmosets to perform spatial self-ordered sequencing tasks in which one cohort of animals performed two and three response sequences on a continuously varying spatial array of response options on a touch-sensitive screen. Inactivation of vlPFC produced a marked disruption of accuracy of sequencing which also exhibited significant error perseveration. There were somewhat contrasting effects of D2 and 5-HT2A-R blockade, with the former producing error perseveration on incorrect trials, though not significantly impairing accuracy overall, and the latter significantly impairing accuracy but not error perseveration. A second cohort of marmosets were directly compared on performance of fixed versus variable spatial arrays. Inactivation of vlPFC again impaired self-ordered sequencing, but only with varying, and not fixed spatial arrays, the latter leading to the consistent use of fewer, preferred sequences. These findings add to evidence that vlPFC is implicated in goal-directed behavior that requires higher-order response heuristics that can be applied flexibly over different (variable), as compared with fixed stimulus exemplars. They also show that dopaminergic and serotonergic chemomodulation has distinctive effects on such performance.SIGNIFICANCE STATEMENT This investigation employing local intracerebral infusions to inactivate the lateral prefrontal cortex (PFC) of the New World marmoset reveals the important role of this region in self-ordered response sequencing in variable but not fixed spatial arrays. These novel findings emphasize the higher order functions of this region, contributing to cognitive flexibility and planning of goal directed behavior. The investigation also reports for the first time somewhat contrasting neuromodulatory deficits produced by infusions of dopamine (DA) D2 and 5-HT2A receptor (R) antagonists into the same region, of possible significance for understanding cognitive deficits produced by anti-psychotic drugs.

Targeting 5-HT2B Receptor Signaling Prevents Border Zone Expansion and Improves Microstructural Remodeling After Myocardial Infarction.

BACKGROUND: Myocardial infarction (MI) induces an intense injury response that ultimately generates a collagen-dominated scar. Although required to prevent ventricular rupture, the fibrotic process is often sustained in a manner detrimental to optimal recovery. Cardiac myofibroblasts are the cells tasked with depositing and remodeling collagen and are a prime target to limit the fibrotic process after MI. Serotonin 2B receptor (5-HT2B) signaling has been shown to be harmful in a variety of cardiopulmonary pathologies and could play an important role in mediating scar formation after MI. METHODS: We used 2 pharmacological antagonists to explore the effect of 5-HT2B inhibition on outcomes after MI and characterized the histological and microstructural changes involved in tissue remodeling. Inducible 5-HT2B ablation driven by Tcf21MCM and PostnMCM was used to evaluate resident cardiac fibroblast- and myofibroblast-specific contributions of 5-HT2B, respectively. RNA sequencing was used to motivate subsequent in vitro analyses to explore cardiac fibroblast phenotype. RESULTS: 5-HT2B antagonism preserved cardiac structure and function by facilitating a less fibrotic scar, indicated by decreased scar thickness and decreased border zone area. 5-HT2B antagonism resulted in collagen fiber redistribution to thinner collagen fibers that were more anisotropic, enhancing left ventricular contractility, whereas fibrotic tissue stiffness was decreased, limiting the hypertrophic response of uninjured cardiomyocytes. Using a tamoxifen-inducible Cre, we ablated 5-HT2B from Tcf21-lineage resident cardiac fibroblasts and saw similar improvements to the pharmacological approach. Tamoxifen-inducible Cre-mediated ablation of 5-HT2B after onset of injury in Postn-lineage myofibroblasts also improved cardiac outcomes. RNA sequencing and subsequent in vitro analyses corroborate a decrease in fibroblast proliferation, migration, and remodeling capabilities through alterations in Dnajb4 expression and Src phosphorylation. CONCLUSIONS: Together, our findings illustrate that 5-HT2B expression in either cardiac fibroblasts or activated myofibroblasts directly contributes to excessive scar formation, resulting in adverse remodeling and impaired cardiac function after MI.

Blockade of 5-HT2 receptors suppresses rate of torque development and motor unit discharge rate during rapid contractions.

Serotonin (5-HT) is a neuromodulator that is critical for regulating the excitability of spinal motoneurons and the generation of muscle torque. However, the role of 5-HT in modulating human motor unit activity during rapid contractions has yet to be assessed. Nine healthy participants (23.7 ± 2.2 yr) ingested 8 mg of the competitive 5-HT2 antagonist cyproheptadine in a double-blinded, placebo-controlled, repeated-measures experiment. Rapid dorsiflexion contractions were performed at 30%, 50%, and 70% of maximal voluntary contraction (MVC), where motor unit activity was assessed by high-density surface electromyographic decomposition. A second protocol was performed where a sustained, fatigue-inducing dorsiflexion contraction was completed before undertaking the same 30%, 50%, and 70% MVC rapid contractions and motor unit analysis. Motor unit discharge rate (P < 0.001) and rate of torque development (RTD; P = 0.019) for the unfatigued muscle were both significantly lower for the cyproheptadine condition. Following the fatigue inducing contraction, cyproheptadine reduced motor unit discharge rate (P < 0.001) and RTD (P = 0.024), whereas the effects of cyproheptadine on motor unit discharge rate and RTD increased with increasing contraction intensity. Overall, these results support the viewpoint that serotonergic effects in the central nervous system occur fast enough to regulate motor unit discharge rate during rapid powerful contractions.NEW & NOTEWORTHY We have shown that serotonin activity in the central nervous system plays a role in regulating human motor unit discharge rate during rapid contractions. Our findings support the viewpoint that serotonergic effects in the central nervous system are fast and are most prominent during contractions that are characterized by high motor unit discharge rates and large amounts of torque development.

Semisynthetic Transformations on (+)-Boldine Reveal a 5-HT2A/2CR Antagonist.

Aporphine alkaloids have shown affinity for serotonin receptors (5-HTRs), and there has been a recent upsurge of interest in aporphines as 5-HT2CR ligands. 1,2,9,10-Tetraoxygenated aporphine alkaloids in particular have demonstrated good affinity for 5-HTRs. In continued efforts to understand the impacts of structural modification of the 1,2,9,10-tetraoxygenated aporphine template on affinity, selectivity, and activity at 5-HT2R subtypes, we used (+)-boldine (8) as a semisynthetic feedstock in the preparation of C-2-alkoxylated (+)-predicentrine analogues. Compound 10n, which contains a benzyloxy group at C-2, has been identified as a novel 5-HT2CR ligand with strong affinity (4 nM) and moderate selectivity versus 5-HT2BR and 5-HT2AR (12-fold and 6-fold, respectively). Compound 10n functions as an antagonist at 5-HT2A and 5-HT2C receptors. Computational experiments indicate that several hydrophobic interactions as well as strong H-bond and salt bridge interactions between the protonated amine moiety in 10n and Asp134 are responsible for the potent 5-HT2CR affinity of this compound. Furthermore, compound 10n displays favorable predicted drug-like characteristics, which is encouraging toward future optimization.

Eupnea and gasping in vivo are facilitated by the activation of 5-HT2A receptors.

Eupnea and gasping in infancy depend on central nervous system (CNS) serotonin (5-hydroxytryptamine; 5-HT). Although previous in vitro preparations have provided some evidence that 5-HT acts through type 2 A receptors (5-HT2A) to facilitate eupnea and gasping, here the hypothesis addressed is that 5-HT2A receptor activation is necessary for eupnea and the proper generation of gasping in vivo. To test this, we administered 2,5-dimethoxy-4-iodoamphetamine (DOI; 0.25 mg/kg i.p.), a 5-HT2A agonist, 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT; 0.25 mg/kg i.p.), a 5-HT1A agonist, or vehicle (saline) to 7-9-day-old tryptophan hydroxylase 2 knockout (TPH2-/-) mice. A second experiment assessed the effect of MDL-11,939 (MDL; 10 mg/kg i.p.), the specific 5-HT2A antagonist, or vehicle (DMSO) on the gasping of wild-type (TPH2+/+) animals. Drugs were given 15 min prior to five episodes of severe hypoxia that elicited gasping. TPH2-/- breathed more slowly but had the same V̇e and V̇e/V̇o2 compared with TPH2+/+. As previously reported, the gasping of TPH2-/- was significantly delayed (P < 0.001) and occurred at a significantly lower frequency compared with TPH2+/+ (P = 0.04). For both genotypes, DOI hastened eupneic frequency but had no effect on V̇e or V̇e/V̇o2. The gasping of TPH2-/-, although unaffected by 8-OH-DPAT, was indistinguishable from the gasping of TPH2+/+ following DOI. In TPH2+/+, application of MDL led to hypoventilation (P = 0.01), a delay in the appearance of gasping (P = 0.005), and reduced gasp frequency (P = 0.05). These data show that, in vivo, 5-HT2A receptors facilitate both eupnea and gasping. As has been shown in vitro, 5-HT2A probably promotes gasping by exciting hypoxia-resistant pacemaker neurons.NEW & NOTEWORTHY Previous in vitro studies suggest that 5-HT2A receptors contribute to eupnea and are necessary for fictive gasping. The current study shows that the impaired gasping displayed by neonatal TPH2-/- mice, deficient in CNS serotonin, is restored by 5-HT2A receptor activation. Following 5-HT2A blockade, wild-type mice hypoventilated and their gasping resembled that of TPH2-/- mice. This study shows that both eupnea and gasping in vivo rely on the activation of 5-HT2A receptors.

Human corticospinal-motoneuronal output is reduced with 5-HT2 receptor antagonism.

Animal models indicate that serotonin (5-HT) release onto motoneurons facilitates motor output, particularly during strong motor activities. However, evidence for 5-HT effects during human movement are limited. This study examined how antagonism of the 5-HT2 receptor, which is a 5-HT receptor that promotes motoneuron excitability, affects human movement. Ten healthy participants (24.2 ± 1.9 yr) ingested 8 mg of cyproheptadine (competitive 5-HT2 antagonist) in a double-blinded, placebo-controlled, repeated-measures design. Transcranial magnetic stimulation (TMS) of the motor cortex was used to elicit motor evoked potentials (MEPs) from biceps brachii. First, stimulus-response curves (90%-160% active motor threshold) were obtained during very weak elbow flexions (10% of maximal). Second, to determine if 5-HT effects are scaled to the intensity of muscle contraction, TMS at a fixed intensity was applied during elbow flexions of 20%, 40%, 60%, 80%, and 100% of maximal. Cyproheptadine reduced the size of MEPs across the stimulus-response curves (P = 0.045). Notably, MEP amplitude was 22.3% smaller for the cyproheptadine condition for the strongest TMS intensity. In addition, cyproheptadine reduced maximal torque (P = 0.045), lengthened the biceps silent period during maximal elbow flexions (P = 0.037), and reduced superimposed twitch amplitude during moderate-intensity elbow flexions (P = 0.035). This study presents novel evidence that 5-HT2 receptors influence corticospinal-motoneuronal output, which was particularly evident when a large number of descending inputs to motoneurons were active. Although it is likely that antagonism of 5-HT2 receptors reduces motoneuron gain to ionotropic inputs, supraspinal mechanisms may have also contributed to the study findings.NEW & NOTEWORTHY Voluntary contractions and responses to magnetic stimulation of the motor cortex are dependent on serotonin activity in the central nervous system. 5-HT2 antagonism decreased evoked potential size to high-intensity stimulation, and reduced torque and lengthened inhibitory silent periods during maximal contractions. We provide novel evidence that 5-HT2 receptors are involved in muscle activation, where 5-HT effects are strongest when a large number of descending inputs activate motoneurons.

Pimavanserin, a 5HT2A receptor inverse agonist, rapidly suppresses Aß production and related pathology in a mouse model of Alzheimer's disease.

Amyloid-ß (Aß) peptide aggregation into soluble oligomers and insoluble plaques is a precipitating event in the pathogenesis of Alzheimer's disease (AD). Given that synaptic activity can regulate Aß generation, we postulated that 5HT2A -Rs may regulate Aß as well. We treated APP/PS1 transgenic mice with the selective 5HT2A inverse agonists M100907 or Pimavanserin systemically and measured brain interstitial fluid (ISF) Aß levels in real-time using in vivo microdialysis. Both compounds reduced ISF Aß levels by almost 50% within hours, but had no effect on Aß levels in 5HT2A -R knock-out mice. The Aß-lowering effects of Pimavanserin were blocked by extracellular-regulated kinase (ERK) and NMDA receptor inhibitors. Chronic administration of Pimavanserin by subcutaneous osmotic pump to aged APP/PS1 mice significantly reduced CSF Aß levels and Aß pathology and improved cognitive function in these mice. Pimavanserin is FDA-approved to treat Parkinson's disease psychosis, and also has been shown to reduce psychosis in a variety of other dementia subtypes including Alzheimer's disease. These data demonstrate that Pimavanserin may have disease-modifying benefits in addition to its efficacy against neuropsychiatric symptoms of Alzheimer's disease. Read the Editorial Highlight for this article on page 560.

A Complex Impact of Systemically Administered 5-HT2A Receptor Ligands on Conditioned Fear.

BACKGROUND: Though drugs binding to serotonergic 5-HT2A receptors have long been claimed to influence human anxiety, it remains unclear if this receptor subtype is best described as anxiety promoting or anxiety dampening. Whereas conditioned fear expressed as freezing in rats is modified by application of 5-HT2A-acting drugs locally into different brain regions, reports on the effect of systemic administration of 5-HT2A receptor agonists and 5-HT2A antagonists or inverse agonists on this behavior remain sparse. METHODS: We assessed the possible impact of systemic administration of 5-HT2A receptor agonists, 5-HT2A receptor inverse agonists, and a selective serotonin reuptake inhibitor (SSRI)-per se or in combination-on the freezing displayed by male rats when re-exposed to a conditioning chamber in which they received foot shocks 7 days earlier. RESULTS: The 5-HT2A receptor agonists psilocybin and 25CN-NBOH induced a reduction in conditioned fear that was countered by pretreatment with 5-HT2A receptor inverse agonist MDL 100907. While both MDL 100907 and another 5-HT2A receptor inverse agonist, pimavanserin, failed to impact freezing per se, both compounds unmasked a robust fear-reducing effect of an SSRI, escitalopram, which by itself exerted no such effect. CONCLUSIONS: The results indicate that 5-HT2A receptor activation is not a prerequisite for normal conditioned freezing in rats but that this receptor subtype, when selectively over-activated prior to expression, exerts a marked fear-reducing influence. However, in the presence of an SSRI, the 5-HT2A receptor, on the contrary, appears to counter an anti-freezing effect of the enhanced extracellular serotonin levels following reuptake inhibition.

Low Basicity as a Characteristic for Atypical Ligands of Serotonin Receptor 5-HT2.

Serotonin receptors are extensively examined by academic and industrial researchers, due to their vital roles, which they play in the organism and constituting therefore important drug targets. Up to very recently, it was assumed that the basic nitrogen in compound structure is a necessary component to make it active within this receptor system. Such nitrogen interacts in its protonated form with the aspartic acid from the third transmembrane helix (D3x32) forming a hydrogen bond tightly fitting the ligand in the protein binding site. However, there are several recent studies that report strong serotonin receptor affinity also for compounds without a basic moiety in their structures. In the study, we carried out a comprehensive in silico analysis of the low-basicity phenomenon of the selected serotonin receptor ligands. We focused on the crystallized representatives of the proteins of 5-HT1B, 5-HT2A, 5-HT2B, and 5-HT2C receptors, and examined the problem both from the ligand- and structure-based perspectives. The study was performed for the native proteins, and for D3x32A mutants. The investigation resulted in the determination of nonstandard structural requirements for activity towards serotonin receptors, which can be used in the design of new nonbasic ligands.

Overcoming Depression with 5-HT2A Receptor Ligands.

Depression is a multifactorial disorder that affects millions of people worldwide, and none of the currently available therapeutics can completely cure it. Thus, there is a need for developing novel, potent, and safer agents. Recent medicinal chemistry findings on the structure and function of the serotonin 2A (5-HT2A) receptor facilitated design and discovery of novel compounds with antidepressant action. Eligible papers highlighting the importance of 5-HT2A receptors in the pathomechanism of the disorder were identified in the content-screening performed on the popular databases (PubMed, Google Scholar). Articles were critically assessed based on their titles and abstracts. The most accurate papers were chosen to be read and presented in the manuscript. The review summarizes current knowledge on the applicability of 5-HT2A receptor signaling modulators in the treatment of depression. It provides an insight into the structural and physiological features of this receptor. Moreover, it presents an overview of recently conducted virtual screening campaigns aiming to identify novel, potent 5-HT2A receptor ligands and additional data on currently synthesized ligands acting through this protein.

Unusual Quadrupedal Locomotion in Rat during Recovery from Lumbar Spinal Blockade of 5-HT7 Receptors.

Coordination of four-limb movements during quadrupedal locomotion is controlled by supraspinal monoaminergic descending pathways, among which serotoninergic ones play a crucial role. Here we investigated the locomotor pattern during recovery from blockade of 5-HT7 or 5-HT2A receptors after intrathecal application of SB269970 or cyproheptadine in adult rats with chronic intrathecal cannula implanted in the lumbar spinal cord. The interlimb coordination was investigated based on electromyographic activity recorded from selected fore- and hindlimb muscles during rat locomotion on a treadmill. In the time of recovery after hindlimb transient paralysis, we noticed a presence of an unusual pattern of quadrupedal locomotion characterized by a doubling of forelimb stepping in relation to unaffected hindlimb stepping (2FL-1HL) after blockade of 5-HT7 receptors but not after blockade of 5-HT2A receptors. The 2FL-1HL pattern, although transient, was observed as a stable form of fore-hindlimb coupling during quadrupedal locomotion. We suggest that modulation of the 5-HT7 receptors on interneurons located in lamina VII with ascending projections to the forelimb spinal network can be responsible for the 2FL-1HL locomotor pattern. In support, our immunohistochemical analysis of the lumbar spinal cord demonstrated the presence of the 5-HT7 immunoreactive cells in the lamina VII, which were rarely 5-HT2A immunoreactive.

Risperidone and 5-HT2A Receptor Antagonists Attenuate and Reverse Cocaine-Induced Hyperthermia in Rats.

BACKGROUND: Cocaine (benzoylmethylecgonine) is one of the most widely used illegal psychostimulant drugs worldwide, and mortality from acute intoxication is increasing. Suppressing hyperthermia is effective in reducing cocaine-related mortality, but a definitive therapy has not yet been found. In this study, we assessed the ability of risperidone to attenuate acute cocaine-induced hyperthermia and delineated the mechanism of its action. METHODS: Rats were injected i.p. with saline, risperidone, ketanserin, ritanserin, haloperidol, or SCH 23 390 before and after injection of cocaine (30 mg/kg) or with WAY-00 635, SB 206 553, or sulpiride before cocaine injection; thereafter, the rectal temperature was measured every 30 minutes for up to 4 hours. In vivo microdialysis was used to reveal the effect of risperidone on cocaine-induced elevation of dopamine (DA), serotonin (5-HT), and noradrenaline concentrations in the anterior hypothalamus. For post-administration experiments, saline or risperidone (0.5 mg/kg) were injected into rats, and cocaine (30 mg/kg) was injected 15 minutes later. For every 30 minutes thereafter, DA, 5-HT, and noradrenaline levels were measured for up to 240 minutes after cocaine administration. RESULTS: Risperidone, 5-HT2A receptor antagonists, and D1 receptor antagonistic drugs prevented and reversed cocaine-induced hyperthermia. In contrast, receptor antagonists for 5-HT1A, 5-HT2B/2C, and D2 did not alter cocaine-induced hyperthermia. Risperidone treatment further attenuated cocaine-induced elevation of DA. CONCLUSIONS: Our results indicate that risperidone attenuates cocaine-induced hyperthermia primarily by blocking the activities of the 5-HT2A and D1 receptors and may be potentially useful for treating cocaine-induced acute hyperthermia in humans.

Synthesis and biological evaluation of thioadatanserin and its dialkylated products as partial 5-HTR1A agonists and 5-HTR2A antagonists for potential use in depression and anxiety disorders.

Thionation of adatanserin hydrochloride (2) with Lawesson's reagent in toluene/triethylamine afforded novel compound, (3r,5r,7r)-N-(2-(4-(pyrimidin-2-yl)piperazin-1-yl)ethyl)adamantane-1-carbothioamide (thioadatanserin, 3) in 84-90% isolated yield. Thioadatanserin underwent a tandem double alkylation with methyl iodide and benzyl bromide in NaH/THF to produce novel dialkylated products 6 and 7 respectively. The single X-ray crystal structure of 7 was determined to be 1-(2-((E- ((3r,5r,7r)-adamantan-1-yl)benzylthio)methylene)amino)ethyl)-1-benzyl-4- (pyrimidin-2-yl)piperazin-1-ium bromide showing that the piperazine ring adopts a chair-like configuration that is not co-planar with the pyrimidine ring. Thioadatanserin emerged as a dual potent partial agonist with activity against 5-HTR1A (EC50 6.7 nM) and antagonist activity against 5-HTR2A (IC50 62.3 nM) and was selective over 5-HTR2C receptor (IC50 > 3333 nM) in the PathHunter® ß-arrestin assays.

Evaluation of anti-depressant effects of phthalazinone-based triple-acting small molecules against 5-HT2A, 5-HT2C, and the serotonin transporter.

Development of highly effective, safe, and fast-acting anti-depressants is urgently required for the treatment of major depressive disorder. It has been suggested that targeting 5-HT2A and 5-HT2C in addition to inhibition of serotonin reuptake may be beneficial in generating anti-depressant agents with better pharmacology and less adverse effects. We have developed phthalazinone-based compounds that potently bind to 5-HT2A, 5-HT2C, and the serotonin transporter. The representative compounds 11j and 11l displayed strong binding affinities against these targets, and showed favorable toxicity profiles as determined by hERG binding and CYP inhibition assays. Furthermore, these compounds presented promising anti-depressant effects comparable to fluoxetine and also synergistic effects with fluoxetine in forced swimming test, which implicates these compounds can be developed to help the treatment of major depressive disorder.

Ritanserin blocks CaV1.2 channels in rat artery smooth muscles: electrophysiological, functional, and computational studies.

CaV1.2 channel blockers or 5-HT2 receptor antagonists constitute effective therapy for Raynaud's syndrome. A functional link between the inhibition of 5-HT2 receptors and CaV1.2 channel blockade in arterial smooth muscles has been hypothesized. Therefore, the effects of ritanserin, a nonselective 5-HT2 receptor antagonist, on vascular CaV1.2 channels were investigated through electrophysiological, functional, and computational studies. Ritanserin blocked CaV1.2 channel currents (ICa1.2) in a concentration-dependent manner (Kr = 3.61 µM); ICa1.2 inhibition was antagonized by Bay K 8644 and partially reverted upon washout. Conversely, the ritanserin analog ketanserin (100 µM) inhibited ICa1.2 by ~50%. Ritanserin concentration-dependently shifted the voltage dependence of the steady-state inactivation curve to more negative potentials (Ki = 1.58 µM) without affecting the slope of inactivation and the activation curve, and decreased ICa1.2 progressively during repetitive (1 Hz) step depolarizations (use-dependent block). The addition of ritanserin caused the contraction of single myocytes not yet dialyzed with the conventional method. Furthermore, in depolarized rings, ritanserin, and to a lesser extent, ketanserin, caused a concentration-dependent relaxation, which was antagonized by Bay K 8644. Ritanserin and ketanserin were docked at a region of the CaV1.2 α1C subunit nearby that of Bay K 8644; however, only ritanserin and Bay K 8644 formed a hydrogen bond with key residue Tyr-1489. In conclusion, ritanserin caused in vitro vasodilation, accomplished through the blockade of CaV1.2 channels, which was achieved preferentially in the inactivated and/or resting state of the channel. This novel activity encourages the development of ritanserin derivatives for their potential use in the treatment of Raynaud's syndrome.