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Glaucoma is a degenerative optic neuropathy in which the degeneration of optic nerve and blindness occur. The main cause is a malfunction of ciliary processes (protrusions of the ciliary bodies) resulting in increased intraocular pressure (IOP). Ocular hypertension (OHT) causes ischemic events leading to retinal ganglion cell (RGC) depletion and blindness. Histaminergic and nitrergic systems are involved in the regulation of IOP. Therefore, we developed novel hybrid compounds that target histamine H 3 receptor (H 3 R) with nitric oxide (NO) releasing features (ST-1989 and ST-2130). After H 3 R binding was proven in vitro, we investigated their effects in two OHT models in New Zealand White rabbits. Compound ST-1989 showed the highest NO elevation, together with antioxidative and anti-inflammatory features partly superior to the co-administered H 3 R antagonist (ciproxifan) and NO donor (molsidomine). This hybrid compound demonstrated IOP reduction in both OHT models induced by intravitreal injection of hypertonic saline and carbomer into the anterior chamber of the eye, respectively. Ocular perfusion and photoreceptor neuroprotection were evaluated in a model of ischemia/reperfusion (I/R) of the ophthalmic artery induced by repeated sub-tenon injections of endothelin-1 (ET-1), twice a week for six weeks. Compound ST-1989 counteracts retinal degeneration reducing ophthalmic artery resistance index and increasing photoreceptor responses, thus rescuing RGCs. Our results indicate that compound ST-1989 is a promising molecule with long-lasting hypotensive effects and good effectiveness in reducing inflammation, oxidative stress, and RGCs apoptosis. In conclusion, these hybrid compounds could be a novel strategy to combat glaucomatous blindness and RGC depletion for ocular diseases involving retinal damage., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Mohammad A. Khanfar reports financial support was provided by Alexander von Humboldt Foundation. Holger Stark reports financial support was provided by German Research Council. Laura Lucarini reports financial support was provided by Fondazione Cassa di Risparmio di Firenze. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Conditioned taste aversion (CTA) is a robust associative learning; liquid deprivation during this conditioning allows researchers to obtain readable measures of associative learning. Recent research suggests that thirst could be a crucial motivator that modulates conditioning and memory extinction processes, highlighting the importance of the body's internal state during learning. Furthermore, the histaminergic system is one of the major modulatory systems controlling several behavioral and neurobiological functions, such as feeding, water intake, and nociception. Therefore, this research aimed to assess the effect of H3 histaminergic receptor activation in the insular cortex (IC) during CTA. For this, we conditioned adult male Wistar rats under two regimens: water deprivation and water ad libitum. A classical CTA protocol was used for water deprivation. Before CTA acquisition, 10 μM R-α-methylhistamine (RAMH), an H3 receptor agonist, was injected into the IC. Results showed that RAMH injections decreased CTA in water-deprived rats without affecting the significant aversion conditioning in rats that were given water ad libitum. Moreover, RAMH accelerated the process of aversive memory extinction under ad libitum water conditions. According to our findings, the degree of liquid satiety differentially affected taste-aversive memory formation, and H3 histamine receptors were more involved under water deprivation conditions during acquisition. However, these receptors modulated the strength of aversive conditioning by altering the rate of aversive memory extinction in the absence of deprivation. In conclusion, histaminergic activity in the IC may influence taste memory dynamics through different mechanisms depending on the degree of liquid satiety or deprivation during conditioning., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 International Brain Research Organization (IBRO). Published by Elsevier Inc. All rights reserved.)

Rationale: The rewarding effect of Methamphetamine (METH) is commonly believed to play an important role in METH use disorder. The altered expression of dopamine D1 receptor (D1R) has been suggested to be essential to the rewarding effect of METH. Notably, D1R could interact with histamine H3 receptors (H3R) by forming a H3R-D1R heteromer (H3R-D1R)., Objectives: This study was designed to specifically investigate the involvement of H3R-D1R in the rewarding effect of METH., Methods: C57BL/6 mice were treated with intraperitoneal injections of a selective H3R antagonist (Thioperamide, THIO; 20 mg/kg), an H1R antagonist (Pyrilamine, PYRI; 10 mg/kg), or microinjections of cytomegalovirus (CMV)-transmembrane domain 5 (TM5) into the nucleus accumbens (NAc). The animal model of Conditioned Place Preference (CPP) was applied to determine the impact of H3R-D1R on the rewarding effect of METH., Results: METH resulted in a significant preference for the drug-associated chamber, in conjunction with increased H3R and decreased D1R expression in both NAc and the ventral tegmental area (VTA). THIO significantly attenuated the rewarding effect of METH, accompanied by decreased H3R and increased D1R expression. In contrast, pyrilamine failed to produce the similar effects. Moreover, the inhibitory effect of THIO on METH-induced CPP was reversed by SKF38393, a D1R agonist. Furthermore, SCH23390, a D1R antagonist, counteracted the ameliorative effect of SKF38393 on THIO. Co-immunoprecipitation (CO-IP) experiments further demonstrated the specific interaction between H3R and D1R in METH CPP mice. The rewarding effect of METH was also significantly blocked by the interruption of CMV-transmembrane domain 5 (TM5), but not CMV-transmembrane domain 7 (TM7) in NAc., Conclusion: These results suggest that modulating the activity of H3R-D1R complex holds promise for regulating METH use disorder and serves as a potential drug target for its treatment., Competing Interests: Declaration of competing interest We declare that we have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The histamine H 3 receptor (H 3 R) is a neurotransmitter receptor that is primarily found in the brain, where it controls the release and synthesis of histamine, as well as the release of other neurotransmitters (e.g. dopamine, serotonin). Notably, 20 H 3 R isoforms are differentially expressed in the human brain as a consequence of alternative gene splicing. The hH 3 R-445, -415, -365 and -329 isoforms contain the prototypical GPCR (7TM) structure, yet exhibit deletions in the third intracellular loop, a structural domain that is pivotal for G protein-coupling, signaling and regulation. To date, the physiological relevance underlying the individual and combinatorial function of hH 3 R isoforms remains poorly understood. Nevertheless, given their significant implication in physiological processes (e.g. cognition, homeostasis) and neurological disorders (e.g. Alzheimer's and Parkinson's disease, schizophrenia), widespread targeting of hH 3 R isoforms by drugs may lead to on-target side effects in brain regions that are unaffected by disease. To this end, isoform- and/or pathway-selective targeting of hH 3 R isoforms by biased agonists could be of therapeutic relevance for the development of region- and disease-specific drugs. Hence, we have evaluated ligand biased signaling at the hH 3 R-445, -415, -365 and -329 isoforms across various Gα i/o -mediated (i.e. [ 35 S]GTPγS accumulation, cAMP inhibition, pERK1/2 activation, pAKT T308/S473 activation) and non Gα i/o -mediated (i.e. β-arrestin2 recruitment) endpoints that are relevant to neurological diseases. Our findings indicate that H 3 R agonists display significantly altered patterns in their degree of ligand bias, in a pathway- and isoform-dependent manner, underlining the significance to investigate GPCRs with multiple isoforms to improve development of selective drugs. SUBJECT CATEGORY: Neuropharmacology., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

A series of scutellarein 7- l -amino acid carbamate-4'-cycloalkylamine propyl ether conjugates were designed and synthesized for the first time as multifunctional agents for Alzheimer's disease (AD) therapy. The designed compounds exhibited more balanced and effective multi-target potency. Among them, compound 11l, l -Valine carbamate derivative of scutellarein cycloheptylamine ether, exhibited the most potent inhibition of electric eel AChE enzymes and human AChE enzymes, with an IC 50 values of 7.04 μM and 9.73 μM, respectively. Moreover, 11l exhibited more potent H 3 R antagonistic activities than clobenpropit, with an IC 50 value of 1.09 nM. Compound 11l not only displayed excellent inhibition of self- and Cu 2+ -induced Aβ 1-42 aggregation (95.48 % and 88.63 % inhibition, respectively) but also induced the disassembly of self- and Cu 2+ -induced Aβ fibrils (80.16 % and 89.30 % disaggregation, respectively). Moreover, 11l significantly reduced tau protein hyperphosphorylation induced by Aβ 25-35 . It exhibited effective antioxidant activity and neuroprotective potency, and inhibited RSL3-induced PC12 cell ferroptosis. Assays of hCMEC/D3 and hPepT1-MDCK cell line permeability indicated that 11l would have optimal blood-brain barrier permeability and intestinal absorption characteristics. In addition, in vivo studies revealed that compound 11l significantly attenuated learning and memory impairment in an AD mouse model. Finally, a pharmacokinetic characterization of 11l indicated favorable druggability and pharmacokinetic properties. Taken together, our results suggest that 11l is a potential candidate for AD treatment and merits further investigation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Many neuropsychiatric disorders can be caused by neurotransmitter dysfunction. Experimental studies have demonstrated that histamine and the harmaline affect physiological processes through interaction with other neurotransmitter systems. The objective of these experiments was to investigate the involvement of the histaminergic system in the effects of harmaline on anxiety- and depressive-related effects in male NMRI mice. Behavioral tests were employed to evaluate anxiety-related symptoms (elevated plus maze; EPM), depressive-like symptoms (forced swim test; FST), and cognitive decline (step-down test). The histamine H3 receptor (H3R) agonist α-methylhistamine dihydrobromide (α-MH; 5 mg/kg, i.p.) had anxiolytic- and depressive-like effects, while the H3R antagonist thioperamide (10 mg/kg, i.p.) showed an antidepressive-like property. The subthreshold dose of α-MH resulted in an increase in the tendency of mice treated with the harmaline (2.5 mg/kg) to remain in the EPM open-arms. A subthreshold dose of thioperamide (5 mg/kg) increased the time spent in the open-arms in mice treated with harmaline (2.5 and 5 mg/kg) while a high dose of harmaline decreased the immobility time. Furthermore, two higher doses of harmaline resulted in a reduction in the number of open-arm entries. Similarly, mice administered with thioperamide and a low dose of harmaline decreased locomotor activity in the EPM. Ultimately, the combined thioperamide and harmaline did not impair memory retrieval of mice. These experiments demonstrate that the histaminergic system is implicated in the anxiety- and depressive-related effects of harmaline. The combination of thioperamide and harmaline is effective in treating anxiety and depression without having an adverse effect on memory formation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Prenatal alcohol exposure can have persistent effects on learning, memory, and synaptic plasticity. Previous work from our group demonstrated deficits in long-term potentiation (LTP) of excitatory synapses on dentate gyrus granule cells in adult offspring of rat dams that consumed moderate levels of alcohol during pregnancy. At present, there are no pharmacotherapeutic agents approved for these deficits. Prior work established that systemic administration of the histaminergic H3R inverse agonist ABT-239 reversed deficits in LTP observed following moderate PAE. The present study examines the effect of a second H3R inverse agonist, SAR-152954, on LTP deficits following moderate PAE. We demonstrate that systemic administration of 1 mg/kg of SAR-152954 reverses deficits in potentiation of field excitatory post-synaptic potentials (fEPSPs) in adult male rats exposed to moderate PAE. Time-frequency analyses of evoked responses revealed PAE-related reductions in power during the fEPSP, and increased power during later components of evoked responses which are associated with feedback circuitry that are typically not assessed with traditional amplitude-based measures. Both effects were reversed by SAR-152954. These findings provide further evidence that H3R inverse agonism is a potential therapeutic strategy to address deficits in synaptic plasticity associated with PAE., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The future of therapy for neurodegenerative diseases (NDs) relies on new strategies targeting multiple pharmacological pathways. Our research led to obtaining the compound AR71 [(E)-3-(3,4,5-trimethoxyphenyl)-1-(4-(3-(piperidin-1-yl)propoxy)phenyl)prop-2-en-1-one], which has high affinity for human H 3 R (K i = 24 nM) and selectivity towards histamine H 1 and H 4 receptors (K i > 2500 nM), and showed anti-inflammatory activity in a model of lipopolysaccharide-induced inflammation in BV-2 cells. The presented tests confirmed its antagonist/inverse agonist activity profile and good metabolic stability while docking studies showed the binding mode to histamine H 1 , H 3 , and H 4 receptors. In in vitro tests, cytotoxicity was evaluated at three cell lines (neuroblastoma, astrocytes, and human peripheral blood mononuclear cells), and a neuroprotective effect was observed in rotenone-induced toxicity. In vivo experiments in a mouse neuropathic pain model demonstrated the highest analgesic effects of AR71 at the dose of 20 mg/kg body weight. Additionally, AR71 showed antiproliferative activity in higher concentrations. These findings suggest the need for further evaluation of AR71's therapeutic potential in treating ND and CNS cancer using animal experimental models.

Alternative splicing significantly enhances the diversity of the G protein-coupled receptor (GPCR) family, including the histamine H 3 receptor (H 3 R). This post-transcriptional modification generates multiple H 3 R isoforms with potentially distinct pharmacological and physiological profiles. H 3 R is primarily involved in the presynaptic inhibition of neurotransmitter release in the central nervous system. Despite the approval of pitolisant for narcolepsy (Wakix ® ) and daytime sleepiness in adults with obstructive sleep apnea (Ozawade ® ) and ongoing clinical trials for other H 3 R antagonists/inverse agonists, the functional significance of the numerous H 3 R isoforms remains largely enigmatic. Recent publicly available RNA sequencing data have confirmed the expression of multiple H 3 R isoforms in the brain, with some isoforms exhibiting unique tissue-specific distribution patterns hinting at isoform-specific functions and interactions within neural circuits. In this review, we discuss the complexity of H 3 R isoforms with a focus on their potential roles in central nervous system (CNS) function. Comparative analysis across species highlights evolutionary conservation and divergence in H 3 R splicing, suggesting species-specific regulatory mechanisms. Understanding the functionality of H 3 R isoforms is crucial for the development of targeted therapeutics. This knowledge will inform the design of more precise pharmacological interventions, potentially enhancing therapeutic efficacy and reducing adverse effects in the treatment of neurological and psychiatric disorders.

G-protein-coupled receptors (GPCRs) transmit downstream signals predominantly via G-protein pathways. However, the conformational basis of selective coupling of primary G-protein remains elusive. Histamine receptors H 2 R and H 3 R couple with G s - or G i -proteins respectively. Here, three cryo-EM structures of H 2 R-G s and H 3 R-G i complexes are presented at a global resolution of 2.6-2.7 Å. These structures reveal the unique binding pose for endogenous histamine in H 3 R, wherein the amino group interacts with E206 5.46 of H 3 R instead of the conserved D114 3.32 of other aminergic receptors. Furthermore, comparative analysis of the H 2 R-G s and H 3 R-G i complexes reveals that the structural geometry of TM5/TM6 determines the primary G-protein selectivity in histamine receptors. Machine learning (ML)-based structuromic profiling and functional analysis of class A GPCR-G-protein complexes illustrate that TM5 length, TM5 tilt, and TM6 outward movement are key determinants of the G s and G i/o selectivity among the whole Class A family. Collectively, the findings uncover the common structural geometry within class A GPCRs that determines the primary G s - and G i/o -coupling selectivity., (© 2024 The Authors. Advanced Science published by Wiley‐VCH GmbH.)

The aims of this work were to evaluate the expression of histamine H 3 receptor (H 3 R) in triple negative breast cancer (TNBC) samples and to investigate the antitumoral efficacy and safety of the LINS01 series of H 3 R antagonists, through in silico, in vitro, and in vivo approaches. Antitumor activity of LINS01009, LINS01010, LINS01022, LINS01023 was assayed in vitro in 4T1 and MDA-MB-231 TNBC cells (0.01-100 μM), and in vivo in 4T1 tumors orthotopically established in BALB/c mice (1 or 20 mg/kg). Additionally, H 3 R expression was assessed in 50 human TNBC samples. We have described a higher H 3 R mRNA expression in basal-like/TNBC tumors vs. matched normal tissue using TCGA Pan-Cancer Atlas data, and a higher H 3 R expression in human tumor samples vs. peritumoral tissue evidenced by immunohistochemistry associated with poorer survival. Furthermore, while all the essayed compounds showed antitumoral properties, LINS01022 and LINS01023 exhibited the most potent antiproliferative effects by: i) inducing cell apoptosis and suppressing cell migration in 4T1 and MDA-MB-231 TNBC cells, and ii) inhibiting cell growth in paclitaxel-resistant 4T1 cells (potentiating the paclitaxel antiproliferative effect). Moreover, 20 mg/kg LINS01022 reduced tumor size in 4T1 tumor-bearing mice, exhibiting a safe toxicological profile and potential for druggability estimated by ADME calculations. We conclude that the H 3 R is involved in the regulation of TNBC progression, offering promising therapeutic potential for the novel LINS01 series of H 3 R antagonists., Competing Interests: Declaration of Competing Interest All authors declare no potential competing interests., (Copyright © 2024. Published by Elsevier Masson SAS.)

The histamine H 3 receptor (H 3 R) regulates as a presynaptic G protein-coupled receptor the release of histamine and other neurotransmitters in the brain, and is consequently a potential therapeutic target for neuronal disorders. The human H 3 R encodes for seven splice variants that vary in the length of intracellular loop 3 and/or the C-terminal tail but are all able to induce heterotrimeric G i protein signaling. The last two decades H 3 R drug discovery and lead optimization has been exclusively focused on the 445 amino acids-long reference isoform H 3 R-445. In this study, we pharmacologically characterized for the first time all seven H 3 R isoforms by determining their binding affinities for reference histamine H 3 receptor agonists and inverse agonists. The H 3 R-453, H 3 R-415, and H 3 R-413 isoforms display similar binding affinities for all ligands as the H 3 R-445. However, increased agonist binding affinities were observed for the three shorter isoforms H 3 R-329, H 3 R-365, and H 3 R-373, whereas inverse agonists such as the approved anti-narcolepsy drug pitolisant (Wakix®) displayed significantly decreased binding affinities for the latter two isoforms. This opposite change in binding affinity of agonist versus inverse agonists on H 3 R-365 and H 3 R-373 is associated with their higher constitutive activity in a cAMP biosensor assay as compared to the other five isoforms. The observed differences in pharmacology between longer and shorter H 3 R isoforms should be considered in future drug discovery programs., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Meichun Gao reports financial support was provided by CSC Chinese scholarship grant. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Steroid-based histamine H 3 receptor antagonists (d-homoazasteroids) were designed by combining distinct structural elements of HTS hit molecules. They were characterized, and several of them displayed remarkably high affinity for H 3 receptors with antagonist/inverse agonist features. Especially, the 17a-aza-d-homolactam chemotype demonstrated excellent H 3 R activity together with significant in vivo H 3 antagonism. Optimization of the chemotype was initiated with special emphasis on the elimination of the hERG and muscarinic affinity. Additionally, ligand-based SAR considerations and molecular docking studies were performed to predict binding modes of the molecules. The most promising compounds ( XXI , XXVIII , and XX ) showed practically no muscarinic and hERG affinity. They showed antagonist/inverse agonist property in the in vitro functional tests that was apparent in the rat in vivo dipsogenia test. They were considerably stable in human and rat liver microsomes and provided significant in vivo potency in the place recognition and novel object recognition cognitive paradigms.

The histamine H 3 receptor, prominently expressed in neurons with a minor presence in glial cells, acts as both an autoreceptor and an alloreceptor, controlling the release of histamine and other neurotransmitters. The receptor impacts various essential physiological processes. Our team's initial investigations had demonstrated that the histamine H 3 receptor antagonists could facilitate nerve regeneration by promoting the histamine H 1 receptors on primary neural stem cells (NSCs) in the traumatic brain injury mouse, which suggested the potential of histamine H 3 receptor as a promising target for treating neurological disorders and promoting nerve regeneration. Pitolisant (PITO) is the only histamine H 3 receptor antagonist approved by the Food and Drug Administration (FDA) for treating narcolepsy. However, there is no report on Pitolisant in neural development or regeneration, and it is urgent to be further studied in strong biological activity models in vitro. The embryonic stem (ES) cells were differentiated into neural cells in vitro, which replicated the neurodevelopmental processes that occur in vivo. It also provided an alternative model for studying neurodevelopmental processes and testing drugs for neurological conditions. Therefore, we aimed to elucidate the regulatory role of Pitolisant in the early differentiation of ES cells into neural cells. Our results demonstrated that Pitolisant could promote the differentiation of ES cells toward NSCs and stimulated the formation of growth cones. Furthermore, Pitolisant was capable of inducing the polarization of NSCs through the cAMP-LKB1-SAD/MARK2 pathway, but had no significant effect on later neuronal maturation. Pitolisant altered mitochondrial morphology and upregulated the levels of mitochondrion-related proteins TOM20, Drp1, and p-Drp1, and reversed the inhibitory effect of Mdivi-1 on mitochondrial fission during the early neural differentiation of ES cells. In addition, Pitolisant induced the increase in cytosolic Ca 2+ . Our study provided an experimental foundation for the potential application of histamine H 3 receptor-targeted modulators in the field of neuroregeneration.

Background: The histamine-3 receptor (H3R) is an auto- and heteroreceptor that inhibits the release of histamine and other neurotransmitters. Post-mortem evidence has found altered H3R expression in patients with psychotic disorders, which may underlie cognitive impairment associated with schizophrenia (CIAS)., Aims: We used positron emission tomography (PET) imaging to compare brain uptake of an H3R selective tracer between patients with schizophrenia and matched controls (healthy individuals). Regions of interest included the dorsolateral prefrontal cortex (DLPFC) and striatum. We explored correlations between tracer uptake and symptoms, including cognitive domains., Methods: A total of 12 patients and 12 matched controls were recruited to the study and were assessed with psychiatric and cognitive rating scales. They received a PET scan using the H3R-specific radioligand [ 11 C]MK-8278 to determine H3R availability., Results: There was no statistically significant difference in tracer uptake between patients and controls in the DLPFC ( t 19  = 0.79, p  = 0.44) or striatum (t 21  = 1.18, p  = 0.25). An exploratory analysis found evidence for lower volume of distribution in the left cuneus (p FWE-corrected  = 0.01). DLPFC tracer uptake was strongly correlated with cognition in controls (trail making test (TMT) A: r  = 0.77, p  = 0.006; TMT B: rho = 0.74, p  = 0.01), but not in patients (TMT A: r  = -0.18, p  = 0.62; TMT B: rho = -0.06, p  = 0.81)., Conclusions: These findings indicate H3R in the DLPFC might play a role in executive function and this is disrupted in schizophrenia in the absence of major alterations in H3R availability as assessed using a selective radiotracer for H3R. This provides further evidence for the role of H3R in CIAS., Competing Interests: Declaration of conflicting interestsThe author(s) declared the following potential conflicts of interest with respect to the research, authorship, and/or publication of this article: AA, MMN, MV and ECO have no conflicting interests to declare. EAR is a full-time employee of Invicro, an imaging centre conducting contract studies for pharmaceutical and biotech partners. As part of his role, EAR has consulted for numerous industry and academic entities. ODH is a part-time employee and stock holder of Lundbeck A/s. He has received investigator-initiated research funding from and/or participated in advisory/speaker meetings organised by Angellini, Autifony, Biogen, Boehringer-Ingelheim, Eli Lilly, Heptares, Global Medical Education, Invicro, Jansenn, Lundbeck, Neurocrine, Otsuka, Sunovion, Recordati, Roche and Viatris/Mylan. ODH has a patent for the use of dopaminergic imaging.

G protein-coupled receptors (GPCRs) are the largest protein family in humans and are important drug targets. Yeast, especially Saccharomyces cerevisiae, is a useful host for modifying the function and stability of GPCRs through protein engineering, which is advantageous for mammalian cells. When GPCRs are expressed in yeast, their function is often impaired. In this study, we performed random mutagenesis using error-prone PCR and then an in vivo screening to obtain mutants that recovered the activity of the human histamine H 3 receptor (H 3 R), which loses its signaling function when expressed in yeast. Four mutations with recovered activity were identified after screening. Three of the mutations were identified near the DRY and NPxxY motifs of H 3 R, which are important for activation and are commonly found in class A GPCRs. The mutants responded exclusively to the yeast YB1 strain harboring G i -chimera proteins, showing retention of G protein specificity. Analysis of one of the mutants with recovered activity, C415R, revealed that it maintained its ligand-binding characteristics. The strategy used in this study may enable the recovery of the activity of other GPCRs that do not function in S. cerevisiae and may be useful in creating GPCRs mutants stabilized in their active conformations., (© 2023. The Author(s).)

H 3 R is becoming an attractive and promising target for epilepsy treatment as well as the discovery of antiepileptics. In this work, a series of 6-aminoalkoxy-3,4-dihydroquinolin-2(1 H )-ones was prepared to screen their H 3 R antagonistic activities and antiseizure effects. The majority of the target compounds displayed a potent H 3 R antagonistic activity. Among them, compounds 2a , 2c , 2h , and 4a showed submicromolar H 3 R antagonistic activity with an IC 50 of 0.52, 0.47, 0.12, and 0.37 μM, respectively. The maximal electroshock seizure (MES) model screened out three compounds ( 2h , 4a , and 4b ) with antiseizure activity. Meanwhile, the pentylenetetrazole (PTZ)-induced seizure test gave a result that no compound can resist the seizures induced by PTZ. Additionally, the anti-MES action of compound 4a fully vanished when it was administrated combined with an H 3 R agonist (RAMH). These results showed that the antiseizure role of compound 4a might be achieved by antagonizing the H 3 R receptor. The molecular docking of 2h , 4a , and PIT with the H 3 R protein predicted their possible binding patterns and gave a presentation that 2h , 4a , and PIT had a similar binding model with H 3 R.

The histamine H3 receptor (H3R) is highly enriched in the spiny projection neurons (SPNs) of the striatum, in both the D1 receptor (D1R)-expressing and D2 receptor (D2R)-expressing populations. A crossantagonistic interaction between H3R and D1R has been demonstrated in mice, both at the behavioral level and at the biochemical level. Although interactive behavioral effects have been described upon coactivation of H3R and D2R, the molecular mechanisms underlying this interaction are poorly understood. Here, we show that activation of H3R with the selective agonist R-(-)-α-methylhistamine dihydrobromide mitigates D2R agonist-induced locomotor activity and stereotypic behavior. Using biochemical approaches and the proximity ligation assay, we demonstrated the existence of an H3R-D2R complex in the mouse striatum. In addition, we examined consequences of simultaneous H3R-D2R agonism on the phosphorylation levels of several signaling molecules using immunohistochemistry. H3R agonist treatment modulated Akt (serine/threonine PKB)-glycogen synthase kinase 3 beta signaling in response to D2R activation via a β-arrestin 2-dependent mechanism in D2R-SPNs but not in D1R-SPNs. Phosphorylation of mitogen- and stress-activated protein kinase 1 and rpS6 (ribosomal protein S6) was largely unchanged under these conditions. As Akt-glycogen synthase kinase 3 beta signaling has been implicated in several neuropsychiatric disorders, this work may help clarify the role of H3R in modulating D2R function, leading to a better understanding of pathophysiology involving the interaction between histamine and dopamine systems., Competing Interests: Conflict of interest Dr Pittenger has served as a consultant and received research funding in the past year for Biohaven Pharmaceuticals, Transcend Therapeutics, Ceruvia Lifesciences, Freedom Biosciences, and Nobilis Therapeutics, and royalties from Oxford University Press, all for work unrelated to the current results. He is an inventor on a patent application related to the use of neurofeedback and of psychedelic drugs for the treatment of obsessive compulsive disorder, also unrelated to this work, Dr Xu reports no competing interests. The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

The human histamine H 3 receptor (hH 3 R) is predominantly expressed in the CNS, where it regulates the synthesis and release of histamine and other neurotransmitters. Due to its neuromodulatory role, the hH 3 R has been associated with various CNS disorders, including Alzheimer's and Parkinson's disease. Markedly, the hH 3 R gene undergoes extensive splicing, resulting in 20 isoforms, of which 7TM isoforms exhibit variations in the intracellular loop 3 (IL3) and/or C-terminal tail. Particularly, hH 3 R isoforms that display variations in IL3 (e.g., hH 3 R-365) are shown to differentially signal via Gα i -dependent pathways upon binding of biased agonists (e.g., immepip, proxifan, imetit). Nevertheless, the mechanisms underlying biased agonism at hH 3 R isoforms remain unknown. Using a structure-function relationship study with a broad range of H 3 R agonists, we thereby explored determinants underlying isoform bias at hH 3 R isoforms that exhibit variations in IL3 (i.e., hH 3 R-445, -415, -365, and -329) in a Gα i -dependent pathway (cAMP inhibition). Hence, we systematically characterized hH 3 R isoforms on isoform bias by comparing various ligand properties (i.e., structural and molecular) to the degree of isoform bias. Importantly, our study provides novel insights into the structural and molecular basis of receptor isoform bias, highlighting the importance to study GPCRs with multiple isoforms to better tailor drugs.

Multitargeting ligands on enzymes and receptors may generate a profile for a potential treatment of cognitive impairment. Considering this, a set of 21 substituted aryl-alkyl-piperazines were designed, prepared and tested for their binding affinities at histamine H 3 and dopamine D 3 receptors (H 3 R and D 3 R, respectively) as well as acetyl- and butyrylcholinesterases (AChE/BChE) as potentially synergistic profile. Initial screening of the compounds at H 3 R and D 3 R was done at 1 or 10 µM and 100 µM at AChE and BChE assays. The most promising compounds were then evaluated in full concentration-response curves to estimate the K i and IC 50 values. Results showed that several compounds were ligands at H 3 R (n = 10), D 3 R (n = 6), AChE (n = 3), and BChE (n = 9). Compounds LINS05006 (K i H 3 R 2.8 µM; D 3 R 0.7 µM; IC 50 BChE 26.3 µM) and LINS05015 (K i H 3 R 1.1 µM; D 3 R 3.1 µM; IC 50 AChE 97.8 µM; BChE 43.7 µM) are highlighted since presented affinity in three different. These results suggest that methylpiperazine moiety led to balanced activity at all three classes of targets, and longer linker provided the best affinities. These compounds presented high ligand efficiency values (LE > 0.3) and may have adequate pharmacokinetic profile as suggested by calculated physicochemical properties., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Objective: Methamphetamine (METH) exposure is commonly believed to result in cognitive impairment. Histamine H3 receptor (H3R) antagonists reportedly have potential applications for treating cognitive impairment accompanied by various neuropsychiatric disorders. The present study aimed to investigate the effect of H3R blockade by Thioperamide (THIO) on METH-induced cognitive impairment and the underlying mechanism., Methods: In Experiment 1, C57BL/6 mice received daily injections of saline or 5 mg/kg METH for 5 consecutive days. The Novel Object Recognition (NOR) and Morris water maze (MWM) tasks were used to assess cognitive functions of mice. H3R protein expression and apoptosis were subsequently measured in the hippocampus. In Experiment 2, HT22 cells were first treated with ddH 2 O or 3 mM METH. The cell survival rate and H3R protein level were subsequently assessed. In Experiment 3, the animals were first treated with saline or 20 mg/kg THIO for 7 days, followed by co-administration of either saline or 5 mg/kg METH for an additional 5 days. The remaining experiments were carried out in the same manner as Experiment 1. In Experiment 4, HT22 cells were pretreated with either ddH 2 O or 5 mM THIO for 2 h, followed by ddH 2 O or 3 mM METH treatment for an additional 12 h. The remaining experiments were carried out in the same manner as Experiment 2. In Experiment 5, the changes in MEK1/2, p-MEK1/2, ERK1/2 and p-ERK1/2 protein levels were examined in the hippocampus of all mice from Experiment 3 and HT22 cells from Experiment 4., Results: METH-treated mice showed significantly worsened NOR and MWM performance, along with markably hippocampal apoptosis. A significantly lower cell survival rate was observed in METH-treated HT22 cells. Increased levels of H3R protein were found in both METH-treated mice and HT22 cells. THIO significantly improved METH-induced cognitive impairment in mice and toxicity in HT22 cells. METH significantly increased the level of p-MEK1/2 and p-ERK1/2 proteins in the hippocampus of mice and HT22 cells, which was reversed by THIO pretreatment., Conclusion: Our findings reveal that H3R blockade by THIO yields a neuroprotective effect against METH-induced cognitive impairment in mice and toxicity in HT22 cells via the raf-MEK-ERK signaling pathway., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Dysregulation in brain neurotransmitters underlies several neuropsychiatric disorders, e.g., autism spectrum disorder (ASD). Also, abnormalities in the extracellular-signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) pathway pave the way for neuroinflammation, neurodegeneration, and altered learning phenotype in ASD. Therefore, the effects of chronic systemic administration of the multiple-targeting antagonist ST-713 at the histamine H3 receptor (H3R) and dopamine D2/D3 receptors (D2/D3R) on repetitive self-grooming, aggressive behaviors, and abnormalities in the MAPK pathway in BTBR T + Itpr3tf/J (BTBR) mice were assessed. The results showed that ST-713 (2.5, 5, and 10 mg/kg, i.p.) mitigated repetitive self-grooming and aggression in BTBR mice (all p < 0.05), and the ameliorative effects of the most promising dose of ST-713 (5 mg/kg, i.p.) on behaviors were completely abrogated by co-administration of the H3R agonist (R)-α-methylhistamine or the anticholinergic drug scopolamine. Moreover, the elevated levels of several MAPK pathway proteins and induced proinflammatory markers such as tumor necrosis factor (TNF-α), interleukin-1β (IL-1β), and IL-6 were significantly suppressed following chronic administration of ST-713 (5 mg/kg, i.p.) (all p < 0.01). Furthermore, ST-713 significantly increased the levels of histamine and dopamine in hippocampal tissue of treated BTBR mice (all p < 0.01). The current observations signify the potential role of such multiple-targeting compounds, e.g., ST-713, in multifactorial neurodevelopmental disorders such as ASD.

The multitarget-directed ligands demonstrating affinity to histamine H 3 receptor and additional cholinesterase inhibitory potency represent a promising strategy for research into the effective treatment of Alzheimer's disease. In this study, a novel series of benzophenone derivatives was designed and synthesized. Among these derivatives, we identified compound 6 with a high affinity for H 3 R ( K i = 8 nM) and significant inhibitory activity toward BuChE (IC 50 = 172 nM and 1.16 µM for eq BuChE and h BuChE, respectively). Further in vitro studies revealed that compound 6 (4-fluorophenyl) (4-((5-(piperidin-1-yl)pentyl)oxy)phenyl)methanone) displays moderate metabolic stability in mouse liver microsomes, good permeability with a permeability coefficient value (P e ) of 6.3 × 10 -6 cm/s, and its safety was confirmed in terms of hepatotoxicity in the HepG2 cell line. Therefore, we investigated the in vivo activity of compound 6 in the Passive Avoidance Test and the Formalin Test. While compound 6 did not show a statistically significant influence on memory and learning, it showed analgesic properties in both acute (ED 50 = 20.9 mg/kg) and inflammatory (ED 50 = 17.5 mg/kg) pain.

Histamine is well known for mediating peripheral inflammation; however, this amine is also found in high concentrations in the brain where its roles are much less known. In vivo chemical dynamics are difficult to measure, thus fundamental aspects of histamine's neurochemistry remain undefined. In this work, we undertake the first in-depth characterization of real time in vivo histamine dynamics using fast electrochemical tools. We find that histamine release is sensitive to pharmacological manipulation at the level of synthesis, packaging, autoreceptors and metabolism. We find two breakthrough aspects of histamine modulation. First, differences in H3 receptor regulation between sexes show that histamine release in female mice is much more tightly regulated than in male mice under H3 or inflammatory drug challenge. We hypothesize that this finding may contribute to hormone-mediated neuroprotection mechanisms in female mice. Second, a high dose of a commonly available antihistamine, the H1 receptor inverse agonist diphenhydramine, rapidly decreases serotonin levels. This finding highlights the sheer significance of pharmaceuticals on neuromodulation. Our study opens the path to better understanding and treating histamine related disorders of the brain (such as neuroinflammation), emphasizing that sex and modulation (of serotonin) are critical factors to consider when studying/designing new histamine targeting therapeutics.

Our previous study showed that H 3 receptor antagonists reduced neuronal apoptosis and cerebral infarction in the acute stage after cerebral ischemia, but through an action independent of activation of histaminergic neurons. Because enhanced angiogenesis facilitates neurogenesis and neurological recovery after ischemic stroke, we herein investigated whether antagonism of H 3 R promoted angiogenesis after brain ischemia. Photothrombotic stroke was induced in mice. We showed that administration of H 3 R antagonist thioperamide (THIO, 10 mg·kg -1 ·d -1 , i.p., from D1 after cerebral ischemia) significantly improved angiogenesis assessed on D14, and attenuated neurological defects on D28 after cerebral ischemia. Compared with wild-type mice, Hrh3 -/- mice displayed more blood vessels in the ischemic boundary zone on D14, and THIO administration did not promote angiogenesis in these knockout mice. THIO-promoted angiogenesis in mice was reversed by i.c.v. injection of H 3 R agonist immepip, but not by H 1 and H 2 receptor antagonists, histidine decarboxylase inhibitor α-fluoromethylhistidine, or histidine decarboxylase gene knockout (HDC -/- ), suggesting that THIO-promoted angiogenesis was independent of activation of histaminergic neurons. In vascular endothelial cells (bEnd.3), THIO (10 -9 -10 -7  M) dose-dependently facilitated cell migration and tube formation after oxygen glucose deprivation (OGD), and H 3 R knockdown caused similar effects. We further revealed that H 3 R antagonism reduced the interaction between H 3 R and Annexin A2, while knockdown of Annexin A2 abrogated THIO-promoted angiogenesis in bEnd.3 cells after OGD. Annexin A2-overexpressing mice displayed more blood vessels in the ischemic boundary zone, which was reversed by i.c.v. injection of immepip. In conclusion, this study demonstrates that H 3 R antagonism promotes angiogenesis after cerebral ischemia, which is independent of activation of histaminergic neurons, but related to the H 3 R on vascular endothelial cells and its interaction with Annexin A2. Thus, H 3 R antagonists might be promising drug candidates to improve angiogenesis and neurological recovery after ischemic stroke., (© 2022. The Author(s), under exclusive licence to Shanghai Institute of Materia Medica, Chinese Academy of Sciences and Chinese Pharmacological Society.)

To discover effective analgesics, we summarize the synthesis, optimization, and pharmacological anti-nociceptive effects of a novel series of benzoxazole derivatives targeting H 3 receptor (H 3 R). The new benzoxazoles were assayed in vitro for histamine H 3 R and H 1 R binding affinity. The best compound 8d (2-methyl-6-(3-(4-methylpiperazin-1-yl)propoxy)benzo[d]oxazole) exhibited high affinity for H 3 R (K i  = 19.7 nM), high selectivity for ten other off-target receptors, and negligible effects on human ether-a-go-go-related gene (hERG, cardiac ion channel). In rodent animals, compound 8d dose-dependently reversed formalin-evoked pain (Phase I, ED 50  = 6.0 mg/kg; Phase II, ED 50  = 7.8 mg/kg) and CCI-induced neuropathic pain (chronic constriction injury, ED 50  = 15.6 mg/kg). Furthermore, compound 8d showed an excellent safety profile in acute toxicity test (LD 50  > 2000 mg/kg) with a therapeutic index (TI = LD 50 /ED 50 ) > 250 and showed a desirable drug-like pharmacokinetic profile. Above characteristics indicate that compound 8d represents a promising candidate analgesic for the treatment of neuropathic pain., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Betahistine and gastrodin are the first-line medications for vestibular disorders in clinical practice, nevertheless, their amelioration effects on vestibular dysfunctions still lack direct comparison and their unexpected extra-vestibular effects remain elusive. Recent clinical studies have indicated that both of them may have effects on the gastrointestinal (GI) tract. Therefore, we purposed to systematically compare both vestibular and GI effects induced by betahistine and gastrodin and tried to elucidate the mechanisms underlying their GI effects. Our results showed that betahistine and gastrodin indeed had similar therapeutic effects on vestibular-associated motor dysfunction induced by unilateral labyrinthectomy. However, betahistine reduced total GI motility with gastric hypomotility and colonic hypermotility, whereas gastrodin did not influence total GI motility with only slight colonic hypermotility. In addition, betahistine, at normal dosages, induced a slight injury of gastric mucosa. These GI effects may be due to the different effects of betahistine and gastrodin on substance P and vasoactive intestinal peptide secretion in stomach and/or colon, and agonistic/anatgonistic effects of betahistine on histamine H1 and H3 receptors expressed in GI mucosal cells and H3 receptors distributed on nerves within the myenteric and submucosal plexuses. Furthermore, treatment of betahistine and gastrodin had potential effects on gut microbiota composition, which could lead to changes in host-microbiota homeostasis in turn. These results demonstrate that gastrodin has a consistent improvement effect on vestibular functions compared with betahistine but less effect on GI functions and gut microbiota, suggesting that gastrodin may be more suitable for vestibular disease patients with GI dysfunction., Competing Interests: Conflict of Interest Statement The authors declare no competing interests., (Copyright © 2022 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Histaminergic (HA) neurons are located in the tuberomamillary nucleus (TMN) of the posterior hypothalamus, from where they project throughout the whole brain to control wakefulness. We examined the effects of N α -oleoylhistamine (OLHA), a non-enzymatic condensation product of oleic acid (OLA) and histamine, on activity of mouse HA neurons in brain slices. OLHA bidirectionally modulated the firing of HA neurons. At 10 nM OLHA inhibited or had no action, whereas at 1 μM it evoked excitatory and inhibitory responses. Inhibition was not seen in presence of the histamine receptor H3 (H 3 R) antagonist clobenpropit and in calcium-free medium. Pre-incubation with a histamine-reuptake blocker prevented the decrease in firing by OLHA. OLHA-evoked increase in firing (EC 50 ∼44 nM) was insensitive to blockers of cannabinoid 1 and 2 receptors and of the capsaicin receptor, but was significantly impaired by the peroxisome proliferator-activated receptor-alpha (PPAR-alpha) antagonist MK886, which suppressed also the rise in intracellular calcium level caused by OLHA. The OLHA-evoked excitation was mimicked by synthetic PPAR-alpha agonists (gemfibrozil and GW7647) and was abolished by the PKA inhibitor H-89. The H 3 R affinity (Ki) for histamine, measured in HEK293 cells with stable expression of human H 3 R, was higher than for OLHA (Ki: 42 vs 310 nM, respectively). Expression of PPAR-alpha was not different between TMN regions of males and females, responses to OLHA did not differ. Molecular modelling of PPAR-alpha bound to either OLHA or OEA showed similar binding energies. These findings shed light on a novel biotransformation product of histamine which may play a role in health and disease., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Conformational biosensors to monitor the activation state of G protein-coupled receptors are a useful addition to the molecular pharmacology assay toolbox to characterize ligand efficacy at the level of receptor proteins instead of downstream signaling. We recently reported the initial characterization of a NanoBRET-based conformational histamine H 3 receptor (H 3 R) biosensor that allowed the detection of both (partial) agonism and inverse agonism on living cells in a microplate reader assay format upon stimulation with H 3 R ligands. In the current study, we have further characterized this H 3 R biosensor on intact cells by monitoring the effect of consecutive ligand injections in time and evaluating its compatibility with photopharmacological ligands that contain a light-sensitive azobenzene moiety for photo-switching. In addition, we have validated the H 3 R biosensor in membrane preparations and found that observed potency values better correlated with binding affinity values that were measured in radioligand competition binding assays on membranes. Hence, the H 3 R conformational biosensor in membranes might be a ready-to-use, high-throughput alternative for radioligand binding assays that in addition can also detect ligand efficacies with comparable values as the intact cell assay.

Many studies involving compounds that enhance histamine release, such as histamine H 3 receptor (H 3 R) antagonists, have shown efficacy in inhibiting weight gain, but none have passed clinical trials. As part of the search for H 3 receptor ligands that have additional properties, the aim of this study is to evaluate the activity in the reduction in weight gain in a rat model of excessive eating, as well as the impact on selected metabolic parameters, and the number and size of adipocytes of two new H 3 R antagonists, KSK-60 and KSK-74, which also exert a significant affinity at the sigma-2 receptor. Compounds KSK-60 and KSK-74 are homologues and the elongation of the distal part of the molecule resulted in an approximate two-fold reduction in affinity at H 3 R, but simultaneously an almost two-fold increase in affinity at the sigma-2 receptor. Animals fed palatable feed and receiving KSK-60 or KSK-74 both at 10 mg/kg b.w. gained significantly less weight than animals in the control obese group. Moreover, KSK-74 significantly compensated for metabolic disturbances that accompany obesity, such as an increase in plasma triglyceride, resistin, and leptin levels; improved glucose tolerance; and protected experimental animals against adipocyte hypertrophy. Furthermore, KSK-74 inhibited the development of inflammation in obesity-exposed adipose tissue. The in vivo pharmacological activity of the tested ligands appears to correlate with the affinity at the sigma-2 receptors; however, the explanation of this phenomenon requires further and extended research.

Human histamine H3 receptor (H3R) was initially described in the brain of rat in 1983 and cloned in 1999. It can be found in the human brain and functions as a regulator of histamine synthesis and release. H3 receptors are predominantly resident in the presynaptic region of neurons containing histamine, where they modulate the synthesis and release of histamine (autoreceptor) or other neurotransmitters such as dopamine, norepinephrine, gamma-aminobutyric acid (GABA), glutamate, acetylcholine and serotonin (all heteroreceptors). The human histamine H3 receptor has twenty isoforms of which eight are functional. H3 receptor expression is seen in the cerebral cortex, neurons of the basal ganglia and hippocampus, which are important for process of cognition, sleep and homoeostatic regulation. In addition, histamine H3R antagonists stimulate insulin release, through inducing the release of acetylcholine and cause significant reduction in total body weight and triglycerides in obese subjects by causing a feeling of satiety in the hypothalamus. The ability of histamine H3R antagonist to reduce diabetes-induced hyperglycaemia is comparable to that of metformin. It is reasonable therefore, to claim that H3 receptor antagonists may play an important role in the therapy of disorders of cognition, the ability to sleep, oxidative stress, inflammation and anomaly of glucose homoeostasis. A large number of H3R antagonists are being developed by pharmaceutical companies and university research centres. As examples of these new drugs, this review will discuss a number of drugs, including the first histamine H3R receptor antagonist produced., (Copyright © 2022. Published by Elsevier Masson SAS.)

The striatum is innervated by histaminergic fibers and expresses a high density of histamine H 3 receptors (H 3 Rs), present on medium spiny neurons (MSNs) and corticostriatal afferents. In this study, in sagittal slices from the rat dorsal striatum, excitatory postsynaptic potentials (EPSPs) were recorded in MSNs after electrical stimulation of corticostriatal axons. The effect of H 3 R activation and blockers of calcium and potassium channels was evaluated with the paired-pulse facilitation protocol. In the presence of the H 3 R antagonist/inverse agonist clobenpropit (1 μM), the H 3 R agonist immepip (1 μM) had no effect on the paired-pulse ratio (PPR), but in the absence of clobenpropit, immepip induced a significant increase in PPR, accompanied by a reduction in EPSP amplitude, suggesting presynaptic inhibition. The blockade of Ca V 2.1 (P/Q-type) channels with ω-agatoxin TK (400 nM) increased PPR and prevented the effect of immepip. The Ca V 2.2 (N-type) channel blocker ω-conotoxin GVIA (1 μM) also increased PPR, but did not occlude the immepip action. Functional K IR 3 channels are present in corticostriatal terminals, and in experiments in which immepip increased PPR, the K IR 3 blocker tertiapin-Q (30 nM) prevented the effect of the H 3 R agonist. These results indicate that the presynaptic modulation by H 3 Rs of corticostriatal synapses involves the inhibition of Ca v 2.1 calcium channels and the activation of K IR 3 potassium channels., (© 2020. Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: Histamine H 3 receptor antagonists and inverse agonists have been extensively developed to treat sleep-wake, neurocognitive, and allied disorders. However, potential adverse effects, including insomnia, hampered the clinical use of these drugs, possibly due to their persistent interaction with the target molecules. The purpose of the present study was to estimate the pharmacokinetics and pharmacodynamics of enerisant, a novel antagonist and inverse agonist for histamine H 3 receptors., Methods: To measure the histamine H 3 receptor occupancy by enerisant, positron emission tomography studies using [ 11 C]TASP457, a specific radioligand for histamine H 3 receptors, were performed in 12 healthy men at baseline and at 2 h after oral administration of enerisant hydrochloride. For three of these subjects, two additional scans were performed at 6 and 26 h after the administration. Relationships between the receptor occupancy by enerisant and its dose and plasma concentrations were then analyzed., Results: Administration of enerisant hydrochloride decreased the radioligand binding in a dose-dependent manner. The estimated receptor occupancy values at 2 h varied as a function of its dose or plasma concentration. The time course of the occupancy showed persistently high levels (> 85%) in the two subjects with higher doses (25 and 12.5 mg). The occupancy was also initially high at 2 h and 6 h with the lower dose of 5 mg, but it decreased to 69.7% at 26 h., Conclusion: The target engagement of enerisant was demonstrated in the brains of living human subjects. The occupancy of histamine H 3 receptors by enerisant at 2 h can be predicted by applying the plasma concentration of enerisant to Hill's plot. The preliminary time-course investigation showed persistently high brain occupancy with high doses of enerisant despite the decreasing plasma concentration of the drug. Five milligrams or less dose would be appropriate for the treatment for narcolepsy with initially high occupancy allowing for effective treatment of narcolepsy, and then the occupancy level would be expected to decrease to a level to avoid this drug's unwanted side effect of insomnia at night, although further research is warranted to confirm the statement since the expected decrease is based on the finding in one subject., Trial Registration: This study was retrospectively registered with ClinicalTrials.gov (NCT04631276) on November 17, 2020., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Several of the drugs currently available for the treatment of premature ejaculation (PE) (e.g., local anesthetics or antidepressants) are associated with numerous safety concerns and exhibit weak efficacy. To date, no therapeutics for PE have been approved in the United States, highlighting the need to develop novel agents with sufficient efficacy and fewer side effects. In this study, we focused on the histamine H 3 receptor (H 3 R) as a potential target for the treatment of PE and evaluated the effects of imetit (an H 3 R/H 4 R agonist), ciproxifan (an H 3 R antagonist), and JNJ-7777120 (an H 4 R antagonist) in vivo. Our in vivo electrophysiological experiments revealed that imetit reduced mechanical stimuli-evoked neuronal firing in anesthetized rats. This effect was inhibited by ciproxifan but not by JNJ-7777120. Subsequently, we evaluated the effect of imetit using a copulatory behavior test to assess ejaculation latency (EL) in rats. Imetit prolonged EL, although this effect was inhibited by ciproxifan. These findings indicate that H 3 R stimulation suppresses mechanical stimuli-evoked neuronal firing in the spinal-penile neurotransmission system, thereby resulting in prolonged EL. To our knowledge, this is the first report to describe the relationship between H 3 R and PE. Thus, H 3 R agonists may represent a novel treatment option for PE.

The histamine H 3 receptor is a favourable target for the treatment of cognitive deficits. Here we report the in vitro and in vivo profile of RGH-235, a new potent, selective, and orally active H 3 receptor antagonist/inverse agonist developed by Gedeon Richter Plc. Radioligand binding and functional assays were used for in vitro profiling. Procognitive efficacy was investigated in rodent cognitive tests, in models of attention deficit hyperactive disorder (ADHD) and in cognitive tests of high translational value (rat touch screen visual discrimination test, primate fixed-foreperiod visual reaction time task). Results were supported by pharmacokinetic studies, neurotransmitter release, sleep EEG and dipsogenia. RGH-235 displayed high affinity to H 3 receptors (K i  = 3.0-9.2 nM, depending on species), without affinity to H 1 , H 2 or H 4 receptors and >100 other targets. RGH-235 was an inverse agonist ([ 35 S] GTPγS binding) and antagonist (pERK1/2 ELISA), showing favourable kinetics, inhibition of the imetit-induced dipsogenia and moderate effects on sleep-wake EEG. RGH-235 stimulated neurotransmitter release both in vitro and in vivo. RGH-235 was active in spontaneously hypertensive rats (SHR), generally considered as a model of ADHD, and revealed a robust pro-cognitive profile both in rodent and primate tests (in 0.3-1 mg/kg) and in models of high translational value (e.g. in a rodent touch screen test and in non-human primates). The multiple and convergent procognitive effects of RGH-235 support the view that beneficial cognitive effects can be linked to antagonism/inverse agonism of H 3 receptors., (Copyright © 2021. Published by Elsevier B.V.)

A series of chemical optimizations, which was guided by in vitro affinity at histamine H 3 receptor (H 3 R), modulation of lipophilicity, ADME properties and preclinical efficacy resulted in the identification of 1-[2-(1-cyclobutylpiperidin-4-yloxy)-6,7-dihydro-4H-thiazolo[5,4-c]pyridin-5-yl]propan-1-one (45 e) as a potent and selective (K i =4.0 nM) H 3 R inverse agonist. Dipsogenia induced by (R)-α-methylhistamine was dose dependently antagonized by 45 e, confirming its functional antagonism at H 3 R. It is devoid of hERG and phospholipidosis issues. Compound 45 e has adequate oral exposures and favorable half-life in both rats and dogs. It has demonstrated high receptor occupancy (ED 80 =0.22 mg/kg) and robust efficacy in object recognition task and, dose dependently increased acetylcholine levels in brain. The sub-therapeutic doses of 45 e in combination with donepezil significantly increased acetylcholine levels. The potent affinity, selectivity, in vivo efficacy and drug like properties together with safety, warrant for further development of this molecule for potential treatment of cognitive disorders associated with Alzheimer's disease., (© 2021 Wiley-VCH GmbH.)

Background: Autism spectrum disorders (ASDs) are a group of neurodevelopmental conditions defined by behavioral deficits in social communication and interactions, mental inflexibility and repetitive behaviors. Converging evidence from observational and preclinical studies suggest that excessive repetitive behaviors in people with ASD may be due to elevated histaminergic H3 receptor signaling in the striatum. We hypothesized that systemic administration of pharmacological histamine H3 receptor antagonists would attenuate the expression of repetitive behaviors in the BTBR T+Itpr3tf/J (BTBR) mouse inbred strain, an established mouse model presenting autism-like repetitive behaviors and novelty-induced hyperactivity. We further aimed to investigate whether agonism of the histamine H3 receptor would be sufficient to induce repetitive behaviors in the C57BL/6J control mouse strain., Methods: Different doses of H3 receptor agonists (i.e., (R)-α-methylhistamine and immethridine) and H3 receptor antagonists/inverse agonists (i.e., ciproxifan and pitolisant) were administered via intraperitoneal (i.p.) injection in male mice to characterize the acute effects of these compounds on ASD-related behavioral readouts., Results: The highly selective H3 receptor agonist immethridine significantly increased the time spent in stereotypic patterns in C57BL/6J mice, but this effect appeared to be driven by general sedative properties of the compound. High doses of pitolisant significantly decreased locomotor hyperactivity in novel environments in BTBR mice, without significant effects on repetitive behaviors., Conclusions: Based on our findings, we conclude that acute H3 receptor manipulation mainly affected general motor activity levels in novel environments. Small changes in stereotyped behaviors were observed but appeared to be driven by altered general activity levels., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Positron emission tomography (PET) provides a unique tool to study the biochemistry of the human brain in vivo. By using PET probes that are binding selectively to certain receptor subtypes, brain PET allows the quantification of receptor levels in various brain areas of human subjects. This approach has the potential to reveal abnormal receptor expressions that may contribute to the physiopathology of some psychiatric and neurological disorders. This approach also has the potential to assist in the drug development process by determining receptor occupancy in vivo allowing selection of proper drug dosage to produce therapeutic effects. Several PET tracers have been developed for histamine H3 receptors (H3R). However, despite the potential of PET to elucidate the role of H3R in vivo, only limited work has been conducted so far. This article reviews the work that has been done in this area. Notably, we will cover the limitations of the first-generation PET radioligand for H3R and present the advantages of novel radioligands that promise an explosion of clinical PET research on the role of H3R in vivo., (© 2021. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

During the past decades, histamine H 3 receptors have received widespread attention in pharmaceutical research due to their involvement in pathophysiology of several diseases such as neurodegenerative disorders. In this context, blocking of these receptors is of paramount importance in progression of such diseases. In the current investigation, novel histamine H 3 receptor ligands were designed by exploiting scaffold-hopping drug-design strategy. We inspected the designed molecules in terms of ADME properties, drug-likeness, as well as toxicity profiles. Additionally molecular docking and dynamics simulation studies were performed to predict binding mode and binding free energy calculations, respectively. Among the designed structures, we selected compound d2 and its demethylated derivative as examples for synthesis and affinity measurement. In vitro binding assays of the synthesized molecules demonstrated that d2 has lower binding affinity (K i  = 2.61 μM) in radioligand displacement assay to hH 3 R than that of demethylated form (K i  = 12.53 μM). The newly designed compounds avoid of any toxicity predictors resulted from extended in silico and experimental studies, can offer another scaffold for histamine H 3 R antagonists for further structure-activity relationship studies., (Copyright © 2021. Published by Elsevier Inc.)

Alzheimeŕs disease (AD) is the most prominent neurodegenerative disorder with high medical need. Protein-protein-interactions (PPI) interactions have a critical role in AD where β-amyloid structures (Aβ) build toxic oligomers. Design of disease modifying multi target directed ligand (MTDL) has been performed, which disable PPI on the one hand and on the other hand, act as procognitive antagonists at the histamine H 3 receptor (H 3 R). The synthetized compounds are structurally based on peptidomimetic amino acid-like structures mainly as keto, diketo-, or acyl variations of a piperazine moiety connected to an H 3 R pharmacophore. Most of them showed low nanomolar affinities at H 3 R and some with promising affinity to Aβ-monomers. The structure-activity relationships (SAR) described offer new possibilities for MTDL with an optimized profile combining symptomatic and potential causal therapeutic approaches in AD., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Aims: One of the therapeutic approaches in the treatment of obesity is the use of histamine H 3 receptor ligands. Histamine plays a significant role in eating behavior because it causes a loss of appetite and is considered to be a satiety signal released during food intake., Material and Methods: Histamine ligands were selected based on the preliminary studies which included determination of intrinsic activity and selected pharmacokinetic parameters. Female Wistar rats were fed palatable feed for 28 days and simultaneously the tested compounds were administered intraperitoneally at a dose of 10 mg/kg b.w./day. Rats' weight was evaluated daily and calories intake was evaluated once per week. At the end of experiment insulin and glucose tolerance tests was performed. Plasma levels of cholesterol, triglycerides, leptin, insulin, glucose, C-peptide and CRP were also determined. In order to rule out false-positive results the influence of tested compounds on spontaneous activity of rats was monitored., Results: Animals fed palatable feed and treated with KSK-61 or KSK-63 compounds showed the slowest weight gain which was comparable to the one observed in control animals. Both compounds with the highest pharmacological activity have also similar pharmacokinetic properties with quite long half-life and high volume of distribution indicating that they can freely cross most biological barriers. Some compounds, especially KSK-63, compensated for metabolic disorders., Conclusion: The presented study proves that search among the active histamine H 3 receptor ligands for the new therapeutic agents to treat obesity is justified. Compounds KSK-61 and KSK-63 can be considered as the leading structures., (Copyright © 2021 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

G-protein-coupled receptors (GPCRs) are dimeric proteins, but the functional consequences of the process are still debated. Active GPCR conformations are promoted either by agonists or constitutive activity. Inverse agonists decrease constitutive activity by promoting inactive conformations. The histamine H 3 receptor (H 3 R) is the target of choice for the study of GPCRs because it displays high constitutive activity. Here, we study the dimerization of recombinant and brain H 3 R and explore the effects of H 3 R ligands of different intrinsic efficacy on dimerization. Co-immunoprecipitations and Western blots showed that H 3 R dimers co-exist with monomers in transfected HEK 293 cells and in rodent brains. Bioluminescence energy transfer (BRET) analysis confirmed the existence of spontaneous H 3 R dimers, not only in living HEK 293 cells but also in transfected cortical neurons. In both cells, agonists and constitutive activity of the H 3 R decreased BRET signals, whereas inverse agonists and GTPγS, which promote inactive conformations, increased BRET signals. These findings show the existence of spontaneous H 3 R dimers not only in heterologous systems but also in native tissues, which are able to adopt a number of allosteric conformations, from more inactive to more active states.

Alzheimer's disease (AD) is a complex and incurable illness that requires the urgent approval of new effective drugs. However, since 2003, no new molecules have shown successful results in clinical trials, thereby making the common "one compound - one target" paradigm questionable. Recently, the multitarget-directed ligand (MTDL) approach has gained popularity, as compounds targeting at least two biological targets may be potentially more effective in treating AD. On the basis of these findings, we designed, synthesized, and evaluated through biological assays a series of derivatives of alicyclic amines linked by an alkoxy bridge to an aromatic lipophilic moiety of [1,1'-biphenyl]-4-carbonitrile. The research results revealed promising biological activity of the obtained compounds toward the chosen targets involved in AD pathophysiology; the compounds showed high affinity (mostly low nanomolar range of K i values) for human histamine H 3 receptors (hH 3 R) and good nonselective inhibitory potency (micromolar range of IC 50 values) against acetylcholinesterase from electric eel (eeAChE) and equine serum butyrylcholinesterase (eqBuChE). Moreover, micromolar/submicromolar potency against human monoamine oxidase B (hMAO B) was detected for some compounds. The study identified compound 5 as a multiple hH 3 R/eeAChE/eqBuChE/hMAO B ligand (5: hH 3 R K i  = 9.2 nM; eeAChE IC 50  = 2.63 µM; eqBuChE IC 50  = 1.30 µM; hMAO B IC 50  = 0.60 µM). Further in vitro studies revealed that compound 5 exhibits a mixed type of eeAChE and eqBuChE inhibition, good metabolic stability, and moderate hepatotoxicity effect on HepG2 cells. Finally, compound 5 showed a beneficial effect on scopolamine-induced memory impairments, as assessed by the passive avoidance test, thus revealing the potential of this compound as a promising agent for further optimization for AD treatment., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

High levels of histamine and histamine receptors (HRs), including H1R~H4R, are found in many different types of tumor cells and cells in the tumor microenvironment, suggesting their involvement in tumor progression. This review summarizes the latest evidence demonstrating the pathophysiological roles of histamine and its cognate receptors in cancer biology. We also discuss the novel therapeutic approaches of selective HR ligands and their potential prognostic values in cancer treatment. Briefly, histamine is highly implicated in cancer development, growth, and metastasis through interactions with distinct HRs. It also regulates the infiltration of immune cells into the tumor sites, exerting an immunomodulatory function. Moreover, the effects of various HR ligands, including H1R antagonists, H2R antagonists, and H4R agonists, on tumor progression in many different cancer types are described. Interestingly, the expression levels of HR subtypes may serve as prognostic biomarkers in several cancers. Taken together, HRs are promising targets for cancer treatment, and HR ligands may offer novel therapeutic potential, alone or in combination with conventional therapy. However, due to the complexity of the pathophysiological roles of histamine and HRs in cancer biology, further studies are warranted before HR ligands can be introduced into clinical settings.

A novel series of 2-styryl-5-hydroxy-4-pyrone derivatives and analogues were designed and synthesized as H 3 receptor antagonism based multitarget-directed ligands (MTDLs) for AD therapy using pharmacophore-combine strategy. The 2-styryl-5-hydroxy-4-pyrone pharmacophore with metal ion chelation, antioxidation, and Aβ aggregation inhibition activities was employed as the "eastern part", and a typical phenoxyalkylamine moiety was used as "central ring + western part" of the H 3 receptor antagonist. The biological evaluation revealed that the majority of the target compounds demonstrated desirable multiple functions. The two most promising compounds 8a and 8b exhibited nanomolar IC 50 values on H 3 receptor antagonism, excellent metal ion chelating capability, more potent ABTS + scavenging activity than Trolox, efficient Aβ self-aggregation and Cu 2+ -induced aggregation inhibitory activities, as well as disaggregation activities against Aβ self/Cu 2+ -induced aggregation., (Copyright © 2021. Published by Elsevier Ltd.)

The histamine H 3 receptor (H 3 R) is considered an attractive drug target for various neurological diseases. We here report the synthesis of UR-NR266, a novel fluorescent H 3 R ligand. Broad pharmacological characterization revealed UR-NR266 as a sub-nanomolar compound at the H 3 R with an exceptional selectivity profile within the histamine receptor family. The presented neutral antagonist showed fast association to its target and complete dissociation in kinetic binding studies. Detailed characterization of standard H 3 R ligands in NanoBRET competition binding using UR-NR266 highlights its value as a versatile pharmacological tool to analyze future H 3 R ligands. The low nonspecific binding observed in all experiments could also be verified in TIRF and confocal microscopy. This fluorescent probe allows the highly specific analysis of native H 3 R in various assays ranging from optical high throughput technologies to biophysical analyses and single-molecule studies in its natural environment. An off-target screening at 14 receptors revealed UR-NR266 as a selective compound.

The role of histamine and acetylcholine in cognitive functions suggests that compounds able to increase both histaminergic and cholinergic neurotransmissions in the brain should be considered as promising therapeutic options. For this purpose, dual inhibitors of histamine H 3 receptors (H 3 R) and cholinesterases (ChEs) have been designed and assessed. In this context, this paper reviews the strategies used to obtain dual H 3 R/ChEs ligands using multitarget design approaches. Hybrid compounds designed by linking tacrine or flavonoid motifs to H 3 R antagonists were obtained with high affinity for both targets, and compounds designed by merging the H 3 R antagonist pharmacophore with known anticholinesterase molecules were also reported. These reports strongly suggest that key modifications in the lipophilic region (including a second basic group) seem to be a strategy to reach novel compounds, allied with longer linker groups to a basic region. Some compounds have already demonstrated efficacy in memory models, although the pharmacokinetic and toxicity profile should be considered when designing further compounds. In conclusion, the key features to be considered when designing novel H 3 R/ChEs inhibitors with improved pharmacological profile were herein summarized., (© 2021 John Wiley & Sons A/S.)

Recent evidence shows that the expression levels of histamine receptor H3 (Hrh3) are upregulated in several types of cancer. However, the role of Hrh3 in non-small cell lung cancer (NSCLC) has not been elucidated. In the present study, we showed that the expression levels of Hrh3 were significantly increased in NSCLC samples, and high levels of Hrh3 were associated with poor overall survival (OS) in NSCLC patients. In five human NSCLC cell lines tested, Hrh3 was significantly upregulated. In NSCLC cell lines H1975, H460, and A549, Hrh3 antagonist ciproxifan (CPX, 10-80 μM) exerted moderate and concentration-dependent inhibition on the cell growth and induced apoptosis, whereas its agonist RAMH (80 μM) reversed these effects. Furthermore, inhibition of Hrh3 by CPX or siRNA retarded the migration and invasion of NSCLC cells through inhibiting epithelial-mesenchymal transition (EMT) progression via reducing the phosphorylation of PI3K/Akt/mTOR and MEK/ERK signaling pathways. In nude mice bearing H1975 cell xenograft or A549 cell xenograft, administration of CPX (3 mg/kg every other day, intraperitoneal) significantly inhibited the tumor growth with increased E-cadherin and ZO-1 expression and decreased Fibronectin expression in tumor tissue. In conclusion, this study reveals that Hrh3 plays an important role in the growth and metastasis of NSCLC; it might be a potential therapeutic target against the lung cancer., (© 2020. CPS and SIMM.)

Adenosine A 1 /A 2A receptors (A 1 R/A 2A R) represent targets in nondopaminergic treatment of motor disorders such as Parkinson's disease (PD). As an innovative strategy, multitargeting ligands (MTLs) were developed to achieve comprehensive PD therapies simultaneously addressing comorbid symptoms such as sleep disruption. Recognizing the wake-promoting capacity of histamine H 3 receptor (H 3 R) antagonists in combination with the "caffeine-like effects" of A 1 R/A 2A R antagonists, we designed A 1 R/A 2A R/H 3 R MTLs, where a piperidino-/pyrrolidino(propyloxy)phenyl H 3 R pharmacophore was introduced with overlap into an adenosine antagonist arylindenopyrimidine core. These MTLs showed distinct receptor binding profiles with overall nanomolar H 3 R affinities ( K i < 55 nM). Compound 4 ( ST-2001 , K i (A 1 R) = 11.5 nM, K i (A 2A R) = 7.25 nM) and 12 ( ST-1992 , K i (A 1 R) = 11.2 nM, K i (A 2A R) = 4.01 nM) were evaluated in vivo . l-DOPA-induced dyskinesia was improved after administration of compound 4 (1 mg kg -1 , i.p. rats). Compound 12 (2 mg kg -1 , p.o. mice) increased wakefulness representing novel pharmacological tools for PD therapy.