H2 Agonist Research Articles

Histamine H3R receptor activation in the dorsal striatum triggers stereotypies in a mouse model of tic disorders

Tic disorders affect ~5% of the population and are frequently comorbid with obsessive-compulsive disorder, autism, and attention deficit disorder. Histamine dysregulation has been identified as a rare genetic cause of tic disorders; mice with a knockout of the histidine decarboxylase (Hdc) gene represent a promising pathophysiologically grounded model. How alterations in the histamine system lead to tics and other neuropsychiatric pathology, however, remains unclear. We found elevated expression of the histamine H3 receptor in the striatum of Hdc knockout mice. The H3 receptor has significant basal activity even in the absence of ligand and thus may modulate striatal function in this knockout model. We probed H3R function using specific agonists. The H3 agonists R-aminomethylhistamine (RAMH) and immepip produced behavioral stereotypies in KO mice, but not in controls. H3 agonist treatment elevated intra-striatal dopamine in KO mice, but not in controls. This was associated with elevations in phosphorylation of rpS6, a sensitive marker of neural activity, in the dorsal striatum. We used a novel chemogenetic strategy to demonstrate that this dorsal striatal activity is necessary and sufficient for the development of stereotypy: when RAMH-activated cells in the dorsal striatum were chemogenetically activated (in the absence of RAMH), stereotypy was recapitulated in KO animals, and when they were silenced the ability of RAMH to produce stereotypy was blocked. These results identify the H3 receptor in the dorsal striatum as a contributor to repetitive behavioral pathology.

Current Knowledge and Perspectives on Histamine H1 and H2 Receptor Pharmacology: Functional Selectivity, Receptor Crosstalk, and Repositioning of Classic Histaminergic Ligands

H1 and H2 histamine receptor antagonists, although developed many decades ago, are still effective for the treatment of allergic and gastric acid-related conditions. This article focuses on novel aspects of the pharmacology and molecular mechanisms of histamine receptors that should be contemplated for optimizing current therapies, repositioning histaminergic ligands for new therapeutic uses, or even including agonists of the histaminergic system in the treatment of different pathologies such as leukemia or neurodegenerative disorders. In recent years, new signaling phenomena related to H1 and H2 receptors have been described that make them suitable for novel therapeutic approaches. Crosstalk between histamine receptors and other membrane or nuclear receptors can be envisaged as a way to modulate other signaling pathways and to potentiate the efficacy of drugs acting on different receptors. Likewise, biased signaling at histamine receptors seems to be a pharmacological feature that can be exploited to investigate nontraditional therapeutic uses for H1 and H2 biased agonists in malignancies such as acute myeloid leukemia and to avoid undesired side effects when used in standard treatments. It is hoped that the molecular mechanisms discussed in this review contribute to a better understanding of the different aspects involved in histamine receptor pharmacology, which in turn will contribute to increased drug efficacy, avoidance of adverse effects, or repositioning of histaminergic ligands.

The Histamine H3 Receptor Differentially Modulates Mitogen-activated Protein Kinase (MAPK) and Akt Signaling in Striatonigral and Striatopallidal Neurons

The basal ganglia have a central role in motor patterning, habits, motivated behaviors, and cognition as well as in numerous neuropsychiatric disorders. Receptors for histamine, especially the H3 receptor (H3R), are highly expressed in the striatum, the primary input nucleus of the basal ganglia, but their effects on this circuitry have been little explored. H3R interacts with dopamine (DA) receptors ex vivo; the nature and functional importance of these interactions in vivo remain obscure. We found H3R activation with the agonist R-(-)-α-methylhistamine to produce a unique time- and cell type-dependent profile of molecular signaling events in the striatum. H3 agonist treatment did not detectably alter extracellular DA levels or signaling through the cAMP/DARPP-32 signaling pathway in either D1- or D2-expressing striatal medium spiny neurons (MSNs). In D1-MSNs, H3 agonist treatment transiently activated MAPK signaling and phosphorylation of rpS6 and led to phosphorylation of GSK3β-Ser9, a novel effect. Consequences of H3 activation in D2-MSNs were completely different. MAPK signaling was unchanged, and GSK3β-Ser9 phosphorylation was reduced. At the behavioral level, two H3 agonists had no significant effect on locomotion or stereotypy, but they dramatically attenuated the locomotor activation produced by the D1 agonist SKF82958. H3 agonist co-administration blocked the activation of MAPK signaling and the phosphorylation of rpS6 produced by D1 activation in D1-MSNs, paralleling behavioral effects. In contrast, GSK3β-Ser9 phosphorylation was seen only after H3 agonist treatment, with no interactive effects. H3R signaling has been neglected in models of basal ganglia function and has implications for a range of pathophysiologies.

In vitro study of histamine and histamine receptor ligands influence on the adhesion of purified human eosinophils to endothelium

It is a well-known fact that histamine is involved in eosinophil-dependent inflammatory responses including cellular chemotaxis and migration. Nevertheless, the relative role of histamine receptors in the mechanisms of eosinophils adhesion to endothelial cells is not known. Therefore the aim of presented study was to examine the effect of selective histamine receptors ligands on eosinophils adhesion to endothelium. For that purpose the highly purified human eosinophils have been isolated from the peripheral blood. The viability and functional integrity of isolated eosinophils have been validated in several tests. Histamine as well as 4-methylhistamine (selective H4 agonist) in concentration-dependent manner significantly increased number of eosinophils that adhere to endothelium. Among the selective histamine receptors antagonist or H1 inverse agonist only JNJ7777120 (histamine H4 antagonist) and thioperamide (dual histamine H3/H4 antagonist) had direct effect on eosinophils adhesion to endothelial cells. Antagonists of H1 (diphenhydramine, mepyramine) H2 (ranitidine and famotidine) and H3 (pitolisant) histamine receptors were ineffective. To the best of our knowledge, this is the first study to demonstrate that histamine receptor H4 plays a dominant role in histamine-induced eosinophils adhesion to endothelium.

A functional study on small intestinal smooth muscles in jejunal atresia.

Aim:The present study was aimed to assess the contractile status of neonatal small intestinal smooth muscle of dilated pre-atretic part of intestinal atresia to resolve debatable issues related to mechanisms of persistent dysmotility after surgical repair.Materials and Methods:A total of 34 longitudinally sectioned strips were prepared from pre-atretic dilated part of freshly excised 8 jejunal atresia type III a cases. Spontaneous as well as acetylcholine- and histamine-induced contractions were recorded in vitro by using organ bath preparations. Chemically evoked contractions were further evaluated after application of atropine (muscarinic blocker), pheniramine (H1 blocker), and lignocaine (neuronal blocker) to ascertain receptors and neuronal involvement. Histological examinations of strips were made by using Masson trichrome stain to assess the fibrotic changes.Results:All 34 strips, except four showed spontaneous contractions with mean frequency and amplitude of 5.49 ± 0.26/min and 24.41 ± 5.26 g/g wet tissue respectively. The response to ACh was nearly twice as compared to histamine for equimolar concentrations (100 μM). ACh (100 μM) induced contractions were attenuated (by 60%) by atropine. Histamine (100 μM)-induced contractions was blocked by pheniramine (0.32 μM) and lignocaine (4 μM) by 74% and 78%, respectively. Histopathological examination showed varying degree of fibrotic changes in muscle layers.Conclusions:Pre-atretic dilated part of jejunal atresia retains functional activity but with definitive histopathologic abnormalities. It is suggested that excision of a length of pre-atretic part and early stimulation of peristalsis by locally acting cholinomimetic or H1 agonist may help in reducing postoperative motility problems in atresia patients.

Histamine H2 Receptor in Blood Cells: A Suitable Target for the Treatment of Acute Myeloid Leukemia.

Acute myeloid leukemia (AML) consists in a cancer of early hematopoietic cells arising in the bone marrow, most often of those cells that would turn into white blood cells (except lymphocytes). Chemotherapy is the treatment of choice for AML but one of the major complications is that current drugs are highly toxic and poorly tolerated. In general, treatment for AML consists of induction chemotherapy and post-remission therapy. If no further post-remission is given, almost all patients will eventually relapse. Histamine, acting at histamine type-2 (H2) receptors on phagocytes and AML blast cells, helps prevent the production and release of oxygen-free radicals, thereby protecting NK and cytotoxic T cells. This protection allows immune-stimulating agents, such as interleukin-2 (IL-2), to activate cytotoxic cells more effectively, enhancing the killing of tumor cells. Based on this mechanism, post-remission therapy with histamine and IL-2 was found to significantly prevent relapse of AML. Alternatively, another potentially less toxic approach to treat AML employs drugs to induce differentiation of malignant cells. It is based on the assumption that many neoplastic cell types exhibit reversible defects in differentiation, which upon appropriate treatment results in tumor reprogramming and the induction of terminal differentiation. There are promissory results showing that an elevated and sustained signaling through H2 receptors is able to differentiate leukemia-derived cell lines, opening the door for the use of H2 agonists for specific differentiation therapies. In both situations, histamine acting through H2 receptors constitutes an eligible treatment to induce leukemic cell differentiation, improving combined therapies.

Effect of histamine receptors agonists or antagonists administered intracerebroventricularly and intrathecally on the blood glucose level in immobilization stress model

Effects of histamine receptors agonists or antagonists administered centrally (supraspinally or spinally) on immobilization stress-induced blood glucose levels were studied in Institute for Cancer Research mice. Mice were pretreated intracerebroventricularly (i.c.v.) or intrathecally (i.t.) with histamine (H1, H2, H3 and H4) receptor agonists or antagonists 10 min prior to immobilization stress (IMO) stimulation for 30 min. We found that the increased blood glucose level induced by IMO was attenuated by the i.c.v. pretreatment with α-methylhistamine (H3 agonist). However, H3 antagonist, H1, H2 and H4 receptor agonists and antagonists did not affect the blood glucose level. Also, i.t. pretreatment with α-methylhistamine or VUF 8430 attenuated the increased blood glucose level induced by IMO. In addition, cetirizine and carcinine enhanced the increased blood glucose level induced by IMO. However, the blood glucose level was not affected by H1 and H2 receptor agonists, H2 and H4 receptor antagonists. Corticosterone and insulin levels were also measured in α-methylhistamine (i.c.v or i.t) and with VUF 8430, carcinine and cetirizine (i.t.) treated group. We found that the corticosterone level was increased only by i.t treatment with α-methylhistamine, whereas the insulin level was attenuated by i.c.v or i.t. treatment with α-methylhistamine and i.t treatment with VUF 8430, carcinine and cetirizine. Our results suggest the active involvement of histamine receptor in blood glucose regulation in IMO.

Morphine modulates the effects of histamine H1 and H3 receptors on seizure susceptibility in pentylenetetrazole-induced seizure model of mice

Histamine regulates release of neurotransmitters such as dopamine, serotonin, gamma-aminobutyric acid (GABA), glutamate and also is involved in several functions in central nervous system (CNS). It has been shown that histamine participates in disorders like seizure. It has been well documented that morphine dose-dependently induces anti or proconvulsant effects. In the current study, we firstly showed that morphine (1mg/kg) exerts anticonvulsant effects which significantly reversed by naltrexone administration. Secondly, we determined seizure threshold for H1 and H3 receptors agonists and antagonists in mouse model of pentylenetetrazole (PTZ)-induced clonic seizures. Our results showed that activation of H1 receptors by 2-(2-Pyridyl)-ethylamine exerts anticonvulsant properties while inhibition of H1 receptors by pyrilamine maleate induced proconvulsant effects. Furthermore, we showed that immepip dihydrobromide, a H3 receptor agonist, increased seizure susceptibility to PTZ whereas thioperamide, a H3 receptor antagonist increased seizure threshold. We also revealed that pretreatment with morphine potently reversed the effects of histaminergic system on seizure threshold suggesting the involvement of opioid system in alteration of seizure threshold by histaminergic drugs.

Histamine and Histamine Receptor Involvement in Sensory‐Motor Integration of Gill Lateral Cell Cilia Activity in the Bivalve Crassostrea viginica

Gill lateral ciliated cells of Crassostrea virginica are innervated by serotonin and dopamine nerves. Motor aspects have been well studied, but the sensory side hasn't. Histamine (HIS) a neurotransmitter in a variety of invertebrates rarely has been reported in bivalves. We quantified HIS in C. virginica and found gill lateral cell cilia beating changed in response to mantle sensory inputs including HIS. The mechanism involves mantle rim sensory tentacles and visceral ganglia. HIS does not alter beating when applied to gill. We hypothesize HIS receptors are present in mantle and can be confirmed by Western Blot (WB). For WB, mantle body and rim lysates were prepared in NP-40 detergent buffer containing protease inhibitor followed by centrifuging to obtain supernatant with solubilized mantle body and rim membrane proteins. H2 receptor immunoreactivity was revealed by 1̊ antibody followed by HRP-conjugated 2̊ antibody. Mantle body and rim H2 receptor protein was resolved by colormetric development using CN/DAB substrate kit. We also did a dose response of HIS H1, H2 and H3 receptor agonists and antagonist at the mantle rim and observed cilia beating in gill. WB showed a 70 kD band corresponding to HIS H2 receptors in mantle body and rim, and H2 agonists and antagonists had the strongest effects on cilia beating. The study shows mantle body and rim have HIS H2 receptors and demonstrates a physiological role of HIS in sensory-motor integration of gill lateral cell cilia activity.

A search for functional histamine H4 receptors in the human, guinea pig and mouse brain.

Histamine H4 receptors are expressed in immune cells, but their potential role in the brain is less clear. Although H4 transcripts have been identified in human and rat brain, the presence of H4 receptors on the protein level has so far not been proven since appropriate antibodies fulfilling the strict criteria for G protein-coupled receptors are missing. Here, we searched for functional H4 receptors in human, guinea pig and mouse cortex. We studied whether H4 receptor activation is associated with increased GTPγS binding and reduced noradrenaline release. The latter two effects have been previously shown for H3 receptors, which, like the H4 receptors, are coupled to G i/o protein. G protein activation was studied using (35)S-GTPγS binding in cortical membranes. The electrically induced (3)H-noradrenaline release was determined in superfused cortical slices. The H4 agonist 4-methylhistamine failed to affect (35)S-GTPγS binding and/or noradrenaline release in human, guinea pig and mouse cortex although an H 3 receptor-mediated increase in (35)S-GTPγS binding and inhibition of noradrenaline release occurred in parallel experiments. In conclusion, functional H4 receptors increasing (35)S-GTPγS binding and/or decreasing noradrenaline release are not found in human, guinea pig and mouse cortex.

Histamine acting on the basolateral amygdala reverts the impairment of aversive memory of rats submitted to neonatal maternal deprivation

Recent findings suggest a role of brain histamine in the regulation of memory consolidation, particularly in one-trial inhibitory avoidance (IA) learning and that disruption in the mother infant relationship i.e. maternal deprivation induces cognitive deficits. We investigate whether histamine itself, and histaminergic compounds given into the basolateral amygdala (BLA) immediately post-training can affect retention (24h after training) of one-trial (IA) in rats submitted to early postnatal maternal deprivation. In all cases, deprived (Dep) animals had lower retention scores than non-deprived controls (N-dep). Histamine induced memory enhancement on its own in N-dep animals and was able to overcome the deleterious effect of Dep. The effects by SKF-91488 is similar to histamine. The H3 agonist, imetit mimetized the enhancing effects of histamine; neither agonist H1 pyridylethylamine nor the H2 dimaprit had any effect. Ranitidine and thioperamide (50nmol) co-infused with histamine (10nmol) fully blocked the restorative effect of histamine on retention in Dep animals. Thioperamide, in addition, blocked the enhancing effect of histamine on memory of the N-dep animals as well. None of the drugs used given into BLA had any effect on open-field or elevated plus-maze behavior in N-dep or Dep rats. Our results are limited to experimental design in rats. Extrapolation i.e. in humans requires further experimentations. The present results suggest that the memory deficit induced by early postnatal maternal deprivation in rats may at least in part be due to an impairment of histamine H3 receptor-mediated mediated mechanisms in the BLA.

The H3 receptor agonist immepip does not affect l-dopa-induced abnormal involuntary movements in 6-OHDA-lesioned rats

The treatment of dyskinesia in Parkinson׳s disease remains poor but H3 receptor agonists have been suggested as a novel pharmacological approach. We examined the effects of the H3 agonist, immepip, in 6-OHDA-lesioned rats exhibiting AIMs (abnormal involuntary movements), a rat analogue of dyskinesia, in response to l-dopa compared to the known anti-dyskinetic agents amantadine, MK-801 and 8-OHDPAT. We then attempted to extend these studies in to dyskinetic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treated common marmosets.Amantadine, MK-801 and 8-OHDPAT all dose-dependently reduced l-dopa-induced axial, lingual and oral (ALO) AIMs in 6-OHDA-lesioned animals accompanied by a reduction in contralateral rotation with higher doses of amantadine and MK-801. By contrast, immepip had no effect on AIMs expression or contralateral rotation. In the MPTP-treated common marmoset exhibiting dyskinesia to l-dopa, immepip alone induced retching and in combination with l-dopa administered subcutaneously or orally induced the rapid onset of retching and vomiting which was not controlled by pretreatment with domperidone. Administration of the unrelated H3 agonist, imetit had the same effect. Despite causing negative side-effects, it appears that both agonists reduced the antiparkinsonian response to l-dopa resulting in reduced dyskinesia.H3 agonists appear unlikely candidates for the treatment of dyskinesia in PD based on lack of evidence of efficacy and potential adverse effects.

Antihistamine Effect of a Pure Bioactive Compound Isolated from Slug (Diplosolenodes occidentalis) Material.

Folklore claims of the therapeutic effect of garden slug (Diplosolenodes occidentalis) extract used to relieve bronchoconstriction in asthmatic individuals were never validated scientifically. The aim of this study was to isolate the pure bioactive compound from slug extract causing this effect. The crude ground material was prepared in ethanol and after filtration, separation by flash column chromatography method was done. The structure was elucidated by data from hydrogen and carbon nuclear magnetic resonance (NMR) profiles. The bioactive compound was assessed for dose-dependent response effects on guinea pig tracheal smooth muscle pre-contracted with histamine. Receptor specificity studies were done by using HTMT dimaleate (H1 agonist). The type of antagonism was also identified. The pure component isolated from garden slug material was identified by spectral studies as glyceryl trilinolenate (GT). It caused dose-dependent relaxation in guinea pig tracheal smooth muscle strips pre-contracted with histamine, it acted via H1 type receptors and showed non-competitive antagonism. Glyceryl trilinolenate produced dose-dependent relaxation in tracheal smooth muscle strips in the presence of the agonist histamine. Glyceryl trilinolenate displayed non-competitive antagonism at H1 receptors in the trachea. This agent was able to alleviate bronchoconstriction in individuals presenting with atopic asthma in rural agricultural areas in Jamaica (verbal communications). It is possible that GT can be useful therapeutically to produce tracheal smooth muscle relaxation in individuals presenting with atopic asthma.

Clobenpropit enhances anti-tumor effect of gemcitabine in pancreatic cancer.

To evaluate the anti-tumor effect of clobenpropit, which is a specific H3 antagonist and H4 agonist, in combination with gemcitabine in a pancreatic cancer cell line. Three kinds of human pancreatic cancer cell lines (Panc-1, MiaPaCa-2, and AsPC-1) were used in this study. Expression of H3 and H4 receptors in pancreatic cancer cells was identified with Western blotting. Effects of clobenpropit on cell proliferation, migration and apoptosis were evaluated. Alteration of epithelial and mesenchymal markers after administration of clobenpropit was analyzed. An in vivo study with a Panc-1 xenograft mouse model was also performed. H4 receptors were present as 2 subunits in human pancreatic cancer cells, while there was no expression of H3 receptor. Clobenpropit inhibited cell migration and increased apoptosis of pancreatic cancer cells in combination with gemcitabine. Clobenpropit up-regulated E-cadherin, but down-regulated vimentin and matrix metalloproteinase 9 in real-time polymerase chain reaction. Also, clobenpropit inhibited tumor growth (gemcitabine 294 ± 46 mg vs combination 154 ± 54 mg, P = 0.02) and enhanced apoptosis in combination with gemcitabine (control 2.5%, gemcitabine 25.8%, clobenpropit 9.7% and combination 40.9%, P = 0.001) by up-regulation of E-cadherin and down-regulation of Zeb1 in Panc-1 xenograft mouse. Clobenpropit enhanced the anti-tumor effect of gemcitabine in pancreatic cancer cells through inhibition of the epithelial-mesenchymal transition process.

Histamine infused into basolateral amygdala enhances memory consolidation of inhibitory avoidance

The role of the basolateral amygdala (BLA) in the consolidation of aversive memory is well established. Here we investigate the involvement of the histaminergic system in BLA on this variable. Rats were chronically implanted with bilateral cannulae in the BLA and after recovery were trained in a one-trial step-down inhibitory avoidance task. Immediately after training histaminergic compounds either alone or in combination were infused through the cannulae. Memory was assessed in test sessions carried out 24 h after the training session. Post-training histamine (1-10 nmol; 0.5 μl/side) enhanced consolidation and the histamine H₃ receptor antagonist thioperamide (50 nmol; 0.5 μl/side) impaired memory consolidation. The effect was shared by the histamine N-methyltransferase inhibitor SKF-91844 (50 nmol; 0.5 μl/side) as well as by the H₃ receptor agonist imetit (10 nmol; 0.5 μl/side). The promnesic action of histamine was unaffected by the H₁ receptor antagonist pyrilamine (50 nmol; 0.5 μl/side). The H1 receptor agonist pyridylethylamine (10 nmol; 0.5 μl/side), the H₂ agonist dimaprit (10 nmol; 0.5 μl/side) and the H₂ antagonist ranitidine (50 nmol; 0.5 μl/side) were ineffective. Histaminergic compounds infused into the BLA had no effect on open-field or elevated plus-maze behaviour. The data show that histamine induces a dose-dependent mnemonic effect in rats and indicate that this reflects a role of endogenous histamine in the BLA mediated by H₃ receptors.

On the role of histamine receptors in regulating pigmentary responses in Oreochromis mossambicus melanophores

Purpose: The present work was carried out to reveal the involvement of histamine receptors at the neuro-melanophore junction of teleost, Oreochromis mossambicus.Methods: The isolated scale melanophores were assayed using the mean melanophore size index and their responses were recorded in presence of various concentrations of histamine along with H1 and H2 receptor specific agonists and antagonist and potentiator compound 48/80.Results: Melanophores showed high sensitivity to histamine and its specific agonists. Histamine caused a dose-dependent pigment aggregation, whereas 2-(2-Pyridyl) ethylamine (PEA), a specific H1R agonist also caused aggregation in a similar manner. Conversely, amthamine, a specific H2R agonist resulted in pigment dispersion. The effects were antagonized by mepyramine; specific H1R antagonist and ranitidine a specific H2R antagonist.Conclusion: It is concluded that O. mossambicus melanophores have both H1 and H2 receptors which mediate melanophore aggregation and dispersion respectively. Compound 48/80 augmented the melanin-aggregating and dispersing effects of PEA and amthamine. It is suggested that the effect of histamine is directly mediated through H1 and H2 receptors, whereas H1Rs may be predominantly involved in the aggregatory responses.

Betahistine for Menière's disease

Betahistine, a histamine H1 agonist and H3 antagonist, has been used for the pharmacotherapy of Meniere's disease for many decades. Earlier its mode of action and efficacy, in particular in low dosages, were the subject of controversy mainly because valid experimental or clinical data were not available. Recent animal studies, however, showed that betahistine increases cochlear blood flow in a sigmoid dose-dependent way, thus supporting the hypothesis that it may act by improving cochlear microcirculation. In terms of its clinical efficacy, two studies demonstrated that a high-dose (at least 48 mg t.i.d.) and long-term treatment (at least six to 12 months) have a significant prophylactic effect on the occurrence of attacks in Meniere's disease.

Involvement of histamine receptors in SAPK/JNK phosphorylation

Histamine is a mediator of inflammation in allergic disease and asthma. Stress activated protein kinases/c-jun N-terminal kinases (SAPK/JNK) are involved in asthma. This study examined the role of histamine receptors on the phosphorylation of SAPK/JNK in splenocytes. C57BL/6 mice splenocytes were treated with histamine (10−4M to 10−11M), and its selective receptor agonists, phorbol 12 myristate 13-acetate (PMA) was used as a positive control, and phosphorylation of SAPK/JNK was determined. Histamine (10−4M–10−8M) inhibited phosphorylation of SAPK/JNK. H1R agonist betahistine (10−5M) decreased SAPK/JNK phosphorylation and H2R agonist amthamine (10−5M) did not show any significant effect. However, H3R agonist methimepip (10−6M) and H4R agonist 4-methyl histamine (10−6M), increased SAPK/JNK phosphorylation.We used TNFα knockout mice to determine if histamine regulated SAPK/JNK phosphorylation via TNFα. While the effects of histamine and H1 agonists were similar to that of wild type mice in inhibiting the phosphorylation of SAPK/JNK, the effects of H3 and H4 agonists differed in TNFα knockout mice splenocytes. Activation of H3 receptors decreased SAPK/JNK phosphorylation in TNFα knockout mice, as opposed to an increase in wild type mice, whereas H4 agonist did not show any significant effect on the phosphorylation of SAPK/JNK. This data showed that histamine acting through H4 receptors caused the phosphorylation of SAPK/JNK via TNFα. The role of H4 receptors in pro-inflammatory response is intriguing.

Development of time-resolved fluorescent based [Eu]-GTP binding assay for selection of human Histamine 3 receptor antagonists/inverse agonist: a potential target for Alzheimer�s treatment

The histamine H3 receptor is an attractive G protein-coupled receptor drug target that regulates neurotransmission in the central nervous system and plays a crucial role in cognitive and homeostatic functions. This receptor exhibits molecular, pharmacological, and functional heterogeneity that affects the preclinical development of effective antagonists. The range of assay technologies like radio isotope based [35S] GTPγS binding assay, luminescent based reporter gene assay (In-direct cAMP measurement) for binding and signaling have been developed in High Throughput Screening (HTS) laboratories for the identification of hit or lead compounds acting on H3 receptor. The [35S] GTPγS binding assay still remains a useful and a simple technique to demonstrate receptor activation and is one of the few functional, cell-free assays that has set the standards in the field of research. However, its radioactive nature imposes clear limitations to its use in regular laboratory practice and in high-throughput experimentation. Herein, we have developed and optimized a membrane based non-radioactive assay using a europium-labeled GTP analogue in which europium-GTP binding can be assayed using time-resolved fluorescence technology. The characterization of H3 agonist or antagonist with HTRF platform has revealed a rank order potency (pEC50 & P K B) comparable to that from isotopic functional studies measured by liquid scintillation counter (LSC). Lastly, the Eu-GTP binding assay has been found to be highly robust (Z' factor 0.84) with high percentage over basal counts. This assay can be utilized as a component of cascade for the screening of H3 receptor ligands.

Docking studies of novel pyrazinopyridoindoles class of antihistamines with the homology modelled H1-receptor

Histamine is an important neurotransmitter as it controls a multitude of physiological functions by activating specific receptors on target cells. It exerts its effects by binding to four different histamine receptors (H1–H4), which all belong to the large family of G protein-coupled receptors (GPCRs). Research and development of H1 ligand has largely focused on antagonists which are used for their anti-allergy effects in the periphery. Recent understanding of the clinical importance of H1 receptors in brain, however, suggests the pharmacotherapeutic potential of H1 agonists in neurodegenerative and neuropsychiatric disorders. Despite the therapeutic importance of the H1 receptor, for many years the molecular features of the H1 receptor protein had been unknown. In view of the recently reported crystal structure of human H1 receptor and in continuation of our work on 3D-pharmacophore on antihistamine H1 and homology model of histamine H1 receptor, docking studies have been carried out on some promising pyrazinopyridoindole class of antihistamine H1, including two outliers, to validate our earlier reported models/hypotheses on H1-receptor, where a good explanation between estimated and observed activities has been obtained. In addition, the docking study also provided insights about the optimal activity of the outliers, for which no explanation was reported previously.