Histamine Stimulation Research Articles

Histamine evokes excitatory response of neurons in the cerebellar dentate nucleus via H2 receptors

Previous studies have shown an excitatory effect of histamine on neurons in two cerebellar nuclei, the fastigial nucleus and the interposed nucleus. Here we investigated action of histamine on the dentate nucleus (DN), another nucleus of the cerebellum, and provided more evidence for motor control by histamine via the cerebellum. Spontaneous unitary discharge of neurons in the DN was extracellularly recorded by use of cerebellar slice preparations. In total 79-recorded neurons, which were from 53 cerebellar slices, 67 neurons (84.8%) had an excitatory response to histamine stimulation, and the rest (15.2%) were not reactive. The histamine-induced excitation of the DN neurons was not blocked by low-Ca2+/high-Mg2+ medium, demonstrating that this effect of histamine was postsynaptic. Triprolidine, an antagonist of histamine H1 receptors, did not block the excitatory effect of histamine, but ranitidine, an antagonist for H2 receptors, blocked the excitatory response to histamine in a concentration-dependent manner. Further, histamine H1 receptor agonist 2-pyridylethylamine did not elicit any response of DN neurons, but H2 receptor agonist dimaprit had an excitatory action on the DN cells and this action was blocked by ranitidine. These results indicate that histamine excites cerebellar DN neurons via histamine H2 receptors. Since the DN receives hypothalamocerebellar histaminergic projections and plays a role in initiation and planning of somatic movement, the postsynaptic excitation of the DN neurons by histamine suggests the possibility that the initiation and planning of movement may be modulated by the histaminergic projections.

Extracellular Ca2+ entry and mobilization of inositol trisphosphate-dependent Ca2+ stores modulate histamine and electrical field stimulation induced contractions of the guinea-pig prostate

This investigation aimed to examine the source of Ca2+ mobilization that leads to the contractile response to either exogenously added histamine (1μM–1mM) or electrical field stimulation (10Hz, 0.5ms, 60V). Removal of extracellular Ca2+ by removal of Ca2+ from the bathing medium reduced histamine (1mM) induced responses by 34% and responses induced by electrical field stimulation by 94%. Similarly, blockade of L-type Ca2+ channels by nifedipine (1μM) reduced histamine (1mM) induced responses by 43% and responses induced by electrical field stimulation by 77%. Application of cyclopiazonic acid (CPA) (10μM) to inhibit Ca2+ reuptake to the sarcoplasmic reticulum enhanced both histamine-induced and electrical field stimulation induced responses to a small degree, while the addition of the inosotol triphosphate (IP3) receptor antagonist, 2-aminophenoxyethane borane (2-APB) (100μM) inhibited histamine induced responses by 70% and electrical field stimulation induced responses by 57%. Ryanodine (1μM) did not affect contractile responses to either histamine or electrical field stimulation, either in the absence or presence of 2-APB (100μM). During both histamine and electrical field stimulation induced contractions, prostate smooth muscle generates IP3 receptor mediated Ca2+ release in conjunction with Ca2+ entry from the extracellular environment. Ryanodine receptors on the other hand, appear not to play a role in this physiological mechanism.

A Study Comparing the Antisecretory Effect of TAK-438, a Novel Potassium-Competitive Acid Blocker, with Lansoprazole in Animals

Proton pump inhibitors (PPIs) are widely used for the treatment of acid-related diseases. However, several medical needs such as suppression of night-time acid secretion and rapid symptom relief remain unmet. In this study, we investigated the effects of 1-[5-(2-fluorophenyl)-1-(pyridin-3-ylsulfonyl)-1H-pyrrol-3-yl]-N-methylmethanamine monofumarate (TAK-438), a novel potassium-competitive acid blocker, on acid secretion in rats and dogs under various conditions, in comparison with the PPI lansoprazole [2-[[[3-methyl-4-(2,2,2-trifluoroethoxy)-2-pyridyl] methyl]sulfinyl]-1H-benzimidazole], to characterize the antisecretory action of TAK-438. TAK-438 showed a more potent and longer-lasting inhibitory effect than lansoprazole on the histamine-stimulated gastric acid secretion in rats and dogs. A pharmacokinetic study in rats showed that TAK-438 accumulated and was retained in the gastric tissue for more than 24 h, unlike that in the plasma. TAK-438 showed significant antisecretory activity with or without cimetidine pretreatment, in contrast to lansoprazole, which did not show antisecretory activity after cimetidine pretreatment in rats. TAK-438 increased the pH of the gastric perfusate to 5.7 in an unstimulated condition, and this effect was maintained in the presence of subsequent histamine stimulation. On the other hand, lansoprazole also increased the pH in an unstimulated condition, but this effect diminished after histamine stimulation. These results indicated that TAK-438 exerted a more potent and longer-lasting antisecretory effect than lansoprazole through high accumulation and slow clearance from the gastric tissue. In addition, TAK-438 was unaffected by the gastric secretory state, unlike PPIs. Therefore, TAK-438 can provide a novel mechanism of action to improve the present PPI-based treatment of acid-related diseases.

ENOS activation mediated by AMPK after stimulation of endothelial cells with histamine or thrombin is dependent on LKB1

Reports on the role of AMP-activated protein kinase (AMPK) in thrombin-mediated activation of endothelial nitric-oxide synthase (eNOS) in endothelial cells have been conflicting. Previously, we have shown that under culture conditions that allow reduction of ATP-levels after stimulation, activation of AMPK contributes to eNOS phosphorylation and activation in endothelial cells after treatment with thrombin. In this paper we examined the signaling pathways mediating phosphorylation and activation of eNOS after stimulation of cultured human umbilical vein endothelial cells (HUVEC) with histamine and the role of LKB1-AMPK in the signaling. In Morgan's medium 199 intracellular ATP was lowered by treatment with histamine or the ionophore A23187 while in medium RMPI 1640 ATP was unchanged after identical treatment. In medium 199 inhibition of Ca + 2 /CaM kinase kinase (CaMKK) by STO-609 only partially inhibited AMPK phosphorylation but after gene silencing of LKB1 with siRNA there was a total inhibition of AMPK phosphorylation by STO-609 after treatment with either histamine or thrombin, demonstrating phosphorylation of AMPK by both upstream kinases, LKB1 and CaMKK. Downregulation of AMPK with siRNA partially inhibited eNOS phosphorylation caused by histamine in cells maintained in medium 199. Downregulation of LKB1 by siRNA inhibited both phosphorylation and activity of eNOS and addition of the AMPK inhibitor Compound C had no further effect on eNOS phosphorylation. When experiments were carried out in medium 1640, STO-609 totally prevented the phosphorylation of AMPK without affecting eNOS phosphorylation. AMPKα2 downregulation resulted in a loss of the integrity of the endothelial monolayer and increased expression of GRP78, indicative of endoplasmic reticular (ER) stress. Downregulation of AMPKα1 had no such effect. The results show that culture conditions affect endothelial signal transduction pathways after histamine stimulation. Under conditions where intracellular ATP is lowered by histamine, AMPK is activated by both LKB1 and CaMKK and, in turn, mediates eNOS phosphorylation in an LKB1 dependent manner. Both AMPKα1 and − α2 are involved in the signaling. Under conditions where intracellular ATP is unchanged after histamine treatment, CaMKK alone activates AMPK and eNOS is phosphorylated and activated independent of AMPK.

A Role for the Ca 2+ Channel TRPML1 in Gastric Acid Secretion, Based on Analysis of Knockout Mice

Mutations in TRPML1, a lysosomal Ca(2+)-permeable TRP channel, lead to mucolipidosis type IV, a neurodegenerative lysosomal storage disease. An unusual feature of mucolipidosis type IV is constitutive achlorhydria. We produced Trpml1(-/-) (null) mice to investigate the requirement for this protein in gastric acid secretion. Trpml1-null mice were generated by gene targeting. The expression of Trpml1 and its role in acid secretion by gastric parietal cells were analyzed using biochemical, histologic, and ultrastructural approaches. Trpml1 is expressed by parietal cells and localizes predominantly to the lysosomes; it was dynamically palmitoylated and dephosphorylated in vivo following histamine stimulation of acid secretion. Trpml1-null mice had significant impairments in basal and histamine-stimulated gastric acid secretion and markedly reduced levels of the gastric proton pump. Histologic and ultrastructural analyses revealed that Trpml1(-/-) parietal cells were enlarged, had multivesicular and multi-lamellated lysosomes, and maintained an abnormal intracellular canalicular membrane. The intralysosomal Ca(2+) content and receptor-mediated Ca(2+) signaling were, however, unaffected in Trpml1(-/-) gastric glands, indicating that Trpml1 does not function in the regulation of lysosomal Ca(2+). Loss of Trpml1 causes reduced levels and mislocalization of the gastric proton pump and alters the secretory canaliculi, causing hypochlorhydria and hypergastrinemia. The lysosomal enlargement and defective intracellular canaliculi formation observed in Trpml1(-/-) parietal cells indicate that Trpml1 functions in the formation and trafficking of the tubulovesicles. This study provides direct evidence for the regulation of gastric acid secretion by a TRP channel; TRPML1 is an important protein in parietal cell apical membrane trafficking.

Phospho-regulated ACAP4-Ezrin Interaction Is Essential for Histamine-stimulated Parietal Cell Secretion

The ezrin-radixin-moesin proteins provide a regulated linkage between membrane proteins and the cortical cytoskeleton and also participate in signal transduction pathways. Ezrin is localized to the apical membrane of parietal cells and couples the protein kinase A activation cascade to the regulated HCl secretion. Our recent proteomic study revealed a protein complex of ezrin-ACAP4-ARF6 essential for volatile membrane remodeling (Fang, Z., Miao, Y., Ding, X., Deng, H., Liu, S., Wang, F., Zhou, R., Watson, C., Fu, C., Hu, Q., Lillard, J. W., Jr., Powell, M., Chen, Y., Forte, J. G., and Yao, X. (2006) Mol. Cell Proteomics 5, 1437-1449). However, knowledge of whether ACAP4 physically interacts with ezrin and how their interaction is integrated into membrane-cytoskeletal remodeling has remained elusive. Here we provide the first evidence that ezrin interacts with ACAP4 in a protein kinase A-mediated phosphorylation-dependent manner through the N-terminal 400 amino acids of ACAP4. ACAP4 locates in the cytoplasmic membrane in resting parietal cells but translocates to the apical plasma membrane upon histamine stimulation. ACAP4 was precipitated with ezrin from secreting but not resting parietal cell lysates, suggesting a phospho-regulated interaction. Indeed, this interaction is abolished by phosphatase treatment and validated by an in vitro reconstitution assay using phospho-mimicking ezrin(S66D). Importantly, ezrin specifies the apical distribution of ACAP4 in secreting parietal cells because either suppression of ezrin or overexpression of non-phosphorylatable ezrin prevents the apical localization of ACAP4. In addition, overexpressing GTPase-activating protein-deficient ACAP4 results in an inhibition of apical membrane-cytoskeletal remodeling and gastric acid secretion. Taken together, these results define a novel molecular mechanism linking ACAP4-ezrin interaction to polarized epithelial secretion.

Electrophysiological characterization of store-operated and agonist-induced Ca2+ entry pathways in endothelial cells

In endothelial cells, agonist-induced Ca(2+) entry takes place via the store-operated Ca(2+) entry pathway and/or via channel(s) gated by second messengers. As cell stimulation leads to both a partial Ca(2+) store depletion as well as the production of second messengers, these two pathways are problematic to distinguish. We showed that passive endoplasmic reticulum (ER) depletion by thapsigargin or cell stimulation by histamine activated a similar Ca(2+)-release-activated Ca(2+) current (CRAC)-like current when 10 mM Ba(2+)/2 mM Ca(2+) was present in the extracellular solution. Importantly, during voltage clamp recordings, histamine stimulation largely depleted the ER Ca(2+) store, explaining the activation of a CRAC-like current (due to store depletion) upon histamine in Ba(2+) medium. On the contrary, in the presence of 10 mM Ca(2+), the ER Ca(2+) content remained elevated, and histamine induced an outward rectifying current that was inhibited by Ni(2+) and KB-R7943, two blockers of the Na(+)/Ca(2+) exchanger (NCX). Both blockers also reduced histamine-induced cytosolic Ca(2+) elevation. In addition, removing extracellular Na(+) increased the current amplitude which is in line with a current supported by the NCX. These data are consistent with the involvement of the NCX working in reverse mode (Na(+) out/Ca(2+) in) during agonist-induced Ca(2+) entry in endothelial cells.

Enhancement of inflammatory mediator release by β2‐adrenoceptor agonists in airway epithelial cells is reversed by glucocorticoid action

Due to their potent bronchodilator properties, beta(2)-adrenoceptor agonists are a mainstay of therapy in asthma. However, the effects of beta(2)-adrenoceptor agonists on inflammation are less clear. Accordingly, we have investigated the effects of beta(2)-adrenoceptor agonists on inflammatory mediator release. Transcription factor activation, and both release and mRNA expression of IL-6 and IL-8 were examined by luciferase reporter assay, elisa and real-time RT-PCR in bronchial human epithelial BEAS-2B cells or primary human bronchial epithelial cells grown at an air-liquid interface. Pre-incubation with beta(2)-adrenoceptor agonists (salbutamol, salmeterol, formoterol) augmented the release and mRNA expression of IL-6 and IL-8 induced by IL-1beta and IL-1beta plus histamine, whereas NF-kappaB-dependent transcription was significantly repressed, and AP-1-dependent transcription was unaffected. These effects were mimicked by other cAMP-elevating agents (PGE(2), forskolin). Enhancement of cytokine release by beta(2)-adrenoceptor agonists also occurred in primary bronchial epithelial cells. Addition of dexamethasone with salmeterol repressed IL-6 and IL-8 release to levels that were similar to the repression achieved in the absence of salmeterol. IL-6 release was enhanced when salmeterol was added before, concurrently or after IL-1beta plus histamine stimulation, whereas IL-8 release was only enhanced by salmeterol addition prior to stimulation. Enhancement of IL-6 and IL-8 release may contribute to the deleterious effects of beta(2)-adrenoceptor agonists in asthma. As increased inflammatory mediator expression is prevented by the addition of glucocorticoid to the beta(2)-adrenoceptor, our data provide further mechanistic support for the use of combination therapies in asthma management.

Histamine induces Th2 activation through the histamine receptor 1 in house dust mite rhinitic but not asthmatic patients

Effects of mast cell-released histamine on smooth muscle and endothelial cells are considered as responsible of immediate symptoms of anaphylaxis. However, little is known about histamine effects on Th2 lymphocytes, which orchestrate the allergic reaction upstream of mast cells. We addressed this question in house dust mite (HDM) allergics, according to the presence of rhinitis or asthma and allergen stimulation. Peripheral blood mononuclear cell from 15 rhinitic and 14 asthmatic HDM-allergic subjects and 16 controls were cultured with Der p 1 or histamine. The effect of Der p 1 on histamine receptor (H1R and H2R) expression was studied. T-cell cytokine production was studied upon Der p 1 or histamine stimulation. The role of H1R in histamine effects was assessed with levocetirizine. H1R and H2R are overexpressed on T cells from asthmatic but not from rhinitic subjects. Der p 1 increases H1R expression on CD4(+) cells from both allergic groups, and decreases it in controls, on CD4(+) and CD8(+) subsets. Der p 1 decreases T-cell H2R expression in asthmatics. Allergen increases IL-4 and IL-13 in both allergic groups. Histamine increases Th2 cytokines in rhinitics only, and levocetirizine abolishes this effect. In asthmatics and controls, histamine decreases T-cell cytokines through a non-H1R dependent pathway. In rhinitis but not in asthma, histamine is able to increase allergic inflammation by increasing Th2 cytokine production in a positive feedback dependent on H1R. This result could explain in part why H1R antagonists, are very efficient in rhinitis, but not in asthma.

Functional Identification of Neural Stem Cell–Derived Oligodendrocytes by Means of Calcium Transients Elicited by Thrombin

Current immunosuppressive treatments for central nervous system demyelinating diseases fail to prevent long-term motor and cognitive decline in patients. Excitingly, glial cell transplantation arises as a promising complementary strategy to challenge oligodendrocytes loss occurring in myelination disorders. A potential source of new oligodendrocytes is the subventricular zone (SVZ) pool of multipotent neural stem cells. However, this approach has been handicapped by the lack of functional methods for identification and pharmacological analysis of differentiating oligodendrocytes, prior to transplantation. In this study, we questioned whether SVZ-derived oligodendrocytes could be functionally discriminated due to intracellular calcium level ([Ca(2+)](i)) variations following KCl, histamine, and thrombin stimulations. Previously, we have shown that SVZ-derived neurons and immature cells can be discriminated on the basis of their selective [Ca(2+)](i) rise upon KCl and histamine stimulation, respectively. Herein, we demonstrate that O4+ and proteolipid protein-positive (PLP+) oligodendrocytes do not respond to these stimuli, but display a robust [Ca(2+)](i) rise following thrombin stimulation, whereas other cell types are thrombin-insensitive. Thrombin-induced Ca(2+) increase in oligodendrocytes is mediated by protease-activated receptor-1 (PAR-1) activation and downstream signaling through G(q/11) and phospholipase C (PLC), resulting in Ca(2+) recruitment from intracellular compartments. This method allows the analysis of functional properties of oligodendrocytes in living SVZ cultures, which is of major interest for the development of effective grafting strategies in the demyelinated brain. Additionally, it opens new perspectives for the search of new pro-oligodendrogenic factors to be used prior grafting.

Endothelial cell ADAMTS-13 and VWF: production, release, and VWF string cleavage

AbstractHuman umbilical vein endothelial cell (HUVEC)–released ADAMTS-13 (a disintegrin and metalloprotease with thrombospondin repeats) and HUVEC-secreted von Willebrand factor (VWF) strings were investigated under static conditions that allow the accumulation and analysis of ADAMTS-13. The latter was released constitutively from HUVECs and cleaved the secreted and cell-anchored VWF strings progressively during 15 minutes in Ca2+/Zn2+-containing buffer. HUVEC ADAMTS13 mRNA expression was approximately 1:100 of VWF monomeric subunit expression. In contrast to multimeric VWF stored within Weibel-Palade bodies and secreted rapidly in response to cell stimulation, ADAMTS-13 was released directly from the Golgi to the cell exterior without an organelle storage site. The constitutive release of ADAMTS-13 continued at the same slow rate regardless of the presence or absence of histamine stimulation of HUVECs. Consequently, the percentage of VWF strings cleaved by ADAMTS-13 at VWF Y1605-M1606 decreased as the rate of VWF string secretion was increased by cell stimulation. Blockade of HUVEC ADAMTS-13 activity by antibodies to different ADAMTS-13 domains made it possible to detect the attachment of ADAMTS-13 all along the lengths of HUVEC-secreted VWF strings. Constitutive ADAMTS-13 released from endothelial cells may contribute to the maintenance of cell surfaces free of hyperadhesive VWF multimeric strings.

Nerve growth factor and receptors are significantly affected by histamine stimulus through H1 receptor in pancreatic carcinoma cells

Nerve growth factor (NGF) as an autocrine or paracrine growth factor plays a critical role in the pathogenesis and progression of human pancreatic cancer. NGF is synthesized as a proform (proNGF) that, when cleaved, releases mature ligand (mNGF). proNGF and mNGF bind to high-affinity tyrosine kinase receptor A (TrkA) and low-affinity receptor p75 to different extents. Histamine is a potent stimulator of NGF in the inflammatory lesion as determined by ELISA. This has generally been attributed to the accumulation of mNGF. To determine the effect of histamine on nerve growth factor/receptor expression in human pancreatic cancer, the present study explored intracellular and extracellular NGF production and p75 and TrkA membrane receptor expression in the PANC-1, KMP-6 and PK-1 cell lines. Histamine enhanced NGF secretion and mRNA expression in PANC-1 and KMP-6 cells, but not in PK-1 cells. proNGF was revealed using Western blotting to be the predominant form of NGF, but was significantly reduced by histamine. p75 receptor binding was increased with histamine treatement, but no significant alteration was observed for TrkA. Proliferating cell nuclear antigen (PCNA), an important indicator of cell proliferation, was significantly reduced by histamine stimulation. H1 and H2 receptors were both observed in the pancreatic cancer cells, and the alterations induced by histamine were counteracted by H1 receptor antagonist pyrilamine; however, the H2 receptor subtype was excluded from this process. These results suggest that histamine induces distinct nerve growth factor/receptor responsiveness via H1 receptor-induced signaling, thus affecting pancreatic cancer cell proliferation.

Histamine Stimulates Interleukin-6 Production through Histamine H1 Receptors in Human Amnion Cells

Background/Aims: Previous studies have stated that maternal allergic diseases are associated with increased risk of preterm labor/delivery, but the underlying mechanisms remain unclear. This study tested the hypothesis that histamine induces interleukin (IL)-6 production in amnion cells. Methods: Using cultured human amnion cells, we examined expression of histamine receptors and effects of histamine on IL-6 production. Results: Reverse transcription-polymerase chain reaction and Western blotting revealed expression of histamine H1 receptor (H1R) and H2 receptor (H2R) in human amnion. Histamine stimulation significantly increased concentrations of IL-6 in conditioned medium, as did tumor necrosis factor-α and IL-1β in positive controls. In addition, the H1R antagonist olopatadine significantly blocked histamine-induced production of IL-6, whereas the H2R antagonist ranitidine did not. Conclusion: Histamine appears to induce IL-6 production through H1R in human amnion cells.

A microfluidic chip for permeability assays of endothelial monolayer

Endothelial cell monolayer (EM), acting as a barrier between blood and tissue, plays an important role in pathophysiological processes. Here we describe a novel microfluidic chip that is applied for convenient and high throughput in vitro permeability assays of EM. The chip included a gradient generator and an array of cell culture chambers. A microporous membrane as a scaffold component was built between a polydimethylsiloxane (PDMS) layer and a glass substrate to grow EM. Cell culture chambers were separated by microchannels and microvalves. The concentration gradient of compound solutions could be generated automatically and affected EM in different chambers. The permeability of EM at different time with histamine stimulation was in situ measured by the fluorescence detection of the leaked tracer. The existence of continuous flow in the channels allowed EM in a dynamic microenvironment and increased the amount of tracer through the EM, comparing to transwell assays. According to the prototype chip, the chip with a bigger array of cell culture chambers could be achieved easily and applied in the high throughput screening for drugs.

Opposing regulation of histamine-induced calcium signaling by sodium selenite and ebselen via alterations of thiol redox status

Elevated blood histamine plays a role in the pathogenesis of atherosclerosis. Calcium signaling mediates histamine action in endothelial cells. Selenium (Se) is a dietary essential trace element for humans. Se compounds in different oxidation states were found to exhibit an opposing effect on the histamine-induced calcium signaling in the ECV304 cell line. When Se in the form of sodium selenite was added in the cell culture, the reactivity of the histamine H 1-receptor was increased as reported in our previous paper. We here show that as a culture supplement, sodium selenite enhanced the activity of selenoprotein thioredoxin reductase (TrxR) and the calcium response to histamine stimulation, which were reversed by treating the cells with gold thioglucose, a nucleophilic drug that selectively modifies thiolate/selenolate groups. Sodium selenite most likely caused a reductive shift in the thiol/disulfide redox balance through increasing TrxR activity. In contrast, when the cells were treated with Se in the form of ebselen, a thiol oxidant with peroxidase-like activity, histamine-induced calcium release and calcium entry were significantly suppressed. This effect appeared related to the thiol-directed modification rather than the peroxidase-like activity of ebselen, because this inhibitory effect was not replicated by increasing cellular peroxidase activity. Thus, the opposing effects of sodium selenite and ebselen on histamine-induced calcium signaling are achieved, at least in part, through their opposite actions in modulating the thiol/disulfide redox state.

Activation of the histaminergic H3 receptor induces phosphorylation of the Akt/GSK‐3β pathway in cultured cortical neurons and protects against neurotoxic insults

Stimulation of histamine H(3) receptors (H(3)R) activates G(i/o)-proteins that inhibit adenylyl cyclase and triggers MAPK and phospholipase A(2). In a previous study, we showed that H(3)R-mediated phosphorylation of Akt at Ser473 occurs in primary cultures of rat cortical neurons, but neither the downstream targets nor the function of such activation were explored. In this report we address these questions. Western blotting experiments showed that H(3)R-mediated activation of Akt in cultured rat cortical neurons was inhibited by LY 294004 and U0126, suggesting that it depends on phosphoinositide-3-kinase and mitogen-activated protein kinase kinase. H(3)R activation phosphorylated, hence inactivated, the Akt downstream effector glycogen synthase kinase-3beta, increased the expression of the antiapoptotic protein Bcl-2 and protected cultured rat and mouse cortical neurons from neurotoxic insults in a dose-dependent manner. All these effects were inhibited by the H(3)R antagonist inverse/agonist thioperamide. Mouse cortical cells expressed H(3)R as revealed by immunostaining experiments, and stimulation of H(3)R phoshorylated Akt and decreased caspase 3 activity. Hence, we uncovered a yet unexplored action of the H(3)R that may help understand the impact of H(3)R signaling in the CNS.

Research on morbidity and relative factors of cough variant asthma among patients with chronic cough syndrome

To study the morbidity of cough variant asthma (CVA) among patients with chronic cough syndrome and its relative risk factors. Patients were recruited with detailed history on their illness. Data were collected on physical examination, chest X-ray, eosinophil cell counts, pulmonary ventilation with histamine stimulating test and bronchi dilation test. According to available data, diagnosis of CVA was confirmed and the relative factors Questionnaire form was completed for each patient. Among 473 patients with chronic cough, 95 (44 male and 51 female) were confirmed to be CVA (20.08%). Analysis of the relative factors suggested that CVA was associated with multiple factors. Morbidity of CVA was associated with season, personal histories on allergy and family history on asthma. CVA could be induced by upper respiratory tract infection, inhale of oil vapor, acrimony air, over-burdened physical exercises etc. For patients with chronic cough symptom, clear diagnosis of CVA, avoid of passable risk factors and timely medical intervention when necessary, would be helpful in controlling clinical courses and improving the prognosis of the disease.

Differential role of β‐catenin in VEGF and histamine‐induced MMP‐2 production in microvascular endothelial cells

Increases in endothelial cell permeability and production of matrix-degrading enzymes are two early steps in the angiogenic process. Factors such as vascular endothelial growth factor (VEGF) and histamine induce the angiogenic process through alterations in both permeability and proteolysis. We hypothesized that beta-catenin acts as a positive regulator of MMP-2 and MT1-MMP transcription following VEGF or histamine stimulation. Rat microvascular endothelial cells were exposed to VEGF or histamine overnight and MMP-2 protein production was assessed by gelatin zymography. Latent MMP-2 protein levels were increased following VEGF and histamine treatment as were MMP-2 mRNA transcript levels. Endothelial cells exposed to VEGF and histamine had an increased level of nuclear beta-catenin, which was sensitive to inhibition of the PI3-kinase signaling pathway. Promoter assays indicated increased transcriptional activity of both MMP-2 and MT1-MMP in endothelial cells co-transfected with luciferase reporter constructs and beta-catenin. Inhibition of beta-catenin signaling with inhibitor of catenin and T cell factor (ICAT) revealed that the VEGF-induced increase in MMP-2 mRNA is beta-catenin dependent. Interestingly, while MMP-2 mRNA levels increased in response to histamine H1 or H2 receptor activation, significantly larger increases were observed in cells co-treated with ICAT and histamine or the histamine receptor agonists, HTMT and dimaprit. While both VEGF and histamine increase nuclear beta-catenin and MMP-2 production, the role of beta-catenin in MMP-2 regulation differs between the two stimuli.

Extraction of membrane cholesterol disrupts caveolae and impairs serotonergic (5-HT2A) and histaminergic (H1) responses in bovine airway smooth muscle: role of Rho-kinase

Some receptors and signaling molecules, such as Rho-kinase (ROCK), localize in caveolae. We asked whether the function of histamine receptors (H(1)) and 5-hydroxytryptamine (serotonin) receptors (5-HT(2A)) in bovine tracheal smooth muscle are modified after caveolae disruption and if so, whether the altered ROCK activity plays a role in this modification. Methyl-beta-cyclodextrin (MbetaCD), used to deplete membrane cholesterol, was shown to disrupt caveolae and diminish sustained contractions to histamine (approximately 80%), 5-HT (100%), alpha-methyl-5-HT (100%), and KCl (approximately 30%). Cholesterol-loaded MbetaCD (CL-MbetaCD) restored the responses to KCl and partially restored the responses to agonists. ROCK inhibition by Y-27632 diminished contractions to histamine (approximately 85%) and 5-HT (approximately 59%). 5-HT or histamine stimulation augmented ROCK activity. These increases were reduced by MbetaCD and partially reestablished by CL-MbetaCD. The increase in intracellular Ca(2+) that was induced by both agonists was reduced by MbetaCD. The presence of caveolin-1 (Cav-1), H1, 5-HT(2A), and ROCK1 was corroborated by immunoblotting of membrane fractions from sucrose gradients and by confocal microscopy. H(1) receptors coimmunoprecipitated with Cav-1 in caveolar and noncaveolar membrane fractions, whereas 5-HT(2A) receptors appeared to be restricted to noncaveolar membrane fractions. We conclude that caveolar and cholesterol integrity are indispensable for the proper functionality of the H(1) and 5-HT(2A) receptors through their Rho/ROCK signaling.

Histamine iontophoresis on the viability of random skin flap in rats

To evaluate the effects of the histamine iontophoresis on the random skin flap viability in rats. Sixty adult male Wistar rats were used. A cranially-based dorsal skin flap measuring 10 x 4 cm was raised and a plastic barrier was placed between the flap and its bed. After the surgical procedure, the animals were randomized into four groups (G1-G4) (n=15 each group) as follows: G1 (control)--sham electrical stimulation, G2 (electrical stimulation)--direct current electrical stimulation, G3 (histamine)--histamine and sham electrical stimulation, and G4 (histamine iontophoresis)--transdermal iontophoresis of histamine. In all groups the procedures were performed immediately after the surgery and on the two subsequent days. The percentage of flap necrosis was measured on the seventh postoperative day. The mean and the respective standard deviation of the percentage of flap necrosis areas were as follows: G1 (control) - 47.87 +/- 9.13%, G2 - 51.49 +/- 8.19%, G3 - 46.33 +/- 8.32% and G4 - 30.82 +/- 11.25%. The G4 group presented a significantly smaller amount of flap necrosis when compared to the other groups (p<0.001). The topical administration of the histamine by iontophoresis was effective to increase the viability of the random skin flaps in rats.