bluuebunny/crossref_metadata_2025 huggingface dataset:
2‐Furoyl‐LIGRL‐NH2, a potent agonist for proteinase‐activated receptor‐2, as a gastric mucosal cytoprotective agent in mice
Authors: Kawabata Atsufumi , Oono Yuko , Yonezawa Daiki , Hiramatsu Kaori , Inoi Naoki , Sekiguchi Fumiko , Honjo Masami , Hirofuchi Michiko , Kanke Toru , Ishiwata Hiroyuki
Published: January 2005

Proteinase‐activated receptor‐2 (PAR2), expressed in capsaicin‐sensitive sensory neurons, plays a protective role in gastric mucosa. The present study evaluated gastric mucosal cytoprotective effect of 2‐furoyl‐LIGRL‐NH2, a novel highly potent PAR2 agonist, in ddY mice and in wild‐type and PAR2‐knockout mice of C57BL/6 background.

Gastric mucosal injury was created by oral administration of HCl/ethanol solution in the mice. The native PAR2‐activating peptide SLIGRL‐NH2, administered intraperitoneally (i.p.) at 0.3–1 μmol kg−1 in combination with amastatin, an aminopeptidase inhibitor, but not alone, revealed gastric mucosal protection in ddY mice, which was abolished by ablation of capsaicin‐sensitive sensory neurons.

I.p. administration of 2‐furoyl‐LIGRL‐NH2 at 0.1 μmol kg−1, without combined treatment with amastatin, exhibited gastric mucosal cytoprotective activity in ddY mice, the potency being much greater than SLIGRL‐NH2 in combination with amastatin. This effect was also inhibited by capsaicin pretreatment.

Oral administration of 2‐furoyl‐LIGRL‐NH2 at 0.003–0.03 μmol kg−1 also protected against gastric mucosal lesion in a capsaicin‐reversible manner in ddY mice.

I.p. 2‐furoyl‐LIGRL‐NH2 at 0.1–0.3 μmol kg−1 caused prompt salivation in anesthetized mice, whereas its oral administration at 0.003–1 μmol kg−1 was incapable of eliciting salivation.

In wild‐type, but not PAR2‐knockout, mice of C57BL/6 background, i.p. administration of 2‐furoyl‐LIGRL‐NH2 caused gastric mucosal protection.

Thus, 2‐furoyl‐LIGRL‐NH2 is considered a potent and orally available gastric mucosal protective agent. Our data also substantiate a role for PAR2 in gastric mucosal protection and the selective nature of 2‐furoyl‐LIGRL‐NH2.

British Journal of Pharmacology (2005) 144, 212–219. doi:10.1038/sj.bjp.0706059
Alpha‐2 adrenoceptor subtypes: are more better?
Authors: Bylund David B
Published: January 2005

The discovery of an additional duplicated alpha‐2 adrenoceptor subtype in the zebrafish raises a pesky nomenclature issue, as well as questions about the functions of the alpha‐2 adrenoceptors in the zebrafish and how many alpha‐2 receptors does an organism really need.

British Journal of Pharmacology (2005) 144, 159–160. doi:10.1038/sj.bjp.0706060
LY294002 and rapamycin co‐operate to inhibit T‐cell proliferation
Authors: Breslin Elaine M , White Paul C , Shore Angharad M , Clement Mathew , Brennan Paul
Published: January 2005

T‐cell proliferation is critical for mounting an effective adaptive immune response. It is regulated by signals through the T‐cell receptor, through co‐stimulation and through cytokines such as interleukin‐2 (IL‐2). Phosphatidylinositol 3‐kinase (PI3K) lies downstream of each of these pathways and has been directly implicated in the regulation of lymphocyte proliferation.

In this study, we have shown that PI3K regulates cyclin D2 and cyclin D3, the first cell cycle proteins induced in T‐cell proliferation, transcriptionally and post‐transcriptionally. In T‐lymphoblasts, LY294002, a PI3K inhibitor, prevents the induction of both D‐type cyclin mRNA and protein, while rapamycin inhibits the induction of protein. Rapamycin inhibits mammalian target of rapamycin (mTOR), which lies downstream of PI3K.

Furthermore, our data show that the combination of LY294002 and rapamycin results in a co‐operative inhibition of T‐cell proliferation. This co‐operation occurs in Kit225 cells stimulated with IL‐2, and also in resting peripheral blood lymphocytes stimulated with antibodies to the T‐cell receptor in the presence and absence of antibodies to CD28.

These data indicate that PI3K regulates T‐cell proliferation in response to diverse stimuli, and suggest that combinations of inhibitors, perhaps isoform‐selective, may be useful as alternative immunosuppressive therapies.

British Journal of Pharmacology (2005) 144, 791–800. doi:10.1038/sj.bjp.0706061
The pore‐forming subunit of the KATP channel is an important molecular target for LPS‐induced vascular hyporeactivity in vitro
Authors: O'Brien Alastair J , Thakur Gita , Buckley James F , Singer Mervyn , Clapp Lucie H
Published: January 2005

ATP‐sensitive K+ (KATP) channel activation is implicated in the vascular hyporeactivity occurring in septic shock. However, channel inhibition with the sulphonylurea receptor (SUR) antagonist, glibenclamide (Glib) fails to reverse lipopolysaccharide (LPS)‐induced vascular hyporeactivity in vitro. We investigated whether inhibitors that act by binding to the KATP channel pore could be effective.

Ring segments of endothelium‐intact rat mesenteric artery were incubated with LPS in culture media for either 6 or 20 h before contractile responses to phenylephrine were assessed in the absence or presence of KATP channel inhibitors.

The pore‐forming subunit inhibitors barium chloride (BaCl2; 300 μM) and PNU‐37883A (1 μM) significantly reversed hyporeactivity at both time points, although less so at 20 h. In contrast, the SUR inhibitors, Glib (10 μM), tolbutamide (Tolb) (1 mM) and PNU‐99963 (1 μM) were ineffective. In LPS‐incubated tissues, Glib and Tolb antagonised contractions to the thromboxane A2 mimetic, U46619 (9,11‐dideoxy‐9α, 11α‐methanoepoxy prostaglandin F2α) (10−7 M), whereas the pinacidil‐derived inhibitor, PNU‐99963, did not.

Contractions to 60 mM KCl were unaffected by LPS at 6 h, but were significantly depressed by LPS at 20 h, suggesting that K+‐channel‐independent pathways contribute to hyporeactivity at the later time point.

The inducible nitric oxide synthase (iNOS) inhibitor, 1400 W (10 μM) and Tolb inhibited the production of nitrite induced by LPS, whereas BaCl2 and PNU‐37883A had no effect.

In conclusion, KATP channels contribute to LPS‐induced vascular hyporeactivity via the iNOS pathway in rat mesenteric artery. The effectiveness of pore inhibitors over SUR inhibitors of the KATP channel suggests altered SUR function following LPS administration, which cannot be explained by thromboxane receptor inhibition.

British Journal of Pharmacology (2005) 144, 367–375. doi:10.1038/sj.bjp.0706065
TNFα reduces tachykinin, PGE2‐dependent, relaxation of the cultured mouse trachea by increasing the activity of COX‐2
Authors: Bachar Ofir , Rose Andrew C , Adner Mikael , Wang Xudong , Prendergast Clodagh E , Kempf Ashley , Shankley Nigel P , Cardell Lars‐Olaf
Published: January 2005

Chronic inflammation is a central feature of asthma. The inflammatory cytokine tumour necrosis factor α (TNFα) has been implicated in this disease, and is known to alter airway smooth muscle functionally.

The aim of this study was to investigate the influence of TNFα on tachykinin‐induced airway relaxation. Mouse tracheae were cultured in the absence and presence of TNFα for 1 or 4 days.

In the absence of TNFα, substance P (SP) and neurokinin A (NKA) induced comparable levels of relaxation in fresh and cultured segments. Functional studies with selective antagonists/inhibitors indicated that the relaxation was mediated by the NK1 receptor coupled to cyclooxygenase (COX)‐2 activation and subsequent release of prostaglandin E2 (PGE2). TNFα attenuated SP‐ and NKA‐induced relaxation in a time‐ and concentration‐dependent manner, decreasing the ability of PGE2 to relax tissues.

Further studies indicated that TNFα elevated COX‐2 activity and that concomitant inhibition of COX‐2 reversed TNFα‐attenuated PGE2 relaxation. Culture with PGE2 decreased SP‐ and PGE2‐mediated relaxation, further implicating the activity of COX‐2 in the attenuation of tachykinin signalling.

Gene expression analysis demonstrated that TNFα increased the expression of smooth muscle COX‐2, PGE2 synthase and EP2 receptor mRNA, and decreased the expression of the EP4 receptor.

Overall, these results show that NK1 receptor‐mediated relaxation induced by PGE2 is attenuated by prolonged TNFα stimulation. Increased COX‐2 activity induced by TNFα appears to be central to this process.

British Journal of Pharmacology (2005) 144, 220–230. doi:10.1038/sj.bjp.0706067
Malonate induces cell death via mitochondrial potential collapse and delayed swelling through an ROS‐dependent pathway
Authors: Fernandez‐Gomez Francisco J , Galindo Maria F , Gómez‐Lázaro Maria , Yuste Victor J , Comella Joan X , Aguirre Norberto , Jordán Joaquín
Published: January 2005

Herein we study the effects of the mitochondrial complex II inhibitor malonate on its primary target, the mitochondrion.

Malonate induces mitochondrial potential collapse, mitochondrial swelling, cytochrome c (Cyt c) release and depletes glutathione (GSH) and nicotinamide adenine dinucleotide coenzyme (NAD(P)H) stores in brain‐isolated mitochondria.

Although, mitochondrial potential collapse was almost immediate after malonate addition, mitochondrial swelling was not evident before 15 min of drug presence. This latter effect was blocked by cyclosporin A (CSA), Ruthenium Red (RR), magnesium, catalase, GSH and vitamin E.

Malonate added to SH‐SY5Y cell cultures produced a marked loss of cell viability together with the release of Cyt c and depletion of GSH and NAD(P)H concentrations. All these effects were not apparent in SH‐SY5Y cells overexpressing Bcl‐xL.

When GSH concentrations were lowered with buthionine sulphoximine, cytoprotection afforded by Bcl‐xL overexpression was not evident anymore.

Taken together, all these data suggest that malonate causes a rapid mitochondrial potential collapse and reactive oxygen species production that overwhelms mitochondrial antioxidant capacity and leads to mitochondrial swelling. Further permeability transition pore opening and the subsequent release of proapoptotic factors such as Cyt c could therefore be, at least in part, responsible for malonate‐induced toxicity.

British Journal of Pharmacology (2005) 144, 528–537. doi:10.1038/sj.bjp.0706069
Prediction of the risk of Torsade de Pointes using the model of isolated canine Purkinje fibres
Authors: Champeroux Pascal , Viaud Karen , El Amrani Abdel Ilah , Fowler John Sinclair Lawrence , Martel Eric , Le Guennec Jean‐Yves , Richard Serge
Published: January 2005

Torsade de Pointes (TdP) is a well‐described major risk associated with various kinds of drugs. However, prediction of this risk is still uncertain both in preclinical and clinical trials. We tested 45 reference compounds on the model of isolated canine Purkinje fibres. Of them, 22 are clearly associated and/or labelled with a risk of TdP, and 13 others are drugs with published clinical evidence of QT prolongation, with only one or two exceptional cases of TdP. The 10 remaining drugs are without reports of TdP and QT prolongation.

The relevance of different indicators such as APD90 increase, reverse use dependency, action potential triangulation or effect on Vmax was evaluated by comparison with available clinical data. Finally, a complex algorithm called TDPscreen™ and based on two subalgorithms corresponding to particular electrophysiological patterns was defined.

This latter algorithm enabled a clear separation of drugs into three groups: (A) drugs with numerous or several reports (>2 cases) of TdP, (B) drugs causing QT prolongation and/or TdP only, the latter at a very low frequency (2 cases), (C) drugs without reports of TdP or QT prolongation.

The use of such an algorithm combined with a database accrued from reference compounds with available clinical data is suggested as a basis for testing new candidate drugs in the early stages of development for proarrhythmic risk prediction.

British Journal of Pharmacology (2005) 144, 376–385. doi:10.1038/sj.bjp.0706070
A comparative study on the acute and long‐term effects of MDMA and 3,4‐dihydroxymethamphetamine (HHMA) on brain monoamine levels after i.p. or striatal administration in mice
Authors: Escobedo Isabel , O'Shea Esther , Orio Laura , Sanchez Veronica , Segura Mireia , de la Torre Rafael , Farre Magi , Green Alfred Richard , Colado Maria Isabel
Published: January 2005

This study investigated whether the immediate and long‐term effects of 3,4‐methylenedioxymethamphetamine (MDMA) on monoamines in mouse brain are due to the parent compound and the possible contribution of a major reactive metabolite, 3,4‐dihydroxymethamphetamine (HHMA), to these changes. The acute effect of each compound on rectal temperature was also determined.

MDMA given i.p. (30 mg kg−1, three times at 3‐h intervals), but not into the striatum (1, 10 and 100 μg, three times at 3‐h intervals), produced a reduction in striatal dopamine content and modest 5‐HT reduction 1 h after the last dose. MDMA does not therefore appear to be responsible for the acute monoamine release that follows its peripheral injection.

HHMA does not contribute to the acute MDMA‐induced dopamine depletion as the acute central effects of MDMA and HHMA differed following i.p. injection. Both compounds induced hyperthermia, confirming that the acute dopamine depletion is not responsible for the temperature changes.

Peripheral administration of MDMA produced dopamine depletion 7 days later. Intrastriatal MDMA administration only produced a long‐term loss of dopamine at much higher concentrations than those reached after the i.p. dose and therefore bears little relevance to the neurotoxicity. This indicates that the long‐term effect is not attributable to the parent compound. HHMA also appeared not to be responsible as i.p. administration failed to alter the striatal dopamine concentration 7 days later.

HHMA was detected in plasma, but not in brain, following MDMA (i.p.), but it can cross the blood–brain barrier as it was detected in the brain following its peripheral injection.

The fact that the acute changes induced by i.p. or intrastriatal HHMA administration differed indicates that HHMA is metabolised to other compounds which are responsible for changes observed after i.p. administration.

British Journal of Pharmacology (2005) 144, 231–241. doi:10.1038/sj.bjp.0706071
A possible pharmacological explanation for quinacrine failure to treat prion diseases: pharmacokinetic investigations in a ovine model of scrapie
Authors: Gayrard Véronique , Picard‐Hagen Nicole , Viguié Catherine , Laroute Valerie , Andréoletti Olivier , Toutain Pierre‐Louis
Published: January 2005

Quinacrine was reported to have a marked in vitro antiprion action in mouse neuroblastoma cells. On compassionate grounds, quinacrine was administered to Creutzfeldt–Jakob disease patients, despite the absence of preclinical in vivo studies to evaluate efficacy. Quinacrine failed to provide therapeutic benefit. The aim of the study was to investigate possible pharmacokinetic and/or pharmacodynamic explanations for the discrepancy between the proven action of quinacrine in vitro and its lack of clinical efficacy.

We conducted in vitro experiments reproducing the culture conditions in which antiprion effects had been previously observed and recalculated the EC50 by determining the actual extracellular (120 nM) and intracellular (6713 nM) quinacrine neuroblastoma concentrations with the reported quinacrine EC50 (300 nM).

A randomized clinical trial in scrapie‐affected ewes confirmed the absence of therapeutic benefit of quinacrine. The in vivo quinacrine exposure was evaluated in a pharmacokinetic investigation in healthy ewes. Cerebrospinal fluid concentrations (<10.6 and 55 nM after administration of therapeutic and toxic quinacrine doses, respectively) were much lower than the quinacrine extracellular neuroblastoma concentrations corresponding to the reported EC50. The total brain tissue concentrations (3556 nM) obtained after a repeated therapeutic dosage regimen were within the range of the intracellular neuroblastoma quinacrine concentrations.

In conclusion, in order to avoid in vivo trials for which failure can be predicted, the measurement in vitro of the antiprion EC50 in both intra‐ and extracellular biophases should be determined. It can then be established if these in vitro antiprion concentrations are achievable in vivo.

British Journal of Pharmacology (2005) 144, 386–393. doi:10.1038/sj.bjp.0706072
Role of basal extracellular Ca2+ entry during 5‐HT‐induced vasoconstriction of canine pulmonary arteries
Authors: Wilson Sean M , Mason Helen S , Ng Lih C , Montague Stephen , Johnston Louise , Nicholson Neil , Mansfield Sarah , Hume Joseph R
Published: January 2005

Measurements of artery contraction, cytosolic [Ca2+], and Ca2+ permeability were made to examine contractile and cytosolic [Ca2+] responses of canine pulmonary arteries and isolated cells to 5‐hydroxytryptamine (5‐HT), and to determine the roles of intracellular Ca2+ release and extracellular Ca2+ entry in 5‐HT responses.

The EC50 for 5‐HT‐mediated contractions and cytosolic [Ca2+] increases was ∼10−7 M and responses were inhibited by ketanserin, a 5‐HT2A‐receptor antagonist.

5‐HT induced cytosolic [Ca2+] increases were blocked by 20 μM Xestospongin‐C and by 2‐APB (IC50=32 μM), inhibitors of InsP3 receptor activation.

5‐HT‐mediated contractions were reliant on release of InsP3 but not ryanodine‐sensitive Ca2+ stores.

5‐HT‐mediated contractions and cytosolic [Ca2+] increases were partially inhibited by 10 μM nisoldipine, a voltage‐dependent Ca2+ channel blocker.

Extracellular Ca2+ removal reduced 5‐HT‐mediated contractions further than nisoldipine and ablated cytosolic [Ca2+] increases and [Ca2+] oscillations. Similar to Ca2+ removal, Ni2+ reduced cytosolic [Ca2+] and [Ca2+] oscillations.

Mn2+ quench of fura‐2 and voltage‐clamp experiments showed that 5‐HT failed to activate any significant voltage‐independent Ca2+ entry pathways, including store‐operated and receptor‐activated nonselective cation channels. Ni2+ but not nisoldipine or Gd3+ blocked basal Mn2+ entry.

Voltage‐clamp experiments showed that simultaneous depletion of both InsP3 and ryanodine‐sensitive intracellular Ca2+ stores activates a current with linear voltage dependence and a reversal potential consistent with it being a nonselective cation channel. 5‐HT did not activate this current.

Basal Ca2+ entry, rather than CCE, is important to maintain 5‐HT‐induced cytosolic [Ca2+] responses and contraction in canine pulmonary artery.

British Journal of Pharmacology (2005) 144, 252–264. doi:10.1038/sj.bjp.0706077
Preclinical pharmacology of lumiracoxib: a novel selective inhibitor of cyclooxygenase‐2
Authors: Esser Ronald , Berry Carol , Du Zhengming , Dawson Janet , Fox Alyson , Fujimoto Roger A , Haston William , Kimble Earl F , Koehler Julie , Peppard Jane , Quadros Elizabeth , Quintavalla Joseph , Toscano Karen , Urban Laszlo , van Duzer John , Zhang Xiaoli , Zhou Siyuan , Marshall Paul J
Published: January 2005

This manuscript presents the preclinical profile of lumiracoxib, a novel cyclooxygenase‐2 (COX‐2) selective inhibitor.

Lumiracoxib inhibited purified COX‐1 and COX‐2 with Ki values of 3 and 0.06 μM, respectively. In cellular assays, lumiracoxib had an IC50 of 0.14 μM in COX‐2‐expressing dermal fibroblasts, but caused no inhibition of COX‐1 at concentrations up to 30 μM (HEK 293 cells transfected with human COX‐1).

In a human whole blood assay, IC50 values for lumiracoxib were 0.13 μM for COX‐2 and 67 μM for COX‐1 (COX‐1/COX‐2 selectivity ratio 515).

Lumiracoxib was rapidly absorbed following oral administration in rats with peak plasma levels being reached between 0.5 and 1 h.

Ex vivo, lumiracoxib inhibited COX‐1‐derived thromboxane B2 (TxB2) generation with an ID50 of 33 mg kg−1, whereas COX‐2‐derived production of prostaglandin E2 (PGE2) in the lipopolysaccharide‐stimulated rat air pouch was inhibited with an ID50 value of 0.24 mg kg−1.

Efficacy of lumiracoxib in rat models of hyperalgesia, oedema, pyresis and arthritis was dose‐dependent and similar to diclofenac. However, consistent with its low COX‐1 inhibitory activity, lumiracoxib at a dose of 100 mg kg−1 orally caused no ulcers and was significantly less ulcerogenic than diclofenac (P<0.05).

Lumiracoxib is a highly selective COX‐2 inhibitor with anti‐inflammatory, analgesic and antipyretic activities comparable with diclofenac, the reference NSAID, but with much improved gastrointestinal safety.

British Journal of Pharmacology (2005) 144, 538–550. doi:10.1038/sj.bjp.0706078
Effects of minocycline on Fas‐mediated fulminant hepatitis in mice
Authors: Chu Heng‐Cheng , Lin Yi‐Ling , Sytwu Huey‐Kang , Lin Shin‐Hua , Liao Ching‐Len , Chao You‐Chen
Published: January 2005

Minocycline has anti‐inflammatory and antiapoptotic effects on cartilage, neurons and periodontal tissues, and both properties are central to the pharmaceutical treatment of liver diseases. We investigated the effects of minocycline on fulminant hepatitis in C57BL/6J mice induced by lethal challenge of the activating anti‐Fas antibody, Jo2.

Intraperitoneal injection of Jo2 (0.6 μg g−1) to mice resulted in fulminant hepatitis, as evidenced by increase of serum alanine/aspartate transaminase activities and histopathological alterations in liver sections, as well as animal death. Nevertheless, mice pretreated with three doses of minocycline (5 mg kg−1) resisted this lethal effect significantly. Minocycline treatment improved the survival kinetics, although to a lesser extent, when mice were challenged simultaneously with Jo2 or even treated 30 min after the lethal challenge.

Jo2‐induced activation of caspase‐3 or ‐9 in liver tissues was inhibited by minocycline pretreatment, and yet the direct addition of minocycline to liver extracts from Jo2‐challenged mice failed to block caspase activation in vitro. Moreover, minocycline efficiently suppressed the release of cytochrome c from mitochondria of the liver tissues from Jo2‐challenged mice. In contrast, caspase‐8 activation and Bid truncation triggered by Jo2 were not diminished by minocycline pretreatment in mouse livers.

Our results suggest that easing of Fas‐triggered fulminant hepatitis by minocycline may involve a mitochondrial apoptotic pathway, probably through preventing cytochrome c release and thereby blocking downstream caspase activation.

British Journal of Pharmacology (2005) 144, 275–282. doi:10.1038/sj.bjp.0706079
TNF regulates leukocyte–endothelial cell interactions and microvascular dysfunction during immune complex‐mediated inflammation
Authors: Norman M Ursula , Lister Karyn J , Yang Yuan H , Issekutz Andrew , Hickey Michael J
Published: January 2005

The aim of this study was to assess directly the role of TNF in immune complex‐induced leukocyte–endothelial cell interactions and microvascular dysfunction.

Intravital microscopy was used to examine immune complex‐induced leukocyte rolling, adhesion and emigration and microvascular permeability in cremasteric postcapillary venules in wild‐type and TNF−/− mice. The reverse passive Arthus (RPA) reaction was used to localize immune complex formation to the cremaster muscle.

In wild‐type mice, immune complex deposition induced a reduction in leukocyte rolling velocity and increases in leukocyte adhesion and emigration. In TNF−/− mice, the immune complex‐induced reduction in leukocyte rolling velocity was significantly attenuated, and leukocyte adhesion and emigration were also significantly reduced relative to responses in wild‐type mice.

The alterations in TNF−/− mice were associated with decreased expression of endothelial P‐selectin and VCAM‐1, and an absence of E‐selectin‐dependent rolling normally seen in wild‐type mice at the peak of the response. In addition, the level of immune complex‐induced microvascular permeability was attenuated in TNF−/− mice.

These findings demonstrate that in immune complex‐induced inflammation, TNF promotes leukocyte rolling and adhesive interactions, and entry of leukocytes into sites of immune complex deposition, in part via the increased expression and/or function of endothelial P‐selectin, E‐selectin and VCAM‐1. In addition, this increase in leukocyte recruitment mediated by TNF correlates directly with an increase in microvascular injury.

British Journal of Pharmacology (2005) 144, 265–274. doi:10.1038/sj.bjp.0706081
Blockade of glutamatergic and GABAergic receptor channels by trimethyltin chloride
Authors: Krüger Katharina , Diepgrond Victoria , Ahnefeld Maria , Wackerbeck Christina , Madeja Michael , Binding Norbert , Musshoff Ulrich
Published: January 2005

Organotin compounds such as trimethyltin chloride (TMT) are among the most toxic of the organometallics. As their main target for toxicity is the central nervous system, the aim of the present study was to investigate the effects of TMT on receptor channels involved in various processes of synaptic transmission.

The Xenopus oocyte expression system was chosen for direct assessment of TMT effects on voltage‐operated potassium channels and glutamatergic and GABAergic receptors, and hippocampal slices from rat brain for analyzing TMT effects on identified synaptic sites.

TMT was found to be ineffective, at 100 μmol l−1, against several potassium‐ and sodium‐operated ion channel functions as well as the metabotropic glutamate receptor.

The functions of the ionotropic glutamate and the GABAA receptor channels were inhibited by TMT in micromolar concentrations. Thus, at a maximum concentration of 100 μmol l−1, around 20–30% of the α‐amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionic acid and GABAA receptor‐mediated ion currents and 35% of the N‐methyl‐D‐aspartate receptor‐mediated ion currents were blocked.

In the hippocampal slice model, the inhibitory effects of TMT were much stronger than expected from the results on the ion channels. Bath application of TMT significantly reduced the amplitudes of evoked excitatory postsynaptic field potentials in a concentration‐dependent and nonreversible manner.

 Induction of long‐term potentiation, recorded from the CA1 dendritic region, was inhibited by TMT and failed completely at a concentration of 10 μmol l−1.

In general, TMT affects the excitatory and inhibitory synaptic processes in a receptor specific manner and is able to disturb the activity within a neuronal network.

British Journal of Pharmacology (2005) 144, 283–292. doi:10.1038/sj.bjp.0706083
Dextromethorphan differentially affects opioid antinociception in rats
Authors: Chen Shiou‐Lan , Huang Eagle Yi‐Kung , Chow Lok‐Hi , Tao Pao‐Luh
Published: January 2005

Opioid drugs such as morphine and meperidine are widely used in clinical pain management, although they can cause some adverse effects. A number of studies indicate that N‐methyl‐D‐aspartate (NMDA) receptors may play a role in the mechanism of morphine analgesia, tolerance and dependence. Being an antitussive with NMDA antagonist properties, dextromethorphan (DM) may have some therapeutic benefits when coadministered with morphine. In the present study, we investigated the effects of DM on the antinociceptive effects of different opioids. We also investigated the possible pharmacokinetic mechanisms involved.

The antinociceptive effects of the μ‐opioid receptor agonists morphine (5 mg kg−1, s.c.), meperidine (25 mg kg−1, s.c.) and codeine (25 mg kg−1, s.c.), and the κ‐opioid agonists nalbuphine (8 mg kg−1, s.c.) and U‐50,488H (20 mg kg−1, s.c.) were studied using the tail‐flick test in male Sprague–Dawley rats. Coadministration of DM (20 mg kg−1, i.p.) with these opioids was also performed and investigated.

The pharmacokinetic effects of DM on morphine and codeine were examined, and the free concentration of morphine or codeine in serum was determined by HPLC.

It was found that DM potentiated the antinociceptive effects of some μ‐opioid agonists but not codeine or κ‐opioid agonists in rats. DM potentiated morphine's antinociceptive effect, and acutely increased the serum concentration of morphine. In contrast, DM attenuated the antinociceptive effect of codeine and decreased the serum concentration of its active metabolite (morphine).

The pharmacokinetic interactions between DM and opioids may partially explain the differential effects of DM on the antinociception caused by opioids.

British Journal of Pharmacology (2005) 144, 400–404. doi:10.1038/sj.bjp.0706086
Piglet saphenous vein contains multiple relaxatory prostanoid receptors: evidence for EP4, EP2, DP and IP receptor subtypes
Authors: Wilson Richard J , Giles Heather
Published: January 2005

Prostaglandin E2 produced endothelium‐independent relaxation of phenylephrine‐ and 5‐HT‐contracted piglet saphenous vein (PSV; pEC50=8.6±0.2; n=6).

The prostanoid EP4 receptor antagonist GW627368X (30–300 nM) produced parallel rightward displacement of PGE2 concentration–effect (E/[A]) curves (pKb=9.2±0.2; slope=1). Higher concentrations of GW627368X did not produce further rightward shifts, revealing the presence of non‐EP4 prostanoid receptors.

In all, 18 other prostanoid receptor agonists relaxed PSV in a concentration‐related manner. Relative potencies of agonists most sensitive to 10 μM GW627368X (and therefore predominantly activating EP4 receptors) correlated well with those at human recombinant EP4 receptors in human embryonic kidney (HEK‐293) cells (r2=0.74).

In the presence of 10 μM GW627368X, the rank order of agonist relative potency matched that of the human recombinant EP2 receptor in Chinese hamster ovary cells (r2=0.72).

Iloprost, cicaprost and PGI2 relaxed PSV maximally and were antagonised by 10 μM GW627368X, demonstrating that they were full EP4 receptor agonists. Residual responses to these compounds in the presence of GW627368X suggested the presence of IP receptors.

BW245C relaxed PSV maximally (pEC50=6.8±0.1). In the presence of 10 μM GW627368X, BW245C produced biphasic E/[A] curves (phase one pEC50=6.6; α=24%; phase two pEC50=5.1; α=112%). Phase two was antagonised by the DP receptor antagonist BW A868C (1 μM), demonstrating that BW245C is an agonist at DP and EP4 receptors.

We conclude that PSV contains EP4, EP2, DP and IP receptors; IP receptor agonists are also porcine EP4 receptor agonists.

British Journal of Pharmacology (2005) 144, 405–415. doi:10.1038/sj.bjp.0706088
Insights into the functional roles of α1‐adrenoceptor subtypes in mouse carotid arteries using knockout mice
Authors: Deighan Clare , Methven Laura , Naghadeh Mustafa M , Wokoma Alexis , Macmillan Joyce , Daly Craig J , Tanoue Akito , Tsujimoto Gozoh , McGrath John C
Published: January 2005

α1‐Adrenoceptor (AR) subtypes in mouse carotid arteries were characterised using a combination of agonist/antagonist pharmacology and knockout (KO) mice.

Phenylephrine (PE) was most potent in the α1B‐KO (pEC50=6.9±0.2) followed by control (pEC50=6.3±0.06) and α1D‐KO (pEC50=5.5±0.07). Both N‐[5‐(4,5‐dihydro‐1H‐imidazol‐2yl)‐2‐hydroxy‐5,6,7,8‐tetrahydronaphthalen‐1‐yl] methanesulphonamide hydrobromide (A‐61603) and 5‐hydroxytryptamine (5‐HT) were more potent in the α1D‐KO (pEC50=7.4±0.27 and 7.4±0.05, respectively) than the control (pEC50=6.9±0.09 and 6.9±0.08, respectively) and equipotent with the control in the α1B‐KO (pEC50=6.7±0.07 and 6.8±0.04). Maximum responses to PE and A‐61603 were reduced in the α1D‐KO compared to control; there was no difference in maximum responses to 5‐HT.

In control arteries, prazosin and 5‐methylurapidil acted competitively with pA2 of 9.6 and 7.5, respectively. BMY7378 produced antagonism only at the highest concentration used (100 nM; pKB 8.3).

Prazosin, 5‐methylurapidil and BMY7378 acted competitively in α1B‐KO carotid arteries with pA2 of 10.3, 7.6 and 9.6, respectively.

In the α1D‐KO, against PE, 5‐methylurapidil produced a pA2 of 8.1. pKB values were calculated for prazosin (10.6) and BMY7378 (7.0). Against A‐61603, 5‐methylurapidil had a pA2 of 8.5, prazosin 8.6, while BMY7378 had no effect.

In conclusion, the α1B‐KO mediates contraction solely through α1D‐ARs and the α1D‐KO through α1A‐ARs. Extrapolating back to the control from the knockout data suggests that all three subtypes could be involved in the responses, but we propose that the α1D‐AR causes the contractile response and that the role of the α1B‐AR is mainly regulatory.

British Journal of Pharmacology (2005) 144, 558–565. doi:10.1038/sj.bjp.0706089
An investigation of whether agonist‐selective receptor conformations occur with respect to M2 and M4 muscarinic acetylcholine receptor signalling via Gi/o and Gs proteins
Authors: Mistry Rajendra , Dowling Mark R , Challiss R A John
Published: January 2005

A range of muscarinic acetylcholine (mACh) receptor agonists (methacholine (MCh), oxotremorine‐M (OXO‐M), oxotremorine (OXO), arecoline (AREC), bethanechol (BETH), pilocarpine (PILO)) have been investigated with respect to their binding to, and activation of, M2 and M4 mACh receptors, recombinantly expressed in Chinese hamster ovary cells, to explore the possibility that these agonists may differentially affect mACh receptor–Gi/o and ‐Gs coupling.

M2/M4 mACh receptor coupling to the adenylyl cyclase/cyclic AMP signalling pathway has been explored in intact cells. Gi/o‐mediated negative coupling to adenylyl cyclase was explored functionally by assessing the ability of the mACh receptor agonists to inhibit forskolin‐stimulated enzymic activity. Following pertussis toxin treatment (100 ng ml−1, 18–20 h) to inactivate Gi/o proteins, each agonist caused a Gs‐mediated enhancement of forskolin‐stimulated adenylyl cyclase activity.

At both M2 and M4 mACh receptors, all agonists tested were more potent in mediating Gi/o‐ versus Gs‐coupled responses. This difference (determined as the pIC50 (Gi/o coupling) minus pEC50 (Gs coupling) value) was greatest for AREC (65–75‐fold) and least for BETH and PILO (10‐fold).

Using apparent binding affinities (pKB), and potency (EC50/IC50) and responsiveness (Emax/Imax) estimates, relative efficacy (erel) values for each agonist with respect to M2 and M4 mACh receptor coupling to Gi/o‐ and Gs‐mediated signalling were also calculated. While the erel values obtained for MCh and OXO‐M in CHO‐m2 cells were similar, OXO‐M behaved as a ‘super‐agonist’ at the M4 mACh receptor giving greater erel values for both Gi/o and Gs coupling relative to MCh.

The experimental data indicate that while interesting differences between agonists with respect to M2/M4 mACh receptor activation and receptor–Gi/o and ‐Gs coupling can be discerned, no clear examples of agonist trafficking of signal have emerged.

British Journal of Pharmacology (2005) 144, 566–575. doi:10.1038/sj.bjp.0706090
Reciprocal changes in endothelium‐derived hyperpolarizing factor‐ and nitric oxide‐system in the mesenteric artery of adult female rats following ovariectomy
Authors: Nawate Satoshi , Fukao Mitsuhiro , Sakuma Ichiro , Soma Takamitsu , Nagai Kazuhiko , Takikawa Osamu , Miwa Soichi , Kitabatake Akira
Published: January 2005

To explore the effects of estrogen on arterial functions, we examined endothelium‐derived hyperpolarizing factor (EDHF)‐ and NO‐mediated responses in isolated mesenteric arteries of female rats, 4 weeks after sham‐operation (CON), ovariectomy (OVX) and OVX plus chronic estrogen treatment (OVX+E2). Tissue levels of connexins‐40, 43 (major components of gap junction), inducible NOS (iNOS), endothelial NOS (eNOS) and eNOS regulator proteins such as calmodulin, heat shock protein 90 (hsp90) and caveolin‐1 were also examined using Western blot.

In OVX, acetylcholine (ACh)‐induced EDHF‐mediated relaxation and membrane hyperpolarization of arterial smooth muscles were reduced, whereas ACh‐induced NO‐mediated relaxation was enhanced, leading to no change in ACh‐induced relaxation.

In OVX, connexin‐40 and 43 were decreased. Tissue levels of eNOS and its positive regulators (calmodulin and hsp90) were unchanged, but that of its negative regulator, caveolin‐1, was decreased. The levels of iNOS in mesenteric artery and aorta and plasma levels of NO metabolites and cholesterol were elevated.

In OVX, contraction of the artery by phenylephrine was reduced, but augmented by nonspecific inhibitor of NOS to the comparable level as that in CON group. The contraction in OVX group unlike that in CON group was augmented by specific iNOS inhibitor, and the difference between contractions in the presence of nonspecific and specific inhibitor as an index of eNOS activity was increased.

In OVX+E2, all these changes were recovered.

In all groups, EDHF‐mediated relaxation was suppressed by 18β‐glycyrrhetinic acid, an inhibitor of gap junction.

These results indicate that estrogen deficiency does not change the diameter of mesenteric artery: it reduces EDHF‐mediated relaxation by decreasing gap junction, whereas it augments NO‐mediated relaxation via an increase in NO release. Increased NO result from increased activity of eNOS subsequent to a decrease in caveolin‐1 and from induction of iNOS. However, excessive NO generation with elevated plasma cholesterol would raise a risk for atherosclerosis.

British Journal of Pharmacology (2005) 144, 178–189. doi:10.1038/sj.bjp.0706091
Vascular effects of ovariectomy and chronic oestrogen treatment in rats: controversy or experimental protocol diversity?
Authors: Chataigneau Thierry , Schini‐Kerth Valérie B
Published: January 2005

Several clinical studies have indicated that oestrogens have protective properties on the cardiovascular system. Although the beneficial effect has been attributable, at least in part, to their ability to stimulate the endothelial fomation of nitric oxide (NO) and endothelium‐derived hyperpolarizing factor (EDHF), the underlying mechanism still remains unclear. In a study from this issue of British Journal of Pharmacology, Nawate et al. have examined the effects of rat ovariectomy and chronic treatment with 17β‐oestradiol on the endothelial function as assessed ex vivo. The data indicate that acetylcholine‐induced endothelium‐dependent relaxations of the isolated mesenteric artery are affected by neither ovariectomy nor chronic hormonal treatment. Despite the maintained endothelium‐dependent relaxation, the contribution of the two major endothelial factors NO and EDHF was changed. Indeed, ovariectomy increased the NO‐mediated component of the relaxation, most likely as a consequence of the downregulation of the physiological allosteric inhibitor of endothelial NO synthase, caveolin‐1. In addition, ovariectomy decreased the EDHF‐mediated component of the relaxation and membrane hyperpolarization of the smooth muscle cells, an effect which might be explained by a concomitant decrease of the expression of the gap junction connexin‐40 and connexin‐43. Furthermore, chronic administration of 17β‐estradiol to ovariectomized rats normalized all these effects. This study provides further experimental evidence indicating that the hormonal status plays a determinant role in the control of the endothelial formation of both NO and EDHF.

British Journal of Pharmacology (2005) 144, 161–163. doi:10.1038/sj.bjp.0706092
Somatostatin‐receptor 2 (sst2)‐mediated effects of endogenous somatostatin on exocrine and endocrine secretion of the rat stomach
Authors: Fykse Vidar , Coy David H , Waldum Helge Lyder , Sandvik Arne Kristian
Published: January 2005

Somatostatin is a potent inhibitor of gastric acid secretion. Its effects are mediated through five specific receptor subtypes (sst1–5), of which sst2 is dominant on the enterochromaffin‐like (ECL) cell and the parietal cell. To study the paracrine mechanisms of somatostatin, the sst2‐specific antagonist PRL‐2903 was used.

Effects of PRL‐2903 on acid secretion and release of histamine were studied in the totally isolated, vascularly perfused rat stomach. Further, the release of histamine and gastrin after bombesin, alone and in combination with PRL‐2903, were studied. Results are presented as mean±standard error of the mean (s.e.m.).

PRL‐2903 concentration‐dependently increased the venous histamine concentration from basal 55.6±7.5 to 367±114 nM at 50 μM PRL‐2903. With 10 μM PRL‐2903, venous histamine output increased from baseline 6.2±0.5 to 20.9±4.9 nmol h−1; P=0.008. The combination of 520 pM gastrin and 10 μM PRL‐2903 increased venous histamine output from 41.7±7.3 nmol h−1 with gastrin alone to 95.2±9.8 nmol h−1; P=0.016. Further, 10 μM PRL‐2903 increased acid output from baseline 8.5±1.8 to 37.4±11 μmol h−1; P=0.017. When combined with 10 μM ranitidine, PRL‐2903 did not significantly stimulate acid secretion. Bombesin/PRL‐2903 increased venous histamine concentration from 50.4±14.8 to 292±64.2 nM; P=0.008, and gastrin concentration from 38.6±13.1 to 95.8±20.3 pM; P=0.037.

Endogenous somatostatin exerts a continuous restraint on histamine and gastrin release from the gastric mucosa and significantly reduces baseline acid secretion.

British Journal of Pharmacology (2005) 144, 416–421. doi:10.1038/sj.bjp.0706094
Blockade of HERG cardiac K+ current by antifungal drug miconazole
Authors: Kikuchi Kan , Nagatomo Toshihisa , Abe Haruhiko , Kawakami Kazunobu , Duff Henry J , Makielski Jonathan C , January Craig T , Nakashima Yasuhide
Published: January 2005

Miconazole, an imidazole antifungal agent, is associated with acquired long QT syndrome and ventricular arrhythmias. Miconazole increases the plasma concentration of QT‐prolonging drugs by inhibiting the hepatic cytochrome P450 metabolic pathway, but whether it has direct effects on cardiac ion channels has not been elucidated.

To determine the mechanism underlying these clinical findings, we investigated the effect of miconazole on human ether‐a‐go‐go‐related gene (HERG) K+ channels.

HERG channels were heterologously expressed in human embryonic kidney 293 (HEK293) cells and whole‐cell currents were recorded using a patch‐clamp technique (23°C).

Miconazole inhibited HERG peak tail current in a concentration‐dependent manner (0.4–40 μM) with an IC50 of 2.1 μM (n=3–5 cells at each concentration, Hill coefficient 1.2). HERG block was not frequency‐dependent. It required channel activation, occurred rapidly, and had very slow dissociation properties.

The activation curve was shifted in a negative direction (V1/2: −9.5±2.3 mV in controls and −15.3±2.4 mV after 4 μM miconazole, P<0.05, n=6). Miconazole did not change other channel kinetics (activation, deactivation, onset of inactivation, recovery from inactivation, steady‐state inactivation).

The S6 domain mutation, F656C, abolished the inhibitory action of miconazole on HERG current indicating that miconazole preferentially binds to an aromatic amino‐acid residue within the pore‐S6 region.

Our findings indicate that miconazole causes HERG channel block by binding to a common drug receptor, and this involves preferential binding to activated channels. Thus, miconazole prolongs the QT interval by direct inhibition of HERG channels.

British Journal of Pharmacology (2005) 144, 840–848. doi:10.1038/sj.bjp.0706095
Positive interaction of the β2‐agonist CHF 4226.01 with budesonide in the control of bronchoconstriction induced by acetaldehyde in the guinea‐pigs
Authors: Rossoni Giuseppe , Manfredi Barbara , Razzetti Roberta , Civelli Maurizio , Bongrani Stefano , Berti Ferruccio
Published: January 2005

Pretreatment of anaesthetized guinea‐pigs with either CHF 4226.01 (8‐hydroxy‐5‐[(1R)‐1‐hydroxy‐2‐[N‐[(1R)‐2‐(p‐methoxyphenyl)‐1‐methylethyl]amino]ethyl] carbostyril hydrochloride), formoterol or budesonide reduced acetaldehyde (AcCHO)‐evoked responses in the lungs with a rank order of potency CHF 4226.01 (ED50 values, from 1.88 to 3.31 pmol) > formoterol (ED50 values, from 3.03 to 5.51 pmol) ≫ budesonide (ED50 values, from 335 to 458 nmol). The duration of action of CHF 4226.01 in antagonizing the airway obstruction elicited by AcCHO was also substantially longer than formoterol (area under the curve) at 10 pmol, 763±58 and 480±34, respectively; P<0.01).

Continuous infusion of a subthreshold dose of AcCHO enhanced the intratracheal pressure (ITP) increases caused by subsequent challenges with substance P (from 9.7±0.8 to 27.5±1.6 cm H2O as a peak, P<0.001). Pretreatment with either CHF 4226.01 or formoterol prevented the sensitizing effect of AcCHO on substance P responses (ED50 values, 2.85 and 6.11 pmol, respectively; P<0.01).

The ED50 value of budesonide (396 nmol) in preventing AcCHO‐evoked ITP increase was reduced when this glucocorticoid was combined with 0.1 pmol CHF 4226.01 (ED50 76 nmol; P<0.001). CHF 4226.01/budesonide was two‐fold more effective (P<0.01) than the formoterol/budesonide combination.

These results suggest that CHF 4226.01/budesonide, by optimizing each other's beneficial potential in the control of pulmonary changes caused by AcCHO in the guinea‐pigs, may represent a new fixed combination in asthma.

British Journal of Pharmacology (2005) 144, 422–429. doi:10.1038/sj.bjp.0706096
Differential action of a protein tyrosine kinase inhibitor, genistein, on the positive inotropic effect of endothelin‐1 and norepinephrine in canine ventricular myocardium
Authors: Chu Li , Zhang Jian‐Xin , Norota Ikuo , Endoh Masao
Published: January 2005

Experiments were carried out in isolated canine ventricular trabeculae and acetoxymethylester of indo‐1‐loaded single myocytes to elucidate the role of protein tyrosine kinase (PTK) in the inotropic effect of endothelin‐1 (ET‐1) induced by crosstalk with norepinephrine (NE). The PTK inhibitor genistein was used as a pharmacological tool.

Genistein but not daidzein inhibited the positive inotropic effect and the increase in Ca2+ transients induced by ET‐1 by crosstalk with NE at low concentrations.

Genistein and daidzein antagonized the negative inotropic effect and the decrease in Ca2+ transients induced by ET‐1 by crosstalk with NE at high concentrations, but genistein did not affect the antiadrenergic effect of carbachol.

Genistein but not daidzein enhanced the positive inotropic effect and the increase in Ca2+ transients induced by NE via β‐adrenoceptors, while the enhancing effect of genistein was abolished by the protein tyrosine phosphatase inhibitor vanadate.

These findings indicate that genistein (1) induces a positive inotropic effect in association with an increase in Ca2+ transients, (2) inhibits the positive inotropic effect of ET‐1 induced by crosstalk with NE, and (3) enhances the positive inotropic effect of NE induced via β‐adrenoceptors by inhibition of PTK. In addition, genistein inhibits the negative inotropic effect of ET‐1 induced by crosstalk with NE through a PTK‐unrelated mechanism. PTK may play a crucial role in the receptor‐mediated regulation of cardiac contractile function in canine ventricular myocardium.

British Journal of Pharmacology (2005) 144, 430–442. doi:10.1038/sj.bjp.0706097
Aldosterone does not mediate angiotensin II‐induced atherosclerosis and abdominal aortic aneurysms
Authors: Cassis Lisa A , Helton Marc J , Howatt Deborah A , King Victoria L , Daugherty Alan
Published: January 2005

We have demonstrated previously that infusion of angiotensin II (AngII) into hyperlipidemic mice augments atherosclerosis and results in the formation of abdominal aortic aneurysms (AAA). The purpose of this study was to determine the role of aldosterone in these AngII‐induced vascular pathologies.

Male apolipoprotein E−/− (apoE) mice were infused with either vehicle or aldosterone (50 or 200 ng kg−1 min−1). Arterial blood pressure was determined throughout the study and serum lipid concentrations and vascular pathology were quantified after 28 days of infusion.

Infusion of aldosterone did not influence body weight or serum cholesterol concentrations. Kidney weight was increased dose‐dependently by aldosterone infusion. Systolic blood pressure was not significantly altered by aldosterone. Plasma aldosterone concentrations were increased dose‐dependently by infusion of aldosterone. However, there was no effect of aldosterone on the extent of atherosclerosis and AAAs were not formed.

Implantation of pellets containing spironolactone (16 mg kg−1 day−1) in AngII‐infused apoE−/− mice (1000 ng kg−1 min−1) had no effect on AngII‐induced elevations in blood pressure. Plasma aldosterone concentration was not influenced by coadministration of spironolactone with AngII. Spironolactone administration did not influence the extent of atherosclerosis. Moreover, spironolactone had no significant effect on AngII‐induced AAA (incidence of AAA formation: 80 versus 70% for vehicle versus spironolactone, respectively; not significant).

These studies demonstrate that the AngII‐induced vascular pathologies of atherosclerosis and AAA formation are not mediated through aldosterone.

British Journal of Pharmacology (2005) 144, 443–448. doi:10.1038/sj.bjp.0706098