bluuebunny/crossref_metadata_2025 huggingface dataset:
Neural modulation of the cyclic electrical and mechanical activity in the rat colonic circular muscle: putative role of ATP and NO
Authors: Plujà Lídia , Fernández Ester , Jiménez Marcel
Published: January 2005

The rat colonic circular muscle displays cyclic episodes of myenteric potential oscillations (MPOs), each of them associated with a spontaneous contraction. Nifedipine 1 μM abolished both MPOs and their associated contractions. TTX (1 μM) increased the amplitude and frequency of spontaneous contractions.

Electrical field stimulation (EFS) induced a non‐adrenergic non‐cholinergic (NANC) inhibitory junction potential (IJP), with two phases: an initial fast hyperpolarization (characterized by IJP amplitude) and a sustained hyperpolarization (characterized by IJP duration).

Sodium nitroprusside (10 μM) hyperpolarized and abolished spontaneous contractions even in presence of TTX or 1 μM apamin. ATP (100 μM) also hyperpolarized and abolished spontaneous contractions but its effects were decreased by TTX and abolished by apamin.

Suramin (100 μM) or apamin reduced the amplitude of the IJPs, but did not affect their duration. Incubation with L‐NOARG (1 mM) reduced the duration but not the amplitude of the IJPs. In presence of L‐NOARG plus suramin or L‐NOARG plus apamin, both duration and amplitude of the IJPs were reduced but a residual IJP could still be recorded.

We conclude that the mechanical and electrical cyclic activity of the rat colonic circular muscle is modulated but not originated by the enteric nervous system and involves L‐type calcium channel activity. EFS induces release of NANC inhibitory neurotransmitters which hyperpolarize and relax smooth muscle cells. Both ATP and NO are involved in IJP generation: ATP is responsible for the first phase of the IJPs involving activation of apamin‐sensitive potassium channels, whereas NO initiates the second phase which is independent of the activation of such channels.

British Journal of Pharmacology (1999) 126, 883–892; doi:10.1038/sj.bjp.0702363
Selective activation of excitation‐contraction coupling pathways by ETA and ETB receptors in guinea‐pig tracheal smooth muscle
Authors: Inui Takashi , Ninomiya Haruaki , Sasaki Yukio , Makatani Maki , Urade Yoshihiro , Masaki Tomoh , Yamamura Takaki
Published: January 2005

Signalling events responsible for endothelinA (ETA) and ETB receptor‐induced contraction were examined in epithelium‐denuded guinea‐pig tracheal smooth muscle strips. Selective stimulation of each subtype was achieved by a combination of ET‐1 (100 nM) and ETA and ETB receptor‐selective antagonists, BQ‐123 (10 μM) and BQ‐788 (3 μM), respectively.

Both ETA and ETB receptors induced long‐lasting contraction that was totally dependent on Ca2+ influx. Stimulation of ETA receptor induced both transient and sustained (Ca2+)i increases whereas that of ETB receptor induced only a sustained increase. Suppression of the transient (Ca2+)i increase by U73122 (3 μM) did not affect the ETA‐induced sustained (Ca2+)i increase and tension development. Stimulation of ETA receptor, but not ETB, induced phosphoinositide breakdown and protein kinase C (PKC). The activated PKC contributed to the contraction by increasing the Ca2+‐sensitivity of the contractile apparatus.

Thus, ETA receptor is coupled both with phospholipase C/Ca2+/PKC signalling and Ca2+ influx pathways whereas ETB receptor was coupled only with the latter.

Stimulation of ETB receptor, but not ETA, caused membrane depolarization measured with a fluorescent indicator, bis‐(1,3 dibutylbarbituric acid)‐trimethine oxonol. Both nifedipine (1 μM) and verapamil (10 μM) abolished ETB‐induced Ca2+ influx and contraction, while they barely affected ETA‐induced responses.

Therefore, the Ca2+ influx pathways activated by each subtype appeared to be completely different; ETA and ETB receptors opens voltage‐independent Ca2+ channels and L‐type voltage‐dependent Ca2+ channels, respectively.

British Journal of Pharmacology (1999) 126, 893–902; doi:10.1038/sj.bjp.0702365
Inhibition of inflammatory actions of aminobisphosphonates by dichloromethylene bisphosphonate, a non‐aminobisphosphonate
Authors: Endo Yasuo , Shibazaki Masahiko , Yamaguchi Kouji , Nakamura Masanori , Kosugi Hiroshi
Published: January 2005

When injected intraperitoneally into mice in doses larger than those used clinically, all the amino derivatives of bisphosphonates (aminoBPs) tested induce a variety of inflammatory reactions such as induction of histidine decarboxylase (HDC, the histamine‐forming enzyme), hypertrophy of the spleen, atrophy of the thymus, hypoglycaemia, ascites and accumulation of exudate in the thorax, and an increase in the number of macrophages and/or granulocytes in the peritoneal cavity of blood. On the other hand, dichloromethylene bisphosphonate (Cl2MBP) a typical non‐aminoBP, has no such inflammatory actions. In the present study, we found that this agent can suppress the inflammatory actions of aminoBPs.

Cl2MBP, when injected into mice before or after injection of 4‐amino‐1‐hydroxybutylidene‐1,1‐bisphosphonic acid (AHBuBP; a typical aminoBP), inhibited the induction of HDC activity by AHBuBP in a dose‐ and time‐dependent manner. The increase in HDC activity induced by AHBuBP was largely suppressed by the injection of an equimolar dose of Cl2MBP. Cl2MBP also inhibited other AHBuBP‐induced inflammatory reactions, as well as the inflammatory actions of two other aminoBPs. However, Cl2MBP did not inhibit the increase in HDC activity induced by lipopolysaccharide (LPS).

We have previously reported that AHBuBP augments the elevation of HDC activity and the production of interleukin‐1β (IL‐1β) that are induced by LPS. These actions of AHBuBP were also inhibited by Cl2MBP.

Based on these results and reported actions of bisphosphonates, the mechanisms underlying the contrasting effects of aminoBPs and Cl2MBP, a non‐aminoBP are discussed. The results suggest that combined administration of Cl2MBP and an aminoBP in patients might be a useful way of suppressing the inflammatory side effects of aminoBPs.

British Journal of Pharmacology (1999) 126, 903–910; doi:10.1038/sj.bjp.0702367
Beta‐adrenoceptor agonist stimulation of pulmonary nitric oxide production in the rabbit
Authors: Adding L Christofer , Agvald Per , Artlich Andreas , Persson Magnus G , Gustafsson Lars E
Published: January 2005

Nitric oxide (NO) is continuously produced in the lung and is present in exhaled air. We examined the effect of β‐adrenoceptor stimulation on the production of pulmonary NO in rabbits.

Exhaled NO was measured by chemiluminescence in anaesthetized and mechanically ventilated rabbits and in buffer‐perfused rabbit lungs.

Intravenous infusions of adrenaline (0.1–10 μg kg−1  min−1) elicited dose‐dependent increases in exhaled NO. The increases in exhaled NO comprised an initial peak followed by a lower plateau level. The increase in exhaled NO was inhibited by propranolol (1 mg kg−1) but not by phentolamine (1 mg kg−1).

Prenalterol, a β1‐adrenoceptor agonist, and terbutaline, a β2‐adrenoceptor agonist, also caused dose‐dependent increases in exhaled NO. However, prenalterol was >100 times more potent than terbutaline.

Infusions of forskolin (0.01–0.03 μmol kg−1  min−1), an adenylate cyclase stimulator, elicited dose‐dependent decreases in blood pressure and concomitant increases in heart rate but caused no alterations in exhaled NO.

Nimodipine, a L‐type calcium channel blocker, antagonized the increases in exhaled NO in response to prenalterol infusions.

The increases in exhaled NO in response to adrenaline and prenalterol were also present in blood‐free, buffer perfused lungs during constant‐flow conditions.

These results demonstrate that pulmonary nitric oxide production can be enhanced by β‐adrenoceptor stimulation. Furthermore, the results indicate that the β‐adrenergic stimulation of pulmonary NO production is not critically dependent on cyclic AMP formation but may require intact calcium‐channels.

British Journal of Pharmacology (1999) 126, 833–839; doi:10.1038/sj.bjp.0702369
Deficiency of nitric oxide in polycation‐induced airway hyperreactivity
Authors: Meurs Herman , Schuurman Fineke E , Duyvendak Michiel , Zaagsma Johan
Published: January 2005

Using a perfused guinea‐pig tracheal tube preparation, we investigated the role of endogenous nitric oxide (NO) in polycation‐induced airway hyperreactivity (AHR) to methacholine. Intraluminal (IL) administration of the NO synthase inhibitor Nω‐nitro‐L‐arginine methyl ester (L‐NAME; 100 μM) caused a 1.8 fold increase in the maximal contractile response (Emax) to IL methacholine compared to control, without an effect on the pEC50 (−log10 EC50). The polycation poly‐L‐arginine (100 μg ml−1, IL) similarly enhanced the Emax for methacholine; however, the pEC50 value was also increased, by one log10 unit. L‐NAME had no effect on the enhanced methacholine response of poly‐L‐arginine‐treated airways, while the enhanced agonist response was completely normalized by the polyanion heparin (25 u ml−1, IL). In addition, the effect of L‐NAME was fully restored in the poly‐L‐arginine plus heparin treated airways. The results indicate that, in addition to enhanced epithelial permeability, a deficiency of endogenous NO contributes to polycation‐induced AHR. The latter finding may represent a novel mechanism of AHR induced by eosinophil‐derived cationic proteins in allergic asthma.

British Journal of Pharmacology (1999) 126, 559–562; doi:10.1038/sj.bjp.0702372
Respective role of lipoxygenase and nitric oxide‐synthase pathways in plasma histamine‐induced macromolecular leakage inconscious hamsters
Authors: Gimeno G , Carpentier P H , Desquand‐Billiald S , Finet M , Hanf R
Published: January 2005

Intravital microscopy technique was used to determine the distribution of a fluorescent plasma marker (fluorescein‐isothiocyanate‐dextran, 150 kD; FD‐150) into venular and interstitial compartments of dorsal skin fold preparations in conscious hamsters.

One mg kg−1 histamine (i.v.) caused a biphasic decrease in venular fluorescence due to FD‐150 extravasation in all organs (general extravasation). Immediately after injection, the venular fluorescence decreased and plateaued in 60 min. Ninety minutes after histamine injection, venular fluorescence further decreased until 180 min. Prior treatment with indomethacin (0.1 mg kg−1, i.v.) did not modify the time‐course of general extravasation but prevented histamine‐induced venule dilatation.

Prior treatment with the 5‐lipoxygenase activating protein (FLAP) inhibitor, 3‐[1‐(p‐chlorobenzyl)‐5‐(isopropyl)‐3‐t‐butylthioindol‐2‐yl]‐2,2‐dimethyl‐propanoic acid sodium (MK‐886)(10 μg kg−1, i.v.), the leukotriene receptor antagonist, benzenemethanol a‐pentyl‐3‐(2‐quinolinylmethoxy) (REV‐5901)(1 mg kg−1, i.v.), or the glutathione‐S‐transferase inhibitor, ethacrynic acid (1 mg kg−1, i.v.), delayed by 60 min the onset of general extravasation caused by 1 mg kg−1 histamine.

Prior treatment with lipoxygenase pathway inhibitors and NG‐nitro‐L‐arginine‐methylester (L‐NAME)(100 mg kg−1, i.v.) abolished the general extravasation and venule dilatation induced by 1 mg kg−1 histamine.

Injection of 1 μg kg−1 (i.v.), of leukotriene‐C4 (LTC4) or ‐D4 (LTD4) induced immediate and sustained general extravasation and reduction in venule diameter, these effects being blocked by REV‐5901.

Histamine (1 mg kg−1, i.v.) induced biphasic decline in mean arterial blood pressure (MAP). An initial phase (from 0 to 60 min) was followed by a late phase beginning 90 min after histamine injection. L‐NAME (100 mg kg−1, i.v.) and aminoguanidine (1 mg kg−1, i.v.) prevented the late phase of histamine‐induced hypotension.

Thus, plasma histamine can trigger both an immediate cysteinyl‐leukotriene (Cys‐LT)‐dependent and a late nitric oxide (NO)‐mediated inflammatory cascade. Although the cyclo‐oxygenase (COX) pathway might account for histamine‐induced venule dilatation, it would not influence histamine‐induced extravasation.

British Journal of Pharmacology (1999) 126, 1801–1809; doi:10.1038/sj.bjp.0702380
Inhibition of the production of endothelium‐derived hyperpolarizing factor by cannabinoid receptor agonists
Authors: Fleming I , Schermer B , Popp R , Busse R
Published: January 2005

The endogenous cannabinoid, anandamide, has been reported to induce an 'endothelium‐derived hyperpolarizing factor (EDHF)‐like' relaxation in vitro. We therefore investigated the effects of cannabinoid CB1 receptor agonists; HU 210, Δ9‐tetrahydrocannabinol (Δ9‐THC) and anandamide, and a CB1 antagonist/inverse agonist, SR 141716A, on nitric oxide (NO) and EDHF‐mediated relaxation in precontracted rings of porcine coronary, rabbit carotid and mesenteric arteries.

In rings of mesenteric artery HU 210 and Δ9‐THC induced endothelium‐ and cyclo‐oxygenase‐independent relaxations which were sensitive to SR 141716A. Anandamide (0.03–30 μM) induced a slowly developing, endothelium‐independent relaxation which was abolished by diclofenac and was therefore mediated by cyclo‐oxygenase product(s). None of the CB1 agonists tested affected the tone of precontracted rings of rabbit carotid or porcine coronary artery.

In endothelium‐intact segments, HU 210, Δ9‐THC and anandamide did not affect NO‐mediated responses but under conditions of continuous NO synthase/cyclo‐oxygenase blockade, significantly inhibited acetylcholine and bradykinin‐induced relaxations which are attributed to the production of EDHF. The effects of HU 210 and Δ9‐THC were not observed when experiments were performed in the presence of SR 141716A suggesting the involvement of the CB1 receptor.

In a patch clamp bioassay of EDHF production, HU 210 decreased the EDHF‐mediated hyperpolarization of detector smooth muscle cells when applied to the donor segment but was without effect on the membrane potential of detector cells. The inhibition of EDHF production was unrelated to alterations in Ca2+‐signalling or cytochrome P450 activity.

These results suggest that the activation of endothelial CB1 receptors appears to be negatively coupled to the production of EDHF.

British Journal of Pharmacology (1999) 126, 949–960; doi:10.1038/sj.bjp.0702381
Effect of cerivastatin sodium, a new inhibitor of HMG‐CoA reductase, on plasma lipid levels, progression of atherosclerosis, and the lesional composition in the plaques of WHHL rabbits
Authors: Shiomi Masashi , Ito Takashi
Published: January 2005

The aim of this study was to examine whether cerivastatin sodium, a new inhibitor of 3‐hydroxy‐3‐methylglutaryl coenzyme A (HMG‐CoA) reductase, affects the lesional composition of spontaneously developed atherosclerosis due to hypercholesterolaemia and delays progression of the lesions.

We administered cerivastatin to 2‐month‐old WHHL rabbits, a low‐density lipoprotein receptor‐deficient animal model, at a dose of 0.6 mg kg−1 day−1 for 32 weeks. We examined the plasma lipid levels, the severity of atherosclerosis, and composition of atherosclerotic lesions. Lesional composition was determined using immunohistostaining for macrophages and smooth muscle cells, and Azan‐Mallory staining for collagen fibres and extracellular lipid deposits.

Compared to the control group, the plasma cholesterol levels were decreased in the treated group by 39% (12.7±0.6 mmol L−1 versus 20.9±1.0 mmol L−1, P<0.001). Atherosclerosis was suppressed by about 37% as measured by the thickness of the aortic lesions (158±13 μm versus 250±15 μm, P<0.001), and by 28% as measured by coronary stenosis (62.7±11.4 versus 86.9±12.2, P<0.05). In the cerivastatin group, regarding the per cent areas of lesional components in the lesion area, the macrophages (21.0±1.5% versus 27.9±1.9%, P<0.01) and extracellular lipid deposits (3.2±0.4% versus 5.1±0.4%, P<0.001) were decreased in the aortic lesions, and the per cent area of macrophages in the coronary lesions was also decreased (4.9±1.4% versus, 11.6±2.4%, P<0.05). The per cent area of smooth muscle cells and collagen fibres did not significantly decrease.

These results indicate that cerivastatin contributed to the plaque stabilization and delayed progression of early atherosclerosis in young WHHL rabbits, in addition to the potent hypolipidemic effects.

British Journal of Pharmacology (1999) 126, 961–968; doi:10.1038/sj.bjp.0702382
Mode of action of ICS 205,930, a novel type of potentiator of responses to glycine in rat spinal neurones
Authors: Chesnoy‐Marchais D
Published: January 2005

The effect of a novel potentiator of glycine responses, ICS 205,930, was studied by whole‐cell recordings from spinal neurones, and compared with that of other known potentiators, in an attempt to differentiate their sites of action.

The ability of ICS 205,930 (0.2 μM) to potentiate glycine responses persisted in the presence of concentrations of Zn2+ (5–10 μM) that were saturating for the potentiating effect of this ion.

Preincubation with 10 μM Zn2+ before application of glycine plus Zn2+ had an inhibitory effect, which did not result from Zn2+ entry into the neurone, since it persisted with either 10 mM internal EGTA or 10 μM internal Zn2+. To test whether the potentiating effects of ICS 205,930 and Zn2+ interact, both compounds were applied without preincubation.

The potentiating effect of ICS 205,930 was similar for responses to glycine and for responses to glycine plus Zn2+, provided the concentrations of agonist were adjusted so as to induce control responses of identical amplitudes.

ICS 205,930 remained able to potentiate glycine responses in the presence of ethanol (200 mM).

ICS 205,930 also retained its potentiating effect in the presence of the anaesthetic propofol (30–90 μM), which strongly potentiated glycine responses but, in contrast with ICS 205,930, also markedly increased the resting conductance.

The anticonvulsant chlormethiazole (50–100 μM) neither potentiated glycine responses nor prevented the effect of ICS 205,930, even though it increased the resting conductance and potentiated GABAA responses.

The mechanism of action of ICS 205,930 appears to be different from those by which Zn2+, propofol or ethanol potentiate glycine responses.

British Journal of Pharmacology (1999) 126, 801–809; doi:10.1038/sj.bjp.0702384
Identification of novel polymorphisms within the promoter region of the human β2 adrenergic receptor gene
Authors: Scott Mark G H , Swan Caroline , Wheatley Amanda P , Hall Ian P
Published: January 2005

By screening the 1470 bp 5′ to the start codon of the human β2 adrenergic receptor gene, we have identified a total of eight polymorphisms (−20 T→C, −47 T→C, −367 T→C, −468 C→G, −654 G→A, −1023 G→A, −1343 A→G and −1429 T→A c.f. β2 adrenergic receptor start codon). Transient transfection of 5′ flanking deletion luciferase reporter constructs demonstrated the majority of activity of the human β2 adrenergic gene 5′ flanking region to be present within a 549 bp fragment immediately upstream from the start codon. Because of linkage disequilibrium, some combinations of polymorphisms were particularly frequent. We transiently transfected COS‐7 cells with luciferase constructs under the control of the 549 bp of 5′ flanking DNA containing the two most frequent extended haplotypes in this region. Luciferase activity was significantly reduced in cells transfected with the ‘mutant’ construct (−20C, −47C, −367C, −468G) c.f. the ‘wild‐type’ construct (−20T, −47T, −367T, −468C). These data suggest that polymorphisms have the potential to alter human β2 adrenergic receptor gene expression.

British Journal of Pharmacology (1999) 126, 841–844; doi:10.1038/sj.bjp.0702385
The effects of phosphodiesterase type 4 inhibitors on tumour necrosis factor‐α and leukotriene B4 in a novel human whole blood assay
Authors: Brideau Christine , Van Staden Carlo , Styhler Angela , Rodger Ian W , Chan Chi‐Chung
Published: January 2005

The aim of this study was to assess the inhibitory activities of phosphodiesterase type 4 (PDE4) inhibitors on tumour necrosis factor‐α (TNF‐α) and leukotriene B4 (LTB4) production in a novel human whole blood assay.

Lipopolysaccharide (LPS) stimulation of human whole blood caused a time dependent increase in TNF‐α and prostaglandin E2 (PGE2) plasma levels. Inhibition of LPS‐induced TNF‐α by the selective PDE4 inhibitor RP73401 was proportionally enhanced with endogenous PGE2 (maximal after 24 h). In contrast, blocking endogenous PGE2 production with indomethacin in blood stimulated with LPS for 24 h decreased the potency of RP73401 to that observed with a 4 h LPS incubation.

Non‐selective and selective PDE4 inhibitors showed greater inhibition of LPS‐induced TNF‐α after 24 h compared to 4 h. Stereoselectivity was only achieved in the 24 h method.

LPS‐stimulation of whole blood for either 30 min or 24 h followed by N‐formyl‐Met‐Leu‐Phe (fMLP) activation resulted in low plasma LTB4 levels. Combination of both treatments resulted in a greater than 7 fold increase in plasma LTB4 levels. Inhibition of the double LPS and fMLP‐activated LTB4 production was observed with non‐selective and PDE4‐selective inhibitors. Their LTB4 inhibitory potencies were similar to that observed in the 24 h LPS‐induced TNF‐α assay. Thus, stimulation of human whole blood with two LPS stimulations followed by fMLP gives rise to both TNF‐α and LTB4 and their inhibition by various compounds can be assessed in the same blood sample.

Calcium ionophore (A23187) stimulation of whole blood resulted in plasma LTB4 levels similar to the double LPS and fMLP method. Inhibition of A23187‐induced LTB4 biosynthesis was also achieved by PDE4‐selective inhibitors as well as the direct 5‐lipoxygenase (5‐LO) inhibitor L‐739,010.

These results confirm the anti‐inflammatory properties of PDE4 inhibitors. Thus, this novel human whole blood can be used to assess the biochemical efficacy of PDE4 inhibitors in human subjects.

British Journal of Pharmacology (1999) 126, 979–988; doi:10.1038/sj.bjp.0702387
Effect of low‐dose treatment with selegiline on dopamine transporter (DAT) expression and amphetamine‐induced dopamine release in vivo
Authors: Lamensdorf Itschak , Porat Shai , Simantov Rabi , Finberg John P M
Published: January 2005

Chronic treatment with low doses of the selective monoamine oxidase (MAO) type B inhibitors selegiline [(−)‐deprenyl] and rasagiline, causes elevation in extracellular level of 3,4‐dihydroxyphenylethylamine (dopamine) in the rat striatum in vivo (Lamensdorf et al., 1996). The present study was carried out to determine whether this effect of selegiline could be the result of an inhibition of the high‐affinity dopamine neuronal transport process.

Changes in activity of the dopamine transporter (DAT) in vivo following selegiline treatment were evaluated indirectly by microdialysis technique in the rat, from the change in striatal dopamine extracellular concentration following systemic amphetamine administration (4 mg kg−1, i.p.). Striatal levels of the DAT molecule were determined by immunoblotting. Uptake of [3H]‐dopamine was determined in synaptosomes from selegiline‐treated animals.

Amphetamine‐induced increase in striatal extracellular dopamine level was attenuated by one day and by chronic (21 days) treatment with selegiline (0.25 mg kg−1, s.c.).

Striatal levels of DAT were elevated after 1 and 21 days treatment with selegiline, but were not affected by clorgyline, rasagiline, nomifensine or amphetamine.

The increase in DAT expression, and attenuation of amphetamine‐induced dopamine release, were not accompanied by a change in [3H]‐dopamine uptake in synaptosomes of selegiline‐treated animals.

The results suggest that a reversible inhibition of dopamine uptake occurs following chronic low dose selegiline treatment in vivo which may be mediated by an increase in endogenous MAO‐B substrates such as 2‐phenylethylamine, rather than by the inhibitor molecule or its metabolites. Increased DAT expression appears to be a special property of the selegiline molecule, since it occurs after one low dose of selegiline, and is not seen with other inhibitors of MAO‐A or MAO‐B. The new DAT molecules formed following selegiline treatment appear not to be functionally active.

British Journal of Pharmacology (1999) 126, 997–1002; doi:10.1038/sj.bjp.0702389
Prostanoid receptors involved in the relaxation of human bronchial preparations
Authors: Norel Xavier , Walch Laurence , Labat Carlos , Gascard Jean‐Pierre , Dulmet Elisabeth , Brink Charles
Published: January 2005

Iloprost and cicaprost (IP‐receptor agonists) induced relaxations in the histamine‐ (50 μM) contracted human bronchial preparations (pD2 values, 6.63±0.12 and 6.86±0.08; Emax values, 90±04 and 65±08% of the papaverine response for iloprost (n=6) and cicaprost (n=3), respectively).

Prostaglandin E2 (PGE2) and misoprostol (EP‐receptor agonist) relaxed the histamine‐contracted human bronchial preparations (pD2 values, 7.13±0.07 and 6.33±0.28; Emax values, 67±04 and 57±08% of the papaverine response for PGE2 (n=14) and misoprostol (n=4), respectively). In addition, both relaxations were inhibited by AH6809 (DP/EP1/EP2‐receptor antagonist; 3 μM; n=5–6).

The PGE2‐induced relaxations of human bronchial preparations were not modified by treatment with AH23848B (TP/EP4‐receptor antagonist; 30 μM; n=4).

The contracted human bronchial preparations were significantly relaxed by prostaglandin D2 (PGD2) or by BW245C a DP‐receptor agonist. However, these responses did not exceed 40% of the relaxation induced by papaverine. In addition, the relaxations induced by PGD2 were significantly inhibited by treatment with a DP‐receptor antagonist BWA868C (0.1 μM; n=3).

These data suggest that the relaxation of human isolated bronchial preparations induced by prostanoids involved IP‐, EP2‐ and to a lesser extent DP‐receptors but not EP4‐receptor.

British Journal of Pharmacology (1999) 126, 867–872; doi:10.1038/sj.bjp.0702392
Role of potassium channels in the antinociception induced by agonists of α2‐adrenoceptors
Authors: Galeotti Nicoletta , Ghelardini Carla , Vinci Maria Cristina , Bartolini Alessandro
Published: January 2005

The effect of the administration of pertussis toxin (PTX) as well as modulators of different subtypes of K+ channels on the antinociception induced by clonidine and guanabenz was evaluated in the mouse hot plate test.

Pretreatment with pertussis toxin (0.25 μg per mouse i.c.v.) 7 days before the hot‐plate test, prevented the antinociception induced by both clonidine (0.08–0.2 mg kg−1, s.c.) and guanabenz (0.1–0.5 mg kg−1, s.c.).

The administration of the KATP channel openers minoxidil (10 μg per mouse, i.c.v.), pinacidil (25 μg per mouse, i.c.v.) and diazoxide (100 mg kg−1, p.o.) potentiated the antinociception produced by clonidine and guanabenz whereas the KATP channel blocker gliquidone (6 μg per mouse, i.c.v.) prevented the α2 adrenoceptor agonist‐induced analgesia.

Pretreatment with an antisense oligonucleotide (aODN) to mKv1.1, a voltage‐gated K+ channel, at the dose of 2.0 nmol per single i.c.v. injection, prevented the antinociception induced by both clonidine and guanabenz in comparison with degenerate oligonucleotide (dODN)‐treated mice.

The administration of the Ca2+‐gated K+ channel blocker apamin (0.5–2.0 ng per mouse, i.c.v.) never modified clonidine and guanabenz analgesia.

At the highest effective doses, none of the drugs used modified animals' gross behaviour nor impaired motor coordination, as revealed by the rota‐rod test.

The present data demonstrate that both KATP and mKv1.1 K+ channels represent an important step in the transduction mechanism underlying central antinociception induced by activation of α2 adrenoceptors.

British Journal of Pharmacology (1999) 126, 1214–1220; doi:10.1038/sj.bjp.0702395
Adenosine 5′‐triphosphate and neuropeptide Y are co‐transmitters in conjunction with noradrenaline in the human saphenous vein
Authors: Racchi Héctor , Irarrázabal Manuel J , Howard Michel , Morán Sergio , Zalaquett Ricardo , Huidobro‐Toro J Pablo
Published: January 2005

Human saphenous veins were used to assess the cooperative participation of adenosine 5‐triphosphate (ATP), neuropeptide Y (NPY), and noradrenaline (NA) in the vasomotor responses elicited following electrical depolarization of the perivascular nerve terminals. Rings from recently dissected human biopsies were mounted to record isometric muscular contractions; the motor activity elicited in the circular muscle layer following electrical depolarization (2.5–20 Hz, 50 V, 0.5 msec) were recorded.

Incubation of the biopsies with either 100 nM tetrodotoxin (TTX) or 1 μM guanethidine abolished the vasomotor response elicited by electrical nerve depolarization. The independent application of either ATP or NA to vein rings induced concentration‐dependent contractions.

Tissue incubation with 30 μM suramin or 10 nM prazosin produced 10 fold rightward displacements of the α,β‐methylene ATP and NA concentration‐response curves respectively. NPY contracted a limited number of biopsies, the vasoconstriction elicited was completely blocked by 1 μM BIBP 3226. A 5 min incubation of the biopsies with 10–100 nM NPY synergized, in a concentration‐dependent fashion, both the ATP and the ATP analogue‐induced contractions. Likewise, tissue preincubation with 10 nM NPY potentiated the vasomotor responses evoked with 20–60 nM NA.

Neither suramin, BIBP 3226, nor prazosin was individually able to significantly modify the derived frequency‐tension curves. In contrast, the co‐application of 30 μM suramin and 10 nM prazosin or 30 μM suramin and 1 μM BIBP 3226, elicited a significant (P<0.01) downward displacement of the respective frequency‐tension curves.

The simultaneous application of the three antagonists–30 μM suramin, 1 μM BIBP 3226 and 10 nM prazosin–caused a significantly greater displacement of the frequency‐tension curve than that achieved in experiments using two of these antagonists.

Electrically‐evoked vasomotor activity is blocked to a larger extent by tissue incubation with 2.5 μM chloroethylclonidine and 30 μM suramin rather than with 10 nM 5 methyl urapidil and 30 μM suramin. As a result, the α1‐adrenoceptor involved in the vasomotor activity has tentatively been associated with the α1B adrenoceptor family subtype.

Results support the physiological role of ATP in sympathetic neurotransmission. The present results are consistent with the working hypothesis that human sympathetic vasomotor reflexes involve the coordinated motor action of ATP, NPY, and NA acting on vascular smooth muscle cells. The present results support the concept of sympathetic co‐transmission in the human saphenous vein.

British Journal of Pharmacology (1999) 126, 1175–1185; doi:10.1038/sj.bjp.0702396
Inhibition by simvastatin, but not pravastatin, of glucose‐induced cytosolic Ca2+ signalling and insulin secretion due to blockade of L‐type Ca2+ channels in rat islet β‐cells
Authors: Yada Toshihiko , Nakata Masanori , Shiraishi Tomoko , Kakei Masafumi
Published: January 2005

Hypercholesterolaemia often occurs in patients with type 2 diabetes, who therefore encounter administration of HMG‐CoA reductase inhibitors. Alteration of pancreatic β‐cell function leading to an impaired insulin secretory response to glucose plays a crucial role in the pathogenesis of type 2 diabetes. Therefore, it is important to examine the effects of HMG‐CoA reductase inhibitors on β‐cell function.

Cytosolic Ca2+ concentration ([Ca2+]i) plays a central role in the regulation of β‐cell function. The present study examined the effects of HMG‐CoA reductase inhibitors on the glucose‐induced [Ca2+]i signalling and insulin secretion in rat islet β‐cells.

Simvastatin, a lipophilic HMG‐CoA reductase inhibitor, at 0.1–3 μg ml−1 concentration‐dependently inhibited the first phase increase and oscillation of [Ca2+]i induced by 8.3 mM glucose in single β‐cells. The less lipophilic inhibitor, simvastatin‐acid, inhibited the first phase [Ca2+]i increase but was two orders of magnitude less potent. The hydrophilic inhibitor, pravastatin (100 μg ml−1), was without effect on [Ca2+]i.

Simvastatin (0.3 μg ml−1), more potently than simvastatin‐acid (30 μg ml−1), inhibited glucose‐induced insulin secretion from islets, whereas pravastatin (100 μg ml−1) had no effect.

Whole‐cell patch clamp recordings demonstrated a reversible inhibition of the β‐cell L‐type Ca2+ channels by simvastatin, but not by pravastatin. Simvastatin also inhibited the [Ca2+]i increases by L‐arginine and KCl, agents that act via opening of L‐type Ca2+ channels.

In conclusion, lipophilic HMG‐CoA reductase inhibitors can inhibit glucose‐induced [Ca2+]i signalling and insulin secretion by blocking L‐type Ca2+ channels in β‐cells, and their inhibitory potencies parallel their lipophilicities. Precaution should be paid to these findings when HMG‐CoA reductase inhibitors are used clinically, particularly in patients with type 2 diabetes.

British Journal of Pharmacology (1999) 126, 1205–1213; doi:10.1038/sj.bjp.0702397
Distinction between surmountable and insurmountable selective AT1 receptor antagonists by use of CHO‐K1 cells expressing human angiotensin II AT1 receptors
Authors: Vanderheyden P M L , Fierens F L P , De Backer J P , Fraeyman N , Vauquelin G
Published: January 2005

CHO‐K1 cells that were stably transfected with the gene for the human AT1 receptor (CHO‐AT1 cells) were used for pharmacological studies of non‐peptide AT1 receptor antagonists.

In the presence of 10 mM LiCl, angiotensin II caused a concentration‐dependent and long‐lasting increase of inositol phosphates accumulation with an EC50 of 3.4 nM. No angiotensin II responses are seen in wild‐type CHO‐K1 cells.

[3H]‐Angiotensin II bound to cell surface AT1 receptors (dissociates under mild acidic conditions) and is subject to rapid internalization.

Non‐peptide selective AT1 antagonists inhibited the angiotensin II (0.1 μM) induced IP accumulation and the binding of [3H]‐angiotensin II (1 nM) with the potency order: candesartan > EXP3174 > irbesartan > losartan. Their potencies are lower in the presence of bovine serum albumin.

Preincubation with the insurmountable antagonist candesartan decreased the maximal angiotensin II induced inositol phosphate accumulation up to 94% and, concomitantly, decreased the maximal binding capacity of the cell surface receptors. These inhibitory effects were half‐maximal for 0.6 nM candesartan and were attenuated by simultaneous preincubation with 1 μM losartan indicating a syntopic action of both antagonists.

Losartan caused a parallel rightward shift of the angiotensin II concentration‐response curves and did not affect the maximal binding capacity. EXP3174 (the active metabolite of losartan) and irbesartan showed a mixed‐type behavior in both functional and binding studies.

Reversal of the inhibitory effect was slower for candesartan as compared with EXP3174 and irbesartan and it was almost instantaneous for losartan, suggesting that the insurmountable nature of selective AT1 receptor antagonists in functional studies was related to their long‐lasting inhibition.

British Journal of Pharmacology (1999) 126, 1057–1065; doi:10.1038/sj.bjp.0702398
Actions of 3‐[2′‐phosphonomethyl[1,1′‐biphenyl]‐3‐yl]alanine (PMBA) on cloned glycine receptors
Authors: Hosie Alastair M , Akagi Hiroyuki , Ishida Michiko , Shinozaki Haruhiko
Published: January 2005

PMBA is a novel antagonist of strychnine‐sensitive glycine receptors in the rat spinal cord, however, its mode of action is unknown. The actions of PMBA on rat glycine receptor α1 and α2 homomers in Xenopus oocytes were studied under two‐electrode voltage‐clamp.

Co‐application of PMBA and glycine to both α1 and α2 homomers yielded inward currents which decayed to a steady‐state. Responses rose slowly to the same steady‐state amplitude following a 2 min pre‐incubation in PMBA. Strychnine, but not picrotoxinin, showed similar antagonism to PMBA. The potency of PMBA was independent of membrane potential between −100 and 0 mV.

When tested against EC50 concentrations of glycine, PMBA was almost equally potent on α1 (IC50, 406±41 nM: Hill coefficient, 1.5±0.2) and α2 (IC50, 539±56 nM; Hill coefficient, 1.4±0.2) homomers.

PMBA (1–10 μM) and strychnine (200 nM) reduced the potency of glycine and the amplitude of the maximal agonist response of α1 and α2 homomers. In 10 μM PMBA, two distinct classes of glycine response were observed on α2, only a single class of responses were observed on α1.

There are similarities in PMBA and strychnine antagonism, although these compounds are structurally distinct. The possibility that PMBA interacts at two binding sites which differ in α1 and α2 subunits is discussed. PMBA may provide a lead structure for novel antagonists with which to investigate structural differences in glycine receptor at α1 and α2 subunits.

British Journal of Pharmacology (1999) 126, 1230–1236; doi:10.1038/sj.bjp.0702402
Inverse agonist activity of pirenzepine at M2 muscarinic acetylcholine receptors
Authors: Daeffler Laurent , Schmidlin Fabien , Gies Jean‐Pierre , Landry Yves
Published: January 2005

The intrinsic properties of muscarinic ligands were studied through their binding properties and their abilities to modulate the GTPase activity of G proteins coupled to muscarinic M2 receptors in pig atrial sarcolemma.

Competition binding experiments were performed with [3H]‐oxotremorine‐M to assess the affinity of receptors coupled to G proteins (R*), with [3H]‐N‐methylscopolamine ([3H]‐NMS) to estimate the affinities of coupled and uncoupled receptors (R*+R) and with [3H]‐NMS in the presence of GppNHp to assess the affinity of uncoupled receptors (R).

The ranking of Ki values for the agonist carbachol was R*<<R*+R<R*+R>R (174, 155, 115 nM), suggesting inverse agonism.

The Vmax of the basal high affinity GTPase activity of pig atrial sarcolemma was increased by mastoparan and decreased by GPAnt‐2 indicating the relevance of this activity to G proteins coupled to receptors (R*). The KM value (0.26–0.33 μM) was not modified by mastoparan or GPAnt‐2.

Carbachol increased the Vmax of GTP hydrolysis (EC50 8.1±0.3 μM), whereas atropine and AF‐DX 116, up to 1 mM, did not modify it. Pirenzepine decreased the Vmax of GTP hydrolysis (EC50 77.5±10.3 μM). This effect was enhanced when KCl was substituted for NaCl (EC50 11.0±0.8 μM) and was antagonized by atropine and AF‐DX 116 (IC50 0.91±0.71 and 197±85 nM).

Pirenzepine is proposed as an inverse agonist and atropine and AF‐DX 116 as neutral antagonists at the muscarinic M2 receptor.

British Journal of Pharmacology (1999) 126, 1246–1252; doi:10.1038/sj.bjp.0702407
Waterton and Wouralia†
Authors: Birmingham A T
Published: January 2005

British Journal of Pharmacology (1999) 126, 1685–1689; doi:10.1038/sj.bjp.0702409
Activation of three types of voltage‐independent Ca2+ channel in A7r5 cells by endothelin‐1 as revealed by a novel Ca2+ channel blocker LOE 908
Authors: Iwamuro Yasushi , Miwa Soichi , Zhang Xiao‐Feng , Minowa Tetsuya , Enoki Taijiro , Okamoto Yasuo , Hasegawa Hiroshi , Furutani Hidekatsu , Okazawa Makoto , Ishikawa Masatsune , Hashimoto Nobuo , Masaki Tomoh
Published: January 2005

We have shown that in addition to voltage‐operated Ca2+ channel (VOC), endothelin‐1 (ET‐1) activates two types of Ca2+‐permeable nonselective cation channel (NSCC) in A7r5 cells: its lower concentrations (1 nM; lower [ET‐1]) activate only an SK&F 96365‐resistant channel (NSCC‐1), whereas its higher concentrations (10 nM; higher [ET‐1]) activate an SK&F 96365‐sensitive channel (NSCC‐2) as well.

We now characterized the effects of a blocker ofCa2+ entry channel LOE 908 on NSCCs and store‐operatedCa2+ channel (SOCC) in A7r5 cells, and using twodrugs, clarified the involvement of these channels in the ET‐1‐inducedincrease in the intracellular free Ca2+ concentrations([Ca2+]i). Whole‐cell recordingsand [Ca2+]i monitoring withfluo‐3 were used.

LOE 908 up to 10 μM had no effect on increases in [Ca2+]i induced by thapsigargin or ionomycin, but SK&F 96365 abolished them.

In the cells clamped at −60 mV, both lower and higher [ET‐1] induced inward currents with linear iv relationships and the reversal potentials of −15.0 mV. Thapsigargin induced no currents.

In the presence of nifedipine, lower [ET‐1] induced a sustained increase in [Ca2+]i, whereas higher [ET‐1] induced a transient peak and a sustained increase. The sustained increases by lower and higher [ET‐1] were abolished by removal of extracellular Ca2+, and they were suppressed by LOE 908 to 0 and 35%, respectively, with the LOE 908‐resistant part being abolished by SK&F 96365.

These results show that LOE 908 is a blocker of NSCCs without effect on SOCC, and that the increase in [Ca2+]i at lower [ET‐1] results from Ca2+ entry through NSCC‐1 in addition to VOC, whereas the increase at higher [ET‐1] involves NSCC‐1, NSCC‐2 and SOCC in addition to VOC.

British Journal of Pharmacology (1999) 126, 1107–1114; doi:10.1038/sj.bjp.0702416
Electrophysiological studies on the postnatal development of the spinal antinociceptive effects of the delta opioid receptor agonist DPDPE in the rat
Authors: Rahman Wahida , Dickenson Anthony H
Published: January 2005

The antinociceptive effects of the delta opioid receptor selective agonist, DPDPE [(D‐Pen2,D‐Pen5)‐enkephalin] was studied in rats aged postnatal day (P) 14, P21, P28 and P56.

Antinociceptive effects of DPDPE were measured as percentage inhibition of the C‐fibre evoked response and post‐discharge of dorsal horn neurones evoked by peripheral electrical stimulation. DPDPE was administered by topical application, akin to intrathecal injection.

DPDPE (0.1–100 μg) produced dose‐related inhibitions at all ages; these inhibitions were reversed by 5 μg of the opioid antagonist naloxone.

The dose‐response curves for C‐fibre evoked response and post‐discharge of the neurones were not different in rats aged P14 and P21. DPDPE was significantly more potent at P14 and P21 compared with its inhibitory effects on these responses at P28 and P56.

DPDPE produced minor inhibitions of the A‐fibre evoked response of the neurones at P14, P21, P28 and P56, suggesting that the inhibitory effects of DPDPE are mediated via presynaptic receptors on the terminals of C‐fibre afferents.

Since spinal delta opioid receptor density changes little over this period, the increased antinociceptive potency of DPDPE in the rat pups compared with the adult is likely to be due to post‐receptor events, or in developmental changes in the actions of other transmitter/receptor systems within the spinal cord.

British Journal of Pharmacology (1999) 126, 1115–1122; doi:10.1038/sj.bjp.0702418
Inhibition of GABA‐gated chloride channels by 12,14‐dichlorodehydroabietic acid in mammalian brain
Authors: Nicholson Russell A , Lees George , Zheng Jian , Verdon Bernard
Published: January 2005

12,14‐dichlorodehydroabietic acid (12,14‐Cl2DHA) reduced GABA‐stimulated uptake of 36Cl− into mouse brain synaptoneurosomes suggesting inhibition of mammalian GABAA receptor function.

12,14‐Cl2DHA did not affect the binding of [3H]‐muscimol to brain membranes but displaced specifically bound [3H]‐EBOB. The inhibitory effect on [3H]‐EBOB binding was not reversible. 12,14‐Cl2DHA reduced the availability of [3H]‐EBOB binding sites (Bmax) without changing the KD of the radioligand for remaining sites. 12,14‐Cl2DHA did not affect the rate of association of [3H]‐EBOB with its chloride channel receptor, but increased the initial rate of [3H]‐EBOB dissociation.

12,14‐Cl2DHA enhanced the incidence of EPSCs when rapidly applied to cultured rat cortical neurones. Longer exposures produced block of IPSCs with marked increases in the frequency of EPSCs and min EPSCs. 12,14‐Cl2DHA also irreversibly suppressed chloride currents evoked by pulses of exogenous GABA in these cells.

Ultimately, 12,14‐Cl2DHA inhibited all synaptic traffic and action currents in current clamped cells indicating that, in contrast to picrotoxinin (which causes paroxysmal bursting), it is not fully selective for the GABAA receptor‐chloride channel complex.

The depolarizing block seen with 12,14‐Cl2DHA in amphotericin‐perforated preparations implicates loss of Ca2+ buffering in the polarity change and this may account for inhibition of spontaneous action potentials.

Our investigation demonstrates that 12,14‐Cl2DHA blocks GABA‐dependent chloride entry in mammalian brain and operates as a non‐competitive insurmountable GABAA antagonist. The mechanism likely involves either irreversible binding of 12,14‐Cl2DHA to the trioxabicyclooctane recognition site or a site that is allosterically coupled to it. We cannot exclude, however, the possibility that 12,14‐Cl2DHA causes localized proteolysis or more extensive conformational change within a critical subunit of the chloride channel.

British Journal of Pharmacology (1999) 126, 1123–1132; doi:10.1038/sj.bjp.0702419
Limited anti‐inflammatory efficacy of cyclo‐oxygenase‐2 inhibition in carrageenan‐airpouch inflammation
Authors: Wallace John L , Chapman Kevin , McKnight Webb
Published: January 2005

Cyclo‐oxygenase‐2 (COX‐2) is expressed at sites of inflammation and is believed to be the major source of inflammation‐associated prostaglandin synthesis. Selective inhibition of COX‐2 has been suggested to produce anti‐inflammatory effects with reduced toxicity in the gastrointestinal tract. We examined the extent to which suppression of COX‐2 led to inhibition of various components of inflammation in the carrageenan‐airpouch model in the rat.

Indomethacin (0.3 mg kg−1), nimesulide (3 mg kg−1) and the selective COX‐2 inhibitor, SC‐58125 (0.3 mg kg−1), significantly suppressed the production of prostaglandin E2 at the site of inflammation. At higher doses, indomethacin (1 mg kg−1) and nimesulide (30 mg kg−1), but not SC‐58125 (up to 10 mg kg−1), significantly inhibited COX‐1 activity (as measured by whole blood thromboxane synthesis).

All three test drugs significantly reduced the volume of exudate in the airpouch, but only at doses greater than those required for substantial (>90%) suppression of COX‐2 activity. Similarly, reduction of leukocyte infiltration was only observed with the doses of indomethacin and nimesulide that caused significant suppression of COX‐1 activity.

SC‐58125 did not significantly affect leukocyte infiltration into the airpouch at any dose tested (up to 10 mg kg−1). A second selective COX‐2 inhibitor, Dup‐697, was also found to suppress exudate PGE2 levels without significant effects on leukocyte infiltration.

These results indicate that selective inhibition of COX‐2 results in profound suppression of PGE2 synthesis in the carrageenan‐airpouch, but does not affect leukocyte infiltration. Exudate volume was only reduced with the highly selective COX‐2 inhibitor when a dose far above that necessary for suppression of COX‐2 activity was used. Inhibition of leukocyte infiltration was observed with indomethacin and nimesulide, but only at doses that inhibited both COX‐1 and COX‐2.

British Journal of Pharmacology (1999) 126, 1200–1204; doi:10.1038/sj.bjp.0702420
Protective effect of cyclosporin A and FK506 from nitric oxide‐dependent apoptosis in activated macrophages
Authors: Hortelano Sonsoles , López‐Collazo Eduardo , Boscá Lisardo
Published: January 2005

Activation of macrophages with lipopolysaccharide (LPS) and low doses of interferon‐γ (IFN‐γ) induced apoptotic death through a nitric oxide‐dependent pathway.

Treatment of cells with the immunosuppressors cyclosporin A (CsA) or FK506 inhibited the activation‐dependent apoptosis.

These drugs decreased the up‐regulation of p53 and Bax characteristic of activated macrophages. Moreover, incubation of activated macrophages with CsA and FK506 contributed to maintain higher levels of Bcl‐2 than in LPS/IFN‐γ treated cells.

The inhibition of apoptosis exerted by CsA and FK506 in macrophages was also observed when cell death was induced by treatment with chemical nitric oxide donors.

Incubation of macrophages with LPS/IFN‐γ barely affected caspase‐1 but promoted an important activation of caspase‐3. Both CsA and FK506 inhibited pathways leading to caspase‐3 activation. Moreover, the cleavage of poly(ADP‐ribose) polymerase, a well established caspase substrate, was reduced by these immunosuppressive drugs.

CsA and FK506 reduced the release of cytochrome c to the cytosol and the activation of caspase‐3 in cells treated with nitric oxide donors.

These results indicate that CsA and FK506 protect macrophages from nitric oxide‐dependent apoptosis and suggest a contribution of the macrophage to innate immunity under conditions of immunosuppression of the host.

British Journal of Pharmacology (1999) 126, 1139–1146; doi:10.1038/sj.bjp.0702422