bluuebunny/crossref_metadata_2025 huggingface dataset:
Block by fluoxetine of volume‐regulated anion channels
Authors: Maertens Chantal , Wei Lin , Voets Thomas , Droogmans Guy , Nilius Bernd
Published: January 2005

We have used the whole‐cell patch clamp technique to study the effect of fluoxetine, a commonly used antidepressant drug, on the volume‐regulated anion channel (VRAC) in calf pulmonary artery endothelial (CPAE) cells. We also examined its effects on other Cl− channels, i.e. the Ca2+‐activated Cl− current (ICl,Ca) and the cystic fibrosis transmembrane conductance regulator (CFTR) to assess the specificity of this compound for VRAC.

At pH 7.4 fluoxetine induced a fast and reversible block of the volume‐sensitive chloride current (ICl,swell), with a Ki value of 6.0±0.5 μM (n=6‐9). The blocking efficiency increased with increasing extracellular pH (Ki=0.32±0.01 μM at pH 8.8, n=3‐9), indicating that the blockade is mediated by the uncharged form of fluoxetine.

Fluoxetine inhibited Ca2+‐activated Cl− currents, ICl,Ca, activated by loading CPAE cells via the patch pipette with 1000 nM free Ca2+ (Ki=10.7±1.6 μM at pH 7.4, n=3‐5). The CFTR channel, transiently transfected in CPAE cells, was also inhibited with a Ki value of 26.9±9.4 μM at pH 7.4 (n=3).

This study describes for the first time the effects of fluoxetine on anion channels. Our data reveal a potent block of VRAC at fluoxetine concentrations close to plasma concentrations. The results suggest a hydrophobic interaction with high affinity between uncharged fluoxetine and volume‐activated chloride channels. Ca2+‐activated Cl− currents and CFTR are also blocked by fluoxetine, revealing a novel characteristic of the drug as a chloride channel modulator.

British Journal of Pharmacology (1999) 126, 508–514; doi:10.1038/sj.bjp.0702314
Cell type‐specific ATP‐activated responses in rat dorsal root ganglion neurons
Authors: Ueno Shinya , Tsuda Makoto , Iwanaga Toshihiko , Inoue Kazuhide
Published: January 2005

The aim of our study is to clarify the relationship between expression pattern of P2X receptors and the cell type of male adult rat (Wistar) dorsal root ganglion (DRG) neurons. We identified the nociceptive cells of acutely dissociated DRG neurons from adult rats type using capsaicin sensitivity.

Two types of ATP‐activated currents, one with fast, the other with slow desensitization, were found under voltage‐clamp conditions. In addition, cells with fast but not slow desensitization responded to capsaicin, indicating that there was a relationship between current kinetics and capsaicin‐sensitivity.

Both types of neurons were responsive to ATP and α, β methylene‐ATP (α,βmeATP). The concentration of α,βmeATP producing half‐maximal activation (EC50) of neurons with fast desensitization was less (11 μM) than that of neurons with slow desensitization (63 μM), while the Hill coefficients were similar. Suramin and pyridoxal‐phosphate‐6‐azophenyl‐2′,4′‐disulphonic acid tetrasodium (PPADS) antagonized α,βmeATP‐induced currents in both types of neurons.

In situ hybridization revealed that small cells of the DRG predominantly expressed mRNAs of P2X3 and medium‐sized cells expressed mRNAs of P2X2 and P2X3. In contrast, both of mRNAs were not detected in large cells of the DRG.

These results suggest that capsaicin‐sensitive, small‐sized DRG neurons expressed mainly the homomeric P2X3 subunit and that capsaicin‐insensitive, medium‐sized DRG neurons expressed the heteromultimeric receptor with P2X2 and P2X3.

British Journal of Pharmacology (1999) 126, 429–436; doi:10.1038/sj.bjp.0702319
Endogenous nitric oxide in the maintenance of rat microvascular integrity against widespread plasma leakage following abdominal laparotomy
Authors: László Ferenc , Whittle Brendan J R
Published: January 2005

The role of nitric oxide (NO) in the maintenance of microvascular integrity during minor surgical manipulation has been evaluated in the rat.

The NO synthase inhibitors, NG‐nitro‐L‐arginine methyl ester (L‐NAME, 5 mg kg−1, s.c.) and NG‐monomethyl‐L‐arginine (L‐NMMA, 50 mg kg−1, s.c.) had no effect on microvascular leakage of radiolabelled albumin over 1 h in the stomach, duodenum, jejunum, colon, lung and kidney in the un‐operated conscious or pentobarbitone‐anaesthetized rat.

In contrast, in anaesthetized rats with a midline abdominal laparotomy (5 cm), L‐NAME (1–5 mg kg−1, s.c.) or L‐NMMA (12.5–50 mg kg−1, s.c.) dose‐dependently increased gastrointestinal, renal and pulmonary vascular leakage, effects reversed by L‐arginine pretreatment (300 mg kg−1, s.c., 15 min). These actions were not observed in anaesthetized rats that had only received a midline abdominal skin incision (5 cm).

Pretreatment with a rabbit anti‐rat neutrophil serum (0.4 ml kg−1, i.p.), 4 h before laparotomy, abolished the plasma leakage induced by L‐NAME in all the organs investigated.

These results indicate that the following abdominal laparotomy, inhibition of constitutive NO synthase provokes vascular leakage in the general microcirculation, by a process that may involve neutrophils. Such effects could thus confound studies on the microvascular actions of NO synthase inhibitors using acute surgically prepared in vivo models. The findings thus suggest that constitutively‐formed NO has a crucial role in the maintenance of acute microvascular integrity following abdominal surgical intervention.

British Journal of Pharmacology (1999) 126, 515–521; doi:10.1038/sj.bjp.0702322
Neurogenic plasma leakage in mouse airways
Authors: Baluk Peter , Thurston Gavin , Murphy Thomas J , Bunnett Nigel W , McDonald Donald M
Published: January 2005

This study sought to determine whether neurogenic inflammation occurs in the airways by examining the effects of capsaicin or substance P on microvascular plasma leakage in the trachea and lungs of male pathogen‐free C57BL/6 mice.

Single bolus intravenous injections of capsaicin (0.5 and 1 μmol kg−1, i.v.) or substance P (1, 10 and 37 nmol kg−1, i.v.) failed to induce significant leakage in the trachea, assessed as extravasation of Evans blue dye, but did induce leakage in the urinary bladder and skin.

Pretreatment with captopril (2.5 mg kg−1, i.v.), a selective inhibitor of angiotensin converting enzyme (ACE), either alone or in combination with phosphoramidon (2.5 mg kg−1, i.v.), a selective inhibitor of neutral endopeptidase (NEP), increased baseline leakage of Evans blue in the absence of any exogenous inflammatory mediator. The increase was reversed by the bradykinin B2 receptor antagonist Hoe 140 (0.1 mg kg−1, i.v.).

After pretreatment with phosphoramidon and captopril, capsaicin increased the Evans blue leakage above the baseline in the trachea, but not in the lung. This increase was reversed by the tachykinin (NK1) receptor antagonist SR 140333 (0.7 mg kg−1, i.v.), but not by the NK2 receptor antagonist SR 48968 (1 mg kg−1, i.v.).

Experiments using Monastral blue pigment as a tracer localized the leakage to postcapillary venules in the trachea and intrapulmonary bronchi, although the labelled vessels were less numerous in mice than in comparably treated rats. Blood vessels of the pulmonary circulation were not labelled.

We conclude that neurogenic inflammation can occur in airways of pathogen‐free mice, but only after the inhibition of enzymes that normally degrade inflammatory peptides. Neurogenic inflammation does not involve the pulmonary microvasculature.

British Journal of Pharmacology (1999) 126, 522–528; doi:10.1038/sj.bjp.0702323
Modulation of ET‐1‐induced contraction of human bronchi by airway epithelium‐dependent nitric oxide release via ETA receptor activation
Authors: Naline Emmanuel , Bertrand Claude , Biyah Keltoum , Fujitani Yasushi , Okada Toshikazu , Bisson Alain , Advenier Charles
Published: January 2005

The purpose of this work was to investigate whether endothelin‐1 (ET‐1) was able to induce the release of an inhibitory factor from the airway epithelium in isolated human bronchi and to identify this mediator as well as the endothelin receptor involved in this phenomenon.

In intact bronchi, ET‐1 induced a concentration‐dependent contraction (−logEC50=7.92±0.09, n=18) which was potentiated by epithelium removal (−logEC50=8.65±0.11, n=17). BQ‐123, an ETA receptor antagonist, induced a significant leftward shift of the ET‐1 concentration‐response curve (CRC). This leftward shift was abolished after epithelium removal.

L‐NAME (3×10−3 M), an inhibitor of nitric oxide (NO) synthase, induced a significant leftward shift of the ET‐1 CRC, and abolished the potentiation by BQ‐123 (10−8 M) of ET‐1‐induced contraction.

In intact preparations, the ETB receptor antagonist BQ‐788 induced only at 10−5 M a slight rightward shift of the ET‐1 CRC. In contrast, in epithelium‐denuded bronchi or in intact preparations in the presence of L‐NAME, BQ‐788 displayed a non‐competitive antagonism toward ET‐1‐induced contraction.

IRL 1620, a selective ETB receptor agonist, induced a contraction of the isolated bronchus (−logEC50=7.94±0.11, n=19). This effect was not modified by epithelium removal or by BQ‐123. BQ‐788 exerted a competitive antagonism against IRL 1620 which was similar in the presence or absence of epithelium.

These results show that ET‐1 exerts two opposite effects on the human airway smooth muscle. One is contractile via ETB‐receptor activation, the other is inhibitory and responsible of NO release which counteracts via ETA‐receptor activation the contraction.

British Journal of Pharmacology (1999) 126, 529–535; doi:10.1038/sj.bjp.0702327
Inhibitory effects of aprindine on the delayed rectifier K+ current and the muscarinic acetylcholine receptor‐operated K+ current in guinea‐pig atrial cells
Authors: Ohmoto‐Sekine Yuki , Uemura Hiroko , Tamagawa Masaji , Nakaya Haruaki
Published: January 2005

In order to clarify the mechanisms by which the class Ib antiarrhythmic drug aprindine shows efficacy against atrial fibrillation (AF), we examined the effects of the drug on the repolarizing K+ currents in guinea‐pig atrial cells by use of patch‐clamp techniques. We also evaluated the effects of aprindine on experimental AF in isolated guinea‐pig hearts.

Aprindine (3 μM) inhibited the delayed rectifier K+ current (IK) with little influence on the inward rectifier K+ current (IK1) or the Ca2+ current. Electrophysiological analyses including the envelope of tails test revealed that aprindine preferentially inhibits IKr (rapidly activating component) but not IKs (slowly activating component).

The muscarinic acetylcholine receptor‐operated K+ current (IK.ACh) was activated by the extracellular application of carbachol (1 μM) or by the intracellular loading of GTPγS. Aprindine inhibited the carbachol‐ and GTPγS‐induced IK.ACh with the IC50 values of 0.4 and 2.5 μM, respectively.

In atrial cells stimulated at 0.2 Hz, aprindine (3 μM) per se prolonged the action potential duration (APD) by 50±4%. The drug also reversed the carbachol‐induced action potential shortening in a concentration‐dependent manner.

In isolated hearts, perfusion of carbachol (1 μM) shortened monophasic action potential (MAP) and effective refractory period (ERP), and lowered atrial fibrillation threshold. Addition of aprindine (3 μM) inhibited the induction of AF by prolonging MAP and ERP.

We conclude the efficacy of aprindine against AF may be at least in part explained by its inhibitory effects on IKr and IK.ACh.

British Journal of Pharmacology (1999) 126, 751–761; doi:10.1038/sj.bjp.0702334
Vasorelaxation and inhibition of the voltage‐operated Ca2+ channels by FK506 in the porcine coronary artery
Authors: Yasutsune Toru , Kawakami Nozomi , Hirano Katsuya , Nishimura Junji , Yasui Hisataka , Kitamura Kenji , Kanaide Hideo
Published: January 2005

Using fura‐2 fluorometry, the effects of FK506, an immunosuppressant, on changes in cytosolic Ca2+ concentrations ([Ca2+]i) and tension were investigated in porcine coronary arterial strips. The effects of FK506 on the activity of voltage‐operated Ca2+ channels were examined by applying a whole cell patch clamp to the isolated smooth muscle cells of porcine coronary artery.

FK506 inhibited the sustained increases in both [Ca2+]i and tension induced by 118 mM K+ depolarization and 100 nM U46619 in a concentration‐dependent manner (1–30 μM). The extent of inhibition of the K+‐induced contraction was greater than that of the U46619‐induced contraction. The increases in [Ca2+]i and tension induced by histamine and endothelin‐1 in the presence of extracellular Ca2+ were also inhibited by 10 μM FK506.

FK506 (10 μM) had no effect on Ca2+ release induced by caffeine or by histamine in the Ca2+‐free solution.

FK506 (10 μM) had no effect on the [Ca2+]i‐tension relationships of the contractions induced by cumulative increases of extracellular Ca2+ during K+ depolarization or stimulation with U46619.

In the patch clamp experiments, FK506 (30 μM) partially inhibited the inward current induced by depolarization pulse from −80 mV to 0 mV.

In conclusion, FK506 induces arterial relaxation by decreasing [Ca2+]i mainly due to the inhibition of the L‐type Ca2+ channels, with no effect on the Ca2+ sensitivity of the contractile apparatus.

British Journal of Pharmacology (1999) 126, 717–729; doi:10.1038/sj.bjp.0702339
Effect of prolonged administration of a urinary kininase inhibitor, ebelactone B on the development of deoxycorticosterone acetate‐salt hypertension in rats
Authors: Ito Hiroshi , Majima Masataka , Nakajima Shin‐ichi , Hayashi Izumi , Katori Makoto , Izumi Tohru
Published: January 2005

The effect of prolonged administration of a carboxypeptidase Y‐like kininase inhibitor, ebelactone B (EB) (2‐ethyl‐3, 11‐dihydroxy‐4, 6, 8, 10, 12‐pentamethyl‐9‐oxo‐6‐tetradecenoic 1, 3‐lactone), on the development of deoxycorticosterone acetate (DOCA)‐salt hypertension was tested.

The systolic blood pressure (SBP) of non‐treated 6‐week‐old Sprague‐Dawley strain rats was gradually increased by DOCA‐salt treatment from 137±2 mmHg (n=11) to 195±7 mmHg at 10 weeks of age.

With daily oral administration of lisinopril (5 mg kg−1, twice a day), which is an inhibitor of angiotensin converting enzyme, a major kininase in plasma, the development of hypertension was not suppressed.

By contrast, administration of EB (5 mg kg−1, twice a day), completely inhibited the development of hypertension (SBP: 146±1 mmHg, n=5, 10 weeks old). The reduced SBP at 10 weeks of age was equal to the SBP before any treatment (142±1 mmHg, n=5).

Direct determination of mean blood pressure (MBP) in conscious, unrestrained rats confirmed that MBP elevation was completely inhibited by EB.

Continuous subcutaneous infusion (5 mg kg−1 day−1) of HOE140, a bradykinin B2 receptor antagonist, restored the elevation of SBP, which was suppressed by EB.

The weights of left ventricle of DOCA‐salt treated rats 10‐weeks‐old (0.36±0.02 g 100 g body weight−1, n=11) was significantly reduced by EB (0.27±0.01, n=5), as were the sodium levels in serum, cerebrospinal fluid and erythrocyte.

These findings suggested that EB is effective in preventing salt‐related hypertension presumably by eliminating sodium retention.

British Journal of Pharmacology (1999) 126, 613–620; doi:10.1038/sj.bjp.0702340
Modulation of noradrenergic neuronal firing by selective serotonin reuptake blockers
Authors: Szabo Steven T , De Montigny Claude , Blier Pierre
Published: January 2005

Using in vivo extracellular unitary recording, the effect of short term (2‐day) and long‐term (21‐day) administration of the selective 5‐HT reuptake inhibitor (SSRI) paroxetine (10 mg kg−1 day−1, s.c. using osmotic minipumps) was examined on the spontaneous firing activity of locus coeruleus noradrenergic neurons. Long‐term but not short‐term treatment significantly decreased firing activity. Thus, it appears that enhancing 5‐HT neurotransmission by sustained SSRI administration leads to a reduction of the firing rate of noradrenergic neurons. The SSRI paroxetine therefore alters the activity of noradrenergic neurons with a delay that is consistent with its therapeutic action in depression and panic disorder.

British Journal of Pharmacology (1999) 126, 568–571; doi:10.1038/sj.bjp.0702343
Electrophysiological examination of the effects of sustained flibanserin administration on serotonin receptors in rat brain
Authors: Rueter Lynne E , Blier Pierre
Published: January 2005

5‐HT1A receptor agonists have proven to be effective antidepressant medications, however they suffer from a significant therapeutic lag before depressive symptoms abate. Flibanserin is a 5‐HT1A receptor agonist and 5‐HT2A receptor antagonist developed to possibly induce a more rapid onset of antidepressant action through its preferential postsynaptic 5‐HT1A receptor agonism.

Flibanserin antagonized the effect of microiontophoretically‐applied DOI in the medial prefrontal cortex (mPFC) following 2 days of administration, indicating antagonism of postsynaptic 5‐HT2A receptors. This reduction in the effect of locally‐applied DOI was no longer present following 7‐day flibanserin administration.

Two‐day flibanserin administration only marginally reduced the firing activity of dorsal raphe (DRN) 5‐HT neurons. Following 7 days of administration, 5‐HT neuronal firing activity had returned to normal and the somatodendritic 5‐HT1A autoreceptors were desensitized.

The responsiveness of postsynaptic 5‐HT1A receptors located on CA3 hippocampus pyramidal neurons and mPFC neurons, examined using microiontophoretically‐applied 5‐HT and gepirone, was unchanged following a 7‐day flibanserin treatment.

As demonstrated by the ability of the 5‐HT1A receptor antagonist WAY 100635 to selectively increase the firing of hippocampal neurons in 2‐ and 7‐day treated rats, flibanserin enhanced the tonic activation of postsynaptic 5‐HT1A receptors in this brain region.

The results suggest that flibanserin could be a therapeutically useful compound putatively endowed with a more rapid onset of antidepressant action.

British Journal of Pharmacology (1999) 126, 627–638; doi:10.1038/sj.bjp.0702344
Effects of vasopressin on the sympathetic contraction of rabbit ear artery during cooling
Authors: García‐Villalón A L , Padilla J , Monge L , Fernández N , Sánchez M A , Gómez B , Diéguez G
Published: January 2005

In order to analyse the effects of arginine‐vasopressin on the vascular contraction to sympathetic nerve stimulation during cooling, the isometric response of isolated, 2‐mm segments of the rabbit central ear (cutaneous) artery to electrical field stimulation (1–8 Hz) was recorded at 37 and 30°C.

Electrical stimulation (37°C) produced frequency‐dependent arterial contraction, which was reduced at 30°C and potentiated by vasopressin (10 pM, 100 pM and 1 nM). This potentiation was greater at 30 than at 37°C and was abolished at both temperatures by the antagonist of vasopressin V1 receptors d(CH2)5 Tyr(Me)AVP (100 nM). Desmopressin (1 μM) did not affect the response to electrical stimulation.

At 37°C, the vasopressin‐induced potentiation was abolished by the purinoceptor antagonist PPADS (30 μM), increased by phentolamine (1 μM) or prazosin (1 μM) and not modified by yohimbine (1 μM), whilst at 30°C, the potentiation was reduced by phentolamine, yohimbine or PPADS, and was not modified by prazosin.

The Ca2+‐channel blockers, verapamil (10 μM) and NiCl2 (1 mM), abolished the potentiating effects of vasopressin at 37°C whilst verapamil reduced and NiCl2 abolished this potentiation at 30°C. The inhibitor of nitric oxide synthesis, L‐NOARG (100 μM), or endothelium removal did not modify the potentiation by vasopressin at 37 and 30°C.

Vasopressin also increased the arterial contraction to the α2‐adrenoceptor agonist BHT‐920 (10 μM) and to ATP (2 mM) at 30 and 37°C, but it did not modify the contraction to noradrenaline (1 μM) at either temperature.

These results suggest that in cutaneous (ear) arteries, vasopressin potentiaties sympathetic vasoconstriction to a greater extent at 30 than at 37°C by activating vasopressin V1 receptors and Ca2+ channels at both temperatures. At 37°C, the potentiation appears related to activation of the purinoceptor component and, at 30°C, to activation of both purinoceptor and α2‐adrenoceptor components of the sympathetic response.

British Journal of Pharmacology (1999) 126, 785–793; doi:10.1038/sj.bjp.0702345
N‐substituted analogues of S‐nitroso‐N‐acetyl‐D,L‐penicillamine: chemical stability and prolonged nitric oxide mediated vasodilatation in isolated rat femoral arteries
Authors: Megson I L , Morton S , Greig I R , Mazzei F A , Field R A , Butler A R , Caron G , Gasco A , Fruttero R , Webb D J
Published: January 2005

Previous studies show that linking acetylated glucosamine to S‐nitroso‐N‐acetyl‐D,L‐penicillamine (SNAP) stabilizes the molecule and causes it to elicit unusually prolonged vasodilator effects in endothelium‐denuded, isolated rat femoral arteries. Here we studied the propanoyl (SNPP; 3 carbon side‐chain), valeryl (SNVP; 5C) and heptanoyl (SNHP; 7C) N‐substituted analogues of SNAP (2C), to further investigate other molecular characteristics that might influence chemical stability and duration of vascular action of S‐nitrosothiols.

Spectrophotometric analysis revealed that SNVP was the most stable analogue in solution. Decomposition of all four compounds was accelerated by Cu(II) and cysteine, and neocuproine, a specific Cu(I) chelator, slowed decomposition of SNHP. Generation of NO from the compounds was confirmed by electrochemical detection at 37°C.

Bolus injections of SNAP (10 μl; 10−8–10−3 M) into the perfusate of precontracted, isolated rat femoral arteries taken from adult male Wistar rats (400–500 g), caused concentration‐dependent, transient vasodilatations irrespective of endothelial integrity. Equivalent vasodilatations induced by SNVP and SNHP were transient in endothelium‐intact vessels but failed to recover to pre‐injection pressures at moderate and high concentrations (10−6–10−3 M) in those denuded of endothelium. This sustained effect (>1 h) was most prevalent with SNHP and was largely reversed by the NO scavenger, haemoglobin.

We suggest that increased lipophilicity of SNAP analogues with longer sidechains facilitates their retention by endothelium‐denuded vessels; subsequent slow decomposition within the tissue generates sufficient NO to cause prolonged vasodilatation. This is a potentially useful characteristic for targeting NO delivery to areas of endothelial damage.

British Journal of Pharmacology (1999) 126, 639–648; doi:10.1038/sj.bjp.0702346
Effects of the prostanoid EP3‐receptor agonists M&B 28767 and GR 63799X on infarct size caused by regional myocardial ischaemia in the anaesthetized rat
Authors: Zacharowski Kai , Olbrich Antje , Otto Mike , Hafner Gerd , Thiemermann Christoph
Published: January 2005

This study investigates the effects of two agonists of the prostanoid EP3‐receptor (M&B 28767 and GR 63799X) on the infarct size caused by regional myocardial ischaemia and reperfusion in the anaesthetized rat.

One hundred and sixty‐seven, male Wistar rats were anaesthetized (thiopentone, 120 mg kg−1, i.p.), ventilated (8–10 ml kg−1, 70 strokes min−1, inspiratory oxygen concentration: 30%; PEEP: 1–2 mmHg) and subjected to occlusion of the left anterior descending coronary artery (LAD, for 7.5, 15, 25, 35, 45 or 60 min) followed by reperfusion (2 h). Infarct size was determined by staining of viable myocardium with a tetrazolium stain (NBT), histological evaluation by light and electron microscopy and determination of the plasma levels of cardiac troponin T.

M&B 28767 (0.5 μg kg−1 min−1, i.v., n=7) or GR 63799X (3 μg kg−1 min−1, i.v., n=7) caused significant reductions in infarct size from 60±3% (25 min ischaemia and 2 h reperfusion; saline‐control, n=8) to 39±6 and 38±4% of the area at risk, without causing a significant fall in blood pressure. Pretreatment of rats with 5‐hydroxydecanoate (5‐HD), an inhibitor of ATP‐sensitive potassium channels, attenuated the cardioprotective effects of both EP3‐receptor agonists. The reduction in infarct size afforded by M&B 28767 was also abolished by glibenclamide and the protein kinase C (PKC) inhibitors staurosporine and chelerythrine.

Thus, M&B 28767 and GR 63799X reduce myocardial infarct size in the rat by a mechanism(s) which involves the activation of PKC and the opening of ATP‐sensitive potassium channels.

British Journal of Pharmacology (1999) 126, 849–858; doi:10.1038/sj.bjp.0702348
Antagonism by acetyl‐RYYRIK‐NH2 of G protein activation in rat brain preparations and of chronotropic effect on rat cardiomyocytes evoked by nociceptin/orphanin FQ
Authors: Berger H , Albrecht E , Wallukat G , Bienert M
Published: January 2005

For the further elucidation of the central functions of nociceptin/orphanin FQ (noc/OFQ), the endogenous ligand of the G protein‐coupled opioid receptor‐like receptor ORL1, centrally acting specific antagonists will be most helpful. In this study it was found that the hexapeptide acetyl‐RYYRIK‐NH2 (Ac‐RYYRIK‐NH2), described in literature as partial agonist on ORL1 transfected in CHO cells, antagonizes the stimulation of [35S]‐GTPγS binding to G proteins by noc/OFQ in membranes and sections of rat brain. The antagonism of the peptide was competitive, of high affinity (Schild constant 6.58 nM), and specific for noc/OFQ in that the stimulation of GTP binding by agonists for the μ‐, δ‐, and κ‐opioid receptor was not inhibited. The hexapeptide also fully inhibited the chronotropic effect of noc/OFQ on neonatal rat cardiomyocytes. It is suggested that Ac‐RYYRIK‐NH2 may provide a promising starting point for in vivo tests for antagonism of the action of noc/OFQ and for the further development of highly active and specific antagonists.

British Journal of Pharmacology (1999) 126, 555–558; doi:10.1038/sj.bjp.0702353
Hyperbaric oxygen increases plasma exudation in rat trachea: involvement of nitric oxide
Authors: Bernareggi M , Radice S , Rossoni G , Oriani G , Chiesara E , Berti F
Published: January 2005

This study investigates the microvascular permeability changes in tracheal tissue of rats exposed to hyperbaric oxygen (HBO).

Rats, following exposure to HBO or ambient air (control animals) for 1.5, 3 and 6 h, were prepared for recording of nitric oxide exhaled (FENO) in air using a chemiluminescence analyser. The level of FENO was not statistically different in the two groups. Plasma exudation, evaluated by measuring the leakage of Evans blue (EB) dye into the tracheal tissue, was significantly elevated (48, 86 and 105% at 1.5, 3 and 6 h, respectively) in HBO‐treated rats.

Plasma exudation in the trachea of control rats was significantly increased (42%, P<0.05) by NG‐nitro‐L‐arginine methyl ester (L‐NAME), whereas it was significantly reduced (31%, P<0.05) in rats exposed to HBO for 3 h.

N‐acetylcysteine (NAC) and flunisolide significantly prevented the increase in plasma leakage in HBO‐treated rats. In contrast, indomethacin was devoid of anti‐exudative activity in these experiments.

Western immunoblot showed a significant increase in the level of inducible nitric oxide synthase (iNOS) protein in the tracheal homogenates of HBO‐treated rats, as compared to basal levels.

These results indicate that nitric oxide (NO) is involved in the maintenance of microvascular permeability in tracheal tissue of rats. The protective effect observed with the steroid seems to support this hypothesis. Furthermore, the beneficial action of NAC underlines that reactive oxygen species participate in the microvascular permeability changes observed in tracheal tissue of rats exposed to HBO.

British Journal of Pharmacology (1999) 126, 794–800; doi:10.1038/sj.bjp.0702354
Suppression of nitric oxide synthase and the down‐regulation of the activation of NFκB in macrophages by resveratrol
Authors: Tsai Shu‐Huei , Lin‐Shiau Shoei‐Yn , Lin Jen‐Kun
Published: January 2005

Resveratrol, naringenin and naringin are naturally occurring flavonoids in grapes and grapefruits. The anti‐inflammatory effects of these flavonoids have been well documented, but the mechanism is poorly characterized. High concentration of NO are produced by inducible NO synthase (iNOS) in inflammation, and the prevention of the expression of iNOS may be an important anti‐inflammatory mechanism. In this study, the effects of these flavonoids on the induction of NO synthase (NOS) in RAW 264.7 cells activated with bacterial lipopolysaccharide (LPS, 50 ng ml−1) were investigated.

Resveratrol was found strongly to inhibit NO generation in activated macrophages, as measured by the amount of nitrite released into the culture medium, and resveratrol strongly reduced the amount of cytosolic iNOS protein and steady state mRNA levels. However, the inhibitory abilities of naringenin were lower, and the inhibitory abilities of naringin were almost negligible.

In electrophoretic mobility shift assays, the activation of NFκB induced by LPS for 1 h was inhibited by resveratrol (30 μM). Furthermore, in immunoblotting analysis, cells treated with LPS plus resveratrol showed an inhibition of phosphorylation as well as degradation of IκBα, and a reduced nuclear content of NFκB subunits.

The flavonoids may be of value for inhibiting the enhanced expression of iNOS in inflammation through down‐regulation of NFκB binding activity.

British Journal of Pharmacology (1999) 126, 673–680; doi:10.1038/sj.bjp.0702357
ATP‐ and glutathione‐dependent transport of chemotherapeutic drugs by the multidrug resistance protein MRP1
Authors: Renes Johan , De Vries Elisabeth G E , Nienhuis Edith F , Jansen Peter L M , Müller Michael
Published: January 2005

The present study was performed to investigate the ability of the multidrug resistance protein (MRP1) to transport different cationic substrates in comparison with MDR1‐P‐glycoprotein (MDR1). Transport studies were performed with isolated membrane vesicles from in vitro selected multidrug resistant cell lines overexpressing MDR1 (A2780AD) or MRP1 (GLC4/Adr) and a MRP1‐transfected cell line (S1(MRP)).

As substrates we used 3H‐labelled derivatives of the hydrophilic monoquaternary cation N‐(4′,4′‐azo‐n‐pentyl)‐21‐deoxy‐ajmalinium (APDA), the basic drug vincristine and the more hydrophobic basic drug daunorubicin. All three are known MDR1‐substrates.

MRP1 did not mediate transport of these substrates per se. In the presence of reduced glutathione (GSH), there was an ATP‐dependent uptake of vincristine and daunorubicin, but not of APDA, into GLC4/Adr and S1(MRP) membrane vesicles which could be inhibited by the MRP1‐inhibitor MK571.

ATP‐ and GSH‐dependent transport of daunorubicin and vincristine into GLC4/Adr membrane vesicles was inhibited by the MRP1‐specific monoclonal antibody QCRL‐3.

MRP1‐mediated daunorubicin transport rates were dependent on the concentration of GSH and were maximal at concentrations 10 mM. The apparent KM value for GSH was 2.7 mM. Transport of daunorubicin in the presence of 10 mM GSH was inhibited by MK571 with an IC50 of 0.4 μM.

In conclusion, these results demonstrate that MRP1 transports vincristine and daunorubicin in an ATP‐ and GSH‐dependent manner. APDA is not a substrate for MRP1.

British Journal of Pharmacology (1999) 126, 681–688; doi:10.1038/sj.bjp.0702360
Effects of angiotensin II receptor blockade on proximal fluid uptake in the rat kidney
Authors: Smart M L , Hiranyachattada S , Harris P J
Published: January 2005

Angiotensin II has a well described dose‐dependent biphasic action on proximal tubule fluid uptake, although the concentration and effect of endogenous luminal angiotensin II remain controversial.

Shrinking split‐droplet micropuncture was used to examine the fluid uptake in response to the luminal application of three AT1 antagonists (losartan, EXP3174, candesartan).

Addition of losartan at 10−8 M decreased fluid uptake rate (Jva) by 17.5±2.2% (P<0.05). Luminal addition of EXP3174 at concentrations between 10−9–10−5 M caused a dose‐dependent decrease in fluid uptake, with a maximum decrease of 41.0±9.5% (P<0.01) at 10−6 M. Candesartan also decreased fluid uptake, by 21.9±4.9% (P<0.05) at 10−8 M and 23.6±5.5% (P<0.05) at 10−5 M.

All three antagonists at a low concentration (10−8 M) decreased fluid uptake. EXP3174 and candesartan at a higher concentration (10−5 M) also decreased fluid uptake in contrast to the previously reported effect of losartan.

We conclude that the endogenous concentration of antiotensin II in the proximal luminal fluid is low and exerts a stimulatory effect on fluid absorption. Losartan at concentrations greater than 10−6 M may have a non‐selective action on fluid uptake.

British Journal of Pharmacology (1999) 126, 697–700; doi:10.1038/sj.bjp.0702366
Pharmacological evidence that inducible nitric oxide synthase is a mediator of delayed preconditioning
Authors: Imagawa J , Yellon D M , Baxter G F
Published: January 2005

Brief periods of myocardial ischaemia preceding a subsequent more prolonged ischaemic period 24–72 h later confer protection against myocardial infarction (‘delayed preconditioning’ or the ‘second window’ of preconditioning). In the present study, we examined the effects of pharmacological modifiers of inducible nitric oxide synthase (iNOS) induction and activity on delayed protection conferred by ischaemic preconditioning 48 h later in an anaesthetized rabbit model of myocardial infarction.

Rabbits underwent a myocardial preconditioning protocol (four 5 min coronary artery occlusions) or were sham‐operated. Forty‐eight hours later they were subjected to a sustained 30 min coronary occlusion and 120 min reperfusion. Infarct size was determined with triphenyltetrazolium staining. In rabbits receiving no pharmacological intervention, the percentage of myocardium infarcted within the risk zone was 43.9±5.0% in sham‐operated animals and this was significantly reduced 48 h after ischaemic preconditioning with four 5 min coronary occlusions to 18.5±5.6% (P<0.01).

Administration of the iNOS expression inhibitor dexamethasone (4 mg kg−1 i.v) 60 min before ischaemic preconditioning completely blocked the infarct‐limiting effect of ischaemic preconditioning (infarct size 48.6±6.1%). Furthermore, administration of aminoguanidine (300 mg kg−1, s.c.), a relatively selective inhibitor of iNOS activity, 60 min before sustained ischaemia also abolished the delayed protection afforded by ischaemic preconditioning (infarct size 40.0±6.0%).

Neither aminoguanidine nor dexamethasone per se had significant effect on myocardial infarct size. Myocardial risk zone volume during coronary ligation, a primary determinant of infarct size in this non‐collateralized species, was not significantly different between intervention groups. There were no differences in systolic blood pressure, heart rate, arterial blood pH or rectal temperature between groups throughout the experimental period.

These data provide pharmacological evidence that the induction of iNOS, following brief periods of coronary occlusion, is associated with increased myocardial tolerance to infarction 48 h later.

British Journal of Pharmacology (1999) 126, 701–708; doi:10.1038/sj.bjp.0702368
Effects of nicorandil as compared to mixtures of sodium nitroprusside and levcromakalim in isolated rat aorta
Authors: Cogolludo Angel L , Pérez‐Vizcaíno Francisco , Fajardo Susan , Ibarra Manuel , Tamargo Juan
Published: January 2005

The contribution of the relaxant mechanisms of nicorandil (NIC) were analysed by comparing its effects with those of sodium nitroprusside (SNP), levcromakalim (LEM) and mixtures (1:10, 1:30 and 1:100) of SNP:LEM in isolated endothelium‐denuded rat aorta.

In rings precontracted with KCl (25 mM), the relative inhibitory potency of the soluble guanylate cyclase inhibitor ODQ and the KATP channel inhibitor glibenclamide (GLI) on SNP:LEM mixtures showed a good correlation with the relative proportion of SNP and LEM in the mixtures. Furthermore, the degree of the inhibition by ODQ and GLI of the effects of the 1:30 SNP:LEM mixture varied as a function of the relative potency of SNP and LEM in KCl‐, noradrenaline‐ (NA) or NA plus nifedipine‐treated arteries.

The inhibitory effects of ODQ, GLI and ODQ plus GLI on NIC‐induced relaxation was similar to that for the 1:30 SNP:LEM mixture in NA plus nifedipine‐contracted arteries, but the inhibition of GLI or ODQ plus GLI was smaller in KCl‐contracted arteries.

In conclusion, the relative importance of activation of the cyclic GMP pathway and KATP channel opening in mixtures of SNP and LEM could be predicted by the proportion of the drugs in the mixtures and by the relative potency of SNP vs LEM in different experimental conditions. Furthermore, the present results suggest that besides these two mechanisms, a third ODQ‐ and GLI‐insensitive mechanism, possibly involving Ca2+ channel blockade, also participates in the relaxant effects of NIC in KCl‐induced contractions.

British Journal of Pharmacology (1999) 126, 1025–1033; doi:10.1038/sj.bjp.0702375
Enhanced role for the opening of potassium channels in relaxant responses to acetylcholine after myocardial ischaemia and reperfusion in dog coronary arteries
Authors: Chan Elsa C H , Woodman Owen L
Published: January 2005

Anaesthetized dogs were subjected to 1 h occlusion of the left circumflex coronary artery followed by 2 h of reperfusion. Relaxant responses were examined in coronary artery rings removed proximal (nonischaemic) or distal (ischaemic) to the site of occlusion.

Relaxant responses to acetylcholine (ACh) were similar in nonischaemic and ischaemic artery rings. In addition ACh‐induced relaxation of nonischaemic and ischaemic artery rings was equally susceptible to inhibition of nitric oxide (NO) synthase using L‐NG‐nitroarginine (L‐NOARG, 10−4 M), or to inhibition of soluble guanylate cyclase using 1H‐[1,2,4]oxadiazolo[4,3‐a]quinoxaline‐1‐one (ODQ, 10−5 M).

In nonischaemic arteries, the relaxation to ACh was unaffected by high K+ (67 mM) but in ischaemic arteries, the maximum relaxation to ACh was significantly reduced from 113±6 to 60±2% (ANOVA, P<0.05). Tetraethylammonium (TEA, 10−3 M), an inhibitor of large conductance calcium activated potassium (BKCa) channels did not inhibit the response to ACh in nonischaemic arteries but in ischaemic arteries TEA significantly shifted the concentration response curve to ACh to the right (pEC50; nonischaemic, 7.07±0.25; ischaemic, 6.54±0.21, P<0.01, ANOVA) without decreasing the maximum relaxation. TEA did not affect the responses to sodium nitroprusside in either nonischaemic or ischaemic arteries.

In conclusion, ischaemia/reperfusion did not change the sensitivity of endothelium‐dependent relaxation to L‐NOARG or ODQ indicating that ischaemia did not affect the contribution of NO or cyclic GMP to ACh‐induced relaxation. However, in ischaemic arteries the opening of the BKCa channels contributed to relaxation caused by ACh whereas TEA had no effect in nonischaemic arteries. The factor responsible for the opening of this potassium channel was a factor other than NO and may be endothelium derived hyperpolarizing factor (EDHF).

British Journal of Pharmacology (1999) 126, 925–932; doi:10.1038/sj.bjp.0702376
Decreased vascular permeability response to substance P in airways of genetically hypertensive rats
Authors: Bakhle Y S , Brogan Juliet D , Bell C
Published: January 2005

The inbred genetically hypertensive strain (GH) of the Otago Wistar rat possesses more sensory neurons containing the neuropeptide substance P (SP) than does its genetically related control normotensive strain.

As SP contributes to airway inflammation by increasing microvascular permeability, we assessed the extravasation of Evans Blue dye in trachea and main bronchus of anaesthetized GH and control rats, in the presence of endogenous (capsaicin‐liberated) or exogenous SP.

Following intravenous administration of either capsaicin (75 μg kg−1) or SP (3.3 nmol kg−1), extravasation of Evans Blue in airways from GH rats was only about 60% of that in airways of control rats. This difference was not gender‐specific and responses to capsaicin were abolished by pretreatment with a selective NK1 receptor antagonist SR 140333 (360 nmol kg−1).

By contrast, the extravasation of dye caused by intravenous 5‐hydroxytryptamine (0.5 μmol kg−1) was similar in magnitude in both GH and control strains.

Falls in systemic arterial blood pressure in response to exogenous SP (0.1–3 nmol kg−1) or acetylcholine (0.2–2 nmol kg−1) were also very similar between strains, but those in response to capsaicin (75 μg kg−1) in the GH rats were about double those in control rats. The hypotensive response to SP was abolished by SR 140333, but that to capsaicin was unaffected.

Our results indicate that the increased peripheral innervation density by SP nerves in GH rats is accompanied by reduced inflammatory responses to SP. This does not involve decreased vasodilator potency of SP and is therefore probably related to altered endothelial responsiveness.

British Journal of Pharmacology (1999) 126, 933–938; doi:10.1038/sj.bjp.0702377
Characterization of high affinity neurotensin receptor NTR1 in HL‐60 cells and its down regulation during granulocytic differentiation
Authors: Choi Se‐Young , Chae Hee‐Don , Park Tae‐Ju , Ha Hyunjung , Kim Kyong‐Tai
Published: January 2005

We investigated responses to neurotensin in human promyelocytic leukaemia HL‐60 cells.

Neurotensin increased the cytosolic calcium concentration ([Ca2+]i) in a concentration‐dependent manner and also produced inositol 1,4,5‐trisphosphate (InsP3).

Among the tested neurotensin analogues, neurotensin 8–13, neuromedin‐N, and xenopsin also increased [Ca2+]i, whereas neurotensin 1–11 and neurotensin 1–8 did not elicit detectable responses.

SR48692, an antagonist of NTR1 neurotensin receptors, blocked the neurotensin‐induced [Ca2+]i increase, whereas levocabastine, which is known as an NTR2 neurotensin receptor antagonist, did not attenuate the neurotensin‐evoked effect.

The expression of NTR1 neurotensin receptors was confirmed by Northern blot analysis and reverse transcriptase‐polymerase chain reaction (RT–PCR).

During 1.25% dimethylsulfoxide (DMSO)‐triggered granulocytic differentiation of HL‐60 cells, the neurotensin‐induced [Ca2+]i rise became gradually smaller and completely disappeared 4 days after treatment with DMSO. The mRNA level for neurotensin receptors was also decreased after differentiation.

The results show that HL‐60 cells express NTR1 neurotensin receptors and suggest that granulocytic differentiation involves transcriptional regulation of the receptors resulting in down‐regulation of the neurotensin‐induced signalling.

British Journal of Pharmacology (1999) 126, 1050–1056; doi:10.1038/sj.bjp.0702378
Effects of the adenylyl cyclase inhibitor SQ22536 on iloprost‐induced vasorelaxation and cyclic AMP elevation in isolated guinea‐pig aorta
Authors: Turcato Sally , Clapp Lucie H
Published: January 2005

The stable prostacyclin analogue, iloprost relaxes a variety of blood vessels and increases cyclic AMP, although the relationship between adenosine 3′ : 5′‐cyclic monophosphate (cyclic AMP) and vasorelaxation remains unclear. We therefore investigated the effect of the adenylyl cyclase inhibitor, 9‐(tetrahydro‐2‐furanyl)‐9H‐purin‐6‐amine (SQ22536) on iloprost‐mediated relaxation and cyclic AMP elevation in endothelium‐denuded aortic strips. Iloprost (1–1000 nM) caused a concentration‐dependent inhibition of phenylephrine (1–6 μM) contractions, the responses being unaffected by pre‐incubation with SQ22536 (100 μM) for 30 min. In other experiments 60 nM iloprost caused a 64% inhibition of phenylephrine contractions concomitant with a 3 fold rise in cyclic AMP. SQ22536 completely abolished the iloprost‐induced elevation in cyclic AMP while having no significant effect on relaxation. Our results therefore strongly suggest that cyclic AMP‐independent pathways are responsible for the vasorelaxant effects of iloprost in guinea‐pig aorta.

British Journal of Pharmacology (1999) 126, 845–847; doi:10.1038/sj.bjp.0702383
Influence of TASP‐V, a novel neuropeptide Y (NPY) Y2 agonist, on nasal and bronchial responses evoked by histamine in anaesthetized pigs and in humans
Authors: Malis Didier‐David , Grouzmann Eric , Morel Denis R , Mutter Manfred , Lacroix Jean‐Silvain
Published: January 2005

In nine anaesthetized pigs we have studied the influence of intranasal or intrabronchial pretreatment with TASP‐V, a neuropeptide Y (NPY) Y2 agonist formed by the attachment of NPY 21–36 to a template‐assembled synthetic peptide (TASP), on the functional responses to subsequent intranasal or intrabronchial histamine challenge.

In a parallel study, subjective and objective nasal airway resistance (NAR) increase following intranasal histamine challenge was evaluated in 11 healthy volunteers after TASP‐V or placebo pretreatment.

In pigs, increase in sphenopalatine blood flow induced by histamine dihydrochloride nasal spray (0.25 mg kg−1 in 3 ml of saline) was significantly reduced by 65% (P<0.05) following intranasal pretreatment with 10 μg kg−1 of TASP‐V. Bronchoconstriction induced by histamine dihydrochloride nebulization (0.5 mg kg−1 in 3 ml of saline) was significantly attenuated by 25 and 55% following aerosolized pretreatment with TASP‐V analogue at 10 and 20 μg kg−1, respectively.

In healthy volunteers, objective increase in NAR and reduction in nasal minimal cross section area (MCSA) induced by intranasal spray of histamine dihydrochloride (15 μg kg−1 in 200 μl of saline) were significantly attenuated by 50% following local pretreatment with 1.275 μg kg−1 of TASP‐V when compared with saline.

It is concluded that intranasal or intrabronchial pretreatment with TASP‐V reduced nasal obstruction and bronchoconstriction evoked by histamine challenge in the pig. In healthy human volunteers, this agent attenuated NAR increase and MCSA reduction induced by intranasal application of histamine.

British Journal of Pharmacology (1999) 126, 989–996; doi:10.1038/sj.bjp.0702388
Cell‐specific coupling of PGE2 to different transduction pathways in arginine vasopressin‐ and glucagon‐sensitive segments of the rat renal tubule
Authors: Aarab Lotfi , Siaume‐Perez Sylvie , Chabardès Danielle
Published: January 2005

The aim of the present study was to investigate the transduction pathways elicited by prostaglandin E2 (PGE2) to inhibit hormone‐stimulated adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) accumulation in the outer medullary collecting duct (OMCD) and medullary thick ascending limb (MTAL) microdissected from the rat nephron.

In the OMCD, 0.3 μM PGE2 and low concentrations of Ca2+ ionophores (10 nM ionomycin or 50 nM A23187) inhibited by about 50% a same pool of arginine vasopressin (AVP)‐stimulated cyclic AMP content through a same process insensitive to Bordetella pertussis toxin (PTX).

Sulprostone, an agonist of the EP1/EP3 subtypes of the PGE2 receptor, decreased AVP‐dependent cyclic AMP accumulation in OMCD and MTAL samples. The concentration eliciting half‐maximal inhibition was of about 50 nM in OMCD and 0.1 nM in MTAL.

In MTAL, 1 nM sulprostone and PGE2 inhibited by about 90% a same pool of AVP‐dependent cyclic AMP content through a PTX‐sensitive, Ca2+‐independent pathway.

In the OMCD, PGE2 decreased by about 50% glucagon‐dependent cyclic AMP synthesis by a process sensitive to PTX and Ca2+‐independent. Sulprostone 1 nM induced the same level of inhibition.

These results demonstrate that PGE2 decrease hormone‐dependent cyclic AMP accumulation through a Gαi‐mediated inhibition of adenylyl cyclase activity in MTAL cells and glucagon‐sensitive cells of the OMCD or through a PTX‐insensitive increase of intracellular Ca2+ concentration in AVP‐sensitive cells of the OMCD.

British Journal of Pharmacology (1999) 126, 1041–1049; doi:10.1038/sj.bjp.0702390