bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of a novel selective and potent phosphinic peptide inhibitor of endopeptidase 3.4.24.16 on neurotensin‐induced analgesia and neuronal inactivation
Authors: Vincent Bruno , Jiracek Jirì , Noble Florence , Loog Mart , Roques Bernard , Dive Vincent , Vincent Jean‐Pierre , Checler Frédéric
Published: August 2006

We have examined a series of novel phosphinic peptides as putative potent and selective inhibitors of endopeptidase 3.4.24.16.

The most selective inhibitor, Pro‐Phe‐Ψ(PO2CH2)‐Leu‐Pro‐NH2 displayed a Ki value of 12 nM towards endopeptidase 3.4.24.16 and was 5540 fold less potent on its related peptidase endopeptidase 3.4.24.15. Furthermore, this inhibitor was 12.5 less potent on angiotensin‐converting enzyme and was unable to block endopeptidase 3.4.24.11, aminopeptidases B and M, dipeptidylaminopeptidase IV and proline endopeptidase.

The effect of Pro‐Phe‐Ψ(PO2CH2)‐Leu‐Pro‐NH2, in vitro and in vivo, on neurotensin metabolism in the central nervous system was examined.

Pro‐Phe‐Ψ(PO2CHH2)‐Leu‐Pro‐NH2 dose‐dependently inhibited the formation of neurotensin 1‐10 and concomittantly protected neurotensin from degradation by primary cultured neurones from mouse embryos.

Intracerebroventricular administration of Pro‐Phe‐Ψ(PO2CH2)‐Leu‐Pro‐NH2 significantly potentiated the neurotensin‐induced antinociception of mice in the hot plate test.

Altogether, our study has established Pro‐Phe‐Ψ(PO2CH2)‐Leu‐Pro‐NH2 as a fully selective and highly potent inhibitor of endopeptidase 3.4.24.16 and demonstrates, for the first time, the contribution of this enzyme in the central metabolism of neurotensin.

British Journal of Pharmacology (1997) 121, 705–710; doi:10.1038/sj.bjp.0701182
Role of Ca2+‐activated K+ channel in epithelium‐dependent relaxation of human bronchial smooth muscle
Authors: Tamaoki J. , Tagaya E. , Isono K. , Kondo M. , Konno K.
Published: August 2006

To elucidate whether K+ channels play a role in the action of epithelium‐dependent bronchodilatation, we studied responses in human bronchial strips in the presence of indomethacin and NG‐nitro‐L‐arginine methylester under isometric conditions, in vitro.

Mechanical removal of the epithelium increased the contractile responses to acetylcholine; the pD2 values increased from 5.0±0.2 to 5.9±0.3 (P<0.001). This potentiation was abolished by iberiotoxin but not by apamin or glibenclamide.

In cascade bioassay, application of the bathing medium from dispersed, bronchial epithelial cells to epithelium‐denuded bronchial strips decreased acetylcholine‐induced contraction by 44±6%. This effect was reduced to 10±3% (P<0.01) when the epithelial cells were pretreated with iberiotoxin, and to 4±1% (P<0.001) when the epithelial cells were incubated with Ca2+‐free medium containing [1,2‐bis (2) aminophenoxy] ethane N,N,N′,N′‐tetraacetic acid‐acetomethoxy ester.

In contrast, the bronchodilator effect of the medium bathing epithelial cells was not altered by the direct addition of iberiotoxin to epithelium‐denuded tissues.

These results suggest that the Ca2+‐activated K+ channel may play a role in the synthesis and/or release of smooth muscle relaxing factor, which is neither nitric oxide nor a cyclo‐oxygenase product, from airway epithelial cells.

British Journal of Pharmacology (1997) 121, 794–798; doi:10.1038/sj.bjp.0701183
Effects of calcium dobesilate on the synthesis of endothelium‐dependent relaxing factors in rabbit isolated aorta
Authors: Ruiz E. , Lorente R. , Tejerina T.
Published: August 2006

Some cardiovascular disturbances which occur in diabetics are a consequence of alterations in vascular contractility as well as in endothelium‐dependent relaxation.

Calcium dobesilate (DOBE) is a drug used in diabetic retinopathy and its mechanism of action is not yet understood.

The aim of this study was to investigate the effects of DOBE on synthesis and release of endothelium‐dependent relaxing factor (EDRF) and endothelium‐dependent hyperpolarizing factor (EDHF) in rabbit isolated aorta.

Endothelium‐dependent relaxation induced by acetylcholine (ACh) (10−8–10−5 M) increased in the presence of DOBE 10−5 M only when vascular endothelium was kept intact.

NG‐nitro‐L‐arginine methyl ester (L‐NAME; 10−8–10−4 M progressively decreased the enhancing effect of DOBE on endothelium‐dependent relaxation whereas it was progressively increased by L‐Arg.

DOBE 10−5 M increased in a non‐significant manner endothelium‐dependent relaxation induced by ACh when the arteries were incubated with both L‐NAME 10−4 M and indomethacin 10−6 M.

DOBE (10−6 M and 10−5 M) was able to scavenge superoxide anion radicals generated by the hypoxanthine/xanthine oxidase reaction.

These results provide evidence that DOBE is able to affect the vascular disorders associated with diabetes mellitus since it enhances the synthesis of endothelium‐dependent relaxing factors.

British Journal of Pharmacology (1997) 121, 711–716; doi:10.1038/sj.bjp.0701184
A study of the neurotoxic effect of MDMA (‘ecstasy’) on 5‐HT neurones in the brains of mothers and neonates following administration of the drug during pregnancy
Authors: Colado M. I. , O'Shea E. , Granados R. , Misra A. , Murray T. K. , Green A. R.
Published: August 2006

It is well established that 3,4‐methylenedioxymethamphetamine (MDMA or ‘ecstasy’) is neurotoxic and produces long term degeneration of cerebral 5‐hydroxytryptamine (5‐HT) nerve terminals in many species. Since MDMA is used extensively as a recreational drug by young people, it is being ingested by many women of child bearing age. We have therefore examined the effect of administering high doses of MDMA to rats during pregnancy on the cerebral content of both the dams and the neonates.

MDMA (20 mg kg−1, s.c.) was injected twice daily on days 14–17 of the gestation period. The initial dose produced a marked hyperthermic response in the dam which was progressively attenuated in both peak height and area under the curve following further doses of the drug. The body weight of the dams decreased during the period of treatment.

There was a modest decrease in litter size (−20%) of the MDMA‐treated dams.

The concentration of 5‐HT and its metabolite 5‐HIAA was decreased by over 65% in the hippocampus and striatum and 40% in the cortex of the dams 1 week after parturition. In contrast, the content of 5‐HT and 5‐HIAA in the dorsal telencephalon of the pups of the MDMA‐treated dams was the same as that seen in tissue from pups born to control animals.

Administration of MDMA (40 mg kg−1, s.c.) to adult rats increased thiobarbituric acid reacting substances (TBARS) in cortical tissue 3 h and 6 h later, indicating increased lipid peroxidation. No increase in TBARS was seen in the cortical tissue of 7–10 day neonates injected with this dose of MDMA 3 h or 6 h earlier.

The data suggest that exposure to MDMA in utero during the maturation phase does not produce damage to 5‐HT nerve terminals in the foetal rat brain, in contrast to the damage seen in the brains of the mothers. This may be due to MDMA being metabolized to free radical producing entities in the adult brain but not in the immature brain or, alternatively, to more effective or more active free radical scavenging mechanisms being present in the immature brain.

British Journal of Pharmacology (1997) 121, 827–833; doi:10.1038/sj.bjp.0701201
Hydrogen peroxide induced responses of cat tracheal smooth muscle cells
Authors: Bauer V , Oike M , Tanaka H , Inoue R , Ito Y
Published: August 2006

The effects of hydrogen peroxide H2O2 (10−6 and 10−3 M) on membrane potential, membrane currents, intracellular calcium concentration, resting muscle tone and contractions elicited by electrical field stimulation (EFS) and carbachol were examined in cat tracheal strips and isolated smooth muscle cells.

H2O2 (10−4 and 10−5 M) enhanced the amplitude of contractions and excitatory junction potentials (e.j.p.) evoked by EFS without changing muscle tone and resting membrane potential of the tracheal smooth muscle, and enhanced the contraction induced by carbachol (10−8 M). At an increased concentration (10−3 M), H2O2 elevated resting muscle tone and marginally hyperpolarized the membrane in the majority of the cells.

In 51 out of 56 cells examined, H2O2 (10−6–10−3 M) elicited an outward current at a holding potential of −40 mV and enhanced the frequency of the spontaneous transient outward current (STOC). In 20 cells the outward current was preceded by a small inward current. In the other cells, H2O2 elicited only an inward current or did not affect the background current.

In Ca2+ free solution the action of H2O2 on the resting muscle tone, STOCs, background current and on the current induced by ramp depolarization was significantly reduced.

H2O2 (10−4 M) increased the intracellular ionized calcium concentration both in the absence and presence of external Ca2+. However, the effect developed faster and was of a higher amplitude in the presence of external Ca2+.

These results suggest that H2O2 increases intracellular Ca2+, with a subsequent augmentation of stimulation‐evoked contractions, and enhances Ca2+ and voltage‐sensitive potassium conductance.
Pharmacological pleiotropism of the human recombinant α1A‐adrenoceptor: implications for α1‐adrenoceptor classification
Authors: Ford Anthony P D W , Daniels Donald V , Chang David J , Gever Joel R , Jasper Jeffrey R , Lesnick John D , Clarke David E
Published: August 2006

Three fully‐defined α1‐adrenoceptors (α1A, α1B and α1D) have been established in pharmacological and molecular studies. A fourth α1‐adrenoceptor, the putative α1L‐adrenoceptor, has been defined in functional but not molecular studies, and has been proposed to mediate contraction of human lower urinary tract tissues; its relationship to the three fully characterized α1‐adrenoceptors is not known.

In the present study, binding affinities were estimated by displacement of [3H]‐prazosin in membrane homogenates of Chinese hamster ovary (CHO‐K1) cells stably expressing the human α1A‐, α1B‐ and α1D‐adrenoceptors and were compared with affinity estimates obtained functionally in identical cells by measuring inhibition of noradrenaline (NA)‐stimulated accumulation of [3H]‐inositol phosphates.

For the α1A‐adrenoceptor, binding studies revealed a pharmacological profile typical for the classically defined α1A‐adrenoceptor, such that prazosin, RS‐17053, WB4101, 5‐methylurapidil, Rec15/2739 and S‐niguldipine all displayed subnanomolar affinity. A different profile of affinity estimates was obtained in inositol phosphates accumulation studies: prazosin, WB4101, 5‐methylurapidil, RS‐17053 and S‐niguldipine showed 10 to 40 fold lower affinity than in membrane binding. However, affinity estimates were not ‘frameshifted’, as tamsulosin, indoramin and Rec15/2739 yielded similar, high affinity estimates in binding and functional assays.

In contrast, results from human α1B‐ and α1D‐adrenoceptors expressed in CHO‐K1 cells gave antagonist affinity profiles in binding and functional assays that were essentially identical.

A concordance of affinity estimates from the functional (inositol phosphates accumulation) studies of the α1A‐adrenoceptor in CHO‐K1 cells was found with estimates published recently from contractile studies in human lower urinary tract tissues (putative α1L‐adrenoceptor). These data show that upon functional pharmacological analysis, the cloned α1A‐adrenoceptor displays pharmacological recognition properties consistent with those of the putative α1L‐adrenoceptor. Why this profile differs from that obtained in membrane binding, and whether it explains the α1L‐adrenoceptor pharmacology observed in many native tissues, requires further investigation.
The selectivity and structural determinants of peptide antagonists at the CGRP receptor of rat, L6 myocytes
Authors: Howitt Stephen G , Poyner David R
Published: August 2006

Potency orders were determined for a series of agonists and antagonists on the calcitonin gene‐related peptide (CGRP) receptor of rat L6 myocytes. The agents tested were all shown to have been active against CGRP, amylin or adrenomedullin receptors.

AC187 had a pIC50 of 6.8±0.10, making it 14 fold less potent as an antagonist than CGRP8–37 (pIC50, 7.95±0.14). Amyline8–37 was equipotent to AC187 (pIC50, 6.6±0.16) and CGRP19–37 was 3 fold less potent than either (pIC50, 6.1±0.24).

[Ala11]‐CGRP8–37 was 6 fold less potent than CGRP8–37, (pIC50, 7.13±0.14), whereas [Ala18]‐CGRP8–37 was approximately equipotent to CGRP8–37 (pIC50, 7.52±0.15). However, [Ala11,Ala18]‐CGRP8–37 was over 300 fold less potent than CGRP8–37 (pIC50, 5.30±0.04).

[Tyr0]‐CGRP28–37, amylin19–37 and adrenomedullin22–52 were inactive as antagonists at concentrations of up to 1 μM.

Biotinyl‐human α‐CGRP was 150 fold less potent than human α‐CGRP itself (EC50 values of 48±17 nM and 0.31±0.13 nM, respectively). At 1 μM, [Cys(acetomethoxy)2,7]‐CGRP was inactive as an agonist.

These results confirm a role for Arg11 in maintaining the high affinity binding of CGRP8–37. Arg18 is of less direct significance for high affinity binding, but it may be important in maintaining the amphipathic nature of CGRP and its analogues.
Lack of involvement of glutamate‐induced excitotoxicity in MPP+ toxicity in striatal dopaminergic terminals: possible involvement of ascorbate
Authors: Matarredona Esperanza R , Santiago Marti , Machado Alberto , Cano Josefina
Published: August 2006

The present study concerns the possible relationship between glutamate excitotoxicity and 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine/1‐methyl‐4‐phenylpyridinium (MPTP/MPP+) neurotoxicity on striatal dopaminergic terminals.

MPP+ neurotoxicity has been studied by means of two MPP+ perfusions separated by 24 h. After the second MPP+ 1 mM perfusion, dopamine extracellular output, measured by microdialysis, was considered to be an index of the dopaminergic neurone damage produced by the first MPP+ 1 mM perfusion.

High concentration (10 mM) of glutamate uptake inhibitor L‐trans‐pyrrolidine‐2,4‐dicarboxylic acid (PDC) stimulated basal release of dopamine and protected against the neurotoxic effect of MPP+.

PDC 10 mM perfusion produced an increase in the extracellular output of glutamate and aspartate, and a decrease in that of ascorbate.

The protective effect against MPP+ toxicity observed with PDC 10 mM was completely abolished when this glutamate uptake inhibitor was co‐perfused with ascorbate 0.5 mM.

These results suggest that glutamate‐induced neurotoxicity is not involved in MPP+ toxicity. The protective effect found with the glutamate uptake inhibitor could be due to a decrease in extracellular ascorbate levels.
Neurochemical actions of the desglycinyl metabolite of remacemide hydrochloride (ARL 12495AA) in mouse brain
Authors: Paul Leach John , Sills Graeme J , Butler Elaine , Forrest Gerard , Thompson George G , Brodie Martin J
Published: August 2006

Remacemide hydrochloride, a recently developed antiepileptic drug, is believed to exert its effects, at least in part, via its desglycinyl metabolite, ARL 12495AA.

We have investigated the effects of ARL 12495AA on several neurochemical parameters in mouse brain. Adult male ICR mice were randomized into two groups and administered ARL 12495AA (0–75 mg kg−1) intraperitoneally, either as a single dose or once daily for 5 days.

Six hours after the final dose, animals were killed and their brains removed. Brain tissues were analysed for concentrations of γ‐aminobutyric acid (GABA), glutamine and glutamate and for the activities of GABA‐transaminase (GABA‐T) and glutamic acid decarboxylase (GAD).

Single dose ARL 12495AA was without effect on any of the parameters investigated.

Repeated ARL 12495AA treatment did not alter brain concentrations of GABA and glutamine, but at a high dose there was a trend toward reduced brain glutamate concentrations (P=0.10).

Repeated administration of ARL 12495AA at a high dose significantly increased GABA‐T activity (P<0.05) and decreased that of GAD (P<0.05). 7 These findings may have relevance to the clinical use of remacemide hydrochloride in human epilepsy.
Characterization of lamotrigine inhibition of Na+ channels in rat hippocampal neurones
Authors: Kuo Chung‐Chin , Lu Lu
Published: August 2006

Lamotrigine (LTG), a new antiepileptic drug, requires long depolarizations to inhibit Na+ currents. This suggests either slow binding of LTG to the fast inactivated state or selective binding of LTG to the slow inactivated state of Na+ channels. To differentiate between these possibilities and to characterize further the action of LTG, we studied the affinity and kinetics of LTG binding to the Na+ channels in acutely dissociated hippocampal neurones of the rat.

LTG inhibited more Na+ currents at more depolarized holding potentials. The inhibitory effect at various holding potentials could be described by one‐to‐one binding curves, which yielded an apparent dissociation constant of ∼7μm for LTG binding to the inactivated channels (KI), and a dissociation constant more than 200 times larger for LTG binding to the resting channels. A similar value of KI (∼9μm) was also derived from the LTG concentration‐dependent shift of the inactivation curve.

The recovery of LTG‐bound inactivated Na+ channels was faster than the recovery of normal (drug‐free) slow inactivated channels. Moreover, the binding kinetics of LTG onto the inactivated channels were faster than the development of the slow inactivated state, and were linearly correlated with LTG concentrations, with a binding rate constant of ∼10,000M−1s−1. These findings suggest that LTG chiefly binds to the fast inactivated state rather than the slow inactivated state.

We conclude that LTG, in therapeutic concentrations and at relatively depolarized membrane potentials, may potently inhibit Na+ currents by slow binding to the fast inactivated state of Na+ channels. Like phenytoin, the slow binding rates may explain why LTG effectively inhibits seizure discharges, yet spares most normal neuronal activities.
Possible mechanisms underlying the midazolam‐induced relaxation of the noradrenaline‐contraction in rabbit mesenteric resistance artery
Authors: Shiraishi Yoshihisa , Ohashi Masuo , Kanmura Yuichi , Yamaguchi Shunichiro , Yoshimura Nozomu , Itoh Takeo
Published: August 2006

The mechanisms underlying the midazolam‐induced relaxation of the noradrenaline (NA)‐contraction were studied by measuring membrane potential, isometric force and intracellular concentration of Ca2+([Ca2+]i) in endothelium‐denuded muscle strips from the rabbit mesenteric resistance artery. The actions of midazolam were compared with those of nicardipine, an L‐type Ca2+‐channel blocker.

Midazolam (30 and 100μm) did not modify either the resting membrane potential or the membrane depolarization induced by 10μm NA.

NA (10μm) produced a phasic, followed by a tonic increase in both [Ca2+]i and force. Midazolam (10–100μm) did not modify the resting [Ca2+]i, but attenuated the NA‐induced phasic and tonic increases in [Ca2+]i and force, in a concentration‐dependent manner. In contrast, nicardipine (0.3μm) attenuated the NA‐induced tonic, but not phasic, increases in [Ca2+]i and force.

In Ca2+‐free solution containing 2mm EGTA, NA (10μm) transiently increased [Ca2+]i and force. Midazolam (10–100μm), but not nicardipine (0.3μm), attenuated this NA‐induced increase in [Ca2+]i and force, in a concentration‐dependent manner. However, midazolam (10 and 30μm), had no effect on the increases in [Ca2+]i and force induced by 10mm caffeine.

In ryanodine‐treated strips, which have functionally lost the NA‐sensitive Ca2+‐ storage sites, NA slowly increased [Ca2+]i and force. Nicardipine (0.3μm) did not modify the resting [Ca2+]i but partly attenuated the NA‐induced increases in [Ca2+]i and force. In the presence of nicardipine, midazolam (100μm) lowered the resting [Ca2+]i and further attenuated the remaining NA‐induced increases in [Ca2+]i and force.

The [Ca2+]i‐force relationship was obtained in ryanodine‐treated strips by the application of ascending concentrations of Ca2+ (0.16–2.6mm) in Ca2+‐free solution containing 100mm K+. NA (10μm) shifted the [Ca2+]i‐force relationship to the left and enhanced the maximum Ca2+‐induced force. Under these conditions, whether in the presence or absence of 10μm NA, midazolam (10 and 30μm) attenuated the increases in [Ca2+]i and force induced by Ca2+ without changing the [Ca2+]i‐force relationship.

It was concluded that, in smooth muscle of the rabbit mesenteric resistance artery, midazolam inhibits the NA‐induced contraction through its inhibitory action on NA‐induced Ca2+ mobilization. Midazolam attenuates NA‐induced Ca2+ influx via its inhibition of both nicardipine‐sensitive and ‐insensitive pathways. Furthermore, midazolam attenuates the NA‐induced release of Ca2+ from the storage sites. This effect contributes to the midazolam‐induced inhibition of the NA‐induced phasic contraction.
Effect of a selective 5‐HT reuptake inhibitor in combination with 5‐HT1A and 5‐HT1B receptor antagonists on extracellular 5‐HT in rat frontal cortex in vivo
Authors: Sharp T , Umbers V , Gartside S E
Published: August 2006

Selective 5‐hydroxytryptamine (5‐HT; serotonin) reuptake inhibitors (SSRIs) cause a greater increase in extracellular 5‐HT in the forebrain when the somatodendritic 5‐HT1A autoreceptor is blocked. Here, we investigated whether blockade of the terminal 5‐HT1B autoreceptor influences a selective 5‐HT reuptake inhibitor in the same way, and whether there is an additional effect of blocking both the 5‐HT1A and 5‐HT1B autoreceptors.

Extracellular 5‐HT was measured in frontal cortex of the anaesthetized rat by use of brain microdialysis. In vivo extracellular recordings of 5‐HT neuronal activity in the dorsal raphe nucleus (DRN) were also carried out.

The selective 5‐HT reuptake inhibitor, paroxetine (0.8 mg kg−1, i.v.), increased extracellular 5‐HT about 2 fold in rats pretreated with the 5‐HT1A receptor antagonist, WAY100635. When administered alone neither paroxetine (0.8 mg kg−1, i.v.) nor WAY100635 (0.1 mg kg−1, i.v.) altered extracellular 5‐HT levels.

Paroxetine (0.8 mg kg−1, i.v.) did not increase 5‐HT in rats pretreated with the 5‐HT1B/D receptor antagonist, GR127935 (1 mg kg−1, i.v.). GR127935 (1 and 5 mg kg−1, i.v.) had no effect on extracellular 5‐HT when administered alone.

Interestingly, paroxetine (0.8 mg kg−1, i.v.) caused the greatest increase in 5‐HT (up to 5 fold) when GR127935 (1 or 5 mg kg−1, i.v.) was administered in combination with WAY100635 (0.1 mg kg−1, i.v.). Administration of GR127935 (5 mg kg−1, i.v.) plus WAY100635 (0.1 mg kg−1, i.v.) without paroxetine, had no effect on extracellular 5‐HT in the frontal cortex.

Despite the lack of effect of GR127935 on 5‐HT under basal conditions, when 5‐HT output was elevated about 3 fold (by adding 1 μM paroxetine to the perfusion medium), the drug caused a dose‐related (1 and 5 mg kg−1, i.v.) increase in 5‐HT.

By itself, GR127935 slightly but significantly decreased 5‐HT cell firing in the DRN at higher doses (2.0–5.0 mg kg−1, i.v.), but did not prevent the inhibition of 5‐HT cell firing induced by paroxetine.

In summary, our results suggest that selective 5‐HT reuptake inhibitors may cause a large increase in 5‐HT in the frontal cortex when 5‐HT autoreceptors on both the somatodendrites (5‐HT1A) and nerve terminals (5‐HT1B) are blocked. This increase is greater than when either set of autoreceptors are blocked separately. The failure of a 5‐HT1B receptor antagonist alone to enhance the effect of the selective 5‐HT reuptake inhibitor in our experiments may be related to a lack of tone on the terminal 5‐HT1B autoreceptor due to a continued inhibition of 5‐HT cell firing. These results are discussed in relation to the use of 5‐HT autoreceptor antagonists to augment the antidepressant effect of selective 5‐HT reuptake inhibitors.
Inhibition of eotaxin‐mediated human eosinophil activation and migration by the selective cyclic nucleotide phosphodiesterase type 4 inhibitor rolipram
Authors: Santamaria Luis F , Maria Palacios Jose , Beleta Jorge
Published: August 2006

The effect of the selective type 4 phosphodiesterase (PDE 4) inhibitor rolipram on human eosinophil activation and migration mediated by eotaxin was investigated.

Studies were performed with human freshly isolated eosinophils from peripheral blood of healthy donors by a magnetic cell separation (MACS) technique to a purity>99%. To test the effect of rolipram, eosinophils were stimulated with recombinant human eotaxin and the cell surface activation markers CD11b and L‐selectin were analysed by flow cytometry. Furthermore, eotaxin mediated eosinophil migration was measured in a transendothelial chemotaxis assay.

Our results indicate that rolipram inhibited eotaxin‐induced CD11b up‐regulation up to 60.6±7.6% at the highest tested dose (10μm), whereas transendothelial chemotaxis was partially inhibited reaching a plateau of approx. 30% at a rolipram concentration of 0.1μm.

We conclude that the selective PDE 4 inhibitor rolipram decreases eotaxin mediated eosinophil activation, an observation that may contribute to elucidate the mechanism by which PDE 4 inhibitors reduce antigen‐induced eosinophil infiltration in different animal models of allergic inflammation.
Selective depolarization of the muscle membrane in frog nerve‐muscle preparations by a chromatographically purified extract of the dinoflagellate Ostreopsis lenticularis
Authors: Meunier Frédéric A , Mercado José A , Molgó Jordi , Tosteson Thomas R , Escalona de Motta Gladys
Published: August 2006

The actions of a chromatographically identified extract of the marine dinoflagellate Ostreopsis lenticularis, named ostreotoxin‐3 (OTX‐3), were studied on frog isolated neuromuscular preparations.

OTX‐3 (1–10μg ml−1) applied to cutaneous pectoris nerve‐muscle preparations depolarized skeletal muscle fibres and caused spontaneous contractions. The depolarization was neither reversed by prolonged washing nor by (+)‐tubocurarine.

OTX‐3 decreased the amplitude of miniature end plate potentials (m.e.p.ps) but did not affect their frequency.

Extracellular recording of compound action potentials revealed that OTX‐3 affected neither excitability nor conduction along intramuscular nerve branches.

End‐plate potentials (e.p.ps) elicited by nerve stimulation were reduced in amplitude by OTX‐3 and even showed reversed polarity in junctions deeply depolarized by the toxin.

Membrane depolarization induced by OTX‐3 was decreased about 70% in muscles pretreated for 30min with 10μm tetrodotoxin. In contrast, muscles pretreated with 5μm μ‐conotoxin GIIIA were completely insensitive to OTX‐3‐induced depolarization.

OTX‐3 did not affect e.p.p. amplitude and the quantal content of e.p.ps in junctions in which muscle depolarization was abolished by μ‐conotoxin GIIIA.

OTX‐3 is a novel type of sodium‐channel activating toxin that discriminates between nerve and skeletal muscle membranes.
P2‐receptor modulation of noradrenergic neurotransmission in rat kidney
Authors: Bohmann Christine , von Kügelgen Ivar , Christian Rump L
Published: August 2006

ATP has previously been shown to act as a sympathetic cotransmitter in the rat kidney. The present study analyses the question of whether postganglionic sympathetic nerve endings in the kidney possess P2‐receptors which modulate noradrenaline release. Rat kidneys were perfused with Krebs‐Henseleit solution containing the noradrenaline uptake blockers cocaine and corticosterone and the α2‐adrenoceptor antagonist rauwolscine. The renal nerves were electrically stimulated, in most experiments by 30 pulses applied at 1 Hz. The outflow of endogenous noradrenaline (or, in some experiments, of ATP and lactate dehydrogenase) as well as the perfusion pressure were measured simultaneously.

The P2‐receptor agonist adenosine‐5′‐O‐(3‐thiotriphosphate) (ATPγS, 3–30 μM) reduced the renal nerve stimulation (RNS)‐induced outflow of noradrenaline (estimated EC50=8 μM). The P2‐receptor antagonist cibacron blue 3GA (30 μM) shifted the concentration‐inhibition curve for ATPγS to the right (apparent pKB value 4.7).

Cibacron blue 3GA (3–30 μM) and its isomer reactive blue 2 (3–30 μM) significantly increased RNS‐induced outflow of noradrenaline in the presence of the P1‐receptor antagonist 8‐(p‐sulphophenyl)theophylline (8‐SPT, 100 μM) by about 70% and 90%, respectively. The P2‐receptor antagonist suramin (30–300 μM) only tended to enhance RNS‐induced outflow of noradrenaline. When the nerves were stimulated by short pulse trains consisting of 6 pulses applied at 100 Hz (conditions under which autoinhibition is inoperative), reactive blue 2 did not affect the RNS‐induced outflow of noradrenaline.

RNS (120 pulses applied at 4 Hz) induced the outflow of ATP but not of the cytoplasmatic enzyme lactate dehydrogenase.

ATPγS (3–30 μM) concentration‐dependently reduced pressor responses to RNS at 1 Hz. Cibacron blue 3GA, reactive blue 2 as well as suramin also reduced pressor responses to RNS (maximally by 50 to 70%).

This study in rat isolated kidney, in which the release of endogenous noradrenaline was measured, demonstrates that renal sympathetic nerves possess prejunctional P2‐receptors that mediate inhibition of transmitter release. These prejunctional P2‐receptors are activated by endogenous ligands, most likely ATP, released upon nerve activity. Both, P2‐receptor agonists and P2‐receptor antagonists reduced pressor responses to RNS either by inhibiting transmitter release or by blocking postjunctional vasoconstrictor P2‐receptors.
Effects of moexiprilat on oestrogen‐stimulated cardiac fibroblast growth
Authors: Grohé C , Kahlert S , Löbbert K , van Eickels M , Stimpel M , Vetter H , Neyses L
Published: August 2006

The effects of 2‐2‐(1‐(ethoxycarbonyl)‐3‐phenylpropyl)‐[amino‐oxopropyl]−6,7‐dimethoxy‐1,2,3,4‐tetrahydroisoquinoline‐3 carboxylic acid (moexiprilat), 17β‐oestradiol (E2), oestrone (ES) and angiotensin II (AII) on growth and activation of oestrogen receptors and the immediate‐early gene egr‐1 were investigated in neonatal rat cardiac fibroblasts of female and male origin.

In BrdU proliferation assays, oestrone (10−7–10−9 M) stimulated cardiac fibroblast growth in a concentration‐dependent fashion (maximum at 10−7 M, 4.0 fold±0.14 in female and 3.1 fold±0.06 in male cells, n=9, P<0.05), while E2 (10−7–10−9 M) had no effect. Moexiprilat (10−7 M) completely inhibited oestrone‐induced cardiac fibroblast growth.

Angiotensin II (10−7 M) induced cardiac fibroblast growth (female 4.1 fold±0.1/male 3.9 fold±0.2; n=9, P<0.05). Angiotensin II induced oestrogen receptor (maximum 21.8 fold at 60 min) and egr‐1 (maximum 47.5 fold at 60 min) expression in a time‐dependent fashion.

In immunoblot experiments, oestrogen activated oestrogen receptor (ES: 12.8 fold±2.0; E2: 14.7 fold±4.9; n=3, P<0.05) and egr‐1 (ES: 5.1 fold,±0.24; E2: 3.8 fold,±0.25; n=3, P<0.05) expression. The induction of oestrogen receptor and egr‐1 protein expression was time‐dependent and inhibited by moexiprilat.

Our results show that oestrone and 17β‐oestradiol reveal a significant difference in their potential to activate cardiac fibroblast growth in female and male cells and that oestrone‐stimulated growth is inhibited by moexiprilat. The inhibition of oestrone‐stimulated cardiac fibroblast growth by moexiprilat may contribute to the beneficial effects seen in postmenopausal women with hypertensive heart disease treated with ACE inhibitors.
Resuscitating effect of melanocortin peptides after prolonged respiratory arrest
Authors: Guarini Salvatore , Bazzani Carla , Bertolini Alfio
Published: August 2006

The resuscitating activity of melanocortin peptides (MSH‐ACTH peptides) was tested in an experimental model of prolonged respiratory arrest.

Anaesthetized, endotracheally intubated rats subjected to a 5 min period of ventilation interruption, invariably died from cardiac arrest within 6–9 min of resumption of ventilation.

When resumption of ventilation was associated with the simultaneous intravenous (i.v.) injection of a melanocortin peptide (α‐MSH or ACTH‐(1–24)) (160 μg kg−1) there was an almost immediate (within 1 min), impressive increase in cardiac output, heart rate, mean arterial pressure (+560% of the before‐treatment value) and pulse pressure (+356% of the before‐treatment value), with full recovery of electroencephalogram after 30–45 min. Blood gases and pH were normalized within 15–60 min after treatment, and all treated animals eventually recovered completely and survived indefinitely (=more than 15 days).

The same response was observed in adrenalectomized animals, as well as in animals pretreated with a β1‐adrenoceptor blocking agent (atenolol, 3 mg kg−1, i.v.), or with an α1‐adrenoceptor blocking agent (prazosin, 0.1 mg kg−1, i.v.), or with an adrenergic neurone blocking agent (guanethidine, 10 mg kg−1, intraperitoneally).

An effect quite similar to that produced by melanocortins was obtained with ouabain (0.1 mg kg−1, i.v.); the antioxidant drug, glutathione (75 mg kg−1, i.v.) also produced 100% resuscitation, but the effect was slower in onset. On the other hand, adrenaline (0.005 mg kg−1, i.v.) was able to resuscitate only 1 out of 8 rats and dobutamine (0.02 mg kg−1, i.v.) resuscitated 4 out of 8 rats; moreover, the effect of both catecholamines was much slower in onset than that of melanocortins and the initial, impressive stimulation of cardiovascular function was absent.

These results show that melanocortin peptides have a resuscitating effect in a pre‐terminal condition produced in rats by prolonged asphyxia. This effect seems primarily due to the restoration of cardiac function, not mediated by catecholamines. These data also suggest that these peptides may have potential therapeutic value in conditions of transient cardiac hypoxia and re‐oxygenation such as occur in coronary artery disease.
Properties of the pore‐forming P2X7 purinoceptor in mouse NTW8 microglial cells
Authors: Chessell I P , Michel A D , Humphrey P P A
Published: August 2006

We have used whole‐cell patch clamping methods to study and characterize the cytolytic P2X7 (P2Z) receptor in the NTW8 mouse microglial cell line.

At room temperature, in an extracellular solution containing 2 mM Ca2+ and 1 mM Mg2+, 2′‐ and 3′‐O‐(4‐benzoylbenzoyl)‐adenosine‐5′‐triphosphate (Bz‐ATP; 300 μM), or ATP (3 mM), evoked peak whole cell inward currents, at a holding potential of −90 mV, of 549±191 and 644±198 pA, respectively. Current‐voltage relationships generated with 3 mM ATP reversed at 4.6 mV and did not display strong rectification.

In an extracellular solution containing zero Mg2+ and 500 μM Ca2+ (low divalent solution), brief (0.5 s) application of these agonists elicited larger maximal currents (909±138 and 1818±218 pA, Bz‐ATP and ATP, respectively). Longer application of ATP (1 mM for 30 s) produced larger, slowly developing, currents which reached a plateau after approximately 15–20 s and were reversible on washing. Under these conditions, in the presence of ATP, ethidium bromide uptake could be demonstrated. Further applictions of 1 mM ATP produced rapid currents of the same magnitude as those observed during the 30 s application. Subsequent determination of concentration‐effect curves to Bz‐ATP, ATP and 2‐methylthio‐ATP yielded EC50 values of 58.3, 298 and 505 μM, respectively. These affects of ATP were antagonized by pyridoxal‐phosphate‐6‐azophenyl‐ 2′, 4′‐disulphonic acid (PPADS; 30 μM) but not suramin (100 μM).

In low divalent solution, repeated application of 1 mM ATP for 1 s produced successively larger currents which reached a plateau, after 8 applications, of 466% of the first application current. PPADS (30 μM) prevented this augmentation, while 5‐(N,N‐hexamethylene)‐amiloride (HMA) (100 μM) accelerated it such that maximal augmentation was observed after only one application of ATP in the presence of HMA. At a bath temperature of 32°C, current augmentation also occurred in normal divalent cation containing solution.

These data demonstrate that mouse microglial NTW8 cells possess a purinoceptor with pharmacological characteristics resembling the P2X7 receptor. We suggest that the current augmentation phenomenon observed reflects formation of the large cytolytic pore characteristic of this receptor. We have demonstrated that pore formation can occur under normal physiological conditions and can be modulated pharmacologically, both positively and negatively.
Stereospecific effects of ketamine enantiomers on canine tracheal smooth muscle
Authors: Pabelick Christina M , Rehder Kai , Jones Keith A , Shumway Robin , Lindahl Sten G E , Warner David O
Published: August 2006

Ketamine is a potent bronchodilator which relaxes airway smooth muscle (ASM). Clinically, ketamine is used as a 1:1 racemic mixture of enantiomers that differ in their analgesic and anaesthetic effects. The aim of this study was to determine whether there was a difference between the enantiomers in their ability to relax isolated ASM and to explore mechanisms responsible for any observed differences.

Canine tracheal smooth muscle strips were loaded with fura‐2 and mounted in a photometric system to measure simultaneously force and [Ca2+]i. Calcium influx was estimated by use of a manganese quenching technique.

In strips stimulated with 0.1 μM ACh (EC50) R(−)‐ketamine (1–100 μM) caused a significantly greater concentration‐dependent decrease in force (P<0.0001) and [Ca2+]i than S(+)‐ketamine (1–100 μM) (P<0.0005). In contrast, there was no significant difference between the enantiomers in their ability to inhibit calcium influx (45% decrease in influx rate for R(−)‐ketamine and 44% for S(+)‐ketamine, P=0.782). In strips contracted with 24 mM isotonic KCl (which activates voltage‐operated calcium channels), the enantiomers modestly decreased force and [Ca2+]i; there was no significant difference between the enantiomers in their effects on force (P=0.425) or [Ca2+]i (P=0.604).

The R(−)‐enantiomer of ketamine is a more potent relaxant of ACh‐induced ASM contraction than the S(+)‐enantiomer. This difference appears to be caused by differential actions on receptor‐operated calcium channels.
In vivo evidence for B1‐receptor synthesis induction by heat stress in the rat
Authors: Lagneux Caroline , Ribuot Christophe
Published: August 2006

A hypotensive effect of intravenously injected [des‐Arg9]‐bradykinin was found in Wistar rats following acute heat stress. This effect was similar to that of intravenously injected bradykinin and was observed 6, 18 and 24h following an increase in rectal temperature at 42°C for 20min (H6, H18 and H24 groups, respectively). In contrast, [des‐Arg9]‐bradykinin had no effect on blood pressure in control or sham conditions, early on (3h) or later on (72h) after heat stress (Ctl, H3 and H72 groups, respectively), while the response to bradykinin was maintained. The hypotension induced by [des‐Arg9]‐bradykinin in groups H6, H18 and H24 was comparable to that induced in rats pretreated with a small amount of endotoxin 24h earlier (LPS group). The hypotensive response in group H24 was totally blunted by [des‐Arg10]‐Hoe 140 (a potent B1 receptor antagonist) infused at a rate of 10μgmin−1. These results suggest that heat stress induces the synthesis of vascular B1 receptors in the rat.
NANC transmitters in the female pig urethra–localization and modulation of release via α2‐adrenoceptors and potassium channels
Authors: Werkström Viktoria , Persson Katarina , Andersson Karl‐Erik
Published: August 2006

To investigate further the release, localization and identity of a non‐nitrergic mediator of smooth muscle relaxation in the female pig urethra, we studied the effects of drugs acting at α2‐adrenoceptors or K+ channels, the effects of capsaicin and chemical sympathectomy, and the actions of several transmitter candidates.

Electrical field stimulation (EFS; frequencies above 12 Hz) of spontaneously contracted smooth muscle strips from the female pig urethra evoked long‐lasting, frequency‐dependent relaxations in the presence of prazosin, scopolamine, and NG‐nitro‐L‐arginine. Treatment with the selective α2‐adrenoceptor agonist UK‐14 304 markedly reduced the relaxations evoked by EFS at all frequencies tested (16–30 Hz). The inhibitory effect of UK‐14 304 was completely antagonized by the α2‐adrenoceptor antagonist rauwolscine. The muscarinic M1 receptor antagonist, pirenzepine, or exogenously administered carbachol, did not have any effects on the electrically evoked relaxations.

Inhibition of high conductance Ca2+ activated K+ channels by iberiotoxin or charybdotoxin significantly enhanced the relaxations evoked by EFS at all frequencies. However, inhibition of voltage‐sensitive K+ channels with 4‐aminopyridine or dendrotoxin‐1, treatment with the ATP‐sensitive K+ channel blocker, glibenclamide, or treatment with the high and low conductance Ca2+ activated K+ channel blockers, tetraethylammonium chloride and apamin, had no effect on the relaxations evoked by EFS.

Electrically evoked relaxations were not affected by adrenergic denervation with 6‐hydroxydopamine (6‐OHDA) at any frequency. However, treatment with 6‐OHDA abolished prazosin‐sensitive electrically induced contractions, and a long‐lasting relaxation was revealed. Treatment with capsaicin, believed to damage selectively a subpopulation of primary afferent fibres, did not affect basal tone or relaxations evoked by EFS.

Exogenously applied vasoactive intestinal polypeptide (VIP), pituitary adenylate cyclase‐activating peptide (PACAP)‐27, PACAP‐38, adenosine, ATP and 5‐hydroxy‐tryptamine caused relaxations of the urethral preparations, whereas prostaglandin E2 and calcitonin gene‐related peptide had no effects. VIP 10‐28, α, β‐methylene‐ATP, reactive blue‐2, suramin or indomethacin did not reduce the electrically‐evoked relaxations at any frequency. However, the relaxations were slightly reduced by trypsin or α‐chymotrypsin.

The present results suggest that the release of the unknown mediator in the female pig urethra can be modulated via α2‐adrenoceptors and high conductance Ca2+ activated K+ channels. Furthermore, the mediator does not appear to be localized to or released from adrenergic or capsaicin‐sensitive sensory nerve‐endings. The identity of the transmitter remains to be established.

British Journal of Pharmacology (1997) 121, 1605–1612; doi:10.1038/sj.bjp.0701308
Involvement of interleukin‐1β, nerve growth factor and prostaglandin E2 in endotoxin‐induced localized inflammatory hyperalgesia
Authors: Safieh‐Garabedian Bared , Kanaan Salim A. , Haddad John J. , Abou Jaoude Pamela , Jabbur Suhayl J. , Saadé Nayef E.
Published: August 2006

Intraplantar endotoxin (ET) injection (1.25 μg) into the hind paw of rats resulted in a localized inflammatory hyperalgesia, as assessed by paw pressure (PP), paw immersion (PI), tail flick (TF) and hot plate (HP) tests.

ET injection resulted in a significant elevation in the levels of interleukin‐1β (IL‐1β) and nerve growth factor (NGF) in the injected foot as compared with the non‐injected foot. This increase was attenuated by intraperitoneal injections of dexamethasone (200 and 400 μg kg−1) and to a lesser extent by indomethacin (2 and 8 mg kg−1).

The tripeptide Lys‐D‐Pro‐Val, which is known to antagonize IL‐1β and prostaglandin E2 (PGE2) reversed mechanical hyperalgesia, as assessed by the PP test, and reduced significantly thermal hyperalgesia, as assessed by the HP and TF tests.

IL‐1ra reversed both mechanical (PP) and thermal (PI) nociceptive thresholds tested on the injected leg and significantly reduced thermal hyperalgesia, as assessed by the HP and TF tests.

A sheep, anti‐mouse NGF antiserum reversed mechanical hyperalgesia (PP test) but had little or no effect on thermal hyperalgesia (PI, HP and TF tests).

Our results indicate the importance of IL‐1β, NGF and prostaglandin E2 (PGE2) in the development of ET induced hyperalgesia and the possible existence of different mechanisms underlying thermal and mechanical as well as central and peripheral hyperalgesia.

British Journal of Pharmacology (1997) 121, 1619–1626; doi:10.1038/sj.bjp.0701313
Role of endothelin receptors, calcium and nitric oxide in the potentiation by endothelin‐1 of the sympathetic contraction of rabbit ear artery during cooling
Authors: García‐Villalón A. L. , Padilla J , Fernández N , Monge L , Gómez B , Diéguez G
Published: August 2006

To examine further the potentiation by endothelin‐1 on the vascular response to sympathetic stimulation, we studied the isometric response of isolated segments, 2 mm long, from the rabbit central ear artery to electrical field stimulation (1–8 Hz), under different conditions, at 37°C and during cooling (30°C).

Electrical stimulation produced frequency‐dependent contraction, which was reduced (about 63% for 8 Hz) during cooling. At 30°C, but not at 37°C, endothelin‐1 (1, 3 and 10 nM) potentiated the contraction to electrical stimulation in a dose‐dependent way (from 43±7% to 190±25% for 8 Hz).

This potentiation by endothelin‐1 was reduced by the antagonist for endothelin ETA receptors BQ‐123 (10 μM) but not by the antagonist for endothelin ETB receptors BQ‐788 (10 μM). The agonist for endothelin ETB receptors IRL‐1620 (0.1 μM) did not modify the contraction to electrical stimulation.

The blocker of L‐type Ca2+ channels verapamil (10 μM l−1) reduced (about 72% for 8 Hz) and the unspecific blocker of Ca2+‐channels NiCl2 (1 mM) practically abolished (about 98%), the potentiating effects of endothelin‐1 found at 30°C.

Inhibition of nitric oxide synthesis with NG‐nitro‐L‐arginine (L‐NOARG, 0.1 mM) increased the contraction to electrical stimulation at 30°C more than at 37°C (for 8 Hz, this increment was 297±118% at 30°C, and 66±15% at 37°C). Endothelium removal increased the contraction to electrical stimulation at 30°C (about 91% for 8 Hz) but not at 37°C. Both L‐NOARG and endothelium removal abolished the potentiating effects of endothelin‐1 on the response to electrical stimulation found at 30°C.

These results in the rabbit ear artery suggest that during cooling, endothelin‐1 potentiates the contraction to sympathetic stimulation, which could be mediated at least in part by increasing Ca2+ entry after activation of endothelin ETA receptors. This potentiating effect of endothelin‐1 may require the presence of an inhibitory tone due to endothelial nitric oxide.

British Journal of Pharmacology (1997) 121, 1659–1664; doi:10.1038/sj.bjp.0701324
The effects of neuroleptic and tricyclic compounds on BKCa channel activity in rat isolated cortical neurones
Authors: Lee K , McKenna F , Rowe I. C. M. , Ashford M. L. J.
Published: August 2006

The actions of several neuroleptic and tricyclic compounds were examined on the large conductance Ca2+‐activated K+ (BKCa) channel present in neurones isolated from the rat motor cortex.

Classical neuroleptic compounds including chlorpromazine and haloperidol applied to the intracellular surface of inside‐out patches produced a concentration‐dependent reduction in BKCa channel activity. Similar effects were observed when these compounds were applied to the extracellular surface of outside‐out patches.

In contrast, the atypical neuroleptic compounds clozapine and sulpiride did not affect BKCa channel activity (100 nM–1 mM) in either inside‐out or outside‐out patches, while 10 μM pimozide produced 73% of the inhibition produced by 10 μM chlorpromazine.

BKCa channel activity was also unaffected by application of structurally related tricyclic compounds including the anti‐cholinesterase tacrine and the anti‐epileptic carbamazepine. The tricyclic antidepressant drug amitriptyline was found to inhibit BKCa channel activity but was much less effective than the classical neuroleptic compounds.

It is concluded that compounds belonging to the classical neuroleptic group of drugs inhibit BKCa channel activity in the rat motor cortex in a structurally‐specific manner. This observation may be of clinical significance as it may contribute to some of the side effects associated with classical neuroleptic drug therapy.

British Journal of Pharmacology (1997) 121, 1810–1816; doi:10.1038/sj.bjp.0701333