bluuebunny/crossref_metadata_2025 huggingface dataset:
Improvement of exercise capacity of rats with chronic heart failure by long‐term treatment with trandolapril
Authors: Yamaguchi Fuminari , Kawana Ken‐ichiro , Tanonaka Kouichi , Kamano Isamu , Igarashi Takahiro , Gen Eigyoku , Fujimoto Yoko , Maki Toshiyuki , Sanbe Atsushi , Nasa Yoshihisa , Takeo Satoshi
Published: January 2005

The effects of long‐term treatment with trandolapril, an angiotensin I‐converting enzyme inhibitor, on exercise capacity of rats with chronic heart failure (CHF) following coronary artery ligation were examined. CHF was developed by 8 weeks after the coronary artery ligation.

The running time of rats with CHF in the treadmill test was shortened to approximately 65% of that of sham‐operated rats (16.3±1.2 vs 25.1±1.6 min, n=7; P<0.05). ATP, creatine phosphate (CP), and lactate contents of the gracilis muscle of rats with CHF were similar to those of sham‐operated rats before running. After running, ATP and CP were decreased and lactate was increased in both rats with CHF and sham‐operated rats. There were no significant differences in the levels of energy metabolites between rats with CHF and sham‐operated rats. The rates of decrease in ATP and CP and rate of increase in lactate in the gracilis muscle of rats with CHF during exercise were greater than those of sham operated rats (2.5, 2.0 and 1.5 fold high, respectively), suggesting wastage of energy during exercise in the animals with CHF.

Myofibrillar Ca2+‐stimulated ATPase (Ca‐ATPase) activity of skeletal muscle of rats with CHF was increased over that of the sham‐operated control (62.03±1.88 vs 52.34±1.19 μmol Pi mg−1 protein h−1 n=7; P<0.05). The compositions of myosin heavy chain (MHC) isoforms of gracilis muscle were altered by CHF; decreases in MHC types I and IIb and an increase in MHC type IIa were found (P<0.05).

Rats with CHF were treated with 1 mg kg−1 day−1 trandolapril from the 2nd to 8th week after surgery. Treatment with trandolapril prolonged the running time, reversed the rates of decrease in ATP and CP and the rate of increase in lactate, and restored the Ca‐ATPase activity (51.11±0.56 μmol Pi mg−1 protein h−1, n=7; P<0.05) and composition ratio of MHC isoforms in the gracilis muscle.

The results suggest that long‐term trandolapril treatment of rats with CHF may restore their ability to utilize energy without wastage and thus improve exercise capacity.

British Journal of Pharmacology (1999) 126, 1585–1592; doi:10.1038/sj.bjp.0702471
Adenosine A3 receptors on human eosinophils mediate inhibition of degranulation and superoxide anion release
Authors: Ezeamuzie Charles I , Philips Elizabeth
Published: January 2005

The role of adenosine A3 receptors on human eosinophil degranulation and superoxide anion (O2−) release was studied in vitro using the complement fragment C5a as the main stimulus and employing a number of selective agonists and antagonists.

In the presence of cytochalasin B (CB), C5a induced a dose‐dependent release of the granular eosinophil peroxidase (EPO), but not O2−, whereas in the absence of CB O2−, but not EPO, was released.

C5a‐induced EPO release was inhibited dose‐dependently by the selective A3 agonist N6‐(3‐iodobenzyl)‐5′‐N‐methylcarbamoyladenosine (IB‐MECA) and to a lesser extent by the less‐selective N6‐2‐(4‐amino‐3‐iodophenyl) ethyladenosine (APNEA). The IC50 (95% CI) for IB‐MECA was 6.8 μM (3.1–12.0 μM). At concentrations up to 100 μM, neither adenosine nor the selective A1 agonist N‐cyclopentyladenosine (CPA) and the selective A2 agonist 2‐[[2‐[4‐(2‐carboxyethyl)phenyl]ethyl]amino]‐N‐ethylcarboxamidoadenosine (CGS 21680) had any significant effect.

The inhibitory effect of IB‐MECA was almost completely abolished by pre‐treatment with 1 μM of the selective A3 antagonist 9‐chloro‐2‐(2‐furyl)‐5‐phenylactylamino[1,2,4]triazolo[1,5‐c]quinazoline (MRS 1220), but not the selective A1 antagonist 1,3‐dipropyly‐8‐cyclopentylxanthine (DPCPX) or the selective A2 antagonist 3,7‐dimethyl‐1‐propargylxanthine (DMPX).

IB‐MECA also significantly inhibited C5a‐induced O2− release with IC50 (95% CI) of 9.5 μM (4.6–13.1 μM) whereas adenosine and the A1 agonist CPA potentiated this effect at low concentrations. The potentiation appeared to be a result of their direct O2− release from these cells, probably mediated via A1 receptors. The inhibition by IB‐MECA was selectively reversed by MRS 1220.

These results show that the A3 receptors on human eosinophils mediate inhibition of both degranulation and O2− release and suggest a therapeutic potential for A3 agonists in diseases such as asthma in which activated eosinophils are involved.

British Journal of Pharmacology (1999) 127, 188–194; doi:10.1038/sj.bjp.0702476
Effects of terbutaline on force and intracellular calcium in slow‐twitch skeletal muscle fibres of the rat
Authors: Ha Tuyen N V , Posterino Giuseppe S , Fryer Martin W
Published: January 2005

The effect of the β2‐adrenoceptor agonist, terbutaline, was investigated on simultaneously measured force and intracellular free calcium ([Ca2+]i) in intact rat soleus muscle fibres, and on contractile protein function and Ca2+ content of the sarcoplasmic reticulum (SR) in skinned fibres.

Terbutaline (10 μM) had no significant effect on either resting force or [Ca2+]i. Exposure to terbutaline increased both the integral of the indo‐1 ratio transient and peak twitch force by 37%.

At sub‐maximal (10 Hz) stimulation frequencies, terbutaline accelerated force relaxation but had highly variable effects on tetanic force amplitude. The corresponding indo‐1 ratio transients were significantly larger, and faster to decay than the controls.

Terbutaline increased tetanic force at near maximal stimulation frequencies (50 Hz) by increasing tetanic [Ca2+]i. Force relaxation was accelerated at this frequency with no significant change in the indo‐1 ratio transient decay rate.

All of terbutaline's effects on force and indo‐1 ratio transients in intact fibres were completely blocked and reversed by ICI 118551 (1 μM).

Mechanically skinned fibres isolated from intact muscles pre‐treated with terbutaline showed no significant changes in SR Ca2+ content, myofilament [Ca2+]i‐sensitivity or maximum force generating capacity.

The results suggest that terbutaline primarily modulates force by altering the amplitude and decay rate of the [Ca2+]i transient via phosphorylation of both the ryanodine receptor (RR) and the SR pump regulatory protein, phospholamban (PLB). The high variability of responses of slow‐twitch muscles to β2‐agonists probably reflects individual differences in basal phosphorylation levels of PLB relative to that of RR.

British Journal of Pharmacology (1999) 126, 1717–1724; doi:10.1038/sj.bjp.0702482
Anandamide‐induced mobilization of cytosolic Ca2+ in endothelial cells
Authors: Mombouli Jean‐Vivien , Schaeffer Gabriela , Holzmann Sigrid , Kostner Gert M , Graier Wolfgang F
Published: January 2005

Experiments were designed to determine whether anandamide affects cytosolic Ca2+ concentrations in endothelial cells and, if so, whether CB1 cannabinoid receptors are involved. To this effect, human umbilical vein‐derived EA.hy926 endothelial cells were loaded with fura‐2 to monitor changes in cytosolic Ca2+ using conventional fluorescence spectrometry methods.

Anandamide induced an increase in Ca2+ in endothelial cells which, in contrast to histamine, developed slowly and was transient. Anandamide caused a concentration‐dependent release of Ca2+ from intracellular stores without triggering capacitative Ca2+ entry, contrary to histamine or the endoplasmic reticulum Ca2+‐ATPase inhibitor thapsigargin.

Anandamide pretreatment slightly reduced the mobilization of Ca2+ from intracellular stores that was evoked by histamine. The mobilization of Ca2+ from intracellular stores evoked by anandamide was impaired by 10 mM caffeine.

Anandamide and histamine each significantly increased NO synthase activity in EA.hy926 cells, as determined by the enhanced conversion of L‐[3H]‐arginine to L‐[3H]‐citruline.

The CB1 cannabinoid receptor antagonist SR141716A (1 μM) only produced a marginal reduction of the mobilization of Ca2+ produced by 5 μM anandamide. However, at 5 μM SR141716A elicited the release of Ca2+ from intracellular stores. This concentration strongly impaired the mobilization of cytosolic Ca2+ evoked by either anandamide, histamine or thapsigargin.

Pretreatment of the cells with either 200 μM phenylmethylsulphonyl fluoride (to inhibit the conversion of anandamide into arachidonic acid) or 400 ng ml−1 pertussis toxin (to uncouple CB1 cannabinoid receptors from Gi/o proteins) had no significant effect on the mobilization of cytosolic Ca2+ evoked by either anandamide, or histamine.

Taken together the results demonstrate that anandamide mobilizes Ca2+ from a caffeine‐sensitive intracellular Ca2+ store that functionally overlaps in part with the internal stores mobilized by histamine. However, a classical CB1 cannabinoid receptor‐mediated and pertussis toxin‐sensitive mechanism does not mediate this novel effect of anandamide in endothelial cells.

The mobilization of cytosolic Ca2+ in endothelial cells may account for the endothelium‐dependent and NO‐mediated vasodilator actions of anandamide. Due to its non‐specific inhibition of Ca2+ signalling in endothelial cells, SR141716A may not be used to assess the physiological involvement of endogenous cannabinoids to endothelium‐dependent control of vascular smooth muscle tone.

British Journal of Pharmacology (1999) 126, 1593–1600; doi:10.1038/sj.bjp.0702483
Pilocarpine modulates the cellular electrical properties of mammalian hearts by activating a cardiac M3 receptor and a K+ current
Authors: Wang Huizhen , Shi Hong , Lu Yanjie , Yang Baofeng , Wang Zhiguo
Published: January 2005

Pilocarpine, a muscarinic acetylcholine receptor (mAChR) agonist, is widely used for treatment of xerostomia and glaucoma. It can also cause many other cellular responses by activating different subtypes of mAChRs in different tissues. However, the potential role of pilocarpine in modulating cardiac function remained unstudied.

We found that pilocarpine produced concentration‐dependent (0.1–10 μM) decrease in sinus rhythm and action potential duration, and hyperpolarization of membrane potential in guinea‐pig hearts. The effects were nearly completely reversed by 1 μM atropine or 2 nM 4DAMP methiodide (an M3‐selective antagonist).

Patch‐clamp recordings in dispersed myocytes from guinea‐pig and canine atria revealed that pilocarpine induces a novel K+ current with delayed rectifying properties. The current was suppressed by low concentrations of M3‐selective antagonists 4DAMP methiodide (2–10 nM), 4DAMP mustard (4–20 nM, an ackylating agent) and p‐F‐HHSiD (20–200 nM). Antagonists towards other subtypes (M1, M2 or M4) all failed to alter the current.

The affinity of pilocarpine (KD) at mAChRs derived from displacement binding of [3H]‐NMS in the homogenates from dog atria was 2.2 μM (65% of the total binding) and that of 4DAMP methiodide was 2.8 nM (70% of total binding), consistent with the concentration of pilocarpine needed for the current induction and for the modulation of the cardiac electrical activity and the concentration of 4DAMP to block pilocarpine effects.

Our data indicate, for the first time, that pilocarpine modulates the cellular electrical properties of the hearts, likely by activating a K+ current mediated by M3 receptors.

British Journal of Pharmacology (1999) 126, 1725–1734; doi:10.1038/sj.bjp.0702486
Mitogenic activation of human prostate‐derived fibromuscular stromal cells by bradykinin
Authors: Walden Paul D , Lefkowitz Gary K , Ittmann Michael , Lepor Herbert , Monaco Marie E
Published: January 2005

Biologically active kinin peptides are released from precursor kininogens by kallikreins. Kinins act on kinin receptors to mediate diverse biological functions including smooth muscle contraction, inflammation, pain and mitogenicity. All components of the kallikrein‐kinin system exist in human male genital secretions suggesting that these molecules participate in physiological and pathophysiological genitourinary function. The objective of this study was to assess the consequences of kinin action on prostate cells.

Primary cultures of prostate secretory epithelial (PE) and prostate fibromuscular stromal (PS) cells were established from human prostate tissue. Transcripts encoding both the human B1 and B2 bradykinin receptor subtypes were detected in human prostate transition‐zone tissue and in cultured cells by RT–PCR. In receptor binding assays, the B1 subtype predominated on PE cell membranes and the B2 subtype predominated on PS cell membranes. In PS cells, but not in PE cells, BK induced significant inositol phosphate accumulation and [3H]‐thymidine uptake. These responses were mediated through the B2 receptor subtype.

The use of signal transduction inhibitors indicated that mitogenic activation by BK occurred through both protein kinase C (PKC) and protein tyrosine kinase dependent mechanisms. PMA (phorbol 12‐myristate 13‐acetate) produced maximal [3H]‐thymidine uptake by PS cells, resulted in cell elongation and caused the α‐actin fibres present in PS smooth muscle cells to became organized into parallel arrays along the length of the elongated cells.

In summary, the prostate contains a functional kallikrein‐kinin system, which could be significant in physiological and pathophysiological prostate function.

British Journal of Pharmacology (1999) 127, 220–226; doi:10.1038/sj.bjp.0702492
Comparison of antagonist potencies at pre‐ and post‐synaptic GABAB receptors at inhibitory synapses in the CA1 region of the rat hippocampus
Authors: Pozza M F , Manuel N A , Steinmann M , Froestl W , Davies C H
Published: January 2005

Synaptic activation of γ‐aminobutyric acid (GABA)B receptors at GABA synapses causes (a) postsynaptic hyperpolarization mediating a slow inhibitory postsynaptic potential/current (IPSP/C) and (b) presynaptic inhibition of GABA release which depresses IPSPs and leads to paired‐pulse widening of excitatory postsynaptic potentials (EPSPs). To address whether these effects are mediated by pharmacologically identical receptors the effects of six GABAB receptor antagonists of widely ranging potencies were tested against each response.

Monosynaptic IPSPBs were recorded in the presence of GABAA, AMPA/kainate and NMDA receptor antagonists. All GABAB receptor antagonists tested depressed the IPSPB with an IC50 based rank order of potency of CGP55679CGP56433=CGP55845A=CGP52432>CGP51176> CGP36742.

Paired‐pulse EPSP widening was recorded as an index of paired‐pulse depression of GABA‐mediated IPSP/Cs. A similar rank order of potency of antagonism of paired‐pulse widening was observed to that for IPSPB inhibition.

Comparison of the IC50 values for IPSPB inhibition and paired‐pulse EPSP widening revealed a close correlation between the two effects in that their IC50s lay within the 95% confidence limits of a correlation line that described IC50 values for inhibition of paired‐pulse EPSP widening that were 7.3 times higher than those for IPSPB inhibition.

Using the compounds tested here it is not possible to assign different subtypes of GABAB receptor to pre‐ and post‐synaptic loci at GABAergic synapses. However, 5–10 fold higher concentrations of antagonist are required to block presynaptic as opposed to postsynaptic receptors when these are activated by synaptically released GABA.

British Journal of Pharmacology (1999) 127, 211–219; doi:10.1038/sj.bjp.0702498
Rate of change of blood concentrations is a major determinant of the pharmacodynamics of midazolam in rats
Authors: Cleton A , Mazee D , Voskuyl R A , Danhof M
Published: January 2005

The objective of this investigation was to characterize quantitatively the influence of the rate of increase in blood concentrations on the pharmacodynamics of midazolam in rats. The pharmacodynamics of midazolam were quantified by an integrated pharmacokinetic‐pharmacodynamic modelling approach.

Using a computer controlled infusion technique, a linear increase in blood concentrations up to 80 ng ml−1 was obtained over different time intervals of 1–6 h, resulting in rates of rise of the blood concentrations of respectively, 1.25, 1.00, 0.87, 0.46, 0.34 and 0.20 ng ml−1 min−1. In one group of rats the midazolam concentration was immediately brought to 80 ng ml−1 and maintained at that level for 4 h. Immediately after the pretreatment an intravenous bolus dose was given to determine the time course of the EEG effect in conjunction with the decline of midazolam concentrations.

The increase in β activity (11.5–30 Hz) of the EEG was used as pharmacodynamic endpoint. For each individual animal the relationship between blood concentration and the EEG effect could be described by the sigmoidal Emax model. After placebo, the values of the pharmacodynamic parameter estimates were Emax=82±5 μV, EC50,u=6.4±0.8 ng ml−1 and Hill factor=1.4±0.1. A bell‐shaped relationship between the rate of change of midazolam concentration and the value of EC50,u was observed with a maximum of 21±5.0 ng ml−1 at a rate of change of 0.46 ng ml−1 min−1; lower values of EC50,u were observed at both higher and lower rates.

The findings of this study show that the rate of change in plasma concentrations is an important determinant of the pharmacodynamics of midazolam in rats.

British Journal of Pharmacology (1999) 127, 227–235; doi:10.1038/sj.bjp.0702499
Cross‐talk between group IIA‐phospholipase A2 and inducible NO‐synthase in rat renal mesangial cells
Authors: Rupprecht Gerhard , Scholz Kirsten , Beck Karl‐Friedrich , Geiger Helmut , Pfeilschifter Josef , Kaszkin Marietta
Published: January 2005

Features of glomerulonephritis are expression of the inducible form of NO synthase (iNOS) as well as expression of the secretory group IIA‐phospholipase A2 (sPLA2) in mesangial cells. Interleukin 1β (IL‐1β) induces both enzymes with a similar time course resulting in an increase in nitrite production and sPLA2‐IIA activity. In this study we investigated the relationship between the formation of NO and sPLA2‐IIA induction in rat renal mesangial cells.

Incubation of mesangial cells with the NO‐donor, spermine‐NONOate, for 24 h induced sPLA2‐IIA mRNA expression and activity, whereas S‐nitroso glutathione alone had only a small stimulatory effect. Stimulation of cells with IL‐1β caused a marked increase in sPLA2‐IIA mRNA and activity that were potentiated 3 fold by both NO donors.

Coincubation of cells with IL‐1β and the NOS inhibitor, L‐NG monomethylarginine (L‐NMMA), caused a dose‐dependent inhibition of cytokine‐induced sPLA2‐IIA mRNA expression and activity.

sPLA2‐IIA activity was not stimulated by 8‐bromo‐cyclic GMP indicating that NO‐induced sPLA2‐IIA induction is independent of cyclic GMP‐mediated signal transduction.

These data show that NO contributes to the expression by cytokines of sPLA2‐IIA and establishes a novel type of interaction between iNOS and sPLA2‐IIA in mesangial cells. This cross‐talk between inflammatory mediators may help to promote and sustain an inflammatory state in the kidney.

British Journal of Pharmacology (1999) 127, 51–56; doi:10.1038/sj.bjp.0702500
Effects of oral propylthiouracil treatment on nitric oxide production in rat aorta
Authors: Grieve David J , Fletcher Sara , Pitsillides Andrew A , Botham Kathleen M , Elliott Jonathan
Published: January 2005

The effects of oral propylthiouracil (PTU) treatment on vascular nitric oxide (NO) production were studied in the rat aorta.

Rats were fed a standard low fat diet with or without 0.1% PTU, for 2 or 4 weeks, or for 2 weeks with additional thyroxine injections. Concentration response curves were then constructed to phenylephrine (PE) in both endothelium‐intact and denuded aortic rings from these animals and after incubation with 0.1 mM L‐NGnitroarginine (L‐NOARG). In addition, expression of nitric oxide synthase (NOS) was analysed in sections of aorta from PTU‐treated and control rats using rabbit polyclonal antibodies to both inducible NOS (iNOS) and endothelial NOS (eNOS).

Oral PTU treatment resulted in a significant reduction in both the maximum response (control, 0.53±0.02; 2 week PTU, 0.20±0.07; 4 week PTU, 0.07±0.02 g mg−1) and vessel sensitivity (EC50 values: control, 9.10×10−8±0.67; 2 week PTU, 7.45×10−7±1.15; 4 week PTU, 9.73×10−7±0.45 M) to PE in endothelium‐intact vessel rings, as compared to controls (P<0.05). Both endothelial removal and incubation with L‐NOARG restored the maximum response after 2, but not 4 weeks, although, in general, vessel sensitivity was not altered by either treatment. Vessels from PTU‐treated rats given thyroxine injections showed no significant differences between any of the dose response curve parameters. Immunohistochemical analysis suggested that labelling for eNOS may be increased after PTU treatment as compared to control animals, whereas iNOS antibody immunoreactivity was not different between the two groups.

These results suggest that the hyporesponsiveness to PE observed after oral PTU treatment is, in part, due to enhanced nitric oxide (NO) production by the endothelium, and demonstrate for the first time that thyroid hormones may play a role in the regulation of eNOS activity in the rat aorta.

British Journal of Pharmacology (1999) 127, 1–8; doi:10.1038/sj.bjp.0702501
Pindolol‐insensitive [3H]‐5‐hydroxytryptamine binding in the rat hypothalamus; identity with 5‐hydroxytryptamine7 receptors
Authors: Clemett Delyth A , Kendall David A , Cockett Mark I , Marsden Charles A , Fone Kevin C F
Published: January 2005

Pindolol‐insensitive [3H]‐5‐hydroxytryptamine ([3H]‐5‐HT) binding to rat hypothalamic membranes was pharmacologically and functionally characterized to resolve whether this procedure selectively labels 5‐HT7 receptors.

Consistent with a previous report, 3 μM and not 100 nM pindolol was required to occupy fully 5‐HT1A and 5‐HT1B receptors. Remaining [3H]‐5‐HT binding was saturable (KD, 1.59±0.21 nM; Bmax, 53.8±3.1 fmol.mg protein−1).

Displacement of [3H]‐5‐HT with metergoline and 5‐CT revealed shallow Hill slopes (<0.5) but seven other compounds had slopes >0.8 and pKi values and the rank order of affinity were significantly correlated (r=0.81 and 0.93, respectively) with published [3H]‐5‐HT binding to rat recombinant 5‐HT7 receptors.

In the presence of pindolol, 5‐HT‐enhanced accumulation of [32P]‐cyclic AMP was unaffected by the 5‐HT4 antagonist RS39604 (0.1 μM) or the 5‐ht6 antagonist Ro 04–6790 (1 μM) but significantly attenuated by mesulergine (250 nM), ritanserin (450 nM) or methiothepin (200 nM) which have high affinity for the 5‐HT7 receptor.

Intracerebroventricular pretreatment with the serotonergic neurotoxin 5,7‐dihydroxytryptamine, 5,7‐DHT, elevated the [3H]‐5‐HT Bmax 2 fold, indicating that the hypothalamic 5‐HT7 receptor is post‐synaptic to 5‐HT nerve terminals and regulated by synaptic 5‐HT levels.

These results suggest that, in the presence of 3 μM pindolol, [3H]‐5‐HT selectively labels hypothalamic binding sites consistent with functional 5‐HT7 receptors.

British Journal of Pharmacology (1999) 127, 236–242; doi:10.1038/sj.bjp.0702503
L‐365,260 inhibits in vitro acid secretion by interacting with a PKA pathway
Authors: Oiry Catherine , Pannequin Julie , Cormier Anne , Galleyrand Jean‐Claude , Martinez Jean
Published: January 2005

The aim of this study was to analyse the antisecretory mechanism of L‐365,260 in vitro in isolated rabbit gastric glands.

We showed that compound L‐365,260, described as a non‐peptide specific competitive CCK‐B receptor antagonist, was able to dose‐dependently inhibit [14C]‐aminopyrine accumulation induced by histamine (10−4 M), carbachol (5×10−5 M), 3‐isobutyl‐1‐methyl‐xanthine (IBMX) (5×10−6 M) and forskolin (5×10−7 M) with similar IC50 values respectively of 1.1±0.6×10−7 M, 1.9±1.2×10−7 M, 4.2±2.0×10−7 M and 4.0±2.8×10−7 M.

We showed that L‐365,260 acted beyond receptor activation and production of intracellular second messengers and that it had no action on the H+/K+‐ATPase.

We found that L‐365,260 inhibited cyclic AMP‐induced [14C]‐aminopyrine accumulation in digitonin‐permeabilized rabbit gastric glands, suggesting that this compound acted, at least in part, as an inhibitor of the cyclic AMP‐dependent protein kinase (PKA) pathway.

British Journal of Pharmacology (1999) 127, 259–267; doi:10.1038/sj.bjp.0702505
Evidence for proteinase‐activated receptor‐2 (PAR‐2)‐mediated mitogenesis in coronary artery smooth muscle cells
Authors: Bretschneider Ellen , Kaufmann Roland , Braun Marina , Wittpoth Michael , Glusa Erika , Nowak Gotz , Schrör Karsten
Published: January 2005

This study investigates, whether in addition to the thrombin receptor (PAR‐1), the proteinase‐activated receptor‐2 (PAR‐2) is present in vascular smooth muscle cells (SMC) and mediates mitogenesis. PAR‐2 is activated by low concentrations of trypsin and the synthetic peptide SLIGRL.

Stimulation of bovine coronary artery SMC by trypsin (2 nM) caused a 3 fold increase in DNLA‐synthesis. A similar effect was observed with 10 nM thrombin. Trypsin‐induced mitogenesis was inhibited by soybean trypsin inhibitor, indicating that the proteolytic activity of the enzyme was required for its mitogenic effect.

The specific PAR‐2‐activating peptide SLIGRL or the PAR1‐activating peptide SFFLRN did not elicit mitogenesis.

When the SMC were exposed to SLIGRL (40 nM), a homologous desensitization of cytosolic Ca2+ mobilization was found after subsequent stimulation with trypsin (40 nM) but not thrombin (15 nM).

Trypsin (2 nM) as well as SLIGRL (100 μM) activated the nuclear factor κB (NFκB) with a maximum response 2 h after stimulation of the SMC. This suggests that both agonists acted via a common receptor, PAR‐2. Maximum activation of NFκB by thrombin (10 nM) was detected after 4–5 h.

These data suggest that PAR‐2 is present in coronary SMC and mediates a mitogenic response. Activation of NFκB via either PAR‐1 or PAR‐2 does not predict mitogenesis.

British Journal of Pharmacology (1999) 126, 1735–1740; doi:10.1038/sj.bjp.0702509
Effects of A1‐adenosine receptor antagonists on purinergic transmission in the guinea‐pig vas deferens in vitro
Authors: Hardy Todd A , Brock James A
Published: January 2005

Intracellularly recorded excitatory junction potentials (e.j.ps) were used to study the effects of adenosine receptor antagonists on neurotransmitter release from postganglionic sympathetic nerve terminals in the guinea‐pig vas deferens in vitro.

The A1 adenosine receptor antagonists, 8‐phenyltheophylline (10 μM) and 8‐cyclopentyl‐1,3‐dipropylxanthine (0.1 μM), increased the amplitude of e.j.ps evoked during trains of 20 stimuli at 1 Hz in the presence, but not in the absence, of the α2‐adrenoceptor antagonist, yohimbine (1 μM) or the non‐selective α‐adrenoceptor antagonist, phentolamine (1 μM).

Adenosine (100 μM) reduced the amplitude of e.j.ps, both in the presence and in the absence of phentolamine (1 μM). This inhibitory effect of adenosine is most likely caused by a reduction in transmitter release as there was no detectable change in spontaneous e.j.p. amplitudes.

In the presence of phentolamine, application of the adenosine uptake inhibitor, S‐(p‐nitrobenzyl)‐6‐thioinosine (0.1 μM), had no effect on e.j.p. amplitudes.

The phosphodiesterase inhibitor, 3‐isobutyl‐1‐methylxanthine (100 μM), significantly increased the amplitudes of all e.j.ps evoked during trains of 20 stimuli at 1 Hz, both in the presence and in the absence of phentolamine (1 μM).

These results suggest that endogenous adenosine modulates neurotransmitter release by an action at prejunctional A1 adenosine receptors only when α2‐adrenoceptors are blocked.

British Journal of Pharmacology (1999) 126, 1761–1768; doi:10.1038/sj.bjp.0702514
Quantitative autoradiographic studies of relaxin binding in rat atria, uterus and cerebral cortex: characterization and effects of oestrogen treatment
Authors: Tan Y Y , Wade J D , Tregear G W , Summers R J
Published: January 2005

The binding characteristics of the relaxin receptor in rat atria, uterus and cortex were studied using a [33P]‐labelled human gene 2 relaxin (B33) and quantitative receptor autoradiography.

The binding kinetics of [33P]‐human gene 2 relaxin (B33) were investigated in slide‐mounted rat atrial sections. The binding achieved equilibrium after 60 min incubation at room temperature (23±1°C) and dissociated slowly. The association and dissociation rate constants were 4.31±0.34×108 M−1 min−1 and 1.55±0.38×10−3 min−1 respectively. Thus, the kinetic dissociation constant was 3.46±0.59 pM.

Binding was saturable to a single population of non‐interacting sites throughout atria, in uterine myometrium and the 5th layer of cerebral cortex. The binding affinities (pKD) of [33P]‐human gene 2 relaxin (B33) were 8.92±0.09 in atrial myocardium and 8.79±0.04 in cerebral cortex of male rats, and 8.79±0.10 in uterine myometrium. Receptor densities in the cerebral cortex and atria were higher than in uterine myometrium, indicating that relaxin also has important roles in non‐reproductive tissues.

In male rats, treatment with 17β‐oestradiol (20 μg in 0.1 ml sesame oil s.c., 18–24 h) significantly decreased the density of relaxin receptors in atria and cerebral cortex. Identical treatment in female rats had no significant effect in atria and cerebral cortex, but it significantly increased the density of relaxin receptors in uterine myometrium.

Relaxin binding was competitively displaced by porcine and rat native relaxins. Porcine native relaxin binds to the relaxin receptor in male rat atria (8.90±0.02), and cerebral cortex (8.90±0.03) and uterine myometrium (8.89±0.03) with affinities not significantly different from human gene 2 (B33) relaxin. Nevertheless, rat relaxin binds to the receptors with affinities (8.35±0.09 in atria, 8.22±0.07 in cerebral cortex and 8.48±0.06 in uterine myometrium) significantly less than human gene 2 (B33) and porcine relaxins.

Quantitative receptor autoradiography is the method of choice for measurement of affinities and densities of relaxin receptor in atria, uterine myometrium and cerebral cortex. High densities were found in all these tissues. 17β‐Oestradiol treatment produced complex effects where it increased the densities of relaxin receptors in uterus but decreased those in atria and cerebral cortex of the male rats, and had no effect on the atria and cerebral cortex of the female rats.

British Journal of Pharmacology (1999) 127, 91–98; doi:10.1038/sj.bjp.0702517
Recombinant human erythropoietin inhibits iNOS activity and reverts vascular dysfunction in splanchnic artery occlusion shock
Authors: Squadrito Francesco , Altavilla Domenica , Squadrito Giovanni , Campo Giuseppe M , Arlotta Mariarita , Quartarone Cristina , Saitta Antonino , Caputi Achille P
Published: January 2005

We investigated the effects of recombinant human erythropoietin (rh‐EPO) in splanchnic artery occlusion (SAO) shock. Sham operated animals were used as controls. Survival rate, mean arterial blood pressure (MAP), serum Tumor Necrosis Factor (TNF‐α), plasma nitrite/nitrate concentrations, red blood cell (RBC) count, blood haemoglobin (Hb), the responsiveness of aortic rings to phenylephrine (PE, 1 nM–10 μM) and the activity of inducible nitric oxide synthase (iNOS) were studied.

SAO shocked rats had a decreased survival rate (0% at 4 h of reperfusion, while sham shocked rats survived more than 4 h), enhanced serum TNF‐α concentrations, increased plasma nitrite/nitrate levels (60±9.5 μM; sham shocked rats=2±0.4 μM), decreased MAP, unchanged RBC count and blood Hb and enhanced iNOS activity in the aorta. Moreover aortic rings from shocked rats showed a marked hyporeactivity to PE.

Rh‐EPO (25, 50 and 100 U 100 g−1, 5 min following the onset of reperfusion) increased survival rate (70% at 4 h of reperfusion with the highest dose), reduced plasma nitrite/nitrate concentrations (10.3±3.3 μM), increased MAP, did not change RBC count and blood Hb, and inhibited iNOS activity in thoracic aortae. Furthermore rh‐EPO, either in vivo or in vitro (10 U for 1 h in the organ bath), restored to control values the hyporeactivity to PE. Finally rh‐EPO inhibited the activity of iNOS in peritoneal macrophages activated with endotoxin.

Our data suggest that rh‐EPO protects against SAO shock by inhibiting iNOS activity.

British Journal of Pharmacology (1999) 127, 482–488; doi:10.1038/sj.bjp.0702521
Effects of GABA on noradrenaline release and vasoconstriction induced by renal nerve stimulation in isolated perfused rat kidney
Authors: Fujimura Shinsei , Shimakage Hironori , Tanioka Hideki , Yoshida Makoto , Suzuki‐Kusaba Mizue , Hisa Hiroaki , Satoh Susumu
Published: January 2005

We examined effects of γ‐aminobutyric acid (GABA) on vasoconstriction and noradrenaline (NA) release induced by electrical renal nerve stimulation (RNS) in the isolated pump‐perfused rat kidney.

RNS (1 and 2 Hz for 2.5 min each, 0.5‐ms duration, supramaximal voltage) increased renal perfusion pressure (PP) and renal NA efflux. GABA (3, 10 and 100 μM) attenuated the RNS‐induced increases in PP by 10–40% (P<0.01) and NA efflux by 10–30% (P<0.01). GABA did not affect exogenous NA (40 and 60 nM)‐induced increases in PP.

The selective GABAB agonist baclofen (3, 10 and 100 μM) also attenuated the RNS‐induced increases in PP and NA efflux, whereas the RNS‐induced responses were relatively resistant to the selective GABAA agonist muscimol (3, 10 and 100 μM).

The selective GABAB antagonist 2‐hydroxysaclofen (50 μM), but not the selective GABAA antagonist bicuculline (50 μM), abolished the inhibitory effects of GABA (10 μM) on the RNS‐induced responses.

The selective α2‐adrenoceptor antagonist rauwolscine (10 nM) enhanced the RNS‐induced responses. GABA (3, 10 and 100 μM) potently attenuated the RNS‐induced increases in PP by 40–60% (P<0.01) and NA efflux by 20–50% (P<0.01) in the presence of rauwolscine.

Prazosin (10 and 30 nM) suppressed the RNS‐induced increases in PP by about 70–80%. Neither rauwolscine (10 nM) nor GABA (10 μM) suppressed the residual prazosin‐resistant PP response.

These results suggest that GABA suppresses sympathetic neurotransmitter release via presynaptic GABAB receptors, and thereby attenuates adrenergically induced vasoconstriction in the rat kidney.

British Journal of Pharmacology (1999) 127, 109–114; doi:10.1038/sj.bjp.0702524
Potentiation of cyclic AMP‐mediated vasorelaxation by phenylephrine in pulmonary arteries of the rat
Authors: Priest R M , Hucks D , Ward J P T
Published: January 2005

α1‐adrenoceptor agonists may potentiate relaxation to β‐adrenoceptor agonists, although the mechanisms are unclear. We compared relaxations induced by β‐adrenoceptor agonists and cyclic AMP‐dependent vasodilators in rat pulmonary arteries constricted with prostaglandin F2α (PGF2α) or the α1‐adrenoceptor agonist phenylephrine (PE). In addition, we examined whether differences were related to cyclic AMP‐ or nitric oxide (NO) and cyclic GMP‐dependent pathways.

Isoprenaline‐induced relaxation was substantially potentiated in arteries constricted with PE compared with PGF2α. Methoxamine was similar to PE, whereas there was no difference between PGF2α and 30 mM KCl. The potentiation was primarily due to a marked increase in the NO‐independent component of relaxation, from 9.1±1.7% for PGF2α to 55.1±4.4% for PE. NO‐dependent relaxation was also enhanced, but to a lesser extent (∼50%). Relaxation to salbutamol was almost entirely NO‐dependent in both groups, and was potentiated ∼50% by PE.

Relaxation to forskolin (activator of adenylate cyclase) was also enhanced in PE constricted arteries. Part of this relaxation was NO‐dependent, but the major effect of PE was to increase the NO‐independent component. Propranolol diminished but did not abolish the potentiation. There was no difference in response to CPT cyclic AMP (membrane permeant analogue) between PE and PGF2α, suggesting that mechanisms distal to the production of cyclic AMP were unchanged.

Relaxation to sodium nitroprusside (SNP) was the same for PE and PGF2α, although relaxation to acetylcholine (ACh) was slightly depressed. This implies that potentiation by PE does not involve the cyclic GMP pathway directly.

Mesenteric arteries constricted with PE did not show potentiation of isoprenaline‐induced relaxation compared to those constricted with PGF2α, suggesting that this effect may be specific to the pulmonary circulation.

These results clearly show that PE potentiates both the NO‐independent and ‐dependent components of cyclic AMP‐mediated relaxation in pulmonary arteries of the rat, although the effect on the former is more profound. We suggest that potentiation of both components is largely due to direct activation of adenylate cyclase via α1‐adrenoceptors, within the smooth muscle and endothelial cells respectively.

British Journal of Pharmacology (1999) 127, 291–299; doi:10.1038/sj.bjp.0702525
Nitric oxide‐dependent relaxation induced by M1 muscarinic receptor activation in the rat small intestine
Authors: Olgart C , Iversen H H
Published: January 2005

The aim of the present study was to investigate whether muscarinic M1 receptor activation induces intestinal relaxation via nerve‐dependent nitric oxide formation.

Mechanical activity in longitudinal segments of rat jejunum was recorded isotonically in organ baths.

The muscarinic M1 receptor agonist 4‐[[[(3‐Chlorophenyl)amino]carbonyl]oxy]‐N,N,N,‐trimethyl‐2‐butyn‐1‐amonium chloride (McN‐A‐343, 10−7–10−4 M) induced a concentration‐dependent relaxation of rat jejunum. Relaxations induced by McN‐A‐343 (10−5 M) were inhibited by the M1 receptor antagonist telenzepine (10−8 M), and enhanced by the M3 receptor antagonist para‐fluoro‐hexahydrosiladifenidol (p‐F‐HHSiD; 3×10−7 M).

The inhibitory responses induced by McN‐A‐343 were abolished by the nitric oxide synthase inhibitors Nω‐nitro‐L‐arginine (L‐NOARG; 10−4 M) and Nω‐monomethyl‐L‐arginine (L‐NMMA; 3×10−5 M), the guanylyl cyclase inhibitor 1H‐[1,2,4]oxadiazolo[4,3,‐a]quinoxalin‐1‐one (ODQ; 10−5 M), and by tetrodotoxin (TTX; 3×10−7 M).

Guanethidine or hexamethonium did not affect inhibitory responses induced by McN‐A‐343.

In conclusion, McN‐A‐343 induces nerve‐dependent, nitrergic relaxations in rat jejunum, via activation of muscarinic M1 receptors. Hence, selective muscarinic M1 receptor agonists or antagonists might offer possibilities for pharmacological manipulation of the NO system.

British Journal of Pharmacology (1999) 127, 309–313; doi:10.1038/sj.bjp.0702529
Pharmacological analysis of CCK2 receptor antagonists using isolated rat stomach ECL cells
Authors: Lindström Erik , Björkqvist Maria , Håkanson Rolf
Published: January 2005

Gastrin stimulates rat stomach ECL cells to secrete histamine and pacreastatin, a chromogranin A (CGA)‐derived peptide. The present report describes the effect of nine cholecystokinin2 (CCK2) receptor antagonists and one CCK1 receptor antagonist on the gastrin‐evoked secretion of pancreastatin from isolated ECL cells.

The CCK2 receptor antagonists comprised three benzodiazepine derivatives L‐740,093, YM022 and YF476, one ureidoacetamide compound RP73870, one benzimidazole compound JB 93182, one ureidoindoline compound AG041R and three tryptophan dipeptoids PD 134308 (CI988), PD135158 and PD 136450. The CCK1 receptor antagonist was devazepide.

A preparation of well‐functioning ECL cells (∼80% purity) was prepared from rat oxyntic mucosa using counter‐flow elutriation. The cells were cultured for 48 h in the presence of 0.1 nM gastrin; they were then washed and incubated with antagonist alone or with various concentrations of antagonist plus 10 nM gastrin (a maximally effective concentration) for 30 min. Gastrin dose‐response curves were constructed in the absence or presence of increasing concentrations of antagonist. The amount of pancreastatin secreted was determined by radioimmunoassay.

The gastrin‐evoked secretion of pancreastatin was inhibited in a dose‐dependent manner. YM022, AG041R and YF476 had IC50 values of 0.5, 2.2 and 2.7 nM respectively. L‐740,093, JB93182 and RP73870 had IC50 values of 7.8, 9.3 and 9.8 nM, while PD135158, PD136450 and PD134308 had IC50 values of 76, 135 and 145 nM. The CCK1 receptor antagonist devazepide was a poor CCK2 receptor antagonist with an IC50 of about 800 nM.

YM022, YF476 and AG041R were chosen for further analysis. YM022 and YF476 shifted the gastrin dose‐response curve to the right in a manner suggesting competitive antagonism, while the effects of AG041R could not be explained by simple competitive antagonism. pKB values were 11.3 for YM022, 10.8 for YF476 and the apparent pKB for AG041R was 10.4.

British Journal of Pharmacology (1999) 127, 530–536; doi:10.1038/sj.bjp.0702538
Characterization of the decrease of extracellular striatal dopamine induced by intrastriatal morphine administration
Authors: Piepponen T Petteri , Mikkola Janne A V , Ruotsalainen Minna , Jonker Daniël , Ahtee Liisa
Published: January 2005

The effect of intrastriatally‐administered morphine on striatal dopamine (DA) release was studied in freely moving rats. Morphine (1, 10 or 100 μM) was given into the striatum by reversed microdialysis, and concentrations of DA and its metabolites 3,4‐dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) were simultaneously measured from the striatal dialysates.

Intrastriatally‐administered morphine significantly and dose‐dependently decreased the extracellular concentration of DA, the concentrations of the acidic DA metabolites were only slightly decreased. The effect of morphine was antagonized by naltrexone (2.25 mg kg−1, s.c.). Pretreatment with a preferential κ‐opioid receptor antagonist, MR2266 [(−)‐5,9 alpha‐diethyl‐2‐(3‐furylmethyl)‐2′‐hydroxy‐6,7‐benzomorphane; 1 mg kg−1, s.c.], had no effect on the decrease of extracellular DA evoked by intrastriatal morphine (100 μM).

Intrastriatal administration of the selective μ‐opioid receptor agonist [D‐Ala2,MePhe4,Gly‐ol5] enkephalin (DAMGO; 1 μM), significantly decreased the extracellular concentration of DA in the striatum.

When the rats were given morphine repeatedly in increasing doses (10–25 mg kg−1, s.c.) twice daily for 7 days and withdrawn for 48 h, the decrease of extracellular DA induced by morphine (100 μM) was significantly less than that seen in saline‐treated controls.

Our results show that besides the well‐known stimulatory effect there is a local inhibitory component in the action of morphine on striatal DA release in the terminal regions of nigrostriatal DA neurones. Tolerance develops to this inhibitory effect during repeated morphine treatment. Furthermore, our results suggest that the effect of intrastriatally‐administered morphine is mediated by the μ‐opioid receptors.

British Journal of Pharmacology (1999) 127, 268–274; doi:10.1038/sj.bjp.0702542
Concentration‐dependent isoflurane effects on depolarization‐evoked glutamate and GABA outflows from mouse brain slices
Authors: Liachenko Serguei , Tang Pei , Somogyi George T , Xu Yan
Published: January 2005

The synaptic concentrations of glutamate and γ‐aminobutyric acid (GABA) are modulated by their release and re‐uptake. The effects of general anaesthetics on these two processes remain unclear. This study evaluates the effects of isoflurane, a clinically important anaesthetic, on glutamate and GABA release and re‐uptake in superfused mouse cerebrocortical slices.

Experiments consisted of two 1.5‐min exposures to 40 mM KCl in 30 min intervals. During the second exposure, different concentrations of isoflurane with and without 0.3 mM L‐trans‐pyrrolidine‐2,4‐dicarboxylic acid (PDC, a competitive inhibitor of glutamate uptake transporter) or 1 mM nipecotic acid (a competitive inhibitor of GABA uptake transporter) were introduced. The ratios of the second to first KCl‐evoked increases in glutamate and GABA were used to determine the isoflurane concentration‐response curves.

The results can be described as a sum of two independent processes, corresponding to the inhibitions of release and re‐uptake, respectively. The EC50 values for the inhibitions of release and re‐uptake were 295±16 and 805±43 μM for glutamate, and 229±13 and 520±25 μM for GABA, respectively. Addition of PDC did not significantly affect glutamate release but shifted the re‐uptake curve to the left (EC50=315±20 μM). Nipecotic acid completely blocked GABA uptake, rendering isoflurane inhibition of GABA re‐uptake undetectable.

Our data suggest that isoflurane inhibits both the release and re‐uptake of neurotransmitters and that the inhibitions occur at different EC50's. For GABA, both EC50's are within the clinical concentration range. The net anaesthetic effect on extracellular concentrations of neurotransmitters, particularly GABA, depends on the competition between inhibition of release and that of re‐uptake.

British Journal of Pharmacology (1999) 127, 131–138; doi:10.1038/sj.bjp.0702543
A study of NPY‐mediated contractions of the porcine isolated ear artery
Authors: Roberts R E , Kendall D A , Wilson V G
Published: January 2005

Enhanced contractions of the porcine isolated ear artery by the α2‐adrenoceptor agonist UK14304 are uncovered by pharmacological manipulation. As both neuropeptide Y (NPY) receptors and α2‐adrenoceptors are negatively‐coupled to adenylyl cyclase in this tissue, we determined whether NPY is also able to produce an enhanced contraction in the same tissue, under the same conditions.

NPY (0.1 μM) produced a small contraction of porcine isolated ear arteries which was 5.1±0.8% of the response to 60 mM KCl (n=14). An enhanced NPY response was uncovered if the tissue was pre‐contracted with 0.1 μM U46619, and relaxed back to baseline with 1–2 μM forskolin before the addition of NPY (49.8±5.3%, n=14).

Forskolin (1 μM) stimulated cyclic AMP accumulation in porcine ear artery segments in the presence of 0.1 μM U46619 and 1 mM isobutylmethylxanthine (IBMX), NPY (0.1 μM) inhibited this response by 40%, but had no effect on basal levels of cyclic AMP.

An enhanced response to 0.1 μM NPY was also obtained after pre‐contraction with 0.1 μM U46619 and relaxation with either SNP (28.9±5.7%, n=14), or dibutyryl cyclic AMP (21.2±4.6%, n=14). This indicates that at least part of the enhanced response to NPY is independent of the agonist's ability to inhibit adenylyl cyclase.

In conclusion, an enhanced contraction to NPY in the porcine isolated ear artery can be obtained by prior pharmacological manipulation. The enhanced responses are mediated through adenylyl cyclase‐dependent and independent pathways similar to those reported for α2‐adrenoceptors in this preparation.

British Journal of Pharmacology (1999) 127, 284–290; doi:10.1038/sj.bjp.0702544
Effect of the neuroprotective compound SR57746A on nerve growth factor synthesis in cultured astrocytes from neonatal rat cortex
Authors: Labie Christophe , Lafon Corinne , Marmouget Corinne , Saubusse Patricia , Fournier Jacqueline , Keane Peter E , Le Fur Gérard , Soubrié Philippe
Published: January 2005

The neurotrophic factor promoting activity of the neuroprotective compound SR57746A was evaluated in primary cultures of neonatal rat cortical astrocytes by studying the synthesis of nerve growth factor (NGF) and brain‐derived neurotrophic factor (BDNF).

A concentration‐ and time‐dependent increase of nerve growth factor mRNA was induced by SR57746A (10 nM–1 μM). In these astrocytes, BDNF mRNA contents were increased to a significant but smaller extent, and β‐actin mRNA showed no variation.

SR57746A (1 μM) induced increases of both de novo protein translation after 6 h of incubation and NGF release into the extracellular medium after 6–24 h.

These effects were preceded by a transient augmentation of junB, c‐fos and c‐jun mRNA contents. These increases of AP‐1 family mRNA were associated with increased nuclear AP‐1 binding activity.

The results show that SR57746A can increase the synthesis and release of NGF in rat cortical astrocytes. Such effects may contribute to the drug's previously described neuroprotective effects.

British Journal of Pharmacology (1999) 127, 139–144; doi:10.1038/sj.bjp.0702545