bluuebunny/crossref_metadata_2025 huggingface dataset:
Inhibition of cyclic GMP hydrolysis with zaprinast reduces basal and cyclic AMP‐elevated L‐type calcium current in guinea‐pig ventricular myocytes
Authors: Ziolo Mark T , Lewandowski Susanne J , Smith Jacquelyn M , Romano Fred D , Wahler Gordon M
Published: March 2003

Cyclic GMP (cGMP) has been shown to be an important modulator of cardiac contractile function. A major component of cGMP regulation of contractility is cGMP‐mediated inhibition of the cardiac calcium current (ICa). An under‐appreciated aspect of cyclic nucleotide signalling is hydrolysis of the cyclic nucleotide (i.e., breakdown by phosphodiesterases (PDEs)). The role of cGMP hydrolysis in regulating ICa has not been studied. Thus the purpose of this study was to investigate if inhibition of cGMP hydrolysis can modulate ICa in isolated guinea‐pig ventricular myocytes.

Zaprinast, a selective inhibitor of cGMP‐specific PDE (PDE5), caused a significant increase in cGMP levels in myocytes, but was without affect on basal or β‐adrenergic stimulated cAMP levels (consistent with its actions as a specific inhibitor of PDE5).

Zaprinast inhibited ICa that was pre‐stimulated with cAMP elevating agents (isoproterenol, a β‐adrenergic agonist; or forskolin, a direct activator of adenylate cyclase). The effect of zaprinast was greatly reduced by KT5823, an inhibitor of cGMP‐dependent protein kinase (PKG).

Zaprinast also significantly inhibited basal ICa when perforated‐patch or whole‐cell recording with physiological pipette calcium concentration (10−7 M) was used. However, this effect was not observed when using standard calcium‐free whole‐cell recording conditions.

These results indicate that inhibition of cGMP hydrolysis can decrease both basal and cAMP‐stimulated ICa. Thus, cGMP hydrolysis may likely be an important step for physiological modulation of ICa. This regulation may also be important in disease states in which cGMP production is increased and PDE5 expression is altered, such as heart failure.

British Journal of Pharmacology (2003) 138, 986–994. doi:10.1038/sj.bjp.0705112
RXR activators molecular signalling: involvement of a PPARα‐dependent pathway in the liver and kidney, evidence for an alternative pathway in the heart
Authors: Ouamrane Laïla , Larrieu Gilberte , Gauthier Béatrice , Pineau Thierry
Published: March 2003

In this study we compared the molecular signalling elicited by rexinoids, selective retinoid X receptor (RXR)‐activators, in several organs (i.e. liver, kidney, heart) and in hepatocytes of various species.

RXR plays the pivotal role of a hetero‐dimerization partner for the members of the class II subset of nuclear receptors which regulate the transcription of numerous target genes, following chemical activation. Several of these selective activators are currently used to treat hyperlipidaemia (fibrates), type II diabetes (glitazones), or skin disorders (retinoic acid). Although these therapeutic pathways are not fully elucidated, receptor activation is considered a pre‐requisite for efficacy. Therefore RXR, which accepts numerous dimeric partners, is considered a worthwhile pharmacological target.

We analysed a number of biochemical and molecular responses to rexinoids which were given orally to mice. Our results showed a prominent involvement of the peroxisome proliferator‐activated receptor (PPARα) as a majority of the observed hepatic and renal regulations were abolished in PPARα‐knockout animals. Therefore we documented the species‐specificity of these rexinoid actions which were reproduced in rat primary hepatocyte cultures but not in cultures of rabbit or human origin. Conversely, we established that the regulation of the pyruvate dehydrogenase kinase (PDK4) gene in the heart, by rexinoids, is independent of PPARα expression.

Our results support the obligatory expression of the active, although quiescent, PPARα to sustain a subset of relevant regulations attributable to rexinoids in the liver and kidney. Their cardiac molecular signalling unveiled an alternate transduction pathway and therefore opens new prospects in the therapeutic potential of rexinoids.

British Journal of Pharmacology (2003) 138, 845–854. doi:10.1038/sj.bjp.0705113
Cannabinoid receptor‐independent inhibition by cannabinoid agonists of the peripheral 5‐HT3 receptor‐mediated von Bezold–Jarisch reflex
Authors: Godlewski Grzegorz , Göthert Manfred , Malinowska Barbara
Published: March 2003

On the basis of previous findings that cannabinoids inhibit the function of human and rat 5‐HT3 receptors in vitro, we investigated whether cannabinoid receptor agonists also modulate the activity of the rat peripheral 5‐HT3 receptors on the terminals of cardiopulmonary afferent C‐fibres in vivo.

In urethane‐anaesthetized rats, pre‐treated intravenously (i.v.) with the CB1 receptor antagonist SR 141716A (3 μmol kg−1) and with the β1/β2 adrenoceptor antagonist propranolol (0.3–0.4 μmol kg−1), bolus injection of the serotonin 5‐HT3 receptor agonist phenylbiguanide (3–10 μg kg−1, i.v.) or the vanilloid VR1 receptor agonist capsaicin (3–10 μg kg−1, i.v.) caused an immediate decrease in heart rate and mean arterial blood pressure (the von Bezold–Jarisch reflex).

The phenylbiguanide‐induced bradycardia was dose‐dependently attenuated by the cannabinoid receptor agonists CP 55,940 (0.1–1 μmol kg−1, i.v.) and WIN 55,212‐2 (0.1–3 μmol kg−1, i.v.) 20 min after injection, but not by the inactive S‐(−)enantiomer of the latter, WIN 55,212‐3 (1 μmol kg−1, i.v.). The inhibition was reversible within 30 min. The extent of inhibition by the highest doses of cannabinoid receptor agonists amounted to about 50%. Both cannabinoid receptor agonists failed to affect the capsaicin‐evoked bradycardia.

In conclusion, our results demonstrate that cannabinoid receptor agonists modulate the von Bezold–Jarisch reflex by inhibiting peripheral serotonin 5‐HT3 receptors in rats in vivo. An analogous mechanism of cannabinoid receptor agonists may be assumed to be involved in other serotonin 5‐HT3 receptor‐mediated responses.

British Journal of Pharmacology (2003) 138, 767–774. doi:10.1038/sj.bjp.0705114
A photosensitive vascular smooth muscle store of nitric oxide in mouse aorta: no dependence on expression of endothelial nitric oxide synthase
Authors: Andrews Karen L , McGuire John J , Triggle Chris R
Published: March 2003

Photorelaxation is the reversible relaxation of vascular smooth muscle (VSM) when irradiated with ultraviolet (UV) light resulting from the release of nitric oxide (NO). In this study we characterize the involvement of endothelial nitric oxide synthase (eNOS) in the photorelaxation response of thoracic aorta from endothelial NOS deficient (−/−) and control (C57BL/6j) mice.

Cirazoline contracted aortae were repeatedly exposed to 30 s of UV light every 3–4 min. Equal levels of photorelaxation (45±2%; n=34) was observed in both strains.

1H‐[1,2,4]‐oxadiazolo[4,3‐a]quinoxalin‐1‐one (ODQ), K+, 2‐(4‐carboxyphenyl)‐4,4,5,5‐tetramethylimidazoline‐1‐oxyl‐3‐oxide (c‐PTIO), 4‐aminopyridine (4‐AP) and ethacrynic acid significantly reduced the photorelaxation response. In C57BL/6j mice diethyldithiocarbamate (DETCA) also reduced photorelaxation.

Control endothelium‐intact and ‐denuded aorta and L‐NAME (100 μM) treated and untreated eNOS (−/−) aortae were repeatedly exposed to UV light for 5 min every 10 min until no photorelaxation response was observed. After 1 h of rest in the dark the vessels showed between 30–70% recovery of the photorelaxation response indicating regeneration of the store in the absence of the endothelium and eNOS.

The results of this study suggest that photorelaxation in mouse aorta VSM results from the release of NO from a stable store of RSNOs, which activates soluble guanylate cyclase (sGC), leading to cGMP‐dependent relaxation that is partially mediated by an increase in KV channel activation and hyperpolarization. In addition, the eNOS isoform is not essential for the formation of the photorelaxation store and a non‐NOS source of NO may be involved in the maintenance of this store.

British Journal of Pharmacology (2003) 138, 932–940. doi:10.1038/sj.bjp.0705115
Functional coupling of the human dopamine D2 receptor with Gαi1, Gαi2, Gαi3 and Gαo G proteins: evidence for agonist regulation of G protein selectivity
Authors: Gazi Lucien , Nickolls Sarah A , Strange Philip G
Published: March 2003

The human dopamine D2long (D2L) receptor was expressed with four different G proteins in Sf9 cells using the baculovirus expression system. When co‐expressed with Gi/Go G proteins (Gi1α, Gi2α, Gi3α, or Goα, plus Gβ1 and Gγ2), the receptor displayed a high‐affinity binding site for the agonists (dopamine and NPA), which was sensitive to GTP (100 μM), demonstrating interaction between the receptor and the different G proteins.

The receptor to G protein ratio (R : G ratio) was evaluated using [3H]‐spiperone saturation binding (R) and [35S]‐GTPγS saturation binding (G). R : G ratios of 1 : 12, 1 : 3, 1 : 14 and 1 : 5 were found for Gi1, Gi2, Gi3, and Go preparations, respectively. However, when R : G ratios of 1 : 2 and 1 : 12 were compared for Gi2 and Go, no difference was found for the stimulation of [35S]‐GTPγS binding.

Several agonists were tested for their ability to stimulate [35S]‐GTPγS binding to membranes co‐expressing the receptor and various G proteins. All the compounds tested showed agonist activity in preparations expressing Gi3 and Go. However, for Gi2 and Gi1 preparations, compounds such as S‐(−)‐3‐PPP and p‐tyramine were unable to stimulate [35S]‐GTPγS binding.

Most of the compounds showed higher relative efficacies (compared to dopamine) and higher potencies in the preparation expressing Go. Comparison of the effects of different agonists in the different preparations showed that each agonist differentially activates the four G proteins.

We conclude that the degree of selectivity of G protein activation by the D2L receptor can depend on the conformation of the receptor stabilised by an agonist.

British Journal of Pharmacology (2003) 138, 775–786. doi:10.1038/sj.bjp.0705116
Anti‐amnesic effect of dimemorfan in mice
Authors: Wang Hui‐Hung , Chien Jyh‐Wei , Chou Yueh‐Ching , Liao Jyh‐Fei , Chen Chieh‐Fu
Published: March 2003

Dimemorfan, an antitussive for more than 25 years, has previously been reported to be a relative high‐affinity ligand at sigma‐1 (σ1) receptor with the Ki value of 151 nM.

To test whether dimemorfan has anti‐amnesic effects similar to a σ1 receptor agonist, this study examined its effects on scopolamine‐ and β‐amyloid peptide‐(25‐35)‐induced amnesia in mice.

Dimemorfan (10–40 mg kg−1, i.p.) administered 30 min before the training trial, immediately after the training trial, or 30 min before the retention test significantly improved scopolamine (1 mg kg−1, i.p.)‐ or β‐amyloid peptide‐(25‐35) (3 nmol mouse−1, i.c.v.)‐induced amnesia in a step‐through passive avoidance test. Dimemorfan (5–40 mg kg−1, i.p.) pretreatment also attenuated scopolamine (8 mg kg−1, i.p.)‐induced amnesia in a water‐maze test. And, these anti‐amnesic effects of dimemorfan, like the putative σ1 receptor agonist (+)‐N‐allylnormetazocine ((+)‐SKF‐10047), were antagonized by a σ receptor antagonist haloperidol (0.25 mg kg−1, i.p.).

These results indicated that dimemorfan has anti‐amnesic effects and acts like a σ1 receptor agonist.

British Journal of Pharmacology (2003) 138, 941–949. doi:10.1038/sj.bjp.0705117
Ocular hypotensive effects of melatonin receptor agonists in the rabbit: further evidence for an MT3 receptor
Authors: Pintor Jesus , Peláez Teresa , Hoyle Charles H V , Peral Assumpta
Published: March 2003

Melatonin is involved in the control of intraocular pressure during the night and day photoperiod. We have investigated the receptor that regulates intraocular pressure in New Zealand white rabbits by means of agonists and antagonists of melatonin receptors.

Melatonin and its analogues: 2‐Phe‐melatonin, 6‐Cl‐melatonin, 2‐I‐melatonin, 5‐ methoxycarbonylamino‐N‐acetyltryptamine (5‐MCA‐NAT) and N‐acetyltryptamine all produced a reduction in intraocular pressure. Dose‐response analysis for these compounds gave pD2 values of 9.3±0.24 for melatonin; 9.0±0.09 for 6‐Cl‐melatonin; 9.0±0.84 for 2‐I‐melatonin; 8.9±0.07 for 5‐MCA‐NAT; 8.7±0.18 for 2‐Phe‐melatonin and 9.4±0.30 for N‐acetyltryptamine (all n=8).

At a dose of 0.5 nmol (in 10 μl) melatonin and the selective melatonin MT3 agonist 5‐MCA‐NAT, induced greater reductions of intraocular pressure (22.8±2.3% and 32.5±1.4%, respectively) than the other compounds.

The melatonin‐receptor antagonists, prazosin, DH‐97 and 4‐P‐PDOT, reversed the effect of 5‐MCA‐NAT in a dose‐dependent manner, with pA2 values of 13.5±0.17 for prazosin, 10.6±0.16 for DH‐97 and 9.4±0.20 for 4‐P‐PDOT (n=8).

Cholinoceptor antagonists (hexamethonium and atropine) and α2‐ and β2‐adrenoceptor antagonists (yohimbine and ICI 118,551) partially reversed the effects produced by melatonin and 5‐MCA‐NAT, suggesting the possible involvement of cholinergic and noradrenergic systems in the hypotensive actions mediated by melatonin agonists. The α1‐adrenoceptor antagonist, corynanthine, had no significant effect.

The strong hypotensive effect of the MT3 agonist, 5‐MCA‐NAT, suggests that this compound may be a useful agent for treating those pathologies where intraocular pressure is abnormally elevated.

British Journal of Pharmacology (2003) 138, 831–836. doi:10.1038/sj.bjp.0705118
Pharmacological characterization of the homomeric and heteromeric UNC‐49 GABA receptors in C. elegans
Authors: Bamber Bruce A , Twyman Roy E , Jorgensen Erik M
Published: March 2003

UNC‐49B and UNC‐49C are γ‐aminobutyric acid (GABA) receptor subunits encoded by the Caenorhabditis elegans unc‐49 gene. UNC‐49B forms a homomeric GABA receptor, or can co‐assemble with UNC‐49C to form a heteromeric receptor. The pharmacological properties of UNC‐49B homomers and UNC‐49B/C heteromers were investigated in Xenopus oocytes.

The UNC‐49 subunits are most closely related to the bicuculline‐ and benzodiazepine‐insensitive RDL GABA receptors of insects. Consistent with this classification, bicuculline (10 μM) did not inhibit, nor did diazepam (10 μM) enhance UNC‐49B homomeric or UNC‐49B/C heteromeric receptors.

The UNC‐49C subunit strongly affects the pharmacology of UNC‐49B/C heteromeric receptors. UNC‐49B homomers were much more picrotoxin sensitive than UNC‐49B/C heteromers (IC50=0.9±0.2 μM and 166±42 μM, respectively). Pentobarbitone enhancement was greater for UNC‐49B homomers compared to UNC‐49B/C heteromers. Propofol (50 μM) slightly enhanced UNC‐49B homomers but slightly inhibited UNC‐49B/C heteromers. Penicillin G (10 mM) inhibited UNC‐49B homomers less strongly than UNC‐49B/C heteromers (30% compared to 53% inhibition, respectively).

Several aspects of UNC‐49 pharmacology were unusual. Picrotoxin sensitivity strongly correlates with dieldrin sensitivity, yet UNC‐49B homomers were highly dieldrin resistant. The enhancing neurosteroid pregnanolone (5β‐pregnan‐3α‐ol‐20‐one; 10 μM) strongly inhibited both UNC‐49 receptors. Alphaxalone (10 μM), another enhancing neurosteroid, did not affect UNC‐49B homomers, but slightly inhibited UNC‐49B/C heteromers.

UNC‐49 subunits and mammalian GABAA receptor α, β, and γ subunit classes all share roughly the same degree of sequence similarity. Thus, although they are most similar to other invertebrate GABA receptors, the UNC‐49 receptors share significant structural and pharmacological overlap with mammalian GABAA receptors.

British Journal of Pharmacology (2003) 138, 883–893. doi:10.1038/sj.bjp.0705119
Different domains of the glucagon and glucagon‐like peptide‐1 receptors provide the critical determinants of ligand selectivity
Authors: Runge S , Wulff B S , Madsen K , Bräuner‐Osborne H , Knudsen L B
Published: March 2003

Glucagon and glucagon‐like peptide‐1 (GLP‐1) are homologous peptide hormones with important functions in glucose metabolism. The receptors for glucagon and GLP‐1 are homologous family B G‐protein coupled receptors. The GLP‐1 receptor amino‐terminal extracellular domain is a major determinant of glucagon/GLP‐1 selectivity of the GLP‐1 receptor. However, the divergent residues in glucagon and GLP‐1 that determine specificity for the GLP‐1 receptor amino‐terminal extracellular domain are not known. Less is known about how the glucagon receptor distinguishes between glucagon and GLP‐1.

We analysed chimeric glucagon/GLP‐1 peptides for their ability to bind and activate the glucagon receptor, the GLP‐1 receptor and chimeric glucagon/GLP‐1 receptors. The chimeric peptide GLP‐1(7–20)/glucagon(15–29) was unable to bind and activate the glucagon receptor. Substituting the glucagon receptor core domain with the GLP‐1 receptor core domain (chimera A) completely rescued the affinity and potency of GLP‐1(7–20)/glucagon(15–29) without compromising the affinity and potency of glucagon. Substituting transmembrane segment 1 (TM1), TM6, TM7, the third extracellular loop and the intracellular carboxy‐terminus of chimera A with the corresponding glucagon receptor segments re‐established the ability to distinguish GLP‐1(7–20)/glucagon(15–29) from glucagon. Corroborant results were obtained with the opposite chimeric peptide glucagon(1–14)/GLP‐1(21–37).

The results suggest that the glucagon and GLP‐1 receptor amino‐terminal extracellular domains determine specificity for the divergent residues in the glucagon and GLP‐1 carboxy‐terminals respectively. The GLP‐1 receptor core domain is not a critical determinant of glucagon/GLP‐1 selectivity. Conversely, the glucagon receptor core domain contains two or more sub‐segments which strongly determine specificity for divergent residues in the glucagon amino‐terminus.

British Journal of Pharmacology (2003) 138, 787–794. doi:10.1038/sj.bjp.0705120
A potent tryptase inhibitor nafamostat mesilate dramatically suppressed pulmonary dysfunction induced in rats by a radiographic contrast medium
Authors: Sendo Toshiaki , Itoh Yoshinori , Goromaru Takeshi , Sumimura Tomoko , Saito Mami , Aki Keisei , Yano Takahisa , Oishi Ryozo
Published: March 2003

Intravenous injection of ioxaglate (4 g iodine kg−1), an iodinated radiographic contrast medium, caused a marked protein extravasation, pulmonary oedema and a decrease in the arterial partial oxygen pressure in rats.

All of these reactions to ioxaglate were reversed by the pretreatment with gabexate mesilate (10 and 50 mg kg−1, 5 min prior to injection) or nafamostat mesilate (3 and 10 mg kg−1), in which the inhibition was complete after injection of nafamostat mesilate (10 mg kg−1).

Both gabexate mesilate and nafamostat mesilate inhibited the activity of purified human lung tryptase, although the latter compound was far more potent than the former.

Ioxaglate enhanced the nafamostat‐sensitive protease activity in the extracellular fluid of rat peritoneal mast cell suspensions.

Tryptase enhanced the permeability of protein through the monolayer of cultured human pulmonary arterial endothelial cells. Ioxaglate, when applied in combination with rat peritoneal mast cells, also produced the endothelial barrier dysfunction. These effects of tryptase and ioxaglate were reversed by nafamostat mesilate.

Consistent with these findings, immunofluorescence morphological analysis revealed that tryptase or ioxaglate in combination with mast cells increased actin stress fibre formation while decreasing VE‐cadherin immunoreactivity. Both of these actions of tryptase and ioxaglate were reversed by nafamostat mesilate.

These findings suggest that tryptase liberated from mast cells plays a crucial role in the ioxaglate‐induced pulmonary dysfunction. In this respect, nafamostat mesilate may become a useful agent for the cure or prevention of severe adverse reactions to radiographic contrast media.

British Journal of Pharmacology (2003) 138, 959–967. doi:10.1038/sj.bjp.0705121
Commentary on the article presented by Murdoch et al.
Authors: Fillion Gilles
Published: March 2003

British Journal of Pharmacology (2003) 138, 731–731. doi:10.1038/sj.bjp.0705122
5‐HT moduline: an endogenous inhibitor of 5‐HT1B/1D‐mediated contraction in pulmonary arteries
Authors: Murdoch R , Morecroft I , MacLean M R
Published: March 2003

5‐HT moduline (5‐HTm) is tetrapeptide (Leu‐Ser‐Ala‐Leu) previously shown to act as a specific endogenous antagonist to central 5‐HT1B/1D receptors. Its effects were investigated in rat and rabbit pulmonary arteries (PAs).

In rabbit PAs, contractile responses to the 5‐HT1B/1D receptor agonist 5‐carboxamidotryptamine (5‐CT) were inhibited by 1 and 10 μM 5‐HTm in a non‐competitive fashion with the maximum contractile response (Emax, per cent of response to 50 mM KCl) being reduced from 65.6±7% (n=6) to 39.7±6.5% (n=6) and 25.2±7.9 (n=4), respectively. The ability of 5‐HTm to inhibit responses to 5‐CT was increased by the aminopeptidase inhibitor bestatin (10 μM).

In the rabbit PAs, the nitric oxide synthase inhibitor, Nω‐nitro‐L‐arginine methylester (L‐NAME) potentiated responses to 5‐CT (Emax: 106±22.5 (n=4)) and this response was also inhibited by 10 μM 5‐HTm (Emax: 38±13% (n=8)).

5‐HTm (10 μM) inhibited responses to 5‐CT in rat PAs, the Emax being reduced from 24.8±4.1% (n=7) to 15.5±3.7% (n=9). 5‐HTm induced relaxation of 5‐CT‐pre‐constricted rat PAs with a pIC50 of 9.0±0.6 (n=9).

In PAs from chronic hypoxic, pulmonary hypertensive rats, the maximum response to 5‐CT was increased to 80±8.5% (n=11). 5‐HTm reduced this response to 34.4±6.3% (n=12). L‐NAME markedly inhibited the ability of 5‐HTm to inhibit responses to 5‐CT (Emax before 5‐HTm: 100.5±16% (n=5), Emax after 5‐HTm: 107±11.3% (n=4)).

In conclusion we show here for the first time that 5‐HTm is a non‐competitive inhibitor of 5‐HT1B/1D receptor‐mediated constriction in PAs. In rat PAs, L‐NAME can inhibit this effect of 5‐HTm.

British Journal of Pharmacology (2003) 138, 795–800. doi:10.1038/sj.bjp.0705123
Interactions of atropine with heterologously expressed and native α3 subunit‐containing nicotinic acetylcholine receptors
Authors: Parker Julie C , Sarkar Deboshree , Quick Michael W , Lester Robin A J
Published: March 2003

Atropine, a classical muscarinic antagonist, has been reported previously to inhibit neuronal nicotinic acetylcholine receptors (nAChRs). In the present study, the action of atropine has been examined on α3β4 receptors expressed heterologously in Xenopus oocytes and native nAChRs in medial habenula neurons.

At concentrations of atropine often used to inhibit muscarinic receptors (1 μM), responses induced by near‐maximal nicotine concentrations (100 μM) at negative holding potentials (−65 mV) are inhibited (14–30%) in a reversible manner in both α4 and α3 subunit‐containing heteromeric nAChRs. Half‐maximal effective concentrations (IC50 values) for atropine inhibition are similar for the four classes of heteromeric receptors studied (4–13 μM).

For α3β4 nAChRs in oocytes, inhibition by atropine (10 μM) is not overcome at higher concentrations of agonist, and is increased with membrane hyperpolarization. These results are consistent with non‐competitive antagonism – possibly ion channel block.

At low concentrations of both nicotine (10 μM) and atropine (<10 μM), potentiation (∼25%) of α3β4 nAChR responses in oocytes is observed. The relative balance between potentiation and inhibition is dependent upon membrane potential.

In rat medial habenula (MHb) neurons, atropine (0.3–3.0 μM) inhibited nicotine‐induced responses in both a concentration and membrane potential‐dependent manner (at −40 mV, IC50=4 μM), similar to the effects on α3β4–nAChRs in oocytes. However, unlike heterologously expressed receptors, potentiation was barely detectable at depolarized membrane potentials using low concentrations of nicotine (3–10 μM). Conversely, the weak agonist, choline (1–3 mM) was observed to augment responses of MHb nAChRs.

British Journal of Pharmacology (2003) 138, 801–810. doi:10.1038/sj.bjp.0705124
Dopamine D2‐like receptor‐mediated opening of K+ channels in opossum kidney cells
Authors: Gomes Pedro , Soares‐da‐Silva Patrício
Published: March 2003

This study examined the effects of dopamine D1‐ and D2‐like receptor activation upon basolateral K+ (IK) currents and changes in membrane potential in opossum kidney (OK) cells.

The addition of amphotericin B (3 μg ml−1) to the apical side resulted in a rapid increase in IK, this effect being markedly inhibited by the addition of the K+ channel blockers barium chloride (1 mM) or glibenclamide (10 μM), but not apamin (1 μM). The K+ channel opener pinacidil increased the amphotericin B‐induced IK. The selective D2‐like receptor agonist quinerolane increased, in a concentration dependent manner (EC50=136 nM), IK across the basolateral membrane, this effect being abolished by pre‐treatment with pertussis toxin (PTX), S‐sulpiride (selective D2‐like receptor antagonist) and glibenclamide. The selective D1‐like receptor agonist SKF 38393 did not change IK. Both H‐89 (PKA inhibitor) and chelerythrine (PKC inhibitor) failed to prevent the stimulatory effect of quinerolane upon IK.

Quinerolane did not change basal levels of cyclic AMP and also failed to affect the forskolin‐induced increase in cyclic AMP levels.

The stimulation of D2‐like receptor was associated with a rapid hyperpolarizing effect, whereas D1‐like receptor activation was accompanied by increases in cell membrane potential. The hyperpolarizing effect of quinerolane (EC50=129 nM) was prevented by pre‐treatment with PTX, S‐sulpiride and glibenclamide.

It is concluded that stimulation of dopamine D2‐like, but not D1‐like, receptors coupled to PTX‐sensitive G proteins of the Gi/o class produce membrane hyperpolarization through opening of KATP channels.

British Journal of Pharmacology (2003) 138, 968–976. doi:10.1038/sj.bjp.0705125
Ajoene, a garlic compound, inhibits protein prenylation and arterial smooth muscle cell proliferation
Authors: Ferri Nicola , Yokoyama Kohei , Sadilek Martin , Paoletti Rodolfo , Apitz‐Castro Rafael , Gelb Michael H , Corsini Alberto
Published: March 2003

Ajoene is a garlic compound with anti‐platelet properties and, in addition, was shown to inhibit cholesterol biosynthesis by affecting 3‐hydroxy‐3‐methyl‐glutaryl coenzyme A (HMG‐CoA) reductase and late enzymatic steps of the mevalonate (MVA) pathway.

MVA constitutes the precursor not only of cholesterol, but also of a number of non‐sterol isoprenoids, such as farnesyl and geranylgeranyl groups. Covalent attachment of these MVA‐derived isoprenoid groups (prenylation) is a required function of several proteins that regulate cell proliferation. We investigated the effect of ajoene on rat aortic smooth muscle cell proliferation as related to protein prenylation.

Cell counting, DNA synthesis, and cell cycle analysis showed that ajoene (1–50 μM) interfered with the progression of the G1 phase of the cell cycle, and inhibited rat SMC proliferation.

Similar to the HMG‐CoA reductase inhibitor simvastatin, ajoene inhibited cholesterol biosynthesis. However, in contrast to simvastatin, the antiproliferative effect of ajoene was not prevented by the addition of MVA, farnesol (FOH), and geranylgeraniol (GGOH). Labelling of smooth muscle cell cellular proteins with [3H]‐FOH and [3H]‐GGOH was significantly inhibited by ajoene.

In vitro assays for protein farnesyltransferase (PFTase) and protein geranylgeranyltransferase type I (PGGTase‐I) confirmed that ajoene inhibits protein prenylation. High performance liquid chromatography (HPLC) and mass spectrometry analyses also demonstrated that ajoene causes a covalent modification of the cysteine SH group of a peptide substrate for protein PGGTase‐I.

Altogether, our results provide evidence that ajoene interferes with the protein prenylation reaction, an effect that may contribute to its inhibition of SMC proliferation.

British Journal of Pharmacology (2003) 138, 811–818. doi:10.1038/sj.bjp.0705126
Desensitization of α2A‐adrenoceptor signalling by modest levels of adrenaline is facilitated by β2‐adrenoceptor‐dependent GRK3 up‐regulation
Authors: Bawa Tasneem , Altememi Ghazi F , Eikenburg Douglas C , Standifer Kelly M
Published: March 2003

Adrenaline (ADR) and noradrenaline (NA) can simultaneously activate inhibitory α2‐ and stimulatory β‐adrenoceptors (AR). However, ADR and NA differ significantly in that ADR is a potent β2‐AR agonist while NA is not. Only recently has the interaction resulting from the simultaneous activation of α2‐ and β2‐AR been examined at the cellular level to determine the mechanisms of α2‐AR regulation following concomitant activation of both α2‐ and β2‐ARs by chronic ADR.

This study evaluates β2‐AR regulation of α2A‐AR signalling following chronic ADR (300 nM) and NA (1 and 30 μM) treatments of BE(2)‐C human neuroblastoma cells that natively express both β2‐ and α2A‐ARs.

Chronic (24 h) treatment with ADR (300 nM) desensitized the response to the α2A‐AR agonist, brimonidine, in BE(2)‐C cells. Addition of the β‐AR antagonist, propranolol, blocked the ADR‐induced α2A‐AR desensitization. Unlike ADR, chronic NA (1 μM) treatment had no effect on the α2A‐AR response. However if NA was increased to 30 μM for 24 h, α2A‐AR desensitization was observed; this desensitization was partially reversed by propranolol.

Chronic ADR (300 nM) treatment reduced α2A‐AR binding levels, contributing to the α2A‐AR desensitization. This decrease was prevented by addition of propranolol during ADR treatment. Chronic NA (30 μM), like ADR, treatment lowered specific binding, whereas 1 μM NA treatment was without effect.

Chronic ADR treatment produced a significant increase in GRK3 levels and this was blocked by propranolol or GRK2/3 antisense DNA treatment. This antisense DNA, common to both GRK2 and GRK3, also blocked chronic ADR‐induced α2A‐AR desensitization and down‐regulation.

Acute (1 h) ADR (300 nM) or NA treatment (1 μM) produced α2A‐AR desensitization. The desensitization produced by acute treatment was β‐AR independent, as it was not blocked by propranolol.

We conclude that chronic treatment with modest levels of ADR produces α2A‐AR desensitization by mechanisms that involve up‐regulation of GRK3 and down‐regulation of α2A‐AR levels through interactions with the β2‐AR.

British Journal of Pharmacology (2003) 138, 921–931. doi:10.1038/sj.bjp.0705127
S‐Nitrosocaptopril: in vitro characterization of pulmonary vascular effects in rats
Authors: Tsui Debbie Y Y , Gambino Agatha , Wanstall Janet C
Published: March 2003

On rat isolated pulmonary arteries, vasorelaxation by S‐nitrosocaptopril (SNOcap) was compared with S‐nitrosoglutathione (GSNO) and nitroprusside, and inhibition by SNOcap of contractions to angiotensin I was compared with the angiotensin converting enzyme (ACE) inhibitor, captopril.

SNOcap was equipotent as a vasorelaxant on main (i.d. 2–3 mm) and intralobar (i.d. 600 μm) pulmonary arteries (pIC50 values: 5.00 and 4.85, respectively). Vasorelaxant responses reached equilibrium rapidly (2–3 min).

Pulmonary vasorelaxant responses to SNOcap, like GSNO, were (i) partially inhibited by the soluble guanylate cyclase inhibitor, ODQ (1H‐(1,2,4) oxadiazolo(4,3‐a)‐quinoxalin‐1‐one; 3 μM) whereas responses to nitroprusside were abolished and (ii) potentiated by hydroxocobalamin (HCOB; NO· free radical scavenger; 100 μM) whereas responses to nitroprusside were inhibited.

The relative potencies for pulmonary vasorelaxation compared with inhibition of platelet aggregation were: SNOcap 7 : 1; GSNO 25 : 1; nitroprusside >2000 : 1.

SNOcap, like captopril, concentration‐dependently and time‐dependently increased the EC50 for angiotensin I but not angiotensin II. The dependence on incubation time was independent of the presence of tissue but differed for SNOcap and captopril. This difference reflected the slow dissociation of SNOcap and instability of captopril, and precluded a valid comparison of the potency of the two drugs. After prolonged incubation (5.6 h) SNOcap was more effective than captopril.

Thus, in pulmonary arteries SNOcap (i) possesses NO donor properties characteristic of S‐nitrosothiols but different from nitroprusside and (ii) inhibits ACE at least as effectively as captopril. These properties suggest that SNOcap could be valuable in the treatment of pulmonary hypertension.

British Journal of Pharmacology (2003) 138, 855–864. doi:10.1038/sj.bjp.0705128
Regulation of intracellular cyclic AMP in skeletal muscle cells involves the efflux of cyclic nucleotide to the extracellular compartment
Authors: Godinho Rosely Oliveira , Costa‐Jr Valter Luiz
Published: March 2003

This report analyses the intracellular and extracellular accumulation of cyclic AMP in primary rat skeletal muscle cultures, after direct and receptor‐dependent stimulation of adenylyl cyclase (AC).

Isoprenaline, calcitonin gene‐related peptide (CGRP) and forskolin induced a transient increase in the intracellular cyclic AMP that peaked 5 min after onset stimulation.

Under stimulation with isoprenaline or CGRP, the intracellular cyclic AMP initial rise was followed by an exponential decline, reaching 46 and 52% of peak levels in 10 min, respectively.

Conversely, the forskolin‐dependent accumulation of intracellular cyclic AMP decreased slowly and linearly, reaching 49% of the peak level in 30 min.

The loss of intracellular cyclic AMP from peak levels, induced by direct or receptor‐induced activation of AC, was followed by an increase in the extracellular cyclic AMP.

This effect was independent on PDEs, since it was obtained in the presence of 3‐isobutyl‐1‐methylxanthine (IBMX).

Besides, in isoprenaline treated cells, the beta‐adrenoceptor antagonist propranolol reduced both intra‐ and extracellular accumulation of cyclic AMP, whereas the organic anion transporter inhibitor probenecid reduced exclusively the extracellular accumulation.

Together our data show that direct or receptor‐dependent activation of skeletal muscle AC results in a transient increase in the intracellular cyclic AMP, despite the continuous presence of the stimulus. The temporal declining of intracellular cyclic AMP was not dependent on the cyclic AMP breakdown but associated to the efflux of cyclic nucleotide to the extracellular compartment, by an active transport since it was prevented by probenecid.

British Journal of Pharmacology (2003) 138, 995–1003. doi:10.1038/sj.bjp.0705130
The immunophilin‐ligands FK506 and V‐10,367 mediate neuroprotection by the heat shock response
Authors: Klettner Alexa , Herdegen Thomas
Published: March 2003

The macrolid FK506 is widely used in transplantation to suppress allograft rejection. FK506 and its derivatives are powerful neuroprotective molecules, but the underlying mechanisms remain to be resolved. We have previously shown that the FK506 mediated neuroprotection against oxygen radicals is independent of the inhibition of calcineurin but depends on de novo protein synthesis.

Here, we have shown that FK506 mediates protection against H2O2, UV‐light or thapsigargin in neuronal cell lines, but not in non‐neuronal cells such as R3T3 fibroblasts. We compared in detail the effect of FK506 on apoptotic features in PC12 cells after H2O2 with V‐10,367 which binds to FKBPs but does not inhibit calcineurin. Both molecules exert the same neuroprotective effect after H2O2 stimulation. FK506, but not V‐10,367, inhibited the cytochrome c release out of the mitochondria and the caspase 3 activation, while both molecules inhibited the cleavage of Poly‐(ADP‐ribose)‐polymerase (Parp) and prevented the expression of p53.

FK506 and V‐10,367 rapidly induced the expression of Hsp70 and Hsp27, but not Hsp90. Their neuroprotective actions could be completely blocked by quercetin, a functional inhibitor of the heat shock proteins.

We conclude that immunophilin‐ligands such as FK506 and V‐10,367 exert their neuroprotection independent of calcineurin through the induction of the heat shock response. The identification of the underlying signal transduction from application of immunophilin ligands to the expression of heat shock proteins represents a novel target cascade for neuroprotection.

British Journal of Pharmacology (2003) 138, 1004–1012. doi:10.1038/sj.bjp.0705132
The vasodilatory action of testosterone: a potassium‐channel opening or a calcium antagonistic action?
Authors: Jones Richard D , Pugh Peter J , Jones T Hugh , Channer Kevin S
Published: March 2003

British Journal of Pharmacology (2003) 138, 733–744. doi:10.1038/sj.bjp.0705141
Expression and function of P2X purinoceptors in rat histaminergic neurons
Authors: Vorobjev Vladimir S , Sharonova Irina N , Haas Helmut L , Sergeeva Olga A
Published: March 2003

The pharmacology of ATP responses and the expression pattern of seven known subunits of the P2X receptor were investigated in individual histaminergic neurons of the tuberomamillary nucleus (TM).

ATP (3–1000 μM) evoked fast non‐desensitizing inward currents in TM neurons. 2‐methylthioATP (2MeSATP) displayed the same efficacy but a lower potency, EC50s 84 μM versus 48 μM, when compared with ATP. Adenosine‐diphosphate (ADP), uridine‐triphosphate (UTP) and alpha beta methylene‐ATP (αβ‐meATP) were inactive.

ATP‐mediated whole cell currents were potentiated by acidification of the recording solution (pH 7.5 and 6.6 were compared).

Single‐cell RT–PCR (scRT–PCR) analysis revealed that the P2X2 receptor is expressed in all PCR‐positive neurons. Each of the P2X1, P2X3, P2X4, P2X5 and P2X6 mRNAs were detected in less than 35% of the cells.

Suramin antagonized ATP responses with an IC50 of 4.2 μM and pyridoxalphosphate‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS, 1 μM) reduced ATP responses to 43% of control, when antagonists were pre‐applied 90s before the agonist. Cibacron blue (3 μM) given together with ATP potentiated control responses by 67%, but inhibited it to 10% after pre‐application.

2′,3′‐O‐(2,4,6‐Trinitrophenyl) adenosine 5′‐triphosphate (TNP–ATP) antagonized ATP responses with an IC50 of 7 μM.

Pharmacological properties of ATP responses together with scRT–PCR data suggest that P2X2 is the major purinoceptor on the soma of TM neurons, however the presence of heteromeric P2X2/5 receptors in some neurons cannot be excluded.

British Journal of Pharmacology (2003) 138, 1013–1019. doi:10.1038/sj.bjp.0705144
Capillary Haemangiomas of the Spinal Cord: Report of Two Cases
Authors: Bozkus H. , Tanriverdi T. , Kizilkiliç O. , Türeci E. , Öz B. , Hancı M.
Published: March 2003

The clinicopathological features of two cases of capillary haemangioma of the spinal cord are described. The presenting symptoms were similar to those of common intradural, intramedullary tumours or tumour-like mass lesions. Radiological features of these two lesions resembled other vascular lesions of the spinal cord. The patients underwent surgery, and recoveries were good. Histologically, the lesions resembled capillary haemangioma of superficial tissues that are composed of lobules of small capillaries with feeding vessels. A fibrous capsule enveloped the lesions. Capillary haemangiomas of the central nervous system are rare. Awareness of their existence may help the surgeon to avoid misdiagnosis and overtreatment of these benign tumour-like lesions.
One-dimensional and zero-dimensional electron systems on liquid helium (Review)
Authors: Kovdrya Yu. Z.
Published: March 2003

A review of the experimental and theoretical research on one-dimensional and zero-dimensional electron systems localized near a liquid helium surface is given. The properties of the electronic states on a flat liquid helium surface, including the surface of thin helium layers, are briefly considered. Ways of realizing one-dimensional and zero-dimensional electron systems and the results of experimental and theoretical studies of their properties are discussed. Experiments on the investigation of localization processes in quasi-one-dimensional electron systems are described. Collective effects in systems of this kind are considered, and the possibilities for the use of low-dimensional electron systems on the surface of liquid helium for creating electronic devices and quantum computers are explored.
Elastic properties and phonon spectra of quasi-two-dimensional VSe2
Authors: Gospodarev I. A. , Eremenko A. V. , Ignatova T. V. , Kamarchuk G. V. , Kolobov I. G. , Minaev P. A. , Syrkin E. S. , Feodosyev S. B. , Fil’ V. D. , Soreau-Leblanc A. , Molinie P. , Faulques E. C.
Published: March 2003

A recently developed method is used to make acoustic measurements of VSe2 single crystals. The components of the elastic constant tensor are determined from the data of these measurements. The experimental data are used to calculate the total and partial phonon densities of VSe2, which permit an explanation of the vibrational spectra of this compound.
Ferroelectric nematic suspension
Authors: Reznikov Yurii , Buchnev Olexander , Tereshchenko Olexander , Reshetnyak Victor , Glushchenko Anatoliy , West John
Published: March 2003

We report on the development of a dilute suspension of ferroelectric particles in a nematic liquid-crystal (LC) host. We found that the submicron particles do not disturb the LC alignment and the suspension macroscopically appears similar to a pure LC with no readily apparent evidence of dissolved particles. The suspension possesses enhanced dielectric anisotropy, and is sensitive to the sign of an applied electric field.
A neurotoxic peripherin splice variant in a mouse model of ALS
Authors: Robertson Janice , Doroudchi Mohammad M. , Nguyen Minh Dang , Durham Heather D. , Strong Michael J. , Shaw Gerry , Julien Jean-Pierre , Mushynski Walter E.
Published: March 2003

Peripherin, a neuronal intermediate filament (nIF) protein found associated with pathological aggregates in motor neurons of patients with amyotrophic lateral sclerosis (ALS) and of transgenic mice overexpressing mutant superoxide dismutase-1 (SOD1G37R), induces the selective degeneration of motor neurons when overexpressed in transgenic mice. Mouse peripherin is unique compared with other nIF proteins in that three peripherin isoforms are generated by alternative splicing. Here, the properties of the peripherin splice variants Per 58, Per 56, and Per 61 have been investigated in transfected cell lines, in primary motor neurons, and in transgenic mice overexpressing peripherin or overexpressing SOD1G37R. Of the three isoforms, Per 61 proved to be distinctly neurotoxic, being assembly incompetent and inducing degeneration of motor neurons in culture. Using isoform-specific antibodies, Per 61 expression was detected in motor neurons of SOD1G37R transgenic mice but not of control or peripherin transgenic mice. The Per 61 antibody also selectively labeled motor neurons and axonal spheroids in two cases of familial ALS and immunoprecipitated a higher molecular mass peripherin species from disease tissue. This evidence suggests that expression of neurotoxic splice variants of peripherin may contribute to the neurodegenerative mechanism in ALS.
Formation of filopodia-like bundles in vitro from a dendritic network
Authors: Vignjevic Danijela , Yarar Defne , Welch Matthew D. , Peloquin John , Svitkina Tatyana , Borisy Gary G.
Published: March 2003

We report the development and characterization of an in vitro system for the formation of filopodia-like bundles. Beads coated with actin-related protein 2/3 (Arp2/3)–activating proteins can induce two distinct types of actin organization in cytoplasmic extracts: (1) comet tails or clouds displaying a dendritic array of actin filaments and (2) stars with filament bundles radiating from the bead. Actin filaments in these bundles, like those in filopodia, are long, unbranched, aligned, uniformly polar, and grow at the barbed end. Like filopodia, star bundles are enriched in fascin and lack Arp2/3 complex and capping protein. Transition from dendritic to bundled organization was induced by depletion of capping protein, and add-back of this protein restored the dendritic mode. Depletion experiments demonstrated that star formation is dependent on Arp2/3 complex. This poses the paradox of how Arp2/3 complex can be involved in the formation of both branched (lamellipodia-like) and unbranched (filopodia-like) actin structures. Using purified proteins, we showed that a small number of components are sufficient for the assembly of filopodia-like bundles: Wiskott-Aldrich syndrome protein (WASP)–coated beads, actin, Arp2/3 complex, and fascin. We propose a model for filopodial formation in which actin filaments of a preexisting dendritic network are elongated by inhibition of capping and subsequently cross-linked into bundles by fascin.
Association of diacylglycerol kinase ζ with protein kinase C α
Authors: Luo Bai , Prescott Stephen M. , Topham Matthew K.
Published: March 2003

Activation of PKC depends on the availability of DAG, a signaling lipid that is tightly and dynamically regulated. DAG kinase (DGK) terminates DAG signaling by converting it to phosphatidic acid. Here, we demonstrate that DGKζ inhibits PKCα activity and that DGK activity is required for this inhibition. We also show that DGKζ directly interacts with PKCα in a signaling complex and that the binding site in DGKζ is located within the catalytic domain. Because PKCα can phosphorylate the myristoylated alanine-rich C-kinase substrate (MARCKS) motif of DGKζ, we tested whether this modification could affect their interaction. Phosphorylation of this motif significantly attenuated coimmunoprecipitation of DGKζ and PKCα and abolished their colocalization in cells, indicating that it negatively regulates binding. Expression of a phosphorylation-mimicking DGKζ mutant that was unable to bind PKCα did not inhibit PKCα activity. Together, our results suggest that DGKζ spatially regulates PKCα activity by attenuating local accumulation of signaling DAG. This regulation is impaired by PKCα-mediated DGKζ phosphorylation.
Drosophila paramyosin is important for myoblast fusion and essential for myofibril formation
Authors: Liu Hongjun , Mardahl-Dumesnil Michelle , Sweeney Sean T. , O'Kane Cahir J. , Bernstein Sanford I.
Published: March 2003

Paramyosin is a major structural protein of thick filaments in invertebrate muscles. Coiled-coil dimers of paramyosin form a paracrystalline core of these filaments, and the motor protein myosin is arranged on the core surface. To investigate the function of paramyosin in myofibril assembly and muscle contraction, we functionally disrupted the Drosophila melanogaster paramyosin gene by mobilizing a P element located in its promoter region. Homozygous paramyosin mutants die at the late embryo stage. Mutants display defects in both myoblast fusion and in myofibril assembly in embryonic body wall muscles. Mutant embryos have an abnormal body wall muscle fiber pattern arising from defects in myoblast fusion. In addition, sarcomeric units do not assemble properly and muscle contractility is impaired. We confirmed that these defects are paramyosin-specific by rescuing the homozygous paramyosin mutant to adulthood with a paramyosin transgene. Antibody analysis of normal embryos demonstrated that paramyosin accumulates as a cytoplasmic protein in early embryo development before assembling into thick filaments. We conclude that paramyosin plays an unexpected role in myoblast fusion and is important for myofibril assembly and muscle contraction.