bluuebunny/crossref_metadata_2025 huggingface dataset:
Pharmacological characterization of the nociceptin receptor mediating hyperalgesia in the mouse tail withdrawal assay
Authors: Calò G , Rizzi A , Marzola G , Guerrini R , Salvadori S , Beani L , Regoli D , Bianchi C
Published: January 2005

The newly discovered neuropeptide nociceptin (NC) has recently been reported to be the endogenous ligand of the opioid‐like orphan receptor. Despite its structural similarity to opioids, when injected intracerebroventricularly (i.c.v.) in the mouse, NC exerts a direct hyperalgesic effect and reverses opioid‐induced analgesia. In the present investigation, these two effects of NC were evaluated under the same experimental conditions; in addition, a pharmacological characterization of the receptor mediating these central effects of NC was attempted.

NC caused a dose dependent (0.1–10 nmol/mouse), naloxone‐insensitive reduction of tail withdrawal latency with a maximal effect of about 50% of the reaction time observed in saline injected mice. In the same range of doses, NC inhibited morphine (1 nmol/mouse) induced analgesia.

The effects of the natural peptide were mimicked by NCNH2 and NC(1–13)NH2 (all tested at 1 nmol/mouse) while 1 nmol NC(1–9)NH2 was found to be inactive either in reducing tail withdrawal latency or in preventing morphine analgesia.

[Phe1ψ(CH2‐NH)Gly2]NC(1–13)NH2 ([F/G]NC(1–13)NH2), which has been shown to antagonize NC effects in the mouse vas deferens, acted as an agonist, mimicking NC effects in both the experimental paradigms. In addition, when NC and [F/G]NC(1–13)NH2 were given together, their effects were additive.

These results demonstrate that both the direct hyperalgesic action and the anti‐morphine effect of NC can be studied under the same experimental conditions in the mouse tail withdrawal assay. Moreover, the pharmacological characterization of the NC functional site responsible for these actions compared with the peripherally active site, indicates the existence of important differences between peripheral and central NC receptors.

British Journal of Pharmacology (1998) 125, 373–378; doi:10.1038/sj.bjp.0702087
Cyclic GMP‐dependent and cyclic GMP‐independent actions of nitric oxide on the renal afferent arteriole
Authors: Trottier Greg , Triggle Chris R , O'Neill Sean K , Loutzenhiser Rodger
Published: January 2005

The effects of exogenous NO and endothelial‐derived NO (EDNO) on the afferent arteriole were investigated in the in vitro perfused hydronephrotic rat kidney. Vessels were pre‐constricted with angiotensin II (0.1–0.3 nM) or KCl (30 mM). NO was infused directly into the renal artery at concentrations ranging from 30–9000 nM. ODQ (10, 30 μM) was administered to examine the effects of guanylyl cyclase inhibition. Kidneys were treated with ibuprofen (10 μM) to avoid actions of prostaglandins.

During angiotensin II‐induced vasoconstriction, NO elicited vasodilation at concentrations of 30–900 nM (EC50=200 nM) and ODQ caused a 10 fold shift in NO‐sensitivity (EC50 1600 nM). During KCl‐induced vasoconstriction, NO elicited a maximal dilation of 82±9% at 9000 nM (EC50 2000 nM) and ODQ had no effect. Thus in the presence of ODQ, the NO concentration‐response curves for KCl‐ and angiotensin II‐induced vasoconstriction were identical (P>0.2).

To assess the possible role of cyclic GMP‐independent mechanisms in the actions of EDNO, we compared the effects of L‐NAME, ODQ and ODQ+L‐NAME on acetylcholine‐induced vasodilation. Angiotensin II reduced afferent arteriolar diameters from 16.7±0.5 to 8.1±0.8 microns and acetylcholine fully reversed this effect (16.9±0.5 microns). ODQ restored the angiotensin II response in the presence of acetylcholine (7.1±0.6 microns) and the subsequent addition of L‐NAME had no further effect (6.8±0.7 microns). Similarly, L‐NAME alone, fully reversed the actions of acetylcholine.

Our findings indicate that exogenous NO is capable of eliciting renal afferent arteriolar vasodilation through both cyclic GMP‐dependent and cyclic GMP‐independent mechanisms. The cyclic GMP‐independent action of NO did not require K+ channel activation, as it could be elicited in the presence of 30 mM KCl. Finally, although cyclic GMP‐independent effects of exogenous NO could be demonstrated in our model, EDNO appears to act exclusively through cyclic GMP.

British Journal of Pharmacology (1998) 125, 563–569; doi:10.1038/sj.bjp.0702090
NK‐3 receptors mediate enhancement of substance P release from capsaicin‐sensitive spinal cord afferent terminals
Authors: Schmid Giovanna , Carità Francesca , Bonanno Giambattista , Raiteri Maurizio
Published: January 2005

The effects of NK‐3 receptor agonists on the release of substance P‐immunoreactivity (SP‐LI) have been investigated using superfused rat spinal cord synaptosomes.

The Ca2+‐dependent overflow of SP‐LI evoked by 35 mM KCl was concentration‐dependently enhanced by senktide (EC50=52 nM; maximal effect=70%) or [MePhe7]NKB (EC50=5.5 nM; maximal effect=125%), both selective agonists at receptors of the NK‐3 type.

The potentiation of the SP‐LI overflow elicited by 100 nM senktide or [MePhe7]NKB was prevented by the NK‐3 receptor antagonist (+)‐SR142801. The antagonist halved, at 10 nM, and almost abolished, at 100 nM, the effect of both agonists. The effect of senktide or [MePhe7]NKB was insensitive to antagonists at NK‐1 or NK‐2 receptors.

Capsaicin (0.1–1 μM) stimulated SP‐LI release in a concentration‐dependent manner from spinal cord synaptosomes. The SP‐LI overflow elicited by 1 μM capsaicin was completely dependent on external Ca2+. Senktide could not affect the capsaicin‐evoked release of SP‐LI.

Senktide failed to potentiate the K+‐evoked overflow of SP‐LI from synaptosomes previously exposed for 15 min in superfusion to capsaicin.

The results show that release‐enhancing NK‐3 receptors are located on axon terminals of capsaicin‐sensitive primary afferent neurones in the spinal cord. Antagonists at NK‐3 receptors might help controlling pain transmission.

British Journal of Pharmacology (1998) 125, 621–626; doi:10.1038/sj.bjp.0702093
Analysis of the 5‐HT1Areceptor involvement in passive avoidance in the rat
Authors: Misane Ilga , Johansson Christina , Ove Ögren Sven
Published: January 2005

The effects of the 5‐HT2A/2Cagonist DOB, the selective 5‐HT1Aagonist NDO 008 (3‐dipropylamino‐5‐hydroxychroman), and the two enantiomers of the selective 5‐HT1Aagonist 8‐OH‐DPAT (R(+)‐8‐OH‐DPAT and S(−)‐8‐OH‐DPAT) were studied in a step‐through passive avoidance (PA) test in the male rat.

The 5‐HT1Aagonists injected prior to training (conditioning) produced a dose‐dependent impairment of PA retention when examined 24 h later. R(+)‐8‐OH‐DPAT was four times more effective than S(−)‐8‐OH‐DPAT to cause an impairment of PA retention. Both NDO 008 and the two enantiomers of 8‐OH‐DPAT induced the serotonin syndrome at the dose range that produced inhibition of the PA response, thus, indicating activation of postsynaptic 5‐HT1Areceptors.

Neither NDO 008 nor R(+)‐8‐OH‐DPAT induced head‐twitches, a behavioural response attributed to stimulation of postsynaptic 5‐HT2Areceptors. In contrast, DOB induced head‐twitches at the 0.01 mg kg−1dose while a 200 times higher dose was required to produce a significant impairment of PA retention.

The impairment of PA retention induced by both NDO 008 and R(+)‐8‐OH‐DPAT was fully blocked by the active S(+)‐ enantiomer of the selective 5‐HT1Aantagonist WAY 100135 and the mixed 5‐HT1A/β‐adrenoceptor antagonist L(−)‐alprenolol. In contrast, the mixed 5‐HT2A/2Cantagonists ketanserin and pirenperone were found to be ineffective. Moreover, the β2‐adrenoceptor antagonist ICI 118551, the β1‐antagonist metoprolol as well as the mixed β‐adrenoceptor blocker D(+)‐alprenolol all failed to modify the deficit of PA retention by NDO 008 and R(+)‐8‐OH‐DPAT. None of the 5‐HT1Aor 5‐HT2A/2Creceptor antagonists tested or the β‐blockers altered PA retention by themselves.

A 3 day pretreatment procedure (200+100+100 mg kg−1) with the tryptophan hydroxylase inhibitor p‐chlorophenylalanine (PCPA) did not alter PA retention and did not prevent the inhibitory action of the 5‐HT1Aagonists, indicating that their effects on PA do not depend on endogenous 5‐HT.

The effects of NDO 008 on PA were also studied using a state‐dependent learning paradigm. NDO 008 was found to produce a disruption of PA when given either prior to training or retention or both prior to training and retention but it failed to affect PA retention when given immediately after training.

These findings indicate that the deficit of passive avoidance retention induced by the 5‐HT1Aagonists is mainly a result of stimulation of postsynaptic 5‐HT1Areceptors but not 5‐HT2Areceptors. The 5‐HT1Areceptor stimulation appears to interfere with learning processes operating at both acquisition and retrieval.

British Journal of Pharmacology(1998)125, 499–509; doi:10.1038/sj.bjp.0702098
Inhibitors of spasmogen‐induced Ca2+ channel suppression in smooth muscle cells from small intestine
Authors: Unno T , Beech D J , Komori S , Ohashi H
Published: January 2005

Whole‐cell patch‐clamp recordings were made from smooth muscle cells isolated from the longitudinal muscle layer of guinea‐pig ileum. Carbachol (acting at muscarinic receptors) or histamine (acting at H1 histamine receptors) suppressed Ca2+ channel current. The effect of either agonist had an initial transient component followed by a sustained component.

Wortmannin inhibited transient and sustained components of carbachol‐induced Ca2+ channel current suppression: half‐effective inhibitory concentrations (IC50) were 1.1 μM and 0.6 μM for the two components respectively. Wortmannin also inhibited the transient phase of carbachol‐induced cationic current (IC50 1.6 μM) and Ca2+‐dependent K+‐current (IC50 1.7 μM). Wortmannin did not appear to produce any direct block of cationic channels or Ca2+ channels.

Intracellular application of the phospholipase inhibitor D609 (tricyclodecan‐9‐ylxanthogenate) inhibited transient and sustained components of histamine action on the Ca2+ channel current: the IC50 was about 130 μM for both components. Carbachol action on Ca2+ channels was also inhibited by D609. D609 had no significant direct blocking effect on Ca2+ channels, cationic channels activated by carbachol, or Ca2+‐activated K+‐current in response to flash‐photolysis of caged‐inositol 1,4,5‐trisphosphate.

Micromolar concentrations of wortmannin and D609 are inhibitors of both components of spasmogen‐induced Ca2+ channel suppression. The data suggest that both components are mediated by a common, or similar, signal transduction element which is a phospholipase C (PLC) or phospholipase D (PLD) isoform.

British Journal of Pharmacology (1998) 125, 667–674; doi:10.1038/sj.bjp.0702112
Effects of mexiletine on ATP sensitive K+ channel of rat skeletal muscle fibres: a state dependent mechanism of action
Authors: Tricarico Domenico , Barbieri Mariagrazia , Franchini Carlo , Tortorella Vincenzo , Conte Camerino Diana
Published: January 2005

The effects of mexiletine were evaluated on the ATP‐sensitive K+ channel (KATP) of rat skeletal muscle fibres using patch clamp techniques. The effects of mexiletine were studied on macropatch currents 20 s (maximally activated), 8 min (early stage of rundown) and 15 min (late stage of rundown) after excision in the absence or in the presence of internal ADP (50–100 μM) or UDP (500 μM). In addition, the effects of mexiletine were tested on single channel.

In the absence of ADP and UDP, mexiletine inhibited the current through maximally activated channels with an IC50 of −5.58±0.3 M. Nucleoside diphosphates shifted the current versus mexiletine concentration relationship to the right on the log concentration axis. UDP (500 μM) was more efficacious than ADP (50–100 μM) in this effect.

At the early stage of rundown, the sensitivity of the channel to mexiletine was reduced and nucleoside diphosphates, particularly UDP, antagonized the effect of mexiletine. At the late stage of rundown, mexiletine did not affect the currents.

At the single channel level, 1 μM mexiletine reduced the mean burst duration by 63% and prolonged the arithmetic mean closed time intervals between the bursts of openings without altering the open time and closed time distributions. Mexiletine did not affect the single channel conductance.

These results show that in skeletal muscle, mexiletine is a state‐dependent KATP channel inhibitor which either acts through the nucleotide binding site or a site allosterically coupled to it.

British Journal of Pharmacology (1998) 125, 858–864; doi:10.1038/sj.bjp.0702117
iNOS contribution to the NMDA‐induced excitotoxic lesion in the rat striatum
Authors: Lecanu L , Verrecchia C , Margaill I , Boulu R G , Plotkine M
Published: January 2005

The aim of this study was to assess whether an excitotoxic insult induced by NMDA may induce an iNOS activity which contributes to the lesion in the rat striatum.

For this purpose, rats were perfused with 10 mM NMDA through a microdialysis probe implanted in the left striatum. Microdialysate nitrite content, striatal Ca‐independent nitric oxide synthase activity and lesion volume were measured 48 h after NMDA exposure in rats treated with dexamethasone (DXM) (3 mg kg−1×4) or aminoguanidine (AG) (100 mg kg−1×4).

A significant increase in microdialysate nitrite content and in the Ca‐independent NOS activity was observed 48 h after NMDA infusion. Both these increases were reduced by DXM and AG. The NMDA‐induced striatal lesion was also reduced by both treatments.

Our results demonstrate that NMDA excitotoxic injury induces a delayed, sustained activation of a Ca‐independent NOS activity. This activity is blocked by DXM and AG, strongly suggesting the involvement of iNOS. The fact that AG and DXM reduce the NMDA‐elicited lesion suggests that iNOS contributes to the brain damage induced by excitotoxic insult.

British Journal of Pharmacology (1998) 125, 584–590; doi:10.1038/sj.bjp.0702119
Effects of propafenone and 5‐hydroxy‐propafenone on hKv1.5 channels
Authors: Franqueza Laura , Valenzuela Carmen , Delpón Eva , Longobardo Mónica , Caballero Ricardo , Tamargo Juan
Published: January 2005

The goal of this study was to analyse the effects of propafenone and its major metabolite, 5‐hydroxy‐propafenone, on a human cardiac K+ channel (hKv1.5) stably expressed in Ltk− cells and using the whole‐cell configuration of the patch‐clamp technique.

Propafenone and 5‐hydroxy‐propafenone inhibited in a concentration‐dependent manner the hKv1.5 current with KD values of 4.4±0.3 μM and 9.2±1.6 μM, respectively.

Block induced by both drugs was voltage‐dependent consistent with a value of electrical distance (referenced to the cytoplasmic side) of 0.17±0.55 (n=10) and 0.16±0.81 (n=16).

The apparent association (k) and dissociation (l) rate constants for propafenone were (8.9±0.9)×106 M−1 s−1 and 39.5±4.2 s−1, respectively. For 5‐hydroxy‐propafenone these values averaged (2.3±0.3)×106 M−1 s−1 and 21.4±3.1 s−1, respectively.

Both drugs reduced the tail current amplitude recorded at −40 mV after 250 ms depolarizing pulses to +60 mV, and slowed the deactivation time course resulting in a ‘crossover’ phenomenon when the tail currents recorded under control conditions and in the presence of each drug were superimposed.

Both compounds induced a small but statistically significant use‐dependent block when trains of depolarizations at frequencies between 0.5 and 3 Hz were applied.

These results indicate that propafenone and its metabolite block hKv1.5 channels in a concentration‐, voltage‐, time‐ and use‐dependent manner and the concentrations needed to observe these effects are in the therapeutical range.

British Journal of Pharmacology (1998) 125, 969–978; doi:10.1038/sj.bjp.0702129
Differential agonist activity of somatostatin and L‐362855 at human recombinant sst4 receptors
Authors: Smalley K S M , Feniuk W , Humphrey P P A
Published: January 2005

The operational characteristics of somatostatin (SRIF) sst4 receptors are poorly understood. In this study, we have characterized human recombinant sst4 receptors expressed in CHO cells (CHOsst4) by radioligand binding and microphysiometry.

Increasing concentrations SRIF or other SRIF receptor ligands inhibited specific [125I]‐Tyr11‐SRIF binding in CHOsst4 cell membranes with respective pIC50 values of SRIF (8.82), L‐362855 (7.40), BIM‐23027 (<5.5) and MK‐678 (<5.5).

These ligands displayed agonist activity, producing concentration‐dependent increases in rates of extracellular acidification (EAR) with pEC50 values of SRIF (9.6) and L‐362855 (8.0), respectively. BIM‐23027 and MK‐678 were at least 1000 times weaker than SRIF. The SRIF maximum was about 40% of that observed with L‐362855.

In the presence of SRIF (0.1–1 nM), concentration‐effect curves to L‐362855 were displaced to the right with a progressive reduction in the L‐362855 maximum.

When cells were only exposed to a single maximally effective concentration of SRIF or L‐362855, there was no difference in the magnitude of the agonist‐induced increase in EAR. However, a second agonist challenge, 30 min later showed that responses to SRIF but not L‐362855 were markedly desensitized.

When concentration‐effect curves to SRIF and L‐362855 were obtained by combining data from cells exposed to only a single agonist concentration, SRIF (pEC50 9.2) was approximately 20 times more potent than L‐362855 (pEC50 8.0) but the maxima were the same. Responses to both SRIF and L‐362855 were abolished by pertussis toxin.

SRIF and L‐362855‐induced increases in EAR were inhibited by N‐ethyl isopropyl amiloride (10 μM) but were not modified by inhibitors of PKC (Go‐6976), MAP kinase (PD‐98059), tyrosine kinase (genistein) or tyrosine phosphatase (sodium orthovanadate).

The results suggest that SRIF‐induced increases in EAR in CHOsst4 cells involved activation of the Na+/H+ antiporter and were mediated via Gi/Go G proteins. Responses to SRIF, but not L‐362855, were subject to marked desensitization which may be a consequence of differential activation of receptor‐effector coupling pathways.

British Journal of Pharmacology (1998) 125, 833–841; doi:10.1038/sj.bjp.0702133
Infarct size‐reducing effect of heat stress and α1 adrenoceptors in rats
Authors: Joyeux Marie , Godin‐Ribuot Diane , Patel Asha , Demenge Pierre , Yellon Derek M , Ribuot Christophe
Published: January 2005

Noradrenaline (NA), which is abundantly released during heat stress (HS), is known to induce both delayed cardioprotection and heat stress protein (HSP) 72 expression by the mediation of α1 adrenoceptors. Therefore, we have investigated the implication of α1 adrenoceptors in HS‐induced resistance to myocardial infarction, in the isolated rat heart model.

Rats were pretreated with prazosin (1 mg kg−1, i.p., Praz) or 5‐methylurapidil (3 mg kg−1, i.v, 5MU) or chloroethylclonidine (3 mg kg−1, i.v., CEC) or vehicle (V) in order to selectively antagonize α1, α1A and α1B adrenoceptors. They were then either heat stressed (42°C for 15 min) or sham anaesthetized. Twenty‐four hours later, their hearts were isolated, retrogradely perfused, and subjected to a 30 min occlusion of the left coronary artery followed by 120 min of reperfusion.

Infarct‐to‐risk ratio was significantly reduced in HS+V (15.4±1.8%) compared to Sham+V (35.7±1.3%) hearts. This effect was abolished in Praz‐treated (29.1±1.6% in HS+Praz vs 34.1±4.0% in Sham+Praz), 5MU‐treated (34.5±2.2% in HS+5MU vs 31.2±2.0% in Sham+5MU) and CEC‐treated (33.4±3.0% in HS+CEC vs 32.4±1.3% in Sham+CEC) groups. Western blot analysis of myocardial HSP72 showed an HS‐induced increase of this protein, which was not modified by Praz, 5MU and CEC pretreatments.

We conclude that both α1A and α1B adrenoceptor subtypes appear to play a role in the heat stress‐induced cardioprotection, independently of the HSP72 level. Further investigations are required to elucidate the precise role of HSPs in this adaptative response.

British Journal of Pharmacology (1998) 125, 645–650; doi:10.1038/sj.bjp.0702137
Influence of receptor number on the stimulation by salmeterol of gene transcription in CHO‐K1 cells transfected with the human β2‐adrenoceptor
Authors: McDonnell J , Latif M L , Rees E S , Bevan N J , Hill S J
Published: January 2005

The β2‐agonist salmeterol is a potent relaxant of airway smooth muscle with a long duration of action. Previous studies of cyclic AMP accumulation, however, have indicated that salmeterol is a low efficacy β2‐agonist when compared to isoprenaline. Here we have compared the properties of salmeterol and isoprenaline as stimulants of gene transcription in CHO‐K1 cells transfected with the human β2‐adrenoceptor to different levels (50 and 310 fmol mg protein−1).

Gene transcription was monitored using a secreted placental alkaline phosphate (SPAP) reporter gene under the transcriptional control of six cyclic AMP response element (CRE) sequences.

In the lower expressing cells (CHO‐β2/6), salmeterol produced a maximal cyclic AMP response that was only 22% that of that obtained with isoprenaline. In contrast in the higher expressing cells (CHO‐β2/4), the two maxima were of similar magnitude.

Salmeterol was a more potent stimulant of gene transcription, producing the same maximal response as isoprenaline in both cell lines. Furthermore, in the CHO‐β2/4 cells, Salmeterol was 50 fold more potent as a stimulant of SPAP secretion than of cyclic AMP accumulation. In contrast, isoprenaline was 24 fold less sensitive as a stimulant of SPAP secretion than of cyclic AMP accumulation. In the presence of serum (10%), the effects of both salmeterol and isoprenaline on gene transcription were augmented.

These data suggest that the low efficacy and/or long duration of action of salmeterol, favours a potent stimulation of gene transcription when compared to more efficacious but shorter‐lived agonists such as isoprenaline.

British Journal of Pharmacology (1998) 125, 717–726; doi:10.1038/sj.bjp.0702139
Weaning‐induced development of δ‐opioid receptors in rat brain: differential effects of guanine nucleotides and sodium upon ligand‐receptor recognition
Authors: Kelly M. D. W. , Hill R. G. , Borsodi A , Toth G , Kitchen I
Published: January 2005

We have previously shown that weaning at day 21 increases δ‐opioid receptor binding in the brain at day 25, which might be due to stimulation of the development of a δ‐opioid receptor subtype or activation of G‐protein coupling processes.

We have addressed the possibility that weaning stimulates coupling of the δ‐receptor by homogenate binding studies with four agonist and one antagonist radioligand in the presence of a GTP analogue and Na+ in brain tissue from weaned and non‐weaned animals.

Saturation studies with three agonist ligands ([3H]‐deltorphin I, [3H]‐S‐Atc‐Ile5,6deltorphin I and [3H]‐R‐Atc‐Ile5,6deltorphin II) showed higher levels of maximal binding in brains from 25‐day weaned than in brains from non‐weaned rats. The magnitude of the effects of GMPPNP and Na+ in decreasing this binding was ligand dependant and in each case was significantly more marked in brains from weaned animals. GMPPNP and Na+ were completely without effect on Bmax for, [3H]‐S‐Atc‐Ile5,6deltorphin I and [3H]‐R‐Atc‐Ile5,6deltorphin II in brains from non‐weaned rats.

[3H]‐Ile5,6deltorphin II and [3H]‐naltrindole showed no differences in labelling between weaned and non‐weaned groups and both groups responded similarly to the effects of GMPPNP and Na+ treatment.

GMPPNP and Na+ had small effects on binding affinity (KD) for some of the agonist radioligands which were similar in both weaned and non‐weaned groups.

Weaning induced increases in binding of δ‐receptors in 25‐day rats can be explained in terms of the way δ‐agonist radioligands recognize the receptor environment.

British Journal of Pharmacology (1998) 125, 979–986; doi:10.1038/sj.bjp.0702142
Effects of prenatal exposure to low concentrations of carbon monoxide on sexual behaviour and mesolimbic dopaminergic function in rat offspring
Authors: Cagiano Raffaele , Ancona Donata , Cassano Tommaso , Tattoli Maria , Trabace Luigia , Cuomo Vincenzo
Published: January 2005

Inhalation of low concentrations of carbon monoxide (CO) by pregnant rats (75 and 150 p.p.m. from day 0 to day 20 of gestation) leads to changes in mesolimbic dopaminergic transmission associated with an impairment of sexual behaviour in male offspring.

Eighty day old males exposed in utero to CO (150 p.p.m.) exhibited a significant increase in mount/intromission latency as well as a significant decrease in mount/intromission frequency. A significant decrease in ejaculation frequency was also found in CO (150 p.p.m.)‐exposed animals.

The acute administration of amphetamine, at a dose (0.5 mg kg−1 s.c.) stimulating copulatory activity in control rats, failed to reduce mount/intromission latency and did not increase mount frequency in 80‐day offspring exposed to CO (150 p.p.m.) during gestation.

These behavioural alterations were paralleled by neurochemical changes (in vivo microdialysis) showing that prenatal CO exposure, at concentrations (150 p.p.m.) that did not affect basal extracellular levels of dopamine in the nucleus accumbens, blunted the amphetamine (0.5 mg kg−1 s.c.)‐induced increase in dopamine release in 80‐day old male rats.

No significant changes in either behavioural or neurochemical parameters were observed in 10‐month old rats exposed prenatally to CO.

Since the alterations in sexual behaviour and dopaminergic transmission have been produced by prenatal exposure to CO levels resulting in maternal blood carboxyhaemoglobin concentrations equivalent to those maintained by human cigarette smokers, the present data further point out the large risk that the smoking mother poses for her offspring.

British Journal of Pharmacology (1998) 125, 909–915; doi:10.1038/sj.bjp.0702143
Systemic anti‐inflammatory effect induced by counter‐irritation through a local release of somatostatin from nociceptors
Authors: Szolcsányi János , Pintér Erika , Helyes Zsuzsanna , Oroszi Gábor , Németh József
Published: January 2005

Neurogenic plasma extravasation evoked by topical application of 1% vv−1 mustard oil on the skin of the acutely denervated rat hindleg (primary reaction) inhibited the development of a subsequent oil‐induced plasma extravasation induced in the skin of the contralateral hindleg by 49.3±7.06% (n=9) and in the conjunctival mucosa due to 0.1% wv−1 capsaicin instillation by 33.5±10.05% (n=6). The primary reaction also inhibited the non‐neurogenic hindpaw oedema evoked by s.c. injection of 5% wv−1 dextran into the chronically denervated hindpaw by 48.0±4.6% (n=5).

Capsaicin injection (100 μg ml−1 in 50 μl, s.c.) into the acutely denervated hindleg caused 56.5±4.0% (n=5) inhibition in the intensity of plasma extravasation elicited by 1% vv−1 mustard oil smearing on the contralateral side. After chronic denervation, subplantar injection of 5% wv−1 dextran elicited a non‐neurogenic inflammatory response with intensive tissue oedema without causing any systemic anti‐inflammatory effect. Bilateral adrenalectomy did not inhibit the mustard oil‐induced anti‐inflammatory effect in the contralateral hindleg.

Pretreating the rats with polyclonal somatostatin antiserum (0.5 ml rat−1, i.v.) or with the somatostatin depleting agent cysteamine (280 mg kg−1, s.c.) prevented the inhibitory action of mustard oil‐induced inflammation on subsequent neurogenic plasma extravasation and strongly diminished the inhibition of non‐neurogenic oedema formation evoked by dextran.

Exogenous somatostatin (10 μg kg−1, i.p.) caused a 30.3±8.3% (n=6) inhibition of plasma extravasation caused by mustard oil smearing on the acutely denervated hindleg and this inhibitory effect was abolished by somatostatin antiserum (0.5 ml rat−1, i.v.). The plasma level of somatostatin‐like immunoreactivity (SST‐LI) increased by 40.03±6.8% (n=6) 10 min after topical application of 1% vv−1 mustard oil on the acutely denervated hindpaws compared to the paraffin oil treated control group. Chronic denervation of the hindlegs or cysteamine (280 mg kg−1, s.c.) pretreatment prevented the mustard oil‐induced elevation of SST‐LI in plasma.

It is concluded that chemical excitation of the capsaicin‐sensitive sensory receptors not only induces local neurogenic plasma extravasation but also inhibits the development of a subsequent inflammatory reaction at remote sites of the body in the rat. A role for somatostatin in this systemic anti‐inflammatory effect is suggested.

British Journal of Pharmacology (1998) 125, 916–922; doi:10.1038/sj.bjp.0702144
Heat stress‐induced B1 receptor synthesis in the rat: an ex vivo study
Authors: Lagneux Caroline , Innocenti‐Francillard Patricia , Godin‐Ribuot Diane , Bader Mickael , Ribuot Christophe
Published: January 2005

This ex vivo study was performed to characterize B1 receptor induction in rats submitted to heat stress. Changes in aortic isometric tension were recorded after a 90 min in vitro incubation with [des‐Arg9]‐bradykinin. B1 receptor mRNA were detected in aorta and heart using RT–PCR technique.

Aortic rings from sham rats did not respond to [des‐Arg9]‐bradykinin. In contrast, this agonist induced a concentration‐dependent relaxation of aortic rings from rats submitted to lipopolysaccharide (LPS) treatment or to heat stress 24 h earlier.

The concentration‐dependent relaxation induced by [des‐Arg9]‐bradykinin on aortic rings from heat‐stressed rats was abolished by [des‐Arg10]‐HOE 140, a selective B1 receptor antagonist.

In endothelium denuded aortic rings from heat‐stress rats, [des‐Arg9]‐bradykinin induced a concentration‐dependent constriction.

Pretreatment of intact aortic rings from heat‐stress rats with the cyclo‐oxygenase inhibitor, diclofenac (1 μM) did not prevent the concentration‐dependent relaxation in response to [des‐Arg9]‐bradykinin. In contrast, NO synthase inhibition with Nω‐nitro‐L‐arginine methyl ester (30 μM) totally prevented the vasorelaxant response.

B1 receptor mRNA were not detected in aorta and heart from sham animals but were present in tissue from heat‐stressed and LPS‐treated rats.

In conclusion, our results suggest that heat stress induces a transcriptional activation of the B1 receptor gene. The induction of B1 receptors leads to an endothelium‐ and NO‐dependent vasorelaxant response to [des‐Arg9]‐bradykinin.

British Journal of Pharmacology (1998) 125, 812–816; doi:10.1038/sj.bjp.0702146
Effects of sulphonylureas on the volume‐sensitive anion channel in rat pancreatic β‐cells
Authors: Best L , Benington S
Published: January 2005

The volume‐sensitive anion conductance in rat pancreatic β‐cells was studied directly using the conventional whole‐cell and perforated patch recording techniques, and indirectly by measuring 3H‐taurine efflux from pre‐loaded, perifused islets.

Using the conventional whole‐cell recording configuration, activation of the outwardly‐rectifying, DIDS‐sensitive conductance was induced by glibenclamide (10 μM) but not by tolbutamide (100 μM) nor by meglitinide (20 μM). A high concentration of glibenclamide (100 μM) caused a voltage‐and time‐dependent inhibition of the conductance. Tolbutamide had a modest inhibitory effect on swelling‐induced inward currents.

In perforated patch recordings, glibenclamide, tolbutamide and meglitinide were all without effect on the conductance, although activation could be induced under these conditions by exposure to a hypotonic bath solution.

The rate of efflux of 3H‐taurine, a marker for activity of the volume‐sensitive anion channel, from preloaded, perifused islets was markedly stimulated by exposure to a hypotonic solution. However, glibenclamide and tolbutamide were both without effect.

Electrical activity of β‐cells in response to glibenclamide or tolbutamide was not inhibited by 4,4′‐dithiocyanatostilbene‐2,2′‐disulphonic acid (DIDS), an inhibitor of the volume‐sensitive anion channel.

It is concluded that activity of the volume‐sensitive anion conductance in rat pancreatic β‐cells is not modulated by the sulphonylurea receptor. The activation of the conductance by glibenclamide in whole‐cell recordings could be the result of a non‐specific interaction of the drug with plasma membrane lipids.

British Journal of Pharmacology (1998) 125, 874–878; doi:10.1038/sj.bjp.0702148
Contrasting effects of an aminobisphosphonate, a potent inhibitor of bone resorption, on lipopolysaccharide‐induced production of interleukin‐1 and tumour necrosis factor α in mice
Authors: Sugawara Shunji , Shibazaki Masahiko , Takada Haruhiko , Kosugi Hiroshi , Endo Yasuo
Published: January 2005

Aminobisphosphonates (aminoBPs), potent inhibitors of bone resorption, have been reported to induce inflammatory reactions such as fever and an increase in acute phase proteins in human patients, and to induce the histamine‐forming enzyme, histidine decarboxylase, in mice. In the present study, we examined the effect of aminoBP, 4‐amino‐1‐hydroxybutylidene‐1,1‐bisphosphonic acid (AHBuBP), on the production of the pro‐inflammatory cytokines, IL‐1 and TNFα, in mice.

Intraperitoneal injection of AHBuBP did not itself produce detectable levels of IL‐1 (α and β) and TNFα in the serum. However, the elevation of serum IL‐1 induced by lipopolysaccharide (LPS) was greatly augmented in mice injected with AHBuBP 3 days before the LPS injection, whereas the LPS‐induced elevation of serum TNFα was almost completely abolished.

Spleen and bone marrow cells taken from mice injected with AHBuBP produced IL‐1β in vitro spontaneously, and the production was augmented following the addition of LPS. Cells that accumulated in the peritoneal cavity in response to AHBuBP produced a particularly large amount of IL‐1β. However, AHBuBP treatment of mice did not lead to an impairment of the in vitro production of TNFα by these three types of cells.

Liposomes encapsulating dichloromethylene bisphosphonate (a non‐amino BP) selectively deplete phagocytic macrophages. When an intraperitoneal injection of these liposomes was given 2 days after an injection of AHBuBP, there was a marked decrease in the LPS‐induced elevation of serum IL‐1 (α and β) (LPS being injected 3 days after the injection of AHBuBP).

These results indicate that AHBuBP has contrasting effects on the in vivo LPS‐induced production of IL‐1 and TNFα in mice, enhancing the production of IL‐1 by phagocytic macrophages and suppressing the production of TNFα, although underling mechanisms remain to be clarified.

British Journal of Pharmacology (1998) 125, 735–740; doi:10.1038/sj.bjp.0702151
Genotypic and phenotypic analysis of the polymorphic thiopurine S‐methyltransferase gene (TPMT) in a European population
Authors: Spire‐Vayron de la Moureyre Catherine , Debuysere Hervé , Mastain Bruno , Vinner Elizabeth , Marez Delphine , Lo Guidice Jean‐Marc , Chevalier Dany , Brique Serge , Motte Kokou , Colombel Jean‐Frédéric , Turck Dominique , Noel Christian , Flipo René‐Marc , Pol Annie , Lhermitte Michel , Lafitte Jean‐Jacques , Libersa Christian , Broly Franck
Published: January 2005

Characterization of allelic variants of the TPMT gene (TPMT) responsible for changes in TPMT activity, and elucidation of the mechanism by which these alleles act, are required because of the clinical importance of this polymorphism for patients receiving thiopurine drugs.

We defined the mutational and allelic spectrum of TPMT in a group of 191 Europeans. Using PCR–SSCP, we screened for mutation the entire coding sequence, the exon‐intron boundaries, the promoter region and the 3′‐flanking region of the gene. Six mutations were detected throughout the ten exons and seven TPMT alleles were characterized. Four of them, TPMT*2, *3A, *3C and *7, harbouring the known mutations, G238C, G460A, A719G or T681G, were nonfunctional and accounted for 0.5, 5.7, 0.8 and 0.3% of the allele totality, respectively.

Within the promoter region, six alleles corresponding to a variable number of tandem repeats (VNTR), were identified. VNTR*V4 and *V5a which harbour four or five repeats of a 17–18 bp unit, were the most frequent (55% and 34%, respectively). The other VNTR alleles, having from five to eight repeats, were rarer.

The TPMT phenotype was correctly predicted by genotyping for 87% of individuals. A clear negative correlation between the total number of repeats from both alleles and the TPMT activity level was observed, indicating that VNTRs contribute to interindividual variations of TPMT activity. Therefore, additional analysis of the promoter region of TPMT can improve the phenotype prediction rate by genotyping.

British Journal of Pharmacology (1998) 125, 879–887; doi:10.1038/sj.bjp.0702152
Properties of neuronal nicotinic acetylcholine receptor mutants from humans suffering from autosomal dominant nocturnal frontal lobe epilepsy
Authors: Bertrand S , Weiland S , Berkovic S F , Steinlein O K , Bertrand D
Published: January 2005

Physiological and pharmacological properties of the human neuronal α4β2 nicotinic AChR and mutants found in patients suffering from autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) were studied.

Investigations of nicotinic AChRs reconstituted in Xenopus oocytes with the control or mutated α4 subunits revealed that both mutation S248F as well as the Leucine insertion (776ins3) result in major but different changes in the physiological and pharmacological properties of the receptors.

Mutation S248F causes a decrease in apparent affinity to ACh of about 7 fold. In addition, this receptor already desensitizes during exposure to agonist concentration 3000 times lower than the control.

776ins3 provokes a 10 fold increase of apparent ACh affinity, an increase in the IC50 caused by prolonged ACh exposures and a slowing down of the response decay.

At saturating ACh concentration cells expressing the S248F mutant display average currents that are about five times smaller than control.

When measured at very low concentration, agonist sensitivities of the control and mutated receptors to ACh, nicotine and epibatidine exhibit differences that match those observed for higher agonist concentrations.

Mutation 776ins3 increases the apparent efficacy to cytisine.

Data presented herein suggest that mutation S248F mainly affects the desensitization properties of the receptor while the leucine insertion (776ins3) increases the probability of transition to the active state. Although these mutations differentially affect the receptor properties they both result in reduced permeability to calcium and enhanced desensitization sensitivity that might account for the ADNFLE phenotype.

British Journal of Pharmacology (1998) 125, 751–760; doi:10.1038/sj.bjp.0702154
Activation of the cloned human NK3 receptor in Chinese Hamster Ovary cells characterized by the cellular acidification response using the Cytosensor microphysiometer
Authors: Jordan R E , Smart D , Grimson P , Suman‐Chauhan N , McKnight A T
Published: January 2005

The aim of the present study was to validate the Cytosensor microphysiometer, a novel system that measures the extracellular acidification rate as a reliable index of the integrated functional response to receptor activation, as a method for studying NK3 receptor pharmacology, and then to use this system to assess the functional activity of novel compounds at this receptor.

The selective NK3 agonist senktide caused reproducible, concentration‐related increases in acidification ratein CHO‐NK3 cells, with a pEC50 value of 8.72±0.11 (n=15). [β‐Ala8]NKA(4–10), the selective NK2 agonist, elicited a much weaker response (pEC50=6.68±0.08, n=4), while the NK1‐selective agonist substance P methylester only caused a very weak response at concentrations 3 μM (n=2). The rank order of potency for the endogenous tachykinins NKB>NKA>substance P (n=3) confirmed the response was mediated by the NK3 receptor. Moreover, the actual potencies obtained were consistent with affinities measured in radioligand binding studies.

The novel compounds PD156319‐121 (0.3–1 μM), PD161182 (10–300 nM), PD168001 (10–100 nM) and PD168073 (10–100 nM) all acted as surmountable antagonists of the senktide‐induced acidification response, with pA2 values of 7.49, 8.67, 9.17 and 9.25 respectively (n=3–5). In comparison the known NK3 antagonist SR142801 (10–100 nM) had a pA2 value of 8.83 (n=8) for the interaction with senktide. Again, these values are consistent with the radioligand binding data.

Amiloride (1 mM) inhibited the senktide‐induced acidification response by 68.3±3.3 (n=4), indicating that the Na+/H+ antiporter plays an important role in this response, and this is consistent with the importance of this antiporter in other acidification responses.

Inhibition of protein kinase C with staurosporine (0.1 μM), or depletion of the intracellular Ca2+ stores with thapsigargin (1 μM), both resulted in a reduction in the maximum response to senktide (63.3±1.7 and 68.9±3.2% respectively, n=3–5), and co‐application of these inhibitors abolished the response (n=3). This strongly suggested that the NK3 receptor was coupling via phospholipase C (PLC), as would be expected, although this could not be confirmed by the use of the putative PLC/PLA2 inhibitor U73122.

In conclusion, we have demonstrated the utility of the Cytosensor in the characterization of functional responses to agonists, and assessment of the affinities of antagonists in CHO cells expressing the human NK3, and have shown that our series of novel compounds are non‐peptide NK3 antagonists of high affinity, as exemplified by PD168073.

British Journal of Pharmacology (1998) 125, 761–766; doi:10.1038/sj.bjp.0702156
The development of overt diabetes in young Zucker Diabetic Fatty (ZDF) rats and the effects of chronic MCC‐555 treatment
Authors: Pickavance L , Widdowson P S , King P , Ishii S , Tanaka H , Williams G
Published: January 2005

Young (6‐week‐old) pre‐diabetic Zucker Diabetic Fatty (ZDF) rats displaying impaired glucose tolerance (IGT), moderate hyperglycaemia and hyperinsulinaemia were treated with the novel thiazolidinedione, MCC‐555, for 28 days, during which time β‐cell failure and progression to overt diabetes occurs.

Treated ZDF rats exhibited consistently lower blood glucose levels than vehicle‐treated diabetic controls, with a delayed rise and lower plateau levels. MCC‐555 maintained plasma insulin levels throughout the treatment period, whereas these fell by 40% in untreated ZDF rats.

The rise in body weight was maintained in MCC‐555‐treated rats, whereas vehicle‐treated rats exhibited blunted body weight gain after 8 weeks of age. Daily food intake was higher in diabetic, as compared to non‐diabetic rats, but treatment did not modify food intake in diabetic rats. Water intake was lower in treated ZDF rats, concomitant with lowering of blood glucose.

The hyperinsulinaemic‐euglycaemic clamp technique was applied to all rats after treatment to examine the effects of MCC‐555 on insulin sensitivity. The glucose infusion rate to maintain normoglycaemia was lower in diabetic than in non‐diabetic rats, demonstrating reduced glucose entry into insulin‐sensitive tissues in diabetic rats. Increased glucose infusion rates were required to maintain euglycaemia in treated diabetic rats, demonstrating increased insulin sensitivity in these animals.

In conclusion, chronic MCC‐555 treatment of young ZDF rats displaying IGT attenuates the development of overt diabetes through improved insulin sensitivity and maintenance of β‐cell function. MCC‐555 may thus be beneficial in humans with IGT, to prevent or delay the progression of diabetes.

British Journal of Pharmacology (1998) 125, 767–770; doi:10.1038/sj.bjp.0702158
The mechanism of the nitric oxide‐mediated enhancement of tert‐butylhydroperoxide‐induced DNA single strand breakage
Authors: Guidarelli Andrea , Clementi Emilio , Sciorati Clara , Cantoni Orazio
Published: January 2005

Caffeine (Cf) enhances the DNA cleavage induced by tert‐butylhydroperoxide (tB‐OOH) in U937 cells via a mechanism involving Ca2+‐dependent mitochondrial formation of DNA‐damaging species (Guidarelli et al., 1997b). Nitric oxide (NO) is not involved in this process since U937 cells do not express the constitutive nitric oxide synthase (cNOS).

Treatment with the NO donors S‐nitroso‐N‐acetyl‐penicillamine (SNAP, 10 μM), or S‐nitrosoglutathione (GSNO, 300 μM), however, potentiated the DNA strand scission induced by 200 μM tB‐OOH. The DNA lesions generated by tB‐OOH alone, or combined with SNAP, were repaired with superimposable kinetics and were insensitive to anti‐oxidants and peroxynitrite scavengers but suppressed by iron chelators.

SNAP or GSNO did not cause mitochondrial Ca2+ accumulation but their enhancing effects on the tB‐OOH‐induced DNA strand scission were prevented by ruthenium red, an inhibitor of the calcium uniporter of mitochondria. Furthermore, the enhancing effects of both SNAP and GSNO were identical to and not additive with those promoted by the Ca2+‐mobilizing agents Cf or ATP.

The SNAP‐ or GSNO‐mediated enhancement of the tB‐OOH‐induced DNA cleavage was abolished by the respiratory chain inhibitors rotenone and myxothiazol and was not apparent in respiration‐deficient cells.

It is concluded that, in cells which do not express the enzyme cNOS, exogenous NO enhances the accumulation of DNA single strand breaks induced by tB‐OOH via a mechanism involving inhibition of complex III.

British Journal of Pharmacology (1998) 125, 1074–1080; doi:10.1038/sj.bjp.0702159
The influence of lobeline on nucleus accumbens dopamine and locomotor responses to nicotine in nicotine‐pretreated rats
Authors: Benwell Maureen E M , Balfour David J K
Published: January 2005

In vivo brain microdialysis was used to investigate the influence of lobeline on dopamine (DA) and dihydroxyphenylacetic acid (DOPAC) overflow in the core of the nucleus accumbens of freely‐moving rats pretreated with nicotine (0.4 mg kg−1, s.c., once per day for 5 days). Locomotion was also recorded.

Lobeline, at doses of 0.7, 4.0 and 10.0 mg kg−1, i.p., failed to elicit any significant changes in extracellular dopamine or dihydroxyphenylacetic acid levels during the 60 min following its administration and did not stimulate locomotor.

The dopamine responses to nicotine (0.4 mg kg−1, s.c.), were abolished (P < 0.01) if the nicotine challenge was administered 10 min but not 60 min, after lobeline doses of 4.0 and 10.0 mg kg−1, i.p., but were unaffected following lobeline at the lowest dose tested (0.7 mg kg−1, i.p.) at either time. The increase in locomotor activity was significantly attenuated (P < 0.01), to a similar extent, when the nicotine was injected 10 min, but not 60 min, after all three doses of lobeline (0.7, 4.0 and 10.0 mg kg−1, i.p.) when compared with the saline‐treated rats.

The results suggest that lobeline is a short‐acting antagonist of the nicotinic AChRs which mediate the effects of nicotine on mesolimbic dopamine activity and locomotor stimulation.

British Journal of Pharmacology (1998) 125, 1115–1119; doi:10.1038/sj.bjp.0702161