bluuebunny/crossref_metadata_2025 huggingface dataset:
The effects of spiradoline (U‐62066E), a κ‐opioid receptor agonist, on neuroendocrine function in man
Authors: Ur Ehud , Wright D Mark , Bouloux Pierre M G , Grossman Ashley
Published: August 2006

Opioid drugs act on specific receptors which are principally classified into μ, δ and κ subtypes. Spiradoline (U‐62066E) is a κ‐selective agent which has been shown to possess potent anti‐nociceptive effects but does not show cross tolerance with morphine.

We have assessed the neuroendocrine effects of spiradoline in healthy volunteers with two doses (1.6 and 4.0 μg kg−1, i.m.) of the compound. Six male non‐smokers aged 19–27 years were studied by use of a randomized, double‐blind three‐limb placebo‐controlled cross‐over design. Blood was taken from an in‐dwelling venous cannula basally and at 15 min intervals for 2 h for determination of serum cortisol, prolactin, growth hormone (GH) and catecholamines.

Psychological function was assessed by the Stanford Sleepiness Scale (SSS) and the Addiction Research Centre Inventory (ARCI) administered before the medication and at 35 min, 1 h 25 min and 2 h afterwards. Cardiovascular variables were recorded at 10 min intervals. Results were analysed by analysis of variance.

Spiradoline showed a significant (P<0.05) dose‐dependent increase in free water clearance, as predicted for a κ‐opioid agonist. It also caused a dose‐dependent stimulation of prolactin, (increment over baseline for higher dose 214%), GH (433%) and cortisol (215%) release (P<0.05). There were no significant drug‐related changes in plasma catecholamines, blood pressure, pulse or psychological variables.

We have therefore confirmed that κ‐opioids increase free‐water clearance and may participate in the stimulation of prolactin and GH release. In contrast to μ and δ‐opioid agonists, this novel κ‐agonist stimulates cortisol release in man.

British Journal of Pharmacology (1997) 120, 781–784; doi:10.1038/sj.bjp.0700971
Renal effects of intracerebroventricularly injected tachykinins in the conscious saline‐loaded rat: receptor characterization
Authors: Ding Yuan Yi , Couture Réjean
Published: August 2006

The effects of intracerebroventricularly (i.c.v.) injected substance P (SP), neurokinin A (NKA) and [MePhe7]neurokinin B (NKB) were investigated on renal excretion of water, sodium and potassium in the conscious saline‐loaded rat. The central effects of [MePhe7]NKB were characterized with selective tachykinin antagonists for NK1 (RP 67580), NK2 (SR 48968) and NK3 (R 820) receptors.

Whereas SP or NKA (65 or 650 pmol) failed to modify the renal responses, [MePhe7]NKB (65–6500 pmol) produced dose‐dependent and long‐lasting (30–45 min) decreases in renal excretion of water (maximal reduction at 65 pmol: from 66.14±7.62 to 21.07±3.79 μl min−1), sodium (maximal reduction at 65 pmol: from 10.19±2.0 to 1.75±0.48 μmol min−1) and potassium (maximal reduction at 65 pmol: from 4.31±1.38 to 0.71±0.27 μmol min−1). While 650 pmol [MePhe7]NKB elevated urinary osmolality, neither 65 pmol nor 6.5 nmol [MePhe7]NKB altered this parameter.

Both the antidiuresis and antinatriuresis induced by [MePhe7]NKB (65 pmol) were significantly blocked by the prior i.c.v. injection of R 820 (1.3 nmol, 5 min earlier), although the potassium excretion was only partially reduced. However, R 820 did not affect the antidiuresis and antinatriuresis elicited by endothelin‐1 (1 pmol, i.c.v.). On its own, R 820 decreased renal potassium excretion with no effect on urinary osmolality and renal excretion of water and sodium. The i.c.v. co‐injection of RP 67580 and SR 48968 (6.5 nmol each, 5 min earlier) failed to modify the renal responses to [MePhe7]NKB in a similar study.

The central effects of [MePhe7]NKB (65 pmol) on renal excretion were blocked by the prior i.v. administration of a linear peptide vasopressin V2 receptor antagonist (50 μg kg−1, 5 min earlier).

These results suggest that the central NK3 receptor, probably located in the hypothalamus, is implicated in the renal control of water and electrolyte homeostasis through the release of vasopressin in the conscious saline‐loaded rat.

British Journal of Pharmacology (1997) 120, 785–796; doi:10.1038/sj.bjp.0700972
Inhibitory action of niflumic acid on noradrenaline‐ and 5‐hydroxytryptamine‐induced pressor responses in the isolated mesenteric vascular bed of the rat
Authors: Criddle D N , Soares de Moura R , Greenwood I A , Large W A
Published: August 2006

The effects of niflumic acid, an inhibitor of calcium‐activated chloride currents, were compared with the actions of the calcium channel blocker nifedipine on noradrenaline‐ and 5‐hydroxytryptamine (5‐HT)‐induced pressor responses of the rat perfused isolated mesenteric vascular bed.

Bolus injections of noradrenaline (1 and 10 nmol) increased the perfusion pressure in a dose‐dependent manner. Nifedipine (1 μm) inhibited the increase in pressure produced by 1 nmol noradrenaline by 31±5%. Niflumic acid (10 and 30 μm) also inhibited the noradrenaline‐induced increase in perfusion pressure and 30 μm niflumic acid reduced the pressor response to 1 nmol noradrenaline by 34±6%.

The increases in perfusion elicited by 5‐HT (0.3 and 3 nmol) were reduced by niflumic acid (10 and 30 μm) in a concentration‐dependent manner and 30 μm niflumic acid inhibited responses to 0.3 and 3 nmol 5‐HT by, respectively, 49±8% and 50±7%. Nifedipine (1 μm) decreased the pressor response to 3 nmol 5‐HT by 44±9%.

In the presence of a combination of 30 μm niflumic acid and 1 μm nifedipine the inhibition of the pressor effects of noradrenaline (10 nmol) and 5‐HT (3 nmol) was not significantly greater than with niflumic acid (30 μm) alone. Thus the effects of niflumic acid and nifedipine were not additive.

In Ca‐free conditions the transient contractions induced by 5‐HT (3 nmol) were not reduced by 30 μm niflumic acid, suggesting that this agent does not inhibit calcium release from the intracellular store or the binding of 5‐HT to its receptor.

Niflumic acid 30 μm did not inhibit the pressor responses induced by KCl (20 and 60 μmol) which were markedly reduced by 1 μm nifedipine. In addition, 1 μm levcromakalim decreased pressor responses produced by 20 μmol KCl. These data suggest that niflumic acid does not block directly calcium channels or activate potassium channels.

It is concluded that niflumic acid selectively reduces a component of noradrenaline‐ and 5‐HT‐induced pressor responses by inhibiting a mechanism which leads to the opening of voltage‐gated calcium channels. Our data suggest that the Ca2+‐activated chloride conductance may play a pivotal role in the activation of voltage‐gated calcium channels in agonist‐induced constriction of resistance blood vessels.

British Journal of Pharmacology (1997) 120, 813–818; doi:10.1038/sj.bjp.0700981
Pharmacological characterization of an α1A‐adrenoceptor mediating contractile responses to noradrenaline in isolated caudal artery of rat
Authors: Lachnit Wilhelm G , Tran Antares M , Clarke David E , Ford Anthony P D W
Published: August 2006

The α1‐adrenoceptor population mediating contraction of caudal artery of rat has been characterized by using quantitative receptor pharmacology.

Cumulative concentration‐effect (E/[A]) curves to noradrenaline (NA) yielded a p[A]50 of 5.56±0.05 (n=16). Prazosin caused concentration‐dependent, parallel, dextral shifts of E/[A] curves to NA yielding a pKb of 8.9 (Schild regression slope=1.0). RS‐17053 (N‐[2‐(2‐cyclopropyl methoxy phenoxy) ethyl]‐5‐chloro‐α,α‐dimethyl −1H‐indole‐ 3‐ethanamine hydrochloride; 10–100 nm), a selective α1A‐adrenoceptor antagonist, produced non‐parallel, biphasic, dextral shifts of E/[A] curves to NA, suggesting the involvement of more than one α1‐adrenoceptor subtype. Analysis of the high affinity component yielded an apparent pA2 value of 9.2±0.3.

A‐61603, a selective agonist at α1A adrenoceptors behaved as a full agonist relative to NA and yielded monophasic E/[A] curves with a p[A50] of 7.59±0.04 (n=15). Pretreatment of tissues with chloroethylclonidine (CEC; 100 μm for 20 min, followed by 40 min washout), which preferentially alkylates α1B‐ and α1D‐adrenoceptors, did not alter E/[A] curves to A‐61603. Prazosin (3–300 nm) caused concentration‐dependent, parallel, dextral shifts of E/[A] curves to A‐61603 yielding a pA2 estimate of 9.2±0.2.

Experiments with α1‐adrenoceptor antagonists of varying subtype selectivities (RS‐17053, SNAP 5089, tamsulosin, 5‐methylurapidil, BMY 7378, HV 723 and REC 15/2739) revealed parallel dextral shifts of E/[A] curves to A‐61603. Schild regression analyses yielded pA2 estimates of 9.2, 9.3, 11.2, 9.0, 6.3, 8.7 and 10.0 for RS‐17053, SNAP 5089, tamsulosin, 5‐methylurapidil, BMY 7378, HV 723 and REC 15/2739, respectively, although deviations from unit slope (possibly reflecting a secondary involvement of another α1‐adrenoceptor) hindered estimations of pKb for some antagonists. The antagonist affinity profile obtained reflects best that described for the α1A‐adrenoceptor.

In conclusion, caudal artery of rat contracts in response to NA via activation of at least two α1‐adrenoceptor subtypes. One of these subtypes displays the pharmacology of the α1A‐adrenoceptor, while the other remains to be defined. Use of the novel selective agonist, A‐61603, allows for limited pharmacological isolation of the α1A‐adrenoceptor permitting characterization of the properties of selective antagonists.

British Journal of Pharmacology (1997) 120, 819–826; doi:10.1038/sj.bjp.0700983
Vasoconstrictor responses via P2X‐receptors are selectively antagonized by NF023 in rabbit isolated aorta and saphenous artery
Authors: Ziyal Ragip , Ziganshin Airat U , Nickel Peter , Ardanuy Ursula , Mutschler Ernst , Lambrecht Günter , Burnstock Geoffrey
Published: August 2006

The effects of NF023, the symmetrical 3′‐urea of 8‐(benzamido)naphthalene‐1,3,5‐trisulphonic acid), and its parent compound suramin were investigated on vasoconstrictor responses to α,β‐methylene ATP in rabbit isolated saphenous artery and vasodilator responses to ATP in noradrenaline‐precontracted rabbit isolated thoracic aorta.

In rabbit isolated saphenous artery, α,β‐methylene ATP‐induced vasoconstrictor responses via P2X‐receptors were concentration‐dependently and competitively antagonised by NF023 (30–300 μm; pA2=5.69±0.04). Suramin (100–1000 μm) also competitively blocked vasoconstrictor responses to α,β‐methylene ATP, albeit with lower potency (pA2=4.79±0.05). In contrast, NF023 (100 μm) did not significantly affect contractile responses to noradrenaline or histamine in the saphenous artery.

In noradrenaline‐precontracted rabbit isolated thoracic aorta preparations, ATP (3–3000 μm) concentration‐dependently induced relaxations via endothelium‐dependent or smooth muscle P2Y‐receptor subtypes. NF023 (30–300 μm) failed to block relaxant responses to ATP at endothelium‐dependent P2Y‐receptors, whereas suramin (100–1000 μm) did antagonise endothelium‐dependent vasodilator responses to ATP. Neither NF023 (100 μm) nor suramin (300 μm) influenced vasorelaxant responses to ATP via endothelium‐independent P2Y‐receptors.

In conclusion, this study outlines the selectivity of NF023 as an effective P2X‐receptor antagonist in rabbit isolated blood vessels without affecting endothelium‐dependent or endothelium‐independent P2Y‐receptor subtypes, adrenoceptors or histamine receptors.

British Journal of Pharmacology (1997) 120, 954–960; doi:10.1038/sj.bjp.0700984
The effects of bradykinin on K+ currents in NG108–15 cells treated with U73122, a phospholipase C inhibitor, or neomycin
Authors: Hildebrandt Jan‐Peter , Plant Timothy D , Meves Hans
Published: August 2006

Bradykinin has multiple effects on differentiated NG108–15 neuroblastoma×glioma cells: it increases Ins(1,4,5)P3 production and intracellular Ca2+ concentration [Ca2+]i, evokes a Ca2+ activated K+ current (IK(Ca)) and inhibits M current (IM). We studied the effect of the aminosteroid U73122 and the antibiotic neomycin, both putative blockers of phospholipase C (PLC), on these four bradykinin effects.

Preincubation with 1 or 5 μm U73122 for 15 min partly suppressed Ins(1,4,5)P3 generation and the increase in [Ca2+]i induced by 1 μm bradykinin. U73122 10 μm caused total and irreversible inhibition. The inactive analogue U73343 was without effect.

Resting levels of Ins(1,4,5)P3 were not affected. However, resting [Ca2+]i was increased by 10 μm U73122, but not by U73343. Individual cells responded to 10 μm U73122 with a small increase in [Ca2+]i, followed in some cells by a large further rise.

Pretreatment of whole‐cell clamped cells with 1 μm U73122 for 30 min reduced the bradykinin‐induced IK(Ca) to a fifth of its normal size. To suppress it totally, a 7–12 min pretreatment with 5 μm U73122 was required. Again, U73343 was without effect.

U73122 and U73343 at concentrations of 5–10 μm irreversibly decreased the holding current (Ih) which at a holding potential of −30 or −20 mV mainly flows through open M channels. The decrease was often preceded by a transient increase.

M current (IM) measured with 1 s pulses, was also decreased by 5–10 μm U73122 and U73343, but short applications of U73122 could cause a small increase. The bradykinin‐induced inhibition of IM was not affected by U73122.

Preincubation with 1 or 3 mm neomycin for 15 min did not affect Ins(1,4,5)P3 generation and the increase in [Ca2+]i induced by bradykinin. Pretreatment with 3 mm neomycin for about 20 min diminished the bradykinin‐induced IK(Ca) to a fifth of its normal size.

The four main conclusions drawn from the results are: (a) U73122 suppresses bradykinin‐induced PLC activation and IK(Ca), but not IM inhibition. (b) This indicates that the transient outward current IK(Ca), but not the decrease of IM in response to bradykinin, is mediated by PLC. (c) U73122 itself inhibits IM and mobilizes Ca2+ from intracellular stores. (d) Externally applied neomycin is not an effective inhibitor of PLC‐mediated signalling pathways in NG108–15 cells.

British Journal of Pharmacology (1997) 120, 841–850; doi:10.1038/sj.bjp.0700991
Evidence for the involvement of 5‐HT4 receptors in the 5‐hydroxytryptamine‐induced pattern of migrating myoelectric complex in sheep
Authors: Plaza Miguel‐Angel , Arruebo María‐Pilar , Murillo María‐Divina
Published: August 2006

The effects induced by 5‐hydroxytryptamine (5‐HT) on gastrointestinal myoelectric activity in conscious sheep were recorded through electrodes chronically implanted and analysed by computer. The 5‐HT receptors and the cholinergic neuronal pathways involved in these actions were investigated.

The intravenous (i.v.) administration of 5‐HT (2, 4 and 8 μg kg−1 min−1, 5 min) induced an antral inhibition concomitant with a duodenal activity front that migrated to the jejunum, followed by a period of intestinal inactivity. This myoelectric pattern closely resembled that observed in the phases III and I of the migrating myoelectric complex (MMC) in sheep. The 0.5 μg kg−1 min−1 dose evoked the same pattern in only two out of the six animals used. Likewise, the 1 μg kg−1 min−1 dose similarly affected four of the six animals. In addition, a transient stimulation was observed in the antrum and jejunum when the two highest doses were used.

The 5‐HT1 antagonist, methiothepin (0.1 mg kg−1), the 5‐HT2 antagonists, ritanserin (0.1 mg kg−1) and ketanserin (0.3 mg kg−1), the 5‐HT3 antagonists, granisetron (0.2 mg kg−1) and ondansetron (0.5 mg kg−1), as well as the 5‐HT4 antagonist, GR113808 (0.2 mg kg−1), did not modify the spontaneous gastrointestinal myoelectric activity. However, the cholinoceptor antagonists, atropine (0.2 mg kg−1) and hexamethonium (2 mg kg−1), inhibited gastrointestinal activity.

When these antagonists were injected i.v. 10 min before 5‐HT (2 or 4 μg kg−1 min−1, 5 min), only GR113808, atropine and hexamethonium were able to modify the 5‐HT‐induced actions, all of them being completely blocked by the three antagonists.

Our data show that 5‐HT initiates a MMC‐like pattern in the gastrointestinal area in sheep through 5‐HT4 receptors. Furthermore, these actions are mediated by cholinergic neural pathways involving muscarinic and nicotinic receptors. However, our results do not indicate a role for either 5‐HT1, 5‐HT2 or 5‐HT3 receptors in the 5‐HT‐induced effects.

British Journal of Pharmacology (1997) 120, 1144–1150; doi:10.1038/sj.bjp.0700997
The site of general anaesthesia and cytochrome P450 oxygenases: similarities defined by straight chain and cyclic alcohols
Authors: LaBella F S , Chen Q ‐M , Stein D , Queen G
Published: August 2006

General anaesthetics disrupt normal cell receptivity and responsiveness while sparing vital respiratory processes. Ultimate elucidation of the molecular basis of general anaesthesia presumes the identification of one or more subcellular components with appropriate sensitivity to the entire array of anaesthetics.

Previously, we showed the universal cellular enzymes, cytochrome P450 mono‐oxygenases, to be sensitive at relevant concentrations to all anaesthetics tested. The potential significance of P450 inhibition by anaesthetics resides in the contribution of this enzyme family, in conjunction with that of cyclo‐oxygenases and lipoxygenases, to the generation from arachidonic acid of lipid second messengers, the eicosanoids.

We have shown that P450 enzymes model the site of general anaesthesia in the tadpole with respect to (a) an absolute sensitivity to increasing chain‐length series of flexible, straight chain primary and secondary alcohols and straight chain diols, (b) an absolute sensitivity to increasing molecular weight series of rigid cyclic alkanols and cyclic alkanemethanols, (c) the points of abrupt change and of reversal (cut‐off) in the linear relationship between increasing anaesthetic potency with increasing carbon chain length, and (d) non‐differentiation between secondary alkanol enantiomers. These findings reveal the P450 enzyme family as the most relevant biomolecular counterpart of the site of general anaesthesia, thus far identified.

British Journal of Pharmacology (1997) 120, 1158–1164; doi:10.1038/sj.bjp.0701006
Attenuation of diet‐induced atherosclerosis in rabbits with a highly selective 15‐lipoxygenase inhibitor lacking significant antioxidant properties
Authors: Sendobry Sandra M. , Cornicelli Joseph A. , Welch Kathryn , Bocan Thomas , Tait Bradley , Trivedi Bharat K. , Colbry Norman , Dyer Richard D. , Feinmark Steven J. , Daugherty Alan
Published: August 2006

15‐Lipoxygenase (15‐LO) has been implicated in the pathogenesis of atherosclerosis because of its localization in lesions and the many biological activities exhibited by its products. To provide further evidence for a role of 15‐LO, the effects of PD 146176 on the development of atherosclerosis in cholesterol‐fed rabbits were assessed. This novel drug is a specific inhibitor of the enzyme in vitro and lacks significant non specific antioxidant properties.

PD 146176 inhibited rabbit reticulocyte 15‐LO through a mixed noncompetitive mode with a Ki of 197 nm. The drug had minimal effects on either copper or 2,2′‐azobis(2‐amidinopropane)hydrochloride (ABAP) induced oxidation of LDL except at concentrations 2 orders higher than the Ki.

Control New Zealand rabbits were fed a high‐fat diet containing 0.25% wt./wt. cholesterol; treated animals received inhibitor in this diet (175 mg kg−1, b.i.d.). Plasma concentrations of inhibitor were similar to the estimated Ki (197 nm). During the 12 week study, there were no significant differences in weight gain, haematocrit, plasma total cholesterol concentrations, or distribution of lipoprotein cholesterol.

The drug plasma concentrations achieved in vivo did not inhibit low‐density lipoprotein (LDL) oxidation in vitro. Furthermore, LDL isolated from PD 146176‐treated animals was as susceptible as that from controls to oxidation ex vivo by either copper or ABAP.

PD 146176 was very effective in suppressing atherogenesis, especially in the aortic arch where lesion coverage diminished from 15±4 to 0% (P<0.02); esterified cholesterol content was reduced from 2.1±0.7 to 0 μg mg−1 (P<0.02) in this region. Immunostainable lipid‐laden macrophages present in aortic intima of control animals were totally absent in the drug‐treated group.

Results of these studies are consistent with a role for 15‐LO in atherogenesis.

British Journal of Pharmacology (1997) 120, 1199–1206; doi:10.1038/sj.bjp.0701007
Nitric oxide production and endothelium‐dependent vasorelaxation induced by wine polyphenols in rat aorta
Authors: Andriambeloson Emile , Kleschyov Andrei L , Muller Bernard , Beretz Alain , Stoclet Jean Claude , Andriantsitohaina Ramaroson
Published: August 2006

The aim of this work was to investigate the mechanism of vasorelaxation induced by red wine polyphenolic compounds (RWPC) and two defined polyphenols contained in wine, leucocyanidol and catechin. The role of the endothelium, especially endothelium‐derived nitric oxide (NO), was also investigated.

Relaxation produced by polyphenols was studied in rat aortic rings with and without functional endothelium, pre‐contracted to the same extent with noradrenaline (0.3 and 0.1 μm, respectively). RWPC and leucocyanidol, but not catechin, produced complete relaxation of vessels with and without endothelium. However, 1000 fold higher concentrations were needed to relax endothelium‐denuded rings compared to those with functional endothelium.

High concentrations of catechin (in the range of 10−1 g l−1) only produced partial relaxation (maximum 30%) and had the same potency in rings with and without endothelium.

The NO synthase inhibitor, Nω‐nitro‐l‐arginine‐methyl‐ester (l‐NAME, 300 μm) completely abolished the endothelium‐dependent but not the endothelium‐independent relaxations produced by all of the polyphenolic compounds.

In contrast to superoxide dismutase (SOD, 100 u ml−1), neither RWPC nor leucocyanidol affected the concentration‐response curve for the NO donor, SIN‐1 (3‐morpholino‐sydnonimine) which also produces superoxide anion (O2−).

In aortic rings with endothelium, RWPC (10−2 g l−1) produced a 7 fold increase in the basal production of guanosine 3′: 5′‐cyclic monophosphate (cyclic GMP) which was prevented by l‐NAME (300 μm).

Electron paramagnetic resonance (e.p.r.) spectroscopy studies with Fe2+‐diethyldithiocarbamate as an NO spin trap demonstrated that RWPC and leucocyanidol increased NO levels in rat thoracic aorta about 2 fold. This NO production was entirely dependent on the presence of the endothelium and was abolished by l‐NAME (300 μm).

These results show that RWPC and leucocyanidol, but not the structurally closely related polyphenol catechin, induced endothelium‐dependent relaxation in the rat aorta. They indicate that this effect results from enhanced synthesis of NO rather than enhanced biological activity of NO or protection against breakdown by O2−. It is concluded that some polyphenols, with specific structure, contained in wine possess potent endothelium‐dependent vasorelaxing activity.

British Journal of Pharmacology (1997) 120, 1053–1058; doi:10.1038/sj.bjp.0701011
The effects of recombinant rat μ‐opioid receptor activation in CHO cells on phospholipase C, [Ca2+]i and adenylyl cyclase
Authors: Smart D , Hirst R A , Hirota K , Grandy D K , Lambert D G
Published: August 2006

The rat μ‐opioid receptor has recently been cloned, yet its second messenger coupling remains unclear. The endogenous μ‐opioid receptor in SH‐SY5Y cells couples to phospholipase C (PLC), increases [Ca2+]i and inhibits adenylyl cyclase (AC). We have examined the effects of μ‐opioid agonists on inositol(1,4,5)trisphosphate (Ins(1,4,5)P3), [Ca2+]i and adenosine 3′: 5′‐cyclic monophosphate (cyclic AMP) formation in Chinese hamster ovarian (CHO) cells transfected with the cloned μ‐opioid receptor.

Opioid receptor binding was assessed with [3H]‐diprenorphine ([3H]‐DPN) as a radiolabel. Ins(1,4,5)P3 and cyclic AMP were measured by specific radioreceptor assays. [Ca2+]i was measured fluorimetrically with Fura‐2.

Scatchard analysis of [3H]‐DPN binding revealed that the Bmax varied between passages. Fentanyl (10 pm–1 μm) dose‐dependently displaced [3H]‐DPN, yielding a curve which had a Hill slope of less than unity (0.6±0.1), and was best fit to a two site model, with pKi values (% of sites) of 9.97±0.4 (27±4.8%) and 7.68±0.07 (73±4.8%). In the presence of GppNHp (100 μm) and Na+ (100 mm), the curve was shifted to the right and became steeper (Hill slope=0.9±0.1) with a pKi value of 6.76±0.04.

Fentanyl (0.1 nm–1 μm) had no effect on basal, but dose‐dependently inhibited forskolin (1 μm)‐stimulated, cyclic AMP formation (pIC50=7.42±0.23), in a pertussis toxin (PTX; 100 ng ml−1 for 24 h)‐sensitive and naloxone‐reversible manner (Ki=1.7 nm). Morphine (1 μm) and [d‐Ala2, MePhe4, gly(ol)5]‐enkephalin (DAMGO, 1 μm) also inhibited forskolin (1 μm)‐stimulated cyclic AMP formation, whilst [d‐Pen2, d‐Pen5], enkephalin (DPDPE, 1 μm) did not.

Fentanyl (0.1 nm–10 μm) caused a naloxone (1 μm)‐reversible, dose‐dependent stimulation of Ins(1,4,5)P3 formation, with a pEC50 of 7.95±0.15 (n=5). PTX (100 ng ml−1 for 24 h) abolished, whilst Ni2+ (2.5 mm) inhibited (by 52%), the fentanyl‐induced Ins(1,4,5)P3 response. Morphine (1 μm) and DAMGO (1 μm), but not DPDPE (1 μm), also stimulated Ins(1,4,5)P3 formation. Fentanyl (1 μm) also caused an increase in [Ca2+]i (80±16.4 nm, n=6), reaching a maximum at 26.8±2.5 s. The increase in [Ca2+]i remained elevated until sampling ended (200 s) and was essentially abolished by the addition of naloxone (1 μm). Pre‐incubation with naloxone (1 μm, 3 min) completely abolished fentanyl‐induced increases in [Ca2+]i.

In conclusion, the cloned μ‐opioid receptor when expressed in CHO cells stimulates PLC and inhibits AC, both effects being mediated by a PTX‐sensitive G‐protein. In addition, the receptor couples to an increase in [Ca2+]i. These findings are consistent with the previously described effector‐second messenger coupling of the endogenous μ‐opioid receptor.

British Journal of Pharmacology (1997) 120, 1165–1171; doi:10.1038/sj.bjp.0701012
Naloxone‐insensitive inhibition of acetylcholine release from parasympathetic nerves innervating guinea‐pig trachea by the novel opioid, nociceptin
Authors: Patel Hema J , Giembycz Mark A , Spicuzza Lucia , Barnes Peter J , Belvisi Maria G
Published: August 2006

The novel peptide, nociceptin and the μ‐opioid agonist [d‐Ala2, N‐Me‐Phe4, Gly5‐ol]‐enkephalin (DAMGO) produced a concentration‐dependent inhibition of electrical field stimulation (EFS)‐evoked release of acetylcholine (ACh) from cholinergic nerves innervating guinea‐pig trachea. The non‐selective opioid receptor antagonist, naloxone, did not antagonize the inhibitory action of nociceptin under conditions where the inhibition of ACh release evoked by DAMGO was completely reversed. It is suggested that DAMGO and nociceptin can inhibit cholinergic, parasympathetic neurotransmission to the airways via the activation of classical (naloxone‐sensitive) and novel (naloxone‐insensitive) opioid receptors, respectively.

British Journal of Pharmacology (1997) 120, 735–736; doi:10.1038/sj.bjp.0701013
The ability of a new hypoglycaemic agent, A‐4166, compared to sulphonylureas, to increase cytosolic Ca2+ in pancreatic β‐cells under metabolic inhibition
Authors: Fujitani Shoji , Okazaki Kyoko , Yada Toshihiko
Published: August 2006

N‐(trans‐4‐isopropylcyclohexanecarbonyl)‐d‐phenylalanine (A‐4166) is a new non‐sulphonylurea oral hypoglycaemic agent which stimulates insulin release by increasing cytosolic Ca2+ concentration ([Ca2+]i) in β‐cells.

We studied comparative effects of A‐4166 and sulphonylureas on [Ca2+]i, measured by dual‐wavelength fura‐2 microfluorometry, in single rat pancreatic β‐cells under normal conditions and conditions where glucose metabolism was inhibited.

A glucokinase inhibitor, mannoheptulose (10 mm), a mitochondrial respiratory inhibitor, KCN (100 μm), and uncouplers, dinitrophenol (DNP, 50 μm) and carbonyl cyanide p‐trifluoromethoxyphenylhydrazone (FCCP, 0.3 μm), were used to abolish glucose‐induced increases in [Ca2+]i in a reversible manner.

Under control conditions, A‐4166 was one order more potent than tolbutamide in increasing [Ca2+]i, and maximal responses were evoked by 30 μm A‐4166 and 300 μm tolbutamide. These equipotent concentrations were employed for the comparative study where glucose metabolism was inhibited.

In the presence of mannoheptulose, [Ca2+]i responses to tolbutamide, but not those to A‐4166, were attenuated in a reversible manner.

KCN, DNP and FCCP inhibited [Ca2+]i responses to tolbutamide to a much greater extent than those to A‐4166. Responses to tolbutamide even at 3.3 times the equipotent concentration (1000 μm) were also markedly attenuated by these inhibitors. Responses evoked by another sulphonylurea, gliclazide, were inhibited by DNP to a larger extent than A‐4166‐induced responses.

The results indicate that A‐4166 acts more effectively than sulphonylureas to increase [Ca2+]i in β‐cells during metabolic inhibition.

British Journal of Pharmacology (1997) 120, 1191–1198; doi:10.1038/sj.bjp.0701017
Evaluation of BTS 67 582, a novel antidiabetic agent, in normal and diabetic rats
Authors: Jones R B , Dickinson K , Anthony D M , Marita A R , Kaul C L , Buckett W R
Published: August 2006

The effect of BTS 67 582, a novel antidiabetic agent, has been evaluated on plasma glucose and plasma insulin in normal and streptozotocin‐induced diabetic rats.

BTS 67 582 (3 to 300 mg kg−1, p.o.) caused a dose‐ and time‐ dependent reduction in plasma glucose and an increase in plasma insulin in both fasted and glucose‐loaded normal rats. The ED50 for the glucose lowering effect of BTS 67 582 in fasted rats was 37.6, 18.4 and 18.5 mg kg−1 at 1, 2 and 4 h after administration respectively.

In streptozotocin‐induced (50 mg kg−1, i.v.) diabetic rats, BTS 67 582 (37–147 mg kg−1, p.o.) caused significant reductions of plasma glucose following a glucose load, whereas glibenclamide (100 mg kg−1, p.o.) was ineffective. BTS 67 582 significantly increased plasma insulin compared to controls whereas glibenclamide did not.

BTS 67 582 did not displace [3H]‐glibenclamide from its binding sites in rat brain, guinea‐pig ventricle or the HIT‐T15 insulinoma β‐cell line. BTS 67 582 does not therefore appear to modulate its action via an effect on the ‘sulphonylurea’ receptor.

In fasted rats, the glucose lowering effect of BTS 67 582 (100 mg kg−1 p.o.) and glibenclamide (1 mg kg−1, p.o.) were antagonized by diazoxide (30 mg kg−1, i.p.). In addition BTS 67 582, like glibenclamide, caused a dose‐dependent rightward shift of cromakalim‐induced relaxation of noradrenaline precontracted rat aortic strips, suggesting the involvement of KATP channels.

In summary, BTS 67 582 produces a blood glucose‐lowering effect in normal and streptozotocin‐induced diabetic rats associated with increased insulin concentrations. This effect appears to be due to a blockade of ATP‐sensitive potassium channel activity via a different binding site to that of glibenclamide.

British Journal of Pharmacology (1997) 120, 1135–1143; doi:10.1038/sj.bjp.0701019
Induction by staurosporine of nitric oxide synthase expression in vascular smooth muscle cells: role of NF‐κB, CREB and C/EBPβ
Authors: Hecker Markus , Preiß Christiane , Schini‐Kerth Valérie B
Published: August 2006

The effect of different protein kinase inhibitors on the expression of the inducible isoform of nitric oxide (NO) synthase (iNOS) was investigated in cultured vascular smooth muscle cells (VSMC) isolated from the rat aorta.

The non‐selective protein kinase C (PKC) inhibitor, staurosporine, but not the more selective PKC inhibitors, calphostin C and Ro 31‐8220, or the tyrosine kinase inhibitors, genistein and erbstatin analogue (erbstatin A), elicited a distinct (up to six fold) up‐regulation of iNOS gene expression in these cells, as demonstrated by a parallel increase in iNOS mRNA and protein abundance as well as an accumulation of nitrite (NO2−) in the conditioned medium. Actinomycin D and cycloheximide inhibited the effect of staurosporine, suggesting an involvement of both DNA transcription and de novo protein synthesis.

Staurosporine also synergistically potentiated the stimulating effect of interleukin‐1β (IL‐1β), but not that of the adenylyl cyclase activator, forskolin, on NO2− production and iNOS protein abundance. Staurosporine, on the other hand, had no effect on the IL‐1β‐mediated increase in iNOS mRNA abundance. The effect of staurosporine on both basal and IL‐1β‐stimulated NO2− production was concentration‐dependent with an apparent maximum at 3 nm. Among the other protein kinase inhibitors tested, only calphostin C also enhanced the stimulant effect of IL‐1β approximately two fold, while genistein, erbstatin A and Ro 31‐8220 inhibited rather than potentiated it.

Staurosporine did not influence basal activity of the transcription factors CREB and nuclear factor κB (NF‐κB), but increased that of C/EBP. Moreover, staurosporine significantly augmented the activation of C/EBP by IL‐1β and forskolin.

These findings suggest that in cultured VSMC a staurosporine‐sensitive protein kinase exists, which is unlikely to be related to PKC, that prevents iNOS gene expression presumably by suppressing basal C/EBP activity. They also indicate that NF‐κB and a member of the C/EBP family of transcription factors, presumably C/EBPβ, act synergistically under basal conditions and possibly also following exposure to IL‐1β in the up‐regulation of iNOS gene expression in these cells. Targeting of the activation of C/EBPβ may thus represent an interesting approach to interfere selectively with the cytokine‐induced over‐production of NO in acute and chronic inflammatory conditions.

British Journal of Pharmacology (1997) 120, 1067–1074; doi:10.1038/sj.bjp.0701026
A comparative study on the rat aorta and mesenteric arterial bed of the possible role of nitric oxide in the desensitization of the vasoconstrictor response to an α1‐adrenoceptor agonist
Authors: Kamata Katsuo , Makino Ayako
Published: August 2006

In thoracic aortic strips with intact endothelium, the first and second (1 h later) dose‐response curves obtained with methoxamine were almost the same.

Methoxamine caused a dose‐dependent increase in perfusion pressure in the rat isolated mesenteric arterial bed, but the second (1 h later) dose‐response curve for methoxamine showed a significant attenuation of the response in comparison with the first.

The attenuation shown by the second dose‐response curve for methoxamine was significantly reduced, but not abolished, in mesenteric arterial beds without endothelium. Incubating endothelium‐intact mesenteric arterial beds with NG‐nitro‐l‐arginine (l‐NOARG) caused a significant, but not complete, reversal of the attenuation shown in the second dose‐response curve.

Incubating the mesenteric arterial bed with capsaicin, tetrodotoxin, indomethacin or with isotonic high k+ (60 mm) plus nicardipine did not affect the above attenuation seen in the second dose‐response curve.

The guanosine 3′:5′‐cyclic monophosphate (cyclic GMP) level in the effluent from the perfused mesenteric arterial bed was significantly increased after the second exposure to methoxamine. This effect was significantly smaller after removal of the endothelium or pretreatment with l‐NOARG.

These results suggest that a desensitization to methoxamine develops rapidly in the mesenteric arterial bed, but not in the aorta, and that release of nitric oxide from the endothelium plays a major role in this desensitization.

British Journal of Pharmacology (1997) 120, 1221–1228; doi:10.1038/sj.bjp.0701031
Characterization of the potassium channels involved in EDHF‐mediated relaxation in cerebral arteries
Authors: Petersson Jesper , Zygmunt Peter M. , Högestätt Edward D.
Published: August 2006

In the presence of NG‐nitro‐l‐arginine (l‐NOARG, 0.3 mm) and indomethacin (10 μm), the relaxations induced by acetylcholine and the calcium (Ca) ionophore A23187 are considered to be mediated by endothelium‐derived hyperpolarizing factor (EDHF) in the guinea‐pig basilar artery.

Inhibitors of adenosine 5′‐triphosphate (ATP)‐sensitive potassium (K)‐channels (KATP; glibenclamide, 10 μm), voltage‐sensitive K‐channels (KV; dendrotoxin‐I, 0.1 μm or 4‐aminopyridine, 1 mm), small (SKCa; apamin, 0.1 μm) and large (BKCa; iberiotoxin, 0.1 μm) conductance Ca‐sensitive K‐channels did not affect the l‐NOARG/indomethacin‐resistant relaxation induced by acetylcholine.

Synthetic charybdotoxin (0.1 μm), an inhibitor of BKCa and KV, caused a rightward shift of the concentration‐response curve for acetylcholine and reduced the maximal relaxation in the presence of l‐NOARG and indomethacin, whereas the relaxation induced by A23187 was not significantly inhibited.

A combination of charybdotoxin (0.1 μm) and apamin (0.1 μm) abolished the l‐NOARG/indomethacin‐resistant relaxations induced by acetylcholine and A23187. However, the acetylcholine‐induced relaxation was not affected by a combination of iberiotoxin (0.1 μm) and apamin (0.1 μm).

Ciclazindol (10 μm), an inhibitor of KV in rat portal vein smooth muscle, inhibited the l‐NOARG/indomethacin‐resistant relaxations induced by acetylcholine and A23187, and the relaxations were abolished when ciclazindol (10 μm) was combined with apamin (0.1 μm).

Human pial arteries from two out of four patients displayed an l‐NOARG/indomethacin‐resistant relaxation in response to substance P. This relaxation was abolished in both cases by pretreatment with the combination of charybdotoxin (0.1 μm) and apamin (0.1 μm), whereas each toxin had little effect alone.

The results suggest that KV, but not KATP and BKCa, is involved in the EDHF‐mediated relaxation in the guinea‐pig basilar artery. The synergistic action of apamin and charybdotoxin (or ciclazindol) could indicate that both KV and SKCa are activated by EDHF. However, a single type of K‐channel, which may be structurally related to KV and allosterically regulated by apamin, could also be the target for EDHF.

British Journal of Pharmacology (1997) 120, 1344–1350; doi:10.1038/sj.bjp.0701032
Effect of short‐term and long‐term treatments with σ ligands on the N‐methyl‐d‐aspartate response in the CA3 region of the rat dorsal hippocampus
Authors: Bergeron Richard , De Montigny Claude , Debonnel Guy
Published: August 2006

Long‐term treatments with the σ ligand haloperidol decrease the density of σ receptors in mammalian CNS. We have shown that σ ligands, such as di(2‐tolyl)guanidin (DTG), potentiate dose‐dependently, with bell‐shaped dose‐response curves, the neuronal response of pyramidal neurones to N‐methyl‐d‐aspartate (NMDA) in the CA3 region of the rat dorsal hippocampus. σ Ligands producing such a potentiation were denoted ‘agonists’. This potentiation was suppressed by low doses of other σ ligands denoted ‘antagonists’. High doses of DTG and JO‐1784 did not modify the NMDA response but acted as ‘antagonists’ by suppressing the potentiation induced by σ ‘agonists’.

Following a 21‐day treatment with haloperidol as well as with high doses of DTG or JO‐1784, after a 48 h washout, the acute administration of σ ‘agonists’ failed to induce any potentiation of the NMDA response. Following a 21 day treatment with a low dose of DTG or JO‐1784, after a 48 h washout, the neuronal response to microiontophoretic applications of NMDA was markedly increased. A 21 day treatment with low or high doses of (+)‐pentazocine, after a 48 h washout, did not produce any change.

Following a two day treatment with a high dose of haloperidol, DTG, JO‐1784 and (+)‐pentazocine, after a 24 h washout, the potentiation of the NMDA response induced by the acute administration of the σ ‘agonists’ was unchanged.

With the minipumps on board, with DTG and JO‐1784, a dose‐dependent enhancement of the NMDA response was seen but no effect was observed in the groups of rats treated at the same doses with haloperidol or (+)‐pentazocine.

The present data suggest that long‐term treatments with σ ‘antagonists’ induce a desensitization of the σ receptors, whereas long‐term treatments with σ ‘agonists’ induce a supersensitivity of the σ receptors.

British Journal of Pharmacology (1997) 120, 1351–1359; doi:10.1038/sj.bjp.0701042
An autoradiographic study of dextromethorphan high‐affinity binding sites in rat brain: sodium‐dependency and colocalization with paroxetine
Authors: Meoni P. , Tortella F. C. , Bowery N. G.
Published: August 2006

The distribution and some pharmacological properties of centrally located dextromethorphan high‐affinity binding sites were investigated by in vitro autoradiography.

Sodium chloride (50 mm) induced a 7 to 12 fold increase in dextromethorphan binding to rat brain in all areas tested. The effect of sodium was concentration‐dependent with a higher dose (120 mm) exerting a smaller effect on binding.

[3H]‐dextromethorphan binding in the presence of sodium was inhibited in the presence of the anticonvulsant phenytoin at a concentration of 100 μm, while the σ ligand (+)‐3‐(‐3‐hydroxyphenyl)‐N‐(1‐propyl)pipendine ((+)‐PPP) had no effect on the binding, suggesting an interaction with the DM2 site.

The distribution of the sodium‐dependent binding identified in this study correlated significantly with the distribution of the selective 5‐HT uptake inhibitor [3H]‐paroxetine, and paroxetine and dextromethorphan mutually displaced their binding at concentrations in the low nanomolar range.

These data show that dextromethorphan and paroxetine share a sodium‐dependent high affinity binding site in rat brain, and suggest that dextromethorphan might interact, in the presence of sodium, with the 5‐HT uptake mechanism in rat brain.

British Journal of Pharmacology (1997) 120, 1255–1262; doi:10.1038/sj.bjp.0701043
β‐Adrenoceptor‐mediated effects in rat cultured thymic epithelial cells
Authors: Kurz Bodo , Feindt Janka , Von Gaudecker Brita , Kranz Andrea , Loppnow Harald , Mentlein Rolf
Published: August 2006

Sympathetic nerves were visualized in sections from rat thymus by immunostaining of tyrosine hydroxylase, the rate‐limiting enzyme of catecholamine biosynthesis, and by glyoxylic acid‐induced fluorescence of catecholamines. Catecholaminergic nerve fibres were detected in close connection to thymic epithelial cells which therefore might be preferred target cells. To evaluate this, rat immunocytochemically defined, cultured thymic epithelial cells were investigated for adrenoceptors and adrenergic effects.

In rat cultured thymic epithelial cells mRNA for β1‐ and β2‐adrenoceptors was detected by reverse transcription‐polymerase chain reaction by use of sequence‐specific primers. Specific, saturable binding to the cultivated cells was observed with the β‐adrenoceptor agonist CGP 12177.

Adrenaline, noradrenaline or the β‐adrenoceptor agonist, isoprenaline, increased intracellular adenosine 3′: 5′‐cyclic monophosphate (cyclic AMP) levels in cultivated thymic epithelial cells dose‐dependently about 25 fold. The pharmacological properties revealed that this response was mediated by receptors of the β1‐ and the β2‐subtypes. The selective β3‐adrenoceptor agonist BRL 37344 had no effect on cyclic AMP levels. The increase in cyclic AMP was downregulated by preincubation with glucocorticoids like dexamethasone or cortisol which also changed the relative importance of β1‐/β2‐adrenoceptors to the response.

Incubation with isoprenaline or the adenylate cyclase activator forskolin decreased basal and serum‐stimulated proliferation of thymic epithelial cells. However, adrenergic stimulation of thymic epithelial cells did not induce interleukin 1 production. Since thymic epithelial cells create a microenvironment which influences the maturation and differentiation of thymocytes to T‐lymphocytes, their observed capacity to respond to catecholamines provides novel evidence for the suggestion that adrenergic stimulation may interfere with the regulation of immune functions.

British Journal of Pharmacology (1997) 120, 1401–1408; doi:10.1038/sj.bjp.0701045
Evidence for a dilator function of 8‐iso prostaglandin F2α in rat pulmonary artery
Authors: Jourdan Karen B. , Evans Timothy W. , Curzen Nicholas P. , Mitchell Jane A.
Published: August 2006

8‐Iso prostaglandin F2α (8‐iso PGF2α) is one of a series of prostanoids formed independently of the cyclo‐oxygenase pathway. It has been shown to be upregulated in many conditions of oxidant stress where its formation is induced by free radical‐catalysed actions on arachidonic acid. As 8‐iso PGF2α is formed in vivo in diseases in which oxidant stress is high such as septic shock, we have assessed the relative potency and efficacy of this compound in pulmonary arteries from control and lipopolysaccharide (LPS)‐treated rats.

Several studies have characterized the contractile actions of 8‐iso PGF2α on various smooth muscle preparations, but its potential dilator actions have not been addressed. Thus these studies examined both the contractile and dilator actions of 8‐iso PGF2α in rat pulmonary artery rings. The thromboxane mimetic U46619, PGE2 sodium nitroprusside (SNP) and acetyl choline (ACh) were used for comparison. Each prostanoid had to be dissolved in ethanol to a maximum concentration of 1×10−2 m. At high concentrations, ethanol directly contracted pulmonary vessels. We were therefore limited by the actions of the vehicle such that we were unable to add prostanoids at concentrations higher than 1×10−4 m. In some cases this meant that maximum responses were not achieved and in these cases the Emax and pD2 values are apparent estimates.

The following rank order of potency was obtained from contractile studies; U46619>8‐iso PGF2α>PGE2, each prostanoid producing concentration‐dependent contractions (10−103×10−4 m, 10−910−4 m, 10−810−4 m, respectively). As has been shown previously for other smooth muscle preparations, the thromboxane receptor (TP) antagonist ICI 192605, (1×10−6, 1×10−5 and 1×10−4 m), inhibited the contractions of 8‐iso PGF2α in a concentration‐dependent fashion.

The nitric oxide synthase inhibitor, NG‐nitro‐l‐arginine methyl ester (l‐NAME; 1×10−4 m), enhanced the contractile function of both 8‐iso PGF2α and PGE2, but had no effect on that caused by U46619. Similarly, l‐NAME inhibited the dilator function of all agents tested except the exogenous nitric oxide (NO) donor SNP, indicating that PGE2 and 8‐iso PGF2α like ACh, act through the release of NO. The specificity of the effects of l‐NAME were confirmed in studies with the inactive enantiomer d‐NAME (1×10−4 m), which did not affect the contractile or the dilator actions of 8‐iso PGF2α. Furthermore, ICI 192605 enhanced the dilator actions of 8‐iso PGF2α, suggesting that the dilator component of 8‐iso PGF2α was achieved via activation of a non‐TP receptor.

Isoprostanes may modulate vascular tone by a direct action on TP receptors to cause contraction and via a distinct receptor leading to the release of NO to cause dilatation.

British Journal of Pharmacology (1997) 120, 1280–1285; doi:10.1038/sj.bjp.0701052