bluuebunny/crossref_metadata_2025 huggingface dataset:
Analysis of the muscarinic receptor subtype mediating inhibition of the neurogenic contractions in rabbit isolated vas deferens by a series of polymethylene tetra‐amines
Authors: Budriesi R , Cacciaguerra S , Di Toro R , Bolognesi M L , Chiarini A , Minarini A , Rosini M , Spampinato S , Tumiatti V , Melchiorre C
Published: January 2005

The pharmacological characteristics of the presynaptic muscarinic receptor subtype, which mediates inhibition of the neurogenic contractions in the prostatic portion of rabbit vas deferens, have been investigated by using a series of polymethylene tetra‐amines, which were selected for their ability to differentiate among muscarinic receptor subtypes.

It was found that all tetra‐amines antagonized McN‐A‐343‐induced inhibition in electrically stimulated rabbit vas deferens in a competitive manner and with affinity values (pA2) ranging between 6.27±0.09 (spirotramine) and 8.51±0.02 (AM170).

Competition radioligand binding studies, using native muscarinic receptors from rat tissues (M1, cortex; M2, heart; M3, submaxillary gland) or from NG 108‐15 cells (M4) and human cloned muscarinic M1‐M4 receptors expressed in CHO‐K1 cells, were undertaken with the same tetra‐amines employed in functional assays. All antagonists indicated a one‐site fit.

The affinity estimates (pKi) of tetra‐amines calculated in binding assays using native receptors were similar to those obtained using cloned receptors. Among these compounds some displayed selectivity between muscarinic receptor subtypes, indicating that they may be valuable tools in receptor characterization. Spirotramine was selective for M1 receptors versus all other subtypes (pKi native: M1, 7.32±0.10; M2, 6.50±0.11; M3, 6.02±0.13; M4, 6.28±0.16; pKi cloned: M1, 7.69±0.08; M2, 6.22±0.14; M3, 6.11±0.16; 6.35±0.11) whereas CC8 is highly selective for M2 receptors versus the other subtypes (pKi native: M1, 7.50±0.04; M2, 9.01±0.12; M3, 6.70±0.08; M4, 7.56±0.04; pKi cloned: M1, 7.90±0.20; M2, 9.04±0.08; M3, 6.40±0.07; M4, 7.40±0.04). Furthermore, particularly relevant for this investigation were tetra‐amines dipitramine and AM172 for their ability to significantly differentiate M1 and M4 receptors.

The apparent affinity values (pA2) obtained for tetra‐amines in functional studies using the prostatic portion of rabbit vas deferens correlated most closely with the values (pKi) obtained at either native or human recombinant muscarinic M4 receptors. This supports the view that the muscarinic receptor mediating inhibition of neurogenic contractions of rabbit vas deferens may not belong to the M1 type but rather appears to be of the M4 subtype.

British Journal of Pharmacology (2001) 132, 1009–1016; doi:10.1038/sj.bjp.0703891
Mechanisms involved in SNP‐induced relaxation and [Ca2+]i reduction in piglet pulmonary and systemic arteries
Authors: Cogolludo Angel L , Pérez‐Vizcaíno Francisco , Zaragozá‐Arnáez Francisco , Ibarra Manuel , López‐López Gustavo , López‐Miranda Visitación , Tamargo Juan
Published: January 2005

We have compared the mechanisms involved in sodium nitroprusside (SNP)‐induced relaxation and [Ca2+]i reduction in isolated piglet pulmonary (PA) and mesenteric (MA) arteries.

SNP (10−8 M−3×10−5 M) evoked a concentration‐dependent relaxation of PA and MA (pD2=6.66±0.06 and 6.74±0.14, respectively) stimulated by noradrenaline, which was markedly reduced by the guanylate cyclase inhibitor ODQ. In fura 2‐incubated PA and MA, SNP produced a parallel reduction in contractile force and in [Ca2+]i, expressed as the ratio of emitted fluorescence at 340 and 380 nm (F340/F380).

The inhibition of the Na+/K+‐ATPase after the incubation in a K+‐free medium or the exposure to ouabain (10−6 M) inhibited SNP‐induced relaxation in MA but not in PA. SNP‐induced relaxation was not attenuated by 80 mM KCl plus nifedipine (10−6 M) but was inhibited by thapsigargin (2×10−6 M; pD2=5.69±0.19 and 5.89±0.19 for PA and MA, respectively).

Pretreatment of PA with thapsigargin and MA with thapsigargin plus ouabain induced a stronger inhibition on the reduction in [Ca2+]i than on the relaxation induced by SNP, indicating the existence of Ca2+‐independent mechanisms.

The activation of the Na+/K+‐ATPase by the addition of KCl after the incubation in a K+‐free medium similarly reduced [Ca2+]i in PA and MA, whereas it relaxed with much less efficacy PA than MA.

We conclude that SNP reduces [Ca2+]i and causes relaxation through the activation of SERCA in PA and SERCA and Na+/K+‐ATPase in MA. However, Ca2+‐independent mechanisms also contribute to SNP‐induced effects.

British Journal of Pharmacology (2001) 132, 959–967; doi:10.1038/sj.bjp.0703894
Alterations in EDHF‐mediated hyperpolarization and relaxation in mesenteric arteries of female rats in long‐term deficiency of oestrogen and during oestrus cycle
Authors: Liu Ming‐Yue , Hattori Yuichi , Fukao Mistuhiro , Sato Atsushi , Sakuma Ichiro , Kanno Morio
Published: January 2005

This study was undertaken to determine whether endothelium‐dependent relaxations are altered in mesenteric arteries from young female rats during oestrus cycle and after castration.

The contractile response to phenylephrine (Phe) was significantly enhanced in arteries from rats subjected to ovariectomy than in those from sham‐operated (control) rats. Treatment of ovariectomized rats with 17β‐oestradiol returned the Phe response to the control level. Arteries from rats at the diestrus stage also exhibited greater contraction in response to Phe. In the presence of 100 μM NG‐nitro‐L‐arginine (L‐NOARG), the enhancement of the Phe contractile response associated with oestrogen deficiency was not observed.

Endothelium‐dependent relaxations elicited by acetylcholine (ACh) in arteries precontracted with Phe were significantly reduced in ovariectomized and diestrus rats regardless of whether endothelium‐derived nitric oxide (NO) was blocked with L‐NOARG. Treatment with 17β‐oestradiol prevented the reduced vascular relaxant response to ACh in ovariectomized rats. The reduction in the ACh responses observed in ovariectomized and diestrus rats was eliminated when 500 nM apamin and 100 nM charybdotoxin were present.

ACh‐induced endothelium‐dependent hyperpolarizations were depressed in arteries from ovariectomized and diestrus rats. The hyperpolarizing response to ACh was significantly improved when ovariectomized rats were treated with 17β‐oestradiol. The resting membrane potentials and pinacidil‐induced hyperpolarizations were unaffected by ovariectomy or the diestrus stage.

These results suggest that oestrogen‐deficient states of both short and long duration reduce the basal release of NO from the endothelium and specifically attenuate endothelium‐dependent hyperpolarization and relaxation transduced by endothelium‐derived hyperpolarizing factor.

British Journal of Pharmacology (2001) 132, 1035–1046; doi:10.1038/sj.bjp.0703899
Pharmacological characterization of muscarinic receptors in dog isolated ciliary and urinary bladder smooth muscle
Authors: Choppin A , Eglen R M
Published: January 2005

The pharmacological characteristics of muscarinic receptors mediating contraction of dog isolated ciliary muscle were determined and compared to those mediating contraction of dog urinary bladder smooth muscle.

(+)‐Cis‐dioxolane induced concentration‐dependent contractions of ciliary muscle (pEC50=7.18±0.07, Emax=453±64 mg, n=19) and urinary bladder isolated smooth muscle (pEC50=6.55±0.07, Emax=11±1 g, n=19). These responses were antagonized by several muscarinic receptor antagonists (pKb values for the ciliary muscle and the bladder smooth muscle, respectively): atropine (8.25±0.14 and 9.21±0.09), pirenzepine (6.31±0.13 and 6.70±0.25), tolterodine (7.97±0.14 and 8.68±0.12), oxybutynin (7.40±0.08 and 7.88±0.12), zamifenacin (6.46±0.19 and 7.69±0.11), S‐secoverine (6.66±0.14 and 8.13±0.07), AQ‐RA 741 (6.16±0.15 and 7.08±0.23), p‐F‐HHSiD (7.10±0.27 and 7.35±0.07) and responses were not antagonized by PD 102807 (up to 100 nM).

In urinary bladder smooth muscle, the profile of antagonist pKB values correlated significantly with pKi values at human recombinant m3 muscarinic receptors, suggesting that M3 muscarinic receptors mediated the response. In the ciliary muscle, a significant (P<0.01) correlation was obtained with human recombinant m3 and m5 receptors.

Darifenacin displayed insurmountable antagonism at receptors in the bladder. At receptors in the ciliary muscle, it exhibited two phases of antagonism, comprising an initial low affinity (pKB<6) component and a high affinity phase (pKB>8).

The role of pigmentation in the atypical behaviour of darifenacin was examined. In blue coloured eyes, darifenacin produced apparent surmountable, competitive antagonism of the responses to (+)‐cis‐dioxolane (pKB=8.76±0.07). The antagonist profile obtained in this tissue suggested the involvement of a site which has the pharmacological attributes of the M5 receptor.

We suggest that the dog urinary bladder contracts in response to M3 muscarinic receptor activation. Contraction of the brown‐eyed dog ciliary muscle is more complex and may include involvement of at least two receptors, possibly the M5 and M3 receptor, whereas blue‐eyed dog ciliary muscle may involve a single population of M5 muscarinic receptors.

British Journal of Pharmacology (2001) 132, 835–842; doi:10.1038/sj.bjp.0703901
Involvement of cannabinoids in the cardioprotection induced by lipopolysaccharide
Authors: Lagneux Caroline , Lamontagne Daniel
Published: January 2005

We have examined the involvement of the endocannabinoid system in the cardioprotection triggered by lipopolysaccharide (LPS). Rats were treated with saline or LPS (10 μg Kg−1). 24 h later, hearts were excised, retrogradely perfused, submitted to a low‐flow ischaemia (0.6 ml min−1) for 90 min and reperfused for 60 min. Some hearts were perfused with either SR 141716A (a cannabinoid CB1, receptor antagonist 1 μM), SR 144528 (a CB2 receptor anagonist μM), NNLA (3 μM) or sodium nitroprusside (1 μM) 5 min before ischaemia and during the ischaemic period. The cardioprotective effects of LPS treatment, in terms of infarction and functional recovery, were not altered by the perfusion of SR 141716A but abolished by both SR 144528 and NNLA. Finally, SR 144528 abolished the beneficial effects of SNP perfusion. Our results suggest an involvement of endocannabinoids, acting through the CB2 receptors, in the cardioprotection triggered by LPS against myocardial ischaemia. This could be attributed to a relationship between cannabinoids and NO.

British Journal of Pharmacology (2001) 132, 793–796; doi:10.1038/sj.bjp.0703902
Activation of type 5 metabotropic glutamate receptors enhances NMDA responses in mice cortical wedges
Authors: Attucci S , Carlà V , Mannaioni G , Moroni F
Published: January 2005

We measured the effects of agonists and antagonists of metabotropic glutamate (mGlu) receptors (types 1 and 5) on NMDA‐induced depolarization of mouse cortical wedges in order to characterize the mGlu receptor type responsible for modulating NMDA responses. We also characterized a number of mGlu receptor agents by measuring [3H]‐inositol phosphate (IP) formation in cortical slices and in BHK cells expressing either mGlu 1 or mGlu 5 receptors.

(S)‐3,5‐dihydroxyphenylglycine (DHPG), an agonist of both mGlu 1 and mGlu 5 receptors, at concentrations ranging from 1 – 10 μM, enhanced up to 105±15% the NMDA‐induced depolarization. Larger concentrations (100 – 300 μM) of the compound were inactive in this test. When evaluated on [3H]‐IP synthesis in cortical slices or in cells expressing either mGlu 1 or mGlu 5 receptors, DHPG responses (1 – 300 μM) increased in a concentration‐dependent manner.

(RS)‐2‐chloro‐5‐hydroxyphenylglycine (CHPG) and (S)‐(+)‐2‐(3′‐carboxybicyclo[1.1.1]pentyl)‐glycine (CBPG), had partial agonist activity on mGlu 5 receptors, with maximal effects reaching approximately 50% that of the full agonists. These compounds, however, enhanced NMDA‐evoked currents with maximal effects not different from those induced by DHPG. Thus the enhancement of [3H]‐IP synthesis and the potentiation of NMDA currents were not directly related.

2‐methyl‐6‐(phenylethynyl)‐pyridine (MPEP, 1 – 10 μM), a selective mGlu 5 receptor antagonist, reduced DHPG effects on NMDA currents. 7‐(hydroxyimino)cyclopropan[b]‐chromen‐1a‐carboxylic acid ethylester (CPCCOEt, 30 μM), a preferential mGlu 1 receptor antagonist, did not reduce NMDA currents.

These results show that mGlu 5 receptor agonists enhance while mGlu 5 receptor antagonists reduce NMDA currents. Thus the use of mGlu 5 receptor agents may be suggested in a number of pathologies related to altered NMDA receptor function.

British Journal of Pharmacology (2001) 132, 799–806; doi:10.1038/sj.bjp.0703904
Protein phosphatase inhibitors facilitate DHPG‐induced LTD in the CA1 region of the hippocampus
Authors: Schnabel R , Kilpatrick I C , Collingridge G L
Published: January 2005

We have shown earlier that activation of metabotropic glutamate (mGlu) receptors using a group I‐specific mGlu receptor agonist, (RS)‐3,5‐dihydroxyphenylglycine (DHPG), can induce long‐term depression (LTD) in the CA1 region of the hippocampus. In an attempt to determine the signal transduction mechanisms involved in this form of synaptic plasticity, we have tested the effects of a range of inhibitors on DHPG‐induced LTD.

In vitro grease‐gap electrophysiological recordings were performed in the rat hippocampal CA1 region.

We have found that DHPG‐induced LTD is resistant to the two potent protein kinase C (PKC) inhibitors, Gö 6976 (10 μM) and Gö 6983 (10 μM), the potent and selective protein kinase A (PKA) inhibitor, KT 5720 (10 μM), and the potent broad spectrum kinase inhibitor, staurosporine (10 μM).

In contrast, non‐selective inhibitors of protein phosphatases (PP1 and PP2A), okadaic acid (1 μM) or calyculin A (1 μM), facilitated DHPG‐induced LTD. However, an inhibitor of protein phosphatase 2B, FK 506 (1 μM), did not influence this process.

The PP1/PP2A protein phosphatase inhibitors, but none of the other agents tested, also inhibited (S)‐α‐methyl‐4‐carboxyphenylglycine (MCPG)‐induced reversal of DHPG‐induced LTD.

These data suggest that activation of neither PKC nor PKA is involved in DHPG‐induced LTD. They do, however, suggest that the process is under regulation by protein phosphorylation and dephosphorylation.

British Journal of Pharmacology (2001) 132, 1095–1101; doi:10.1038/sj.bjp.0703905
The endogenous cannabinoid agonist, anandamide stimulates sensory nerves in guinea‐pig airways
Authors: Tucker R C , Kagaya M , Page C P , Spina D
Published: January 2005

The endogenous cannabinoid agonist, anandamide produced a modest contractile response in guinea‐pig isolated bronchus compared with the vanilloid receptor agonist capsaicin. The contractile response to both anandamide and capsaicin was inhibited by the vanilloid receptor antagonist, capsazepine. Furthermore, the NK2‐selective antagonist, SR48968 but not the NK1‐selective antagonist, SR140333 inhibited contractile responses to anandamide.

The contractile response to anandamide was abolished in tissues desensitized by capsaicin. However, anandamide failed to cross‐desensitize the contractile response to capsaicin.

The contractile response to anandamide was not significantly altered in the presence of the CB1 receptor antagonist, SR141716A, nor the amidase inhibitor, phenylmethylsulphonyl fluoride (PMSF) but was significantly increased in the presence of the neutral endopeptidase inhibitor, thiorphan.

The cannabinoid agonist, CP55,940 failed to significantly attenuate the excitatory non‐adrenergic non‐cholinergic (eNANC) response in guinea‐pig airways. In contrast, the ORL1 receptor agonist, nociceptin, significantly inhibited this response.

The results demonstrate that anandamide induces a modest contractile response in guinea‐pig isolated bronchus that is dependent upon the activation of vanilloid receptors on airway sensory nerves. However, cannabinoid receptors do not appear to play a role in this regard, nor in regulating the release of neuropeptides from airway sensory nerves under physiological conditions.

British Journal of Pharmacology (2001) 132, 1127–1135; doi:10.1038/sj.bjp.0703906
Effects of the endogenous opioid peptide, endomorphin 1, on supraoptic nucleus oxytocin and vasopressin neurones in vivo and in vitro
Authors: Doi Naomi , Brown Colin H , Cohen Helena Delgado , Leng Gareth , Russell John A
Published: January 2005

We investigated the actions of the endogenous opioid tetra‐peptide endomorphin 1, a selective μ‐opioid receptor agonist, on oxytocin and vasopressin cell activity in vivo and in vitro.

The activity of antidromically‐identified supraoptic nucleus cells were recorded from urethane‐anaesthetized female rats. The firing rates of both oxytocin and vasopressin cells were reduced by intracerebroventricular endomorphin 1 (5–100 pmol); this inhibition was prevented by intravenous naloxone (5 mg kg−1).

A second group of rats was infused intracerebroventricularly with endomorphin 1 (27 pmol min−1) over 5 days. The firing rates of oxytocin and vasopressin cells in endomorphin 1 pre‐treated rats were similar to those of endomorphin 1 naïve rats, indicating tolerance to the inhibitory effects of endomorphin 1. Intravenous naloxone induced similar modest and transient increases in the firing rate of oxytocin cells in endomorphin 1 pre‐treated rats and endomorphin 1 naïve rats, indicating that endomorphin 1, unlike the μ‐opioid alkaloid agonist, morphine, does not induce μ‐opioid dependence in these cells.

In vitro, whole‐cell current clamp recordings were made from supraoptic nucleus cells in superfused coronal hypothalamic slices from young female rats. Endomorphin 1 (100 nM) inhibited the firing rate of oxytocin cells but had no significant effect on vasopressin cells at up to 10 μM. Inhibition of oxytocin cells was reversed by naloxone, and remained when synaptic transmission was blocked by superfusion with low Ca2+/Co2+‐containing medium.

Thus, endomorphin 1 directly inhibits oxytocin cells but inhibits vasopressin cells by indirect actions. Chronic endomorphin 1 administration induces μ‐opioid tolerance in oxytocin and vasopressin cells but not μ‐opioid dependence in oxytocin cells.

British Journal of Pharmacology (2001) 132, 1136–1144; doi:10.1038/sj.bjp.0703911
Comparative effects of frovatriptan and sumatriptan on coronary and internal carotid vascular haemodynamics in conscious dogs
Authors: Carel Ivan , Ghaleh Bijan , Edouard Alain , Dubois‐Rande Jean‐Luc , Parsons Andrew A , Giudicelli Jean‐François , Berdeaux Alain
Published: January 2005

The effects of frovatriptan and sumatriptan on internal carotid and coronary vascular haemodynamics were investigated and compared in conscious dogs.

Frovatriptan and sumatriptan (0.1–100 μg kg−1) induced a transient increase in external coronary artery diameter (eCOD) of up to 2.9±1.2 and 1.8±0.6%, respectively (both P<0.05). This was followed by a prolonged and dose‐dependent decrease in eCOD of up to −5.2±1.2 and −5.3±0.9% (both P<0.05), with ED50 values of 86±21 and 489±113 μmol kg−1, respectively. In contrast, only a decrease in the external diameter of the internal carotid artery was observed (−6.0±0.6 and −6.2±1.4%, both P<0.05, and ED50 values of 86±41 and 493±162 μmol kg−1, respectively). Frovatriptan was thus 5.7 fold more potent than sumatriptan at the level of both large coronary and internal carotid arteries.

After endothelium removal by balloon angioplasty in coronary arteries, the initial dilatation induced by the triptans was abolished and delayed constriction enhanced.

The selective antagonist for the 5‐HT1B receptors SB224289 dose‐dependently blocked the effects of sumatriptan on large coronary and internal carotid arteries whereas the selective antagonist for the 5‐HT1D receptors BRL15572 did not affect any of these effects.

In conclusion, frovatriptan and sumatriptan initially dilate and subsequently constrict large coronary arteries in the conscious dog, whereas they directly constrict the internal carotid artery. The vascular endothelium modulates the effects of these triptans on large coronary arteries. Finally, 5‐HT1B but not 5‐HT1D receptors are primarily involved in canine coronary and internal carotid vasomotor responses to sumatriptan.

British Journal of Pharmacology (2001) 132, 1071–1083; doi:10.1038/sj.bjp.0703917
Cannabinoid inhibition of the capsaicin‐induced calcium response in rat dorsal root ganglion neurones
Authors: Millns Paul J , Chapman Victoria , Kendall David A
Published: January 2005

Cannabinoids have marked inhibitory effects on somatosensory processing, which may arise from actions at both peripheral and central cannabinoid receptors. Here, the effect of a synthetic cannabinoid agonist HU210 on capsaicin‐evoked responses in adult rat dorsal root ganglion (DRG) neurones was studied. The vanilloid capsaicin produced a concentration‐related increase in intracellular calcium in DRG neurones, which was significantly inhibited by HU210 (1 μM). The cannabinoid CB1 receptor antagonist SR141716A (1 μM) had no effect alone and did not influence the response to capsaicin but significantly reversed the inhibitory effect of HU210. These data indicate that DRG CB1 receptors are functional and can inhibit nociceptive responses.

British Journal of Pharmacology (2001) 132, 969–971; doi:10.1038/sj.bjp.0703919
Xenopus tropicalis oocytes as an advantageous model system for the study of intracellular Ca2+ signalling
Authors: Marchant Jonathan S , Parker Ian
Published: January 2005

The purpose of this study was to compare oocytes from the pipid frogs Xenopus tropicalis and Xenopus laevis, with respect to their utility for studying Ca2+ signalling mechanisms and for expression of heterologous proteins.

We show that X. tropicalis oocytes possess an intracellular Ca2+ store that is mobilized by inositol (1,4,5) trisphosphate (IP3). Ca2+ signalling is activated by endogenous lysophosphatidic acid receptors and cytosolic Ca2+ activates a plasma membrane chloride conductance. The spatiotemporal organization of cytosolic Ca2+ signals, from the microscopic architecture of elementary Ca2+ ‘puffs’ to the macroscopic patterns of Ca2+ spiking are closely similar to the local and global patterns of Ca2+ release previously characterized in oocytes from X. laevis.

By injecting X. tropicalis oocytes with cDNA encoding an ER‐targeted fluorescent protein construct, we demonstrate the capacity of the X. tropicalis oocyte to readily express heterologous proteins. The organization of ER is polarized across the oocyte, with the IP3‐releaseable store targeted within an ∼8 μm wide band that circumscribes the cell.

We conclude that the X. tropicalis oocyte shares many of the characteristics that have made oocytes of X. laevis a favoured system for studying Ca2+ signalling mechanisms. Moreover, X. tropicalis oocytes display further practical advantages in terms of imaging depth, Ca2+ signal magnitude and electrical properties. These further enhance the appeal of X. tropicalis as an experimental system, in addition to its greater amenability to transgenic approaches.

British Journal of Pharmacology (2001) 132, 1396–1410; doi:10.1038/sj.bjp.0703922
Potential anxiolytic‐ and antidepressant‐like effects of MPEP, a potent, selective and systemically active mGlu5 receptor antagonist
Authors: Tatarczyńska Ewa , Kłodzińska Aleksandra , Chojnacka‐Wójcik Ewa , Pałucha Agnieszka , Gasparini Fabrizio , Kuhn Rainer , Pilc Andrzej
Published: January 2005

Several lines of evidence suggest a crucial involvement of glutamate in the mechanism of action of anxiolytic and/or antidepressant drugs. The involvement of group I mGlu receptors in anxiety and depression has also been proposed. Given the recent discovery of a selective and brain penetrable mGlu5 receptor antagonists, the effect of 2‐methyl‐6‐(phenylethynyl)‐pyridine (MPEP), i.e. the most potent compound described, was evaluated in established models of anxiety and depression.

Experiments were performed on male Wistar rats or male Albino Swiss or C57BL/6J mice. The anxiolytic‐like effects of MPEP was tested in the conflict drinking test and the elevated plus‐maze test in rats as well as in the four‐plate test in mice. The antidepressant‐like effect was estimated using the tail suspension test in mice and the behavioural despair test in rats.

MPEP (1 – 30 mg kg−1) induced anxiolytic‐like effects in the conflict drinking test and the elevated plus‐maze test in rats as well as in the four‐plate test in mice. MPEP had no effect on locomotor activity or motor coordination. MPEP (1 – 20 mg kg−1) did shorten the immobility time in a tail suspension test in mice, however it was inactive in the behavioural despair test in rats.

These data suggest that selective mGlu5 receptor antagonists may play a role in the therapy of anxiety and/or depression, further studies are required to identify the sites and the mechanism of action of MPEP.

British Journal of Pharmacology (2001) 132, 1423–1430; doi:10.1038/sj.bjp.0703923
Effects of anti‐oestrogens and β‐estradiol on calcium uptake by cardiac sarcoplasmic reticulum
Authors: Dodds Marnie L , Kargacin Margaret E , Kargacin Gary J
Published: January 2005

Tamoxifen and a group of structurally similar non‐steroidal, triphenolic compounds inhibit the oestrogen receptor. In addition to this action, these anti‐oestrogens are known to inhibit some types of plasma membrane ion channels and other proteins through mechanisms that do not appear to involve their interactions with the estrogen receptor but could be the result of their effect on membrane lipid structure or fluidity.

We studied the effects of β‐estradiol and three anti‐oestrogens (tamoxifen, 4‐hydroxytamoxifen and clomiphene) on Ca2+ uptake into sarcoplasmic reticulum (SR) vesicles isolated from canine cardiac ventricular tissue.

The antiestrogens all inhibit SR Ca2+ uptake in a concentration‐dependent manner (order of potency: tamoxifen > 4‐hydroxytamoxifen clomiphene). Although these compounds rapidly inhibit net Ca2+ uptake they do not have a similar rapid effect on the ATPase activity of the SR Ca pump. β‐estradiol has no effect on Ca2+ uptake nor does it alter the inhibitory action of tamoxifen on the SR.

The differences in the effects of β‐estradiol and the anti‐oestrogens on cardiac SR Ca2+ uptake do not correlate with differences in the ways in which these compounds have been reported to interact with membrane lipids. Our results are consistent, however, with direct effects on a membrane protein (possibly an SR Cl− or K+ channel).

British Journal of Pharmacology (2001) 132, 1374–1382; doi:10.1038/sj.bjp.0703924
Nerve evoked P2X receptor contractions of rat mesenteric arteries; dependence on vessel size and lack of role of L‐type calcium channels and calcium induced calcium release
Authors: Gitterman D P , Evans R J
Published: January 2005

Contractile responses to short trains of nerve stimulation have been characterized in small, medium and large arteries from the rat mesenteric circulation (5th – 6th, 2nd – 3rd and 1st order, respectively). In addition, sources of calcium for smooth muscle contraction have been investigated.

Nerve stimulation (10 pulses at 10 Hz) evoked reproducible contractions. The P2 receptor antagonist suramin (100 μM) reduced constrictions by 65.3±7.4, 82.7±3.3 and 3.1±6.1% in small, medium and large arteries respectively. The α‐adrenoceptor antagonist prazosin (0.1 μM) reduced responses by 32.6±2.6, 27.0±1.5 and 97.0±1.9% respectively.

The L‐type calcium channel antagonist nifedipine (1 μM) reduced nerve‐evoked contractions by 2.8±3.3, 10.0±3.7 and 13.5±2.7% in small, medium and large arteries respectively. When the adrenergic component of contraction was blocked by prazosin (0.1 μM) nifedipine reduced responses by 4.6±7.9, 14.3±2.0 and 3.0±1.9% respectively.

Contractile responses to exogenous α,β‐meATP were unaffected by the depletion of calcium stores with cyclopiazonic acid (30 μM). This indicates that mobilization of calcium from internal stores is not required for P2X receptor mediated smooth muscle contraction.

We conclude that for neurogenic responses, the P2X receptor mediated component of constriction dominates in small mesenteric arteries (3rd  – 6th order) while in large arteries (1st order) noradrenaline mediates contraction. For P2X receptor mediated responses all the calcium required for smooth muscle contraction enters the cell directly through P2X receptor channels.

British Journal of Pharmacology (2001) 132, 1201–1208; doi:10.1038/sj.bjp.0703925
Pharmacokinetics of pefloxacin and its interaction with cyclosporin A, a P‐glycoprotein modulator, in rat blood, brain and bile, using simultaneous microdialysis
Authors: Tsai Tung‐Hu
Published: January 2005

In vivo microdialysis with HPLC was used to investigate the pharmacokinetics of pefloxacin and its interaction with cyclosporin A. Microdialysis probes were inserted into the jugular vein/right atrium, the striatum and the bile duct of male Sprague‐Dawley rats. Biological fluid sampling thereby allowed the simultaneous determination of pefloxacin levels in blood, brain and bile.

Following pefloxacin administration, the brain‐to‐blood coefficient of distribution was 0.036. This was calculated by dividing the area under the concentration curve (AUC) of pefloxacin in brain by its AUC in blood (k=AUCbrain/AUCblood).

When the P‐glycoprotein cyclosporin A (10 mg kg−1) was co‐administered with pefloxacin (10 mg kg−1), the AUC and the mean residence time in rat blood did not differ significantly (P>0.05). Similarly, the pharmacokinetics of pefloxacin in rat brain was not affected by the presence of cyclosporin A.

The AUC of unbound pefloxacin in bile was significantly greater than that in blood. The disposition of pefloxacin in rat bile shows a slow elimination phase following a peak concentration 30 min after pefloxacin administration (10 mg kg−1, i.v.). The bile‐to‐blood coefficient of distribution (k=AUCbile/AUCblood) was 1.53.

The results indicated that pefloxacin was able to penetrate the blood‐brain barrier and that the concentration in bile was greater than that in the blood, suggesting active biliary excretion of pefloxacin. Current data obtained from rats show no significant impact of cyclosporin A on the pharmacokinetics of pefloxacin in rat blood and brain when administered by concomitant i.v. bolus.

British Journal of Pharmacology (2001) 132, 1310–1316; doi:10.1038/sj.bjp.0703927
Morphological and functional in vitro and in vivo characterization of the mouse corpus cavernosum
Authors: Mizusawa Hiroya , Hedlund Petter , Håkansson Anders , Alm Per , Andersson Karl‐Erik
Published: January 2005

In normal mice, the distribution of adrenergic, cholinergic, some peptidergic, and neuronal nitric oxide synthase (nNOS)‐containing nerves were investigated. Functional in vitro correlates were obtained. An in vivo model was developed in which erectile haemodynamics in response to drugs or nerve‐stimulation were studied.

Immunoreactivities for vesicular acetylcholine transporter protein (VAChT), nNOS‐, and vasoactive intestinal polypeptide (VIP), co‐existed in nerve fibres and terminal varicosities. Immunoreactivities for neuropeptide Y (NPY) and tyrosine hydroxylase (TH) were found in the same nerve structures.

Chemical sympathectomy abolished TH‐ and NPY‐IR nerve structures in cavernous smooth muscle bundles. The distribution of calcitonin gene‐related peptide (CGRP)‐, nNOS‐, VAChT‐ and VIP‐IR nerve structures was unchanged.

In endothelial cells of the central and helicine arteries, veins and venules, intense immunoreactivity for endothelial NOS (eNOS) was observed. No distinct eNOS‐IR cells were found lining the cavernous sinusoids.

In vitro, nerve‐induced relaxations were verified, and endothelial NO/cyclic GMP‐mediated relaxant responses were established. VIP and CGRP had small relaxant effects. A functioning adenylate cyclase/cyclic AMP pathway was confirmed.

Neuronal excitatory responses were abolished by prazosin, or forskolin. VIP and CGRP counteracted contractions, whereas NPY and scopolamine enhanced excitatory responses.

In vivo, erectile responses were significantly attenuated by L‐NAME (50 mg kg−1) and facilitated by sildenafil (200 μg kg−1).

It is concluded that the mouse is a suitable model for studies of erectile mechanisms in vitro and in vivo.

British Journal of Pharmacology (2001) 132, 1333–1341; doi:10.1038/sj.bjp.0703938
[125I]‐(Pyr1)Apelin‐13 is a novel radioligand for localizing the APJ orphan receptor in human and rat tissues with evidence for a vasoconstrictor role in man
Authors: Katugampola Sidath D , Maguire Janet J , Matthewson Stewart R , Davenport Anthony P
Published: January 2005

We have determined the binding characteristics of [125I]‐(Pyr1)Apelin‐13, a putative ligand for the APJ orphan receptor in human cardiovascular and rat tissue and investigated the functional properties of (Pyr1)Apelin‐13 in human saphenous vein.

The binding of [125I]‐(Pyr1)Apelin‐13 to sections of human heart tissue was time dependent and rapid at 23°C. Data were fitted to a single site model with an association rate constant (kobs) of 0.115 min−1. [125I]‐(Pyr1)Apelin‐13 also dissociated from a single site with a dissociation rate constant of 0.0105 min−1.

In saturation binding experiments [125I]‐(Pyr1)Apelin‐13 bound to human left ventricle with a KD value of 0.35±0.08 nM, Bmax of 4.3±0.9 fmol mg−1 protein with a Hill slope of 0.97±0.04 and to the right atria with a KD of 0.33±0.09 nM, Bmax of 3.1±0.6 fmol mg−1 protein and a Hill slope of 0.93±0.05.

[125I]‐(Pyr1)Apelin‐13 binding sites were localized using autoradiography to human cardiovascular tissue, including coronary artery, aorta and saphenous vein grafts. In rat tissue a high density of receptors were localized to the molecular layer of the rat cerebellum, rat lung, rat heart and low levels in the rat kidney cortex.

(Pyr1)Apelin‐13 potently contracted human saphenous vein with a pD2 value of 8.4±0.2 (n=8). The maximum response elicited by the peptide was 22.6±6% of 100 mM KCl.

We provide the first evidence of APJ receptor expression, relative densities and functional properties of (Pyr1)Apelin‐13 in human cardiovascular tissue.

British Journal of Pharmacology (2001) 132, 1255–1260; doi:10.1038/sj.bjp.0703939
Vascular endothelial growth factor and the in vivo increase in plasma extravasation in the hamster cheek pouch
Authors: Félétou Michel , Staczek Joanna , Duhault Jacques
Published: January 2005

The purpose of this study in the hamster cheek pouch was to determine whether or not vascular endothelial growth factor (VEGF) induced changes in plasma extravasation and if so, the mechanism(s) involved.

The cheek pouch microcirculatory bed of the anaesthetized hamster was directly observed under microscope and the number of vascular leakage sites, as shown by fluorescein isothiocyanate (FITC‐dextran, 150 kD) extravasation, was counted. Drugs and VEGF were applied topically. VEGF from 0.05 to 0.5 μg ml−1 (1.2 to 12 nM) produced a dose‐dependent increase in the number of microvascular leakage sites from virtually none in basal conditions to up to 250 in some pouches. The effects of VEGF (0.1 μg ml−1 or 2.4 nM) were blocked in a concentration‐dependent manner by the non‐specific heparin growth factor antagonist TBC‐1635 (0.1, 1 and 3μM). The placenta growth factor (PlGF‐1: 0.1 and 0.5 μg ml−1 or 3.4 and 17 nM) did not increase plasma extravasation, per se, but abolished the effects of VEGF (2.4 nM).

The increases in microvascular leakage produced by VEGF (2.4 nM) were partially but significantly (P<0.05) inhibited by genistein (5 and 10 μM, up to 33% inhibition), LY 294002 (30 μM, 41%), bisindolylmaleimide (1 μM, 65%) and virtually abolished by indomethacin (3 μM, 88%) and L‐nitro‐arginine (10 μM, 95%), these drugs being inhibitors of tyrosine kinase, phosphatidylinositol‐3‐kinase, protein kinase C, cyclo‐oxygenase and nitric oxide synthase respectively. None of these inhibitors, at the concentration tested, induced alone an increase in plasma extravasation.

These results indicate that the VEGF‐induced plasma extravasation may involve the stimulation of VEGF‐R2 (Flk‐1/KDR) and the activation of phosphatidylinositol‐3‐kinase and protein kinase C. The production of both nitric oxide and prostaglandin is required to observe an increase in vascular leakage.

British Journal of Pharmacology (2001) 132, 1342–1348; doi:10.1038/sj.bjp.0703941
Role of nitric oxide synthases in the infarct size‐reducing effect conferred by heat stress in isolated rat hearts
Authors: Arnaud Claire , Laubriet Aline , Joyeux Marie , Godin‐Ribuot Diane , Rochette Luc , Demenge Pierre , Ribuot Christophe
Published: January 2005

Nitric oxide (NO) donors are known to induce both delayed cardioprotection and myocardial heat stress protein (HSP) expression. Moreover, heat stress (HS), which also protects myocardium against ischaemic damages, is associated with a NO release. Therefore, we have investigated the implication of NO in HS‐induced resistance to myocardial infarction, in the isolated rat heart model.

Rats were divided in six groups (n=10 in each group), subjected or not to heat stress (42°C internal temperature, 15 min) and treated or not with nitro‐L‐arginine‐methylester (L‐NAME) a non‐selective inhibitor of NO synthase isoforms, or L‐N6‐(1‐imino‐ethyl)lysine (L‐NIL), a selective inhibitor of the inducible NO synthase. Twenty‐four hours after heat stress, 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 (18.7±1.6%) compared to Sham (33.0±1.7%) hearts. This effect was abolished in L‐NAME‐treated (41.7±3.1% in HS+L‐NAME vs 35.2±3.0% in Sham+L‐NAME) and L‐NIL‐treated (36.1±3.4% in HS+L‐NIL vs 42.1±4.6% in Sham+L‐NIL) groups. Immunohistochemical analysis of myocardial HSP 27 and 72 showed an HS‐induced increase of these proteins, which was not modified by L‐NAME pretreatment.

We conclude that NO synthases, and in particular the inducible isoform, appear to play a role in the heat stress‐induced cardioprotection, independently of HSP 27 and 72 levels. Further investigations are required to elucidate the precise role of HSPs in this adaptive response.

British Journal of Pharmacology (2001) 132, 1845–1851; doi:10.1038/sj.bjp.0703942
Inducible nitric oxide synthase (iNOS) activity promotes ischaemic skin flap survival
Authors: Kane Anthony J , Barker Jane E , Mitchell Geraldine M , Theile David R B , Romero Rosalind , Messina Aurora , Wagh Milind , Fraulin Frankie O G , Morrison Wayne A , Stewart Alastair G
Published: January 2005

We have examined the role of nitric oxide (NO) in a model of functional angiogenesis in which survival of a skin flap depends entirely on angiogenesis to provide an arterial blood supply to maintain tissue viability.

The different effects of nitric oxide synthase (NOS) inhibitors on rat skin flap survival appeared to be explained on the basis of their NOS isoform selectivity. Skin flap survival was decreased by iNOS‐selective (inducible NOS) inhibitors, S‐methyl‐isothiourea, aminoguanidine and aminoethylthiorea; unaffected by the non‐selective inhibitor nitro‐imino‐L‐ornithine; and enhanced by the cNOS (constitutive NOS, that is endothelial NOS (eNOS) and neuronal NOS (nNOS)) inhibitor, nitro‐L‐arginine methyl ester.

Skin flap survival was reduced in mice with targeted disruption of the iNOS gene (iNOS knockout mice), and the administration of nitro‐L‐arginine methyl ester significantly increased flap survival in iNOS knockout mice (P<0.05).

iNOS immunoreactivity was identified in mast cells in the angiogenic region. Immunoreactive vascular endothelial growth factor (VEGF) and basic fibroblast growth factor were also localized to mast cells.

The combination of interferon‐γ and tumour necrosis factor‐α induced NO production and increased VEGF levels in mast cells cultured from bone marrow of wild‐type, but not iNOS KO mice.

The increased tissue survival associated with the capacity for iNOS expression may be related to iNOS‐dependent enhancement of VEGF levels and an ensuing angiogenic response. Our results provide both pharmacological and genetic evidence that iNOS activity promotes survival of ischaemic tissue.

British Journal of Pharmacology (2001) 132, 1631–1638; doi:10.1038/sj.bjp.0703944
Evidence for functionally active protease‐activated receptor‐4 (PAR‐4) in human vascular smooth muscle cells
Authors: Bretschneider E , Kaufmann R , Braun M , Nowak G , Glusa E , Schrör K
Published: January 2005

This study investigates, whether in addition to the protease‐activated receptor‐1 (PAR‐1), PAR‐4 is present in vascular smooth muscle cells (SMC) of the human saphenous vein and whether this receptor is functionally active. PAR‐1 and PAR‐4 are stimulated by thrombin and by the synthetic peptides SFLLRN and GYPGQV, respectively.

mRNAs for both, PAR‐1 and PAR‐4, were detected in the SMC by using reverse transcriptase polymerase chain reaction (RT – PCR).

Treatment of the SMC with GYPGQV (200 μM) resulted in a transient increase in free intracellular calcium. This calcium signal was completely abolished after a preceding challenge with thrombin (10 nM), indicating homologous receptor desensitization.

Stimulation of the SMC with 10 nM thrombin or 200 μM SFLLRN caused a time‐dependent activation of the extracellular signal‐regulated kinases‐1/2 (ERK‐1/2) with a maximum at 5 min. In contrast, 100 nM thrombin as well as 200 μM of GYPGQV induced a prolonged phosphorylation of ERK‐1/2 with a maximum at 60 min. These data suggest that PAR‐1 and PAR‐4 are activated by thrombin at distinct concentrations and with distinct kinetics.

GYPGQV stimulated [3H]‐thymidine incorporation in SMC. At 500 μM, the peptide increased DNA synthesis 2.5 fold above controls. A comparable mitogenic effect was obtained after stimulation of the SMC by 10 nM thrombin or 100 μM SFLLRN, respectively.

These data indicate that a functionally active PAR‐4 is present in SMC and, in addition to PAR‐1, might contribute to thrombin‐induced mitogenesis.

British Journal of Pharmacology (2001) 132, 1441–1446; doi:10.1038/sj.bjp.0703947
A nitric oxide‐dopamine link pathway in organum vasculosum laminae terminalis of rat brain exerts control over blood pressure
Authors: Chang C‐P , Pan S‐P , Lin M‐T
Published: January 2005

Experiments were carried out to explore the possible role played by the nitric oxide (NO) and dopamine (DA) system in the organum vasculosum laminae terminalis (OVLT) of rat brain in arterial pressure regulation.

Intracerebroventricular (ICV) administration of NO donors such as hydroxylamine or sodium nitro‐prusside (SNP) caused an up to 59 mmHg decrease in blood pressure (BP) and a decrease in DA release (measured by nafion coated carbon fibre electrodes in combination with voltammetry) in the OVLT. In contrast, ICV administration of NG‐nitro‐L‐arginine methyl ester (L‐NAME; a constitutive NO synthase inhibitor) or 7‐nitroindazol (a neuronal NO synthase inhibitor) caused an up to 98 mmHg increase in BP and an increase in DA release in the OVLT.

Intra‐OVLT injection of amphetamine (0.1 – 0.3 mg), SKF 38393 (a DA D1 receptor agonist; 0.01 – 0.03 mg), or apomorphine (a DA D2,3 receptor agonist; 0.01 – 0.03 mg) caused an increase in BP. On the other hand, intra‐OVLT injection of SCH23390 (a DA D1 receptor antagonist; 0.005 – 0.020 mg) or haloperidol (0.005 – 0.020 mg) caused a decrease in BP.

The pressor effects induced by intra‐OVLT administration of L‐NAME were attenuated by pretreatment with intra‐OVLT injection of haloperidol, SCF23390, or 6‐hydroxydopamine. In the contrast, the hydroxylamine‐, 8‐Br‐cGMP‐ or SNP‐induced depressor effects were attenuated by pretreatment with intra‐OVLT injection of amphetamine, SKF 38393 or apomorphine.

The data suggest that activation of a NO‐DA link pathway within the OVLT of rat brain exerts control over blood pressure.

British Journal of Pharmacology (2001) 132, 1524–1530; doi:10.1038/sj.bjp.0703948
Glucocorticoids inhibit lipopolysaccharide‐induced up‐regulation of arginase in rat alveolar macrophages
Authors: Klasen Stefanie , Hammermann Rainer , Fuhrmann Margarita , Lindemann Dirk , Beck Karl‐Friedrich , Pfeilschifter Josef , Racké Kurt
Published: January 2005

As arginase by limiting nitric oxide (NO) synthesis may play a role in airway hyperresponsiveness and glucocorticoids are known to induce the expression of arginase I in hepatic cells, glucocorticoid effects on arginase in alveolar macrophages (AMΦ) were studied.

Rat AMΦ were cultured in absence or presence of test substances. Thereafter, nitrite accumulation, arginase activity, and the expression pattern of inducible NO synthase, arginase I and II mRNA (RT – PCR) and proteins (immunoblotting) were determined.

Lipopolyssacharides (LPS, 20 h) caused an about 2 fold increase in arginase activity, whereas interferon‐γ (IFN‐γ), like LPS a strong inducer of NO synthesis, had no effect.

Dexamethasone decreased arginase activity by about 25% and prevented the LPS‐induced increase. Mifepristone (RU‐486) as partial glucocorticoid receptor agonist inhibited LPS‐induced increase by 45% and antagonized the inhibitory effect of dexamethasone.

Two different inhibitors of the NF‐κB‐pathway also prevented LPS‐induced increase in arginase activity.

Rat AMΦ expressed mRNA and protein of arginase I and II, but arginase I expression was stronger. Arginase I mRNA and protein was not affected by IFN‐γ, but increased by LPS and this effect was prevented by dexamethasone. Both, LPS and IFN‐γ enhanced the levels of arginase II mRNA and protein, effects also inhibited by dexamethasone. As IFN‐γ did not affect total arginase activity, arginase II may represent only a minor fraction of total arginase activity.

In rat AMΦ glucocorticoids inhibit LPS‐induced up‐regulation of arginase activity, an effect which may contribute to the beneficial effects of glucocorticoids in the treatment of inflammatory airway diseases.

British Journal of Pharmacology (2001) 132, 1349–1357; doi:10.1038/sj.bjp.0703951
SB‐334867‐A: the first selective orexin‐1 receptor antagonist
Authors: Smart D , Sabido‐David C , Brough S J , Jewitt F , Johns A , Porter R A , Jerman J C
Published: January 2005

The pharmacology of various peptide and non‐peptide ligands was studied in Chinese hamster ovary (CHO) cells stably expressing human orexin‐1 (OX1) or orexin‐2 (OX2) receptors by measuring intracellular calcium ([Ca2+]i) using Fluo‐3AM. Orexin‐A and orexin‐B increased [Ca2+]i in CHO‐OX1 (pEC50=8.38±0.04 and 7.26±0.05 respectively, n=12) and CHO‐OX2 (pEC50=8.20±0.03 and 8.26±0.04 respectively, n=8) cells. However, neuropeptide Y and secretin (10 pM – 10 μM) displayed neither agonist nor antagonist properties in either cell‐line. SB‐334867‐A (1‐(2‐Methyylbenzoxanzol‐6‐yl)‐3‐[1,5]naphthyridin‐4‐yl‐urea hydrochloride) inhibited the orexin‐A (10 nM) and orexin‐B (100 nM)‐induced calcium responses (pKB=7.27±0.04 and 7.23±0.03 respectively, n=8), but had no effect on the UTP (3 μM)‐induced calcium response in CHO‐OX1 cells. SB‐334867‐A (10 μM) also inhibited OX2 mediated calcium responses (32.7±1.9% versus orexin‐A). SB‐334867‐A was devoid of agonist properties in either cell‐line. In conclusion, SB‐334867‐A is a non‐peptide OX1 selective receptor antagonist.

British Journal of Pharmacology (2001) 132, 1179–1182; doi:10.1038/sj.bjp.0703953