bluuebunny/crossref_metadata_2025 huggingface dataset:
Evidence for a 5‐HT3 receptor involvement in the facilitation of peristalsis on mucosal application of 5‐HT in the guinea pig isolated ileum
Authors: Tuladhar B. R. , Kaisar M. , Naylor R. J.
Published: January 2005

The 5‐HT receptor involved in the effect of mucosal application of 5‐HT to facilitate peristalsis was investigated in the isolated guinea pig ileum.

An application of 5‐HT (3–100 μM) to the mucosal surface (by inclusion of 5‐HT in the Krebs‐Henseleit solution passing through the lumen of the ileum) caused a concentration related facilitation of peristalsis characterized by a reduction in the peristaltic threshold.

Peristalsis was not modified by methiothepine (0.1 μM), ritanserin (0.1 μM), ondansetron (5 μM), granisetron (1 μM) or SB 204070 (0.1 μM) administered alone to the mucosal surface.

The concentration–response curve to mucosally applied 5‐HT was not altered by the mucosally applied 5‐HT1/2 receptor antagonist methiothepine (0.1 μM), the 5‐HT2 receptor antagonist ritanserin (0.1 μM) or the 5‐HT4 receptor antagonist SB 204070 (0.1 μM). However, the mucosally applied 5‐HT3 receptor antagonists ondansetron (5 μM) and granisetron (1 μM) shifted the response curves to mucosally applied 5‐HT to the right in a parallel and surmountable manner. The pD2 values in the absence and presence of ondansetron were 5.42±0.07 and 4.12±0.10, respectively, (n=6) and that of granisetron were 5.45±0.12 and 4.50±0.10 respectively, (n=5).

Serosally applied ondansetron (5 μM) or granisetron (1 μM) had no effect on the concentration–response curve to mucosally applied 5‐HT. However, the serosally applied ondansetron and granisetron antagonised the facilitatory effect of serosally applied 5‐HT (10 μM) when administered in the presence of serosally applied SB 204070 (0.1 μM).

It is concluded that the facilitatory effect of mucosally applied 5‐HT to reduce the peristaltic threshold in the guinea pig ileum is mediated via a 5‐HT3 receptor located on the mucosal and not the serosal side of the ileum.

British Journal of Pharmacology (1997) 122, 1174–1178; doi:10.1038/sj.bjp.0701503
ZM241385 is an antagonist of the facilitatory responses produced by the A2A adenosine receptor agonists CGS21680 and HENECA in the rat hippocampus
Authors: Cunha Rodrigo A. , Dolores Constantino M. , Alexandre Ribeiro J.
Published: January 2005

In the present study, we investigated the ability of a recently introduced non‐xanthine A2A receptor antagonist, ZM241385 (4‐(2‐[7‐amino‐2‐(2‐furyl{1,2,4}‐triazolo{2,3‐a{1,3,5}triazin‐5‐yl‐aminoethyl)phenol) to displace binding of the prototypical A2A adenosine receptor agonist [3H]CGS21680 (2‐[4‐(2‐p‐carboxyethyl)phenylamino]‐5′‐N‐ethylcarboxamidoadenosine) and to modify the facilitatory responses caused by the A2A selective agonists, CGS21680 and HENECA (2‐hexynl‐5′‐N‐ethylcarboxamidoadenosine) in rat hippocampal preparations.

ZM241385 was nearly equipotent to displace [3H]CGS21680 (30 nM) binding to hippocampal (Ki of 0.52 nM) and to striatal membranes (Ki of 0.35 nM), whereas HENECA was a more potent displacer of [3H]CGS21680 binding to striatal (Ki of 4.5 nM) than to hippocampal membranes (Ki of 19 nM).

HENECA (3–30 nM) was equipotent with CGS21680 to facilitate veratridine‐evoked [3H]acetylcholine release from superfused hippocampal synaptosomes and ZM241385 (20 nM) inhibited the facilitatory effects of both HENECA (30 nM) and CGS21680 (30 nM); this antagonism was mimicked by CSC (250 nM).

In contrast, CGS21680 (10–30 nM) was more potent than HENECA (10–30 nM) to facilitate synaptic transmission in Schaffer fibres/CA1 pyramid synapses of hippocampal slices and the facilitatory effect of CGS21680 (10 nM) was blocked by ZM241385 (20 nM) whereas CSC (250 nM) caused a 40% attenuation of this CGS21680‐induced facilitation.

These results indicate that ZM241385 is the first A2A antagonist with equal potency to displace [3H]CGS21680 binding to striatal and limbic regions, and with general efficiency to antagonize HENECA‐ or CGS21680‐mediated facilitatory responses in the hippocampus.
Cardiovascular effects of CPU‐23, a novel L‐type calcium channel blocker with a unique molecular structure
Authors: Dong Hui , Earle Mary L. , Jiang Yanfen , Loutzenhiser Kathy A. , Triggle Christopher R.
Published: January 2005

The cardiovascular effects of CPU‐23 (1‐{1‐[(6‐methoxy)‐naphth‐2‐yl]}‐ethyl‐2‐(1‐piperidinyl)‐acetyl‐6,7‐dimethoxy‐1,2,3,4‐tetrahydroisoquinoline), a cleavage product of tetrandrine, were investigated using the whole cell perforated patch‐clamp technique, in vitro tension measurements and in vivo haemodynamic recordings.

CPU‐23 (1 and 10 μM) dose‐dependently reduced concentration–response curves for KCl and phenylephrine (PE) in the rat tail artery; inhibition of KCl‐induced contraction was much more potent than for PE. At the same concentrations, CPU‐23 inhibited the inward Ba2+ currents in single smooth muscle cells isolated from the rat tail artery, while CPU‐23 (10 μM) produced 95% vasorelaxation of the rat middle cerebral artery preconstricted with BayK 8644.

CPU‐23 (10 and 30 μM) inhibited the noradrenaline‐induced phasic contraction of the rat tail artery in the absence of extracellular Ca2+ from 40% of control to 23% and 14%, respectively (P<0.01) and tonic contraction of the artery after addition of Ca2+ (2 mM) from 100% of control to 83% and 75%, respectively (P<0.01). In the presence of extracellular Ca2+ the PE‐induced contraction was reduced by CPU‐23 (30 and 100 μM) to 27% and 37%, respectively.

The haemodynamic profile of CPU‐23 in the rat was very similar to diltiazem. At 5 mg kg−1 CPU‐23 induced a rapid onset and long‐lasting decrease in left ventricular systolic pressure (LVSP), maximal velocity of pressure increase (dP/dtmax), systolic blood pressure (SBP), diastolic blood pressure (DBP) and heart rate (HR). When haemodynamic actions of CPU‐23, verapamil, diltiazem and nifedipine were compared at equidepressor doses, the order of potency for reducing LVSP and dP/dtmax was verapamil > CPU‐23 = diltiazem > nifedipine and the order of potency for decreasing HR was verapamil = CPU‐23 = diltiazem > nifedipine.

These data indicate that CPU‐23 is a novel calcium channel blocker with unique molecular structure, which exerts antihypertensive and cardiac depressant effects due primarily to its action on L‐type voltage‐gated calcium channels.
Characterization of 5‐HT receptors on human pulmonary artery and vein: functional and binding studies
Authors: Cortijo Julio , Martí‐Cabrera Miguel , Bernabeu Eva , Domènech Teresa , Bou Josep , Fernández Andrés G. , Beleta Jorge , Palacios José M. , Morcillo Esteban J.
Published: January 2005

This study aimed to investigate the 5‐hydroxytryptamine (5‐HT) receptors mediating contraction of ring preparations isolated from human pulmonary arteries and veins. In functional studies, the responses to 5‐HT, sumatriptan, ergotamine, serotonin‐O‐carboxymethyl‐glycyl‐tyrosinamide (SCMGT), α‐methyl 5‐HT (α‐Me) and 2‐methyl 5‐HT (2‐Me) were studied with WAY100635, GR127935, ritanserin, zacopride and SB204070 as antagonists.

All agonists produced concentration‐dependent contractions of human pulmonary artery and vein preparations. The order of potency (−log EC50 values) was ergotamine (6.88)>5‐HT (6.41)SCMGT (6.20)=sumatriptan (6.19) α‐Me (6.04) in the artery, and ergotamine (7.84)>5‐HT (6.96)>sumatriptan (6.60)=α‐Me (6.56)>SCMGT (6.09) in the vein. The potency of each agonist, except for SCMGT, was greater in vein than in artery preparations. Contractile responses to 5‐HT were similar in intact and endothelium‐denuded preparations but responses to sumatriptan were enhanced in artery rings without endothelium.

GR127935 (1 nM to 0.5 μM) produced an unsurmountable antagonism of the response to 5‐HT, sumatriptan, ergotamine and SCMGT. Ritanserin (1 nM to 1 μM) also reduced the maximum contractile responses to 5‐HT, ergotamine and α‐Me in artery and vein preparations without affecting those to sumatriptan and SCMGT. In endothelium‐denuded preparations, surmountable antagonism of sumatriptan by GR127935 (in the presence of ritanserin) and of α‐Me by ritanserin (in the presence of GR127935) allowed for the calculation of the apparent pKB values of GR127935 (9.17±0.11 in artery and 9.11±0.05 in vein) and ritanserin (8.82±0.09 in artery and 8.98±0.12 in vein).

WAY100635 (1 nM to 1 μM), zacopride (1 nM to 1 μM), or SB204070 (1 nM) did not significantly alter the concentration‐response curves for 5‐HT, sumatriptan, ergotamine, SCMGT or 2‐Me in human pulmonary artery or vein thus indicating that 5‐HT1A, 5‐HT3 and 5‐HT4 receptors are presumably not involved in the contractile response to these agonists.

Binding studies using selective radioligands for different 5‐HT receptors could not detect the presence of 5‐HT1A receptor binding in human pulmonary blood vessels whereas the 5‐HT1B/1D radioligand [3H]‐5‐CT significantly labelled a population of specific binding sites in both vessel types. The presence of 5‐HT2A receptors could also be inferred from the level of binding of [3H]‐ketanserin to membranes obtained from human pulmonary vessels, although significance could not be reached for arteries. 5‐HT4 specific receptor binding was scarce in veins and absent in the case of arteries.

These findings indicate that the human pulmonary artery and vein have a mixed functional population of 5‐HT1B/1D and 5‐HT2A receptors mediating the contractile response to 5‐HT which is consistent with results of the binding studies.
Tumour necrosis factor‐α expression and cell recruitment in Sephadex particle‐induced lung inflammation: effects of dexamethasone and cyclosporin A
Authors: Williams Cara M. M. , Smith Lance , Flanagan Brian F. , Steve Clegg L. , Coleman John W.
Published: January 2005

Tumour necrosis factor‐α (TNF‐α) is a cytokine with diverse properties consistent with a possible role in inflammatory disease. We investigated whether TNF‐α is induced during the progression of lung inflammation elicited by a particulate non‐antigenic stimulus, and whether pharmacological control of TNF‐α expression influences recruitment of specific inflammatory cell types.

A single intravenous injection of Sephadex particles into rats led to extensive granulomatous inflammation in lung alveolar and bronchial tissue that peaked in intensity after 24–72 h. Mononuclear cells were the principal component of granulomas, but neutrophils and eosinophils were also abundant. Numbers of mononuclear cells, neutrophils and eosinophils recovered by bronchoalveolar lavage (BAL) peaked at 72 h, 48 h and 72 h, respectively.

Messenger RNA encoding TNF‐α was induced in lung epithelial cells, lung granulomas and BAL cells 6 h after Sephadex administration and remained elevated for 72 h before declining to baseline by 7 days. In BAL cell populations TNF‐α protein was localized to mononuclear cells at all times points pre‐ and post‐Sephadex administration.

Treatment of rats with dexamethasone significantly reduced the Sephadex‐induced recruitment of mononuclear cells, neutrophils and eosinophils into the bronchoalveolar cavity, and significantly reduced TNF‐α mRNA expression by BAL cells.

Treatment of rats with cyclosporin A was without effect on Sephadex‐induced elevations of mononuclear cell numbers and expression of TNF‐α, but did reduce significantly recruitment of neutrophils and eosinophils to BAL cell populations.

These results show that a sequential asthma‐like recruitment of neutrophils, eosinophils and mononuclear cells into lung tissue can be induced by single exposure to a non‐antigenic stimulus. Pharmacological and histological studies reveal that mononuclear cell mobilization relates closely to induced TNF‐α expression, whereas mobilization of neutrophils and eosinophils appears secondary to expression of the cytokine.

British Journal of Pharmacology (1997) 122, 1127–1134; doi:10.1038/sj.bjp.0701515
Effect of mycophenolic acid on TNFα‐induced expression of cell adhesion molecules in human venous endothelial cells in vitro
Authors: Hauser Ingeborg A. , Johnson David R. , Thévenod Frank , Goppelt‐Strübe Margarete
Published: January 2005

Mycophenolic acid (MPA) is an inhibitor of inosine‐5′‐monophosphate dehydrogenase and therefore interferes with cellular GTP biosynthesis. Recently, MPA has been used as an antiproliferative and immunosuppressive agent. In the present study, the effect of MPA on the expression of the endothelial cell adhesion molecules (CAMs), intercellular (I) CAM‐1, vascular (V) CAM‐1 and endothelial (E)‐selectin, was investigated in tumour necrosis factor‐α (TNFα)‐activated cultured human venous endothelial cells (EC).

Surface expression of CAMs was measured by flow cytometry and mRNA expression by Northern blot analysis. Transcriptional activation of CAMs by the nuclear factor NF‐κB was determined by an electromobility shift assay. The function of CAMs was studied by a static adhesion assay with human monocyte‐like undifferentiated U937 cells.

Pretreatment of TNFα‐ (5  ng ml−1, 12 h) activated EC with MPA (10 μM, 24 h) increased the binding of U937 cells, which had not been treated with MPA, by ∼amp;2 fold. MPA‐pretreatment of EC did not affect TNFα‐induced surface expression of ICAM‐1. However, VCAM‐1 and E‐selectin were increased 2–3 fold and remained elevated up to 24 h, by which time TNFα‐activated control EC had returned to baseline levels of expression. The effect of MPA on the surface expression of CAMs was half‐maximal at ∼amp;1 μM and required 12 h of pretreatment. Guanosine (0.3 mM), a precursor of GTP, did not prevent the effect of MPA on the expression of CAMs in TNFα‐activated EC.

Kinetics of mRNA expression of CAMs mirrored protein expression: mRNA for ICAM‐1 was unaffected, whereas TNFα‐induced mRNA expression for E‐selectin and VCAM‐1 was prolonged and increased by MPA. This effect was not due to increased transcription mediated by the nuclear transcription factor NF‐κB. However, half‐life for E‐selectin mRNA was increased 10 fold by MPA, whereas ICAM‐1 mRNA half‐life was unchanged.

The data demonstrate that apart from its antiproliferative effects on lymphocytes, MPA enhances TNFα‐induced VCAM‐1 and E‐selectin surface expression on EC by selectively increasing the mRNA‐stability of these cell adhesion molecules. This effect of MPA on EC appears to be independent from inhibition of inosine‐5′‐monophosphate dehydrogenase.
The regulation of mitogenesis and apoptosis in response to the persistent stimulation of α1‐adrenoceptors: a possible role of 15‐lipoxygenase
Authors: Nishio Eisuke , Watanabe Yasuhiro
Published: January 2005

Activation of α1‐adrenoceptor stimulation regulates eicosanoid metabolism and growth in vascular smooth muscle cells (VSMCs). The purpose of this study was to investigate the functional implications of lipoxygenase pathway in α1‐adrenoceptor‐stimulated VSMCs growth through mutually exclusive biological functions, that is cell proliferation and cell death.

Phenylephrine (10 μM), a specific α1‐adrenoceptor agonist, enhanced [3H]‐thymidine incorporation by 300% above basal. Nordihydroguaiaretic acid (NDGA), a lipoxygenase inhibitor, caused 36 and 50% decrease in phenylephrine (10 μM)‐stimulated [3H]‐thymidine incorporation at concentrations of 1 μM and 10 μM respectively.

Inversely, treatment of phenylephrine (10 μM)‐stimulated VSMCs with NDGA induced DNA fragmentation in a dose‐dependent fashion. The level of induction of DNA fragmentation by NDGA was 225, 319 and 406% above the phenylephrine (10 μM)‐level at concentrations of 0.1 μM, 1 μM and 10 μM, respectively. This induction of DNA fragmentation was partially prevented by exogenous 15‐hydroxyeicosatetraenoic acid (15‐HETE). The inhibition of apoptosis was 53 and 63% at concentrations of 5 μM and 10 μM of 15HETE, respectively, as compared with phenylephrine (10 μM) in the presence of NDGA (10 μM).

Furthermore, we performed the time‐course analysis of Bcl‐2 protein expression in phenylephrine (10 μM)‐stimulated VSMCs. The expression of Bcl‐2 protein disappeared after a 2 h incubation in the presence of NDGA (10 μM), but remained stable after a 2 h incubation period in the absence of NDGA (10 μM).

These results suggest that the lipoxygenase pathway is involved in cell proliferation by preventing apoptosis through the level of Bcl‐2 protein expression.
Influence of olfactory bulbectomy and subsequent imipramine treatment on 5‐hydroxytryptaminergic presynapses in the rat frontal cortex: behavioural correlates
Authors: Grecksch Gisela , Zhou Dan , Franke Christian , Schröder Ulrike , Sabel Bernhard , Becker Axel , Huether Gerald
Published: January 2005

Alterations of 5‐hydroxytryptaminergic mechanisms are thought to play a special role in the pathogenesis of depression and antidepressant treatments are assumed to restore these changes.

We have used one of the most reliable models of depression, the olfactory bulbectomized rat to study the long term consequences of this manipulation and of subchronic imipramine treatment on two parameters of 5‐hydroxytryptaminergic presynapses, 5‐hydroxytryptamine (5‐HT) transporter density and tryptophan hydroxylase apoenzyme concentration, in the frontal cortex as well as on active avoidance learning several weeks after bulbectomy.

The Bmax value of [3H]‐paroxetine binding and the concentration of the 5‐HT synthesizing enzyme were both significantly elevated in the frontal cortex of bulbectomized rats compared to sham‐operated controls.

Imipramine treatment, either by daily injections or by subcutaneous implantation of slow release imipramine‐containing polymers reduced the elevated tryptophan hydroxylase apoenzyme levels in the frontal cortex of bulbectomized, but not of sham‐operated control rats and restored the deficient learning performance of bulbectomized rats.

Both effects were more pronounced after continuous drug administration by imipramine‐releasing polymers compared to daily i.p. injections.

These findings indicate that bulbectomy leads to a compensatory 5‐hydroxytryptaminergic hyperinnervation of the frontal cortex. Chronic antidepressant treatment seems to attenuate the increased output of the 5‐hydroxytryptaminergic projections in the frontal cortex through the destabilization of the rate limiting enzyme of 5‐HT synthesis of the 5‐hydroxytryptaminergic nerve endings in this brain region.
Analogues of γ‐aminobutyric acid (GABA) and trans‐4‐aminocrotonic acid (TACA) substituted in the 2 position as GABAC receptor antagonists
Authors: Chebib Mary , Vandenberg Robert J. , Johnston Graham A. R.
Published: January 2005

γ‐Aminobutyric acid (GABA) and trans‐4‐aminocrotonic acid (TACA) have been shown to activate GABAC receptors. In this study, a range of C2, C3, C4 and N‐substituted GABA and TACA analogues were examined for activity at GABAC receptors.

The effects of these compounds were examined by use of electrophysiological recording from Xenopus oocytes expressing the human ρ1 subunit of GABAC receptors with the two‐electrode voltage‐clamp method.

trans‐4‐Amino‐2‐fluorobut‐2‐enoic acid was found to be a potent agonist (KD=2.43 μm). In contrast, trans‐4‐amino‐2‐methylbut‐2‐enoic acid was found to be a moderately potent antagonist (IC50=31.0 μm and KB=45.5 μm). These observations highlight the possibility that subtle structural substitutions may change an agonist into an antagonist.

4‐Amino‐2‐methylbutanoic acid (KD=189 μm), 4‐amino‐2‐methylenebutanoic acid (KD=182 μm) and 4‐amino‐2‐chlorobutanoic acid (KD=285 μm) were weak partial agonists. The intrinsic activities of these compounds were 12.1%, 4.4% and 5.2% of the maximal response of GABA, respectively. These compounds more effectively blocked the effects of the agonist, GABA, giving rise to KB values of 53 μm and 101 μm, respectively.

The sulphinic acid analogue of GABA, homohypotaurine, was found to be a potent partial agonist (KD=4.59 μm, intrinsic activity 69%).

It was concluded that substitution of a methyl or a halo group in the C2 position of GABA or TACA is tolerated at GABAC receptors. However, there was dramatic loss of activity when these groups were substituted at the C3, C4 and nitrogen positions of GABA and TACA.

Molecular modelling studies on a range of active and inactive compounds indicated that the agonist/competitive antagonist binding site of the GABAC receptor may be smaller than that of the GABAA and GABAB receptors. It is suggested that only compounds that can attain relatively flat conformations may bind to the GABAC receptor agonist/competitive antagonist binding site.
Human vascular kinin receptors of the B2 type characterized by radioligand binding
Authors: Gessi S. , Rizzi A. , Calò G. , Agnello G. , Jorizzo G. , Mollica G. , Borea P. A. , Regoli D.
Published: January 2005

The human umbilical vein responds to bradykinin (BK) with contractions that are mediated by B2 receptors. In the present study, the corresponding vascular smooth muscle B2 binding sites have been investigated.

[3H]‐BK, a full agonist labelled ligand, was used to demonstrate a single binding site giving a Kd value of 0.51±0.02 nM and a Bmax of 24±1 fmol mg−1 protein. Scatchard plots were linear (r=0.98) in the 0.05–5 nM range of concentrations. Non‐specific binding was found to be 30% of total binding.

Competition binding curves gave the following order of potency for various B2 receptor agonists: BK‐[Hyp3]‐BKLys‐BK>>[Aib7]‐BK>>>[desArg9]‐BK, which is typical of B2 receptors. There was no binding to B1 receptors since the selective B1 receptor ligand, Lys‐[desArg9]BK was inactive up to 10 μM (n=4).

Characterization of the binding site with antagonists, performed with three chemically distinct series of peptide and non‐peptide compounds, revealed a high affinity of Hoe 140 (D‐Arg‐[Hyp3,Thi5,D‐Tic7,Oic8]‐BK) (Ki 0.17 nM; n=4) which was more potent that FR 173657 ([(E)‐3‐(6‐acetamido‐3‐pyridyl)‐N‐[N‐[2,4‐dichloro‐3‐[(2‐methyl‐8‐quinolinyl)oxymethyl]phenyl]‐N‐methylaminocarbonylmethyl] acrylamide]) (Ki 1.94 nM; n=4), D‐Arg‐[Hyp3,D‐Phe7,Leu8]‐BK (Ki 256 nM; n=4) and Win 64338 (phosphonium, [[4[[2[[bis(cyclohexylamino)methylene]amino]‐3‐(2‐naphthalenyl)‐1‐oxopropyl]amino]phenyl]methyl]tributyl, chloride, monohydrochloride) (Ki 1,450 nM; n=4).

The present study describes and characterises B2 receptor binding sites in the vascular smooth muscle of the human umbilical vein. The binding assay appears to be suitable for studying new agonists or antagonists designed to activate or block the B2 receptor class that mediate the majority of the physiopathological effects of kinins in man.
Glucose‐lowering effect of BTS 67 582
Authors: Page T. , Bailey C. J.
Published: January 2005

The hypoglycaemic effect of BTS 67 582 (1,1‐dimethyl‐2(2‐morpholinophenyl) guanidine fumarate) was studied in normal rats.

BTS 67 582 (100 mg kg−1, p.o.) acutely lowered basal plasma glucose concentrations: onset within 1 h, maximum decrease of >40% at 2–3 h, and partial return to euglycaemia by 5 h. Plasma insulin concentrations were increased: onset within 30 min, maximum increase 3 fold at 1–2 h; returning to normal by 5 h.

BTS 67 582 (100 mg kg−1) increased (by 56%) the rate of disappearance of plasma glucose during an intravenous glucose tolerance test, accompanied by a 51% increase in insulin concentrations.

During hyperglycaemic clamp studies BTS 67 582 (100 mg kg−1) increased glucose utilization 3 fold. This was associated with a 3 fold increase in insulin concentrations, even in the presence of adrenaline at a dosage which inhibits glucose‐induced insulin release.

When the insulin‐releasing effect of BTS 67 582 (100 mg kg−1) was inhibited by infusion of somatostatin, there was no effect on glycaemia.

Insulin‐dependent diabetic BB/S rats, which do not produce endogenous insulin, showed no effect of BTS 67 582 (100 mg kg−1) on plasma glucose concentrations in the presence or absence of exogenous insulin.

The results demonstrate an acute hypoglycaemic effect of BTS 67 582 which appears to result mainly from its potent insulin‐releasing action.
Spinal pharmacology of tactile allodynia in diabetic rats
Authors: Calcutt Nigel A. , Chaplan Sandra R.
Published: January 2005

Rats develop tactile allodynia to stimulation of the plantar surface of the hindpaw with von Frey filaments within days of the onset of streptozotocin‐induced diabetes. This is prevented by insulin and alleviated by systemic lignocaine, but the aetiology is unknown.

Using indwelling lumbar intrathecal catheters to deliver pharmacological agents, we have investigated whether tactile allodynia in streptozotocin‐diabetic rats is dependent on mechanisms associated with spinal sensitization, by assessing the efficacy of agents that inhibit specific components of spinal nociceptive processing.

Dose‐dependent inhibition of tactile allodynia in diabetic rats was noted with the N‐type calcium channel antagonist SNX 239, the α2‐adrenoceptor agonist dexmedetomidine, the μ‐opioid receptor agonist morphine, the N‐methyl‐D‐aspartate (NMDA) receptor antagonist AP5 and the non‐NMDA receptor antagonist NBQX.

No effect on tactile allodynia was noted after intrathecal administration of the nitric oxide synthase inhibitor NG‐nitro‐L‐arginine methyl ester (L‐NAME), the cyclo‐oxygenase inhibitor ketorolac, the L‐type calcium channel inhibitor diltiazem or any vehicle.

These data suggest that the tactile allodynia of diabetic rats involves spinal glutamatergic pathways but is not associated with spinal release of nitric oxide or prostaglandins.
Cortistatin increase of a potassium conductance in rat locus coeruleus in vitro
Authors: Connor Mark , Ingram Susan L. , Christie MacDonald J.
Published: January 2005

In this study we examined the effects of cortistatin, a putative endogenous ligand for somatostatin (SRIF) receptors, on the membrane properties of rat locus coeruleus (LC) neurones in vitro, by use of intracellular and whole cell patch clamp recording. We have compared the actions of cortistatin with those of SRIF and the SRIF analogue d‐Phe‐Cys‐Tyr‐d‐Trp‐Orn‐Thr‐Pen‐Thr‐NH2 (CTOP).

When LC neurones were voltage clamped to −60 mV, application of cortistatin caused an outward current in all cells examined (n=44), with a pEC50 of 6.62. SRIF also caused an outward current in all cells examined (n=43), with a pEC50 of 6.93.

The outward currents caused by cortistatin in 2.5 mm extracellular K+ reversed polarity at −106 mV, very close to the predicted K+ reversal potential of −105 mV. Increasing extracellular K+ to 10.5 mm resulted in a shift of the reversal potential of +38 mV, a shift consistent with a K+ conductance. The conductance activated by cortistatin showed mild inward rectification.

Continuous application of a high concentration of SRIF (1 μm) resulted in a decrease of the outward current to a steady level of 49% of the maximum response, with a t1/2 of 131 s. Application of a high concentration of cortistatin (3 μm) during the desensitized portion of the SRIF response did not result in any further outward current. Continuous application of a high concentration of cortistatin (10 μm) resulted in a decrease of the outward current to a steady level of 42% of the maximum response with a t1/2 of 114 s. Application of a high concentration of SRIF (3 μm) during the desensitized portion of the cortistatin response produced only a small outward current.

Continuous application of cortistatin (3 μm) also resulted in a decrease of the outward current (by 43%, t1/2 of 136 s) and application of a high concentration of CTOP (10 μm) during the desensitized portion of the cortistatin response did not produce any outward current. Continuous application of a high concentration of CTOP (10 μm) resulted in a decrease of the outward current to a steady level of 70% of the maximum response with a t1/2 of 143 s. Application of a high concentration of cortistatin (3 μm) during the desensitized portion of the CTOP response did not result in any further outward current.

The actions of cortistatin (300 nm–10 μm) were not affected by the opioid antagonist naloxone (10 μm). Application of met‐enkephalin during the desensitized portion of the response to a high concentration of cortistatin (3 μm) produced an outward current similar to that produced by met‐enkephalin application alone.

Thus cortistatin efficaciously activates an inwardly rectifying K+ conductance in LC neurones. These actions appear to be mediated by a population of SRIF receptors, at which CTOP is also an agonist. Cortistatin does not appear to be a ligand for μ‐opioid receptors in rat LC neurons.
UTP induces vascular responses in the isolated and perfused canine epicardial coronary artery via UTP‐preferring P2Y receptors
Authors: Matsumoto Takako , Nakane Tokio , Chiba Shigetoshi
Published: January 2005

Vasoconstrictor responses of the isolated and perfused canine epicardial coronary artery to uridine 5′‐triphosphate (UTP) were analysed pharmacologically.

At basal perfusion pressure, UTP induced vasoconstriction in a dose‐related manner and the vasoconstriction was sometimes followed by a slight vasodilatation at large doses (more than 10 nmol). The rank order of potency for vasoconstriction was UTP=UDP>ATP>TTPITP>> UMP. At raised perfusion pressure by 20 mm KCl, the vasoconstriction was not changed and a small vasodilatation was induced at large doses. The rank order of potency for vasodilatation was induced at large doses. The rank order of potency for vasodilatation was ATP>>ITPUDP>UTPTTP. The maximal vasodilator response to UTP was much less than that to ATP. UMP did not induce vasodilatation.

The P2X receptor agonist and desensitizing agent α,β‐methylene ATP (1 μm) and the P2 receptor antagonist suramin (100 μm) inhibited the vasoconstrictor responses to ATP but not those to UTP and UDP. The P2 receptor antagonist reactive blue 2 (30 μm) did not inhibit the vascular responses to UTP.

UTP (200 μm) desensitized the vasoconstrictor responses to UTP, but not either the vasodilator responses to UTP or the vasoconstrictor responses to ATP and UDP. UDP (200 μm) did not desensitize the vascular responses to UTP.

Preincubating the UDP stock solution and arterial preparation with hexokinase (10 and 1 uml−1, respectively) did not change the vasoconstrictor responses to UDP.

The Ca channel blocker diltiazem (1 μm) inhibited the vasoconstrictor responses to UTP but not those to ATP and UDP. Incubation in a Ca2+‐free solution containing 1 mm EGTA inhibited the vascular responses to ATP, UTP and UDP.

Removal of the endothelium by an intraluminal injection of saponin (1 mg) inhibited the vasodilator responses to UTP. Indomethacin, a cyclo‐oxygenase inhibitor (1 μm), inhibited the vasodilator responses to UTP, but NG‐nitro‐l‐arginine, a nitric oxide synthase inhibitor (300 μm), did not have an inhibitory effect.

The results suggest that (1) UTP induces vasoconstriction via UTP‐preferring P2Y receptors on the smooth muscle and vasodilatation via receptors different from those mediating the vasoconstriction induced by UTP and mediating the vasodilatation by ATP on the endothelium, through mainly the release of prostacyclin in the canine epicardial coronary artery; (2) UDP induces vasoconstriction via UDP‐preferring P2Y receptors; and (3) L‐type Ca ion channels are involved in the vasoconstriction induced by UTP, but not in that induced by UDP.
Risperidone inhibits 5‐hydroxytryptaminergic neuronal activity in the dorsal raphe nucleus by local release of 5‐hydroxytryptamine
Authors: Hertel Peter , Nomikos George G. , Svensson Torgny H.
Published: January 2005

The effects of risperidone on brain 5‐hydroxytryptamine (5‐HT) neuronal functions were investigated and compared with other antipsychotic drugs and selective receptor antagonists by use of single cell recording and microdialysis in the dorsal raphe nucleus (DRN).

Administration of risperidone (25–400 μg kg−1, i.v.) dose‐dependently decreased 5‐HT cell firing in the DRN, similar to the antipsychotic drug clozapine (0.25–4.0 mg kg−1, i.v.), the putative antipsychotic drug amperozide (0.5–8.0 mg kg−1, i.v.) and the selective α1‐adrenoceptor antagonist prazosin (50–400 μg kg−1, i.v.).

The selective α2‐adrenoceptor antagonist idazoxan (10–80 μg kg−1, i.v.), in contrast, increased the firing rate of 5‐HT neurones in the DRN, whereas the D2 and 5‐HT2A receptor antagonists raclopride (25–200 μg kg−1, i.v.) and MDL 100,907 (50–400 μg kg−1, i.v.), respectively, were without effect. Thus, the α1‐adrenoceptor antagonistic action of the antipsychotic drugs might, at least partly, cause the decrease in DRN 5‐HT cell firing.

Pretreatment with the selective 5‐HT1A receptor antagonist WAY 100,635 (5.0 μg kg−1, i.v.), a drug previously shown to antagonize effectively the inhibition of 5‐HT cells induced by risperidone, failed to prevent the prazosin‐induced decrease in 5‐HT cell firing. This finding argues against the notion that α1‐adrenoceptor antagonism is the sole mechanism underlying the inhibitory effect of risperidone on the DRN cells.

The inhibitory effect of risperidone on 5‐HT cell firing in the DRN was significantly attenuated in rats pretreated with the 5‐HT depletor PCPA (p‐chlorophenylalanine; 300 mg kg−1, i.p., day−1 for 3 consecutive days) in comparison with drug naive animals.

Administration of risperidone (2.0 mg kg−1, s.c.) significantly enhanced 5‐HT output in the DRN.

Consequently, the reduction in 5‐HT cell firing by risperidone appears to be related to increased availability of 5‐HT in the somatodendritic region of the neurones leading to an enhanced 5‐HT1A autoreceptor activation and, in turn, to inhibition of firing, and is probably only to a minor extent caused by its α1‐adrenoceptor antagonistic action.
Characterization of β‐adrenoceptors in urinary bladder: comparison between rat and rabbit
Authors: Oshita Masafumi , Hiraoka Yasuko , Watanabe Yoshinari
Published: January 2005

β‐Adrenoceptor subtypes in rat and rabbit urinary bladder were investigated in functional experiments by use of several agonists and antagonists.

All agonists tested produced concentration‐dependent relaxation, but the relative potencies varied between both species: BRL  37344 (pD2:8.0)>isoprenaline (7.3)>adrenaline (6.7)=noradrenaline (6.6) in rat bladder, and isoprenaline (8.7)=adrenaline (8.5)>noradrenaline (7.7)=BRL  37344 (7.4) in rabbit bladder.

The relaxation response to isoprenaline in rat bladder was relatively resistant to propranolol and ICI  118551, and the slopes of Schild plot for both antagonists were different from unity. The apparent pKB values estimated by single concentrations of propranolol (1, 10 μm) and ICI  118551(10 μm) were 6.6 and 5.4, respectively.

On the other hand, the relaxation response to isoprenaline in rabbit bladder was antagonized by lower concentrations (1 nm–100 nm) of propranolol and ICI  118551 in a competitive manner, resulting in pA2 values of 8.7 and 8.6, respectively.

These results suggest species‐heterogeneity of β‐adrenoceptors in urinary bladder; β3 and β2 subtypes in rat and β2 subtype in rabbit.
Vasorelaxant effects induced by the antiangiogenic drug linomide in aortic and saphenous vein preparations of the rabbit
Authors: Amerini Sandra , Filippi Sandra , Parenti Astrid , Ledda Fabrizio , Ziche Marina
Published: January 2005

Linomide (N‐phenylmethyl‐1,2‐dihydro‐4‐hydroxyl‐1‐methyl‐2‐oxoquinoline‐3‐carboxamide) inhibits vascular proliferation and has been proposed as an antiangiogenic drug. We have investigated the vascular effect of linomide in rabbit aortic and saphenous vein ring preparations and in rat cultured vascular smooth muscle cells (VSMCs).

Linomide (25–300 μg ml−1) did not alter the basal tone of the preparations. The drug induced a concentration‐dependent relaxant effect in aortic rings with endothelium, preconstricted by noradrenaline (NA), 5‐hydroxytryptamine (5‐HT) and by the thromboxane mimetic U46619.

The degree of relaxation induced by linomide was significantly reduced by exposure to the cyclo‐oxygenase inhibitors indomethacin (3 μm) and acetylsalicylic acid (500 μm), and was not influenced by pretreatment with the nitric oxide synthase inhibitor NG‐nitro‐l‐arginine methyl ester (l‐NAME) (100 μm) in aortic rings with endothelium, preconstricted with NA.

Endothelium removal significantly reduced the relaxant response to linomide in aortic ring preparations.

A concentration‐dependent relaxant response was observed also in rabbit saphenous vein preparations deprived of endothelium and preconstricted either by NA or U46619. The degree of relaxation obtained in a high potassium solution was consistently smaller than that observed in NA‐pretreated venous preparations.

The vasorelaxant effect of linomide was consistently blunted by the adenylate cyclase inhibitor SQ 22536 (50 μm), both in intact aortic rings and in those deprived of endothelium.

In rat cultured vascular smooth muscle cells, linomide (100–200 μg ml−1) induced a significant increase in cyclic AMP levels, which was blocked by exposure to 50 μm SQ 22536.

In endothelium‐deprived aortic ring preparations, the linomide‐induced relaxant effect was greatly reduced in high potassium medium (KCl=25 mm). Pretreatment with the ATP potassium channel inhibitor glibenclamide (3 μm) significantly reduced the linomide‐induced relaxation.

The results show that linomide possesses a vasorelaxant effect which is attributable to both endothelium‐dependent and ‐independent properties. While the former component of the drug's activity is apparently due to the release of a prostanoid from endothelial cells, the endothelium‐independent mechanism involved in linomide relaxation is linked to cyclic AMP accumulation and to ATP‐sensitive potassium channel activation in VSMCs.
Dopamine stimulation of cardiac β‐adrenoceptors: the involvement of sympathetic amine transporters and the effect of SKF38393
Authors: Habuchi Yoshizumi , Tanaka Hideo , Nishio Manabu , Yamamoto Taku , Komori Toshiaki , Morikawa Junichiro , Yoshimura Manabu
Published: January 2005

Mechanisms underlying β‐adrenoceptor stimulation by dopamine were examined on guinea‐pig Langendorff‐perfused hearts and isolated cells from the right atrium, by using the chronotropic effects and the enhancement of L‐type Ca2+ current (ICa,L) in the presence of prazosin as indicators of β‐adrenoceptor stimulation. Dopamine‐induced overflow of noradrenaline (NA) concentrations was measured by high‐performance liquid chromatography.

Dopamine caused positive chronotropic effects with an EC50 of 2.5 μm and induced NA overflow with a similar EC50 (1.3 μm). The chronotropic effect of dopamine was abolished by bisoprolol (1 μm).

The effects of dopamine were maintained during prolonged application, whereas the effects of tyramine faded with time. Dopamine (3 μm) restored the chronotropic effects and the NA release suppressed by pretreatment with tyramine, suggesting a de novo synthesis of NA during the exposure to dopamine.

Dopamine (3 μm)‐induced NA release was not affected by tetrodotoxin, ω‐conotoxin, rauwolscine, ICI118551 or sulpiride, but was inhibited by desipramine, a NA uptake inhibitor (IC50 ∼1 μm). It was also not affected by GBR12909 and bupropion, dopamine uptake inhibitors in the central nervous system.

SKF38393, a D1 receptor partial agonist, potently inhibited the 3 μm dopamine‐induced release of NA (IC50 ∼0.1 μm). D1 receptors are not involved in the DA‐induced release of NA, since SCH23390 (3 μm), a potent D1 antagonist, inhibited the NA release only slightly, and dihydrexidine (1 μm) and chloro‐APB (1 μm), full D1 agonists, caused no significant NA release.

SKF38393 inhibited tyramine‐induced overflow of NA, and potentiated the field stimulation‐induced NA release. SKF38393 and desipramine retarded the decay of the stimulation‐induced tachycardia in a similar manner. These results indicate that SKF38393 is a potent monoamine transport inhibitor and a useful tool for the functional evaluation of indirectly‐acting sympathomimetic agonists in the heart. In the presence of SKF38393 (10 μm), dopamine at 1 μm showed no chronotropic effect.

Voltage clamp experiments with isolated atrial cells revealed that dopamine is a weak partial agonist. The EC50 for ICa,L stimulation by dopamine was high (13 μm). As a result, dopamine at 1 μm did not affect ICa,L. Bisoprolol abolished the stimulation of ICa,L by dopamine (30 μm), and dihydrexidine (1 μm) did not affect ICa,L.

It was concluded that the cardiac effects of dopamine at clinically relevant concentrations (<1 μm) result almost exclusively from the indirect effect of β adrenoceptor stimulation, involving the release of NA from sympathetic nerve terminals. The roles of the direct stimulation of β adrenoceptors by dopamine at these concentrations and the stimulation of postjunctional D1 receptors seem negligible. The desipramine‐ and SKF38393‐sensitive monoamine transporter mediates the release of NA.
Role of mast cells, neutrophils and nitric oxide in endotoxin‐induced damage to the neonatal rat colon
Authors: Brown J F , Chafee K A , Tepperman B L
Published: January 2005

The mechanisms involved in mediating bacterial endotoxin lipopolysaccharide (LPS)‐induced injury in the colon of neonatal rat pups aged 10–12 days was examined.

Administration of LPS (3 mg kg−1, i.p.) caused a time‐related increase in the plasma concentration of rat mast cell protease‐II (RMCP‐II) which was attenuated dose‐dependently, by the non‐selective mast cell stabilizer doxantrazole (0.05–5 mg kg−1, i.p.). The selective connective tissue mast cell stabilizer ketotifen (5–25 mg kg−1, i.p.) was without effect at the lower dose and had only a limited inhibitory effect at the higher dose.

In addition, doxantrazole (5 mg kg−1, i.p.) inhibited mast cell degranulation in response to LPS in sections of neonatal rat colon, but ketotifen (5 mg kg−1, i.p.) was without effect.

The increase in plasma RMCP‐II concentration in response to LPS treatment preceded increases in tissue myeloperoxidase (MPO) activity, inducible nitric oxide synthase (iNOS) activity and tissue lipid peroxidation. These events were all attenuated by pretreatment with doxantrazole (5 mg kg−1, i.p.), antineutrophil serum (100 μl kg−1, i.p.), dexamethasone (2 mg kg−1, i.p.) and the selective iNOS inhibitor, aminoguanidine (25 mg kg−1, i.p.).

In addition, lipid peroxidation was inhibited by pre‐administration of the antioxidant enzymes superoxide dismutase (2000 u kg−1, i.p.) and catalase (2000 u kg−1, i.p.), the xanthine oxidase inhibitor allopurinol (100 mg kg−1, i.p.) and the peroxyl scavenger deferoxamine (10 mg kg−1, i.p.), suggesting the involvement of reactive oxygen metabolites in the colonic injury.

These findings suggest that the sequence of events resulting in colonic damage in the neonatal rat following administration of LPS include mast cell degranulation, neutrophil infiltration, elevation in iNOS activity and subsequent lipid peroxidation.

British Journal of Pharmacology (1998) 123, 31–38; doi:10.1038/sj.bjp.0701576
ω‐Conotoxin GVIA‐resistant neurotransmitter release from postganglionic sympathetic nerves in the guinea‐pig vas deferens and its modulation by presynaptic receptors
Authors: Smith Amanda B , Cunnane Tom C
Published: January 2005

Intracellular recording techniques were used to study neurotransmitter release mechanisms in postganglionic sympathetic nerve terminals in the guinea‐pig isolated vas deferens.

Recently, a component of action potential‐evoked release which is insensitive to high concentrations of the selective N‐type calcium channel blocker ω‐conotoxin GVIA termed ‘residual release’ has been described. Under these conditions, release of the neurotransmitter ATP evoked by trains of low frequency stimuli is abolished, but at higher frequencies a substantial component of release is revealed.

‘Residual release’ was studied with trains of 5 or 10 stimuli at stimulation frequencies of 10, 20 and 50 Hz. The α2‐adrenoceptor agonist clonidine (30–100 nM) inhibited ‘residual release’, the degree of inhibition being most marked at the beginning of a train.

The α2‐adrenoceptor antagonist yohimbine (1 μM) induced a marked increase in ‘residual release’ which was dependent on both the frequency of stimulation and the number of stimuli in a train.

Prostaglandin E2 (30 nM) and neuropeptide Y (100 nM) caused a rapid inhibition of ‘residual release’ at all stimulation frequencies examined.

4‐Aminopyridine (100 μM) induced a powerful potential of ‘residual release’ and could reverse the inhibition of ω‐conotoxin GVIA.

‘Residual release’ was modulated through presynaptic α2‐adrenoceptors suggesting that (i) residual release of ATP is subject to α‐autoinhibition through the co‐release of noradrenaline, (ii) noradrenaline release can be triggered by calcium channels other than the N‐type and (iii) when presynaptic receptors are activated, inhibition of transmitter release can occur by mechanisms other than modulation of calcium‐entry through N‐type calcium channels in postganglionic sympathetic nerves. Prostaglandin E2 and neuropeptide Y also modulated neurotransmitter release.

British Journal of Pharmacology (1998) 123, 167–172; doi:10.1038/sj.bjp.0701577
Inhibitory and facilitatory presynaptic effects of endothelin on sympathetic cotransmission in the rat isolated tail artery
Authors: Mutafova‐Yambolieva Violeta N , Westfall David P
Published: January 2005

The present study was undertaken to determine the modulatory effects of the endothelin peptides on the neurogenically‐induced release of endogenous noradrenaline (NA) and the cotransmitter adenosine 5′‐triphosphate (ATP) from the sympathetic nerves of endothelium‐free segments of the rat isolated tail artery. The electrical field stimulation (EFS, 8 Hz, 0.5 ms, 3 min) evoked overflow of NA and ATP, in the absence of endothelins, was 0.035±0.002 pmol mg−1 tissue and 0.026±0.002 pmol mg−1 tissue, respectively.

Endothelin‐1 (ET‐1; 1–30 nM) significantly reduced the EFS evoked overflow of both NA and ATP. The maximum inhibitory effect was produced by a peptide concentration of 10 nM, the amount of NA overflow being 0.020±0.002 pmol mg−1 and that of ATP overflow 0.015±0.001 pmol mg−1. Higher peptide concentrations (100 and 300 nM) reversed the EFS‐evoked overflow of NA to control levels and that of ATP to above control levels. The inhibitory effect of ET‐1 (10 nM) was resistant to the selective ETA receptor antagonist cyclo‐D‐Trp‐D‐Asp(ONa)‐Pro‐D‐Val‐Leu (BQ‐123) but was prevented by ETB receptor desensitization with sarafotoxin S6c (StxS6c) or by ETB receptor blockade with N, cis‐2,6‐dimethylpiperidinocarbonyl‐L‐gmethylleucyl‐D‐1‐methoxycarbonyltryptophanyl‐D‐norleucine (BQ‐788).

StxS6c, upon acute application, exerted a dual effect on transmitter release. At concentrations of 0.001–0.3 nM the peptide significantly reduced the EFS‐evoked NA overflow, whereas at concentrations of 1–10 nM it caused a significant increase in the evoked overflow of both ATP and NA. Both the maximum inhibitory effect of StxS6c at a concentration of 0.003 nM (approximately 85% reduction of NA overflow and 40% of ATP overflow) and the maximum facilitatory effect of the peptide at a concentration of 3 nM (approximately 400% increase of ATP overflow and 200% of NA overflow) were completely antagonized by either BQ‐788 or by StxS6c‐induced ETB receptor desensitization.

ET‐3 (10–100 nM) did not affect the EFS evoked overflow of either ATP or NA, but at a concentration of 300 nM significantly potentiated the release of both transmitters (0.118± 0.02 pmol mg−1 tissue ATP overflow and 0.077±0.004 pmol mg−1 NA overflow). This effect was prevented either by BQ‐123 or by BQ‐788.

In summary, the endothelin peptides exerted both facilitatory and inhibitory effects on the neurogenically‐induced release of the sympathetic cotransmitters ATP and NA in the rat tail artery. Both transmitters were modulated in parallel indicating that the endothelins do not differentially modulate the release of NA and ATP in this tissue.

British Journal of Pharmacology (1998) 123, 136–142; doi:10.1038/sj.bjp.0701579