bluuebunny/crossref_metadata_2025 huggingface dataset:
Molecular identification and pharmacological characterization of adenosine receptors in the guinea‐pig colon
Authors: Kadowaki Makoto , Takeda Masahiro , Tokita Kenichi , Hanaoka Kaori , Tomoi Masaaki
Published: January 2005

The aim of this study is to elucidate the role of adenosine in the motor function of the guinea‐pig distal colon.

To determine whether adenosine A1 receptors and A2B receptors are expressed in the guinea‐pig colon, we employed the reverse transcription‐polymerase chain reaction (RT–PCR). The gene expression of A1 receptor and A2B receptor was found for the first time in the guinea‐pig proximal and distal colon.

Adenosine A1 agonist N6‐cyclopentyladenosine (CPA), and A1/A2 agonist 5′‐N‐ethylcarboxamidoadenosine (NECA) concentration‐dependently inhibited neurogenic responses to electrical field stimulation (EC50=1.07×10−8 and 2.12×10−8 M) in the longitudinal muscle, but A2A agonist 2‐p‐(2‐carboxyethyl)phenylethylamino‐5′‐N‐ethycarboxamido‐adenosine (CGS21680) had only a slight inhibitory effect (25.9%, 1 μM). A1 antagonist 8‐cyclopentyl‐1,3‐dipropylxanthine (DPCPX, 10 nM: A1 selective concentration) antagonized responses to CPA and NECA. Furthermore, the affinity order of antagonists at inhibiting the effect NECA was: DPCPX>8‐phenyltheophylline (8‐PT: A1/A2 antagonist).

In the presence of tetrodotoxin (TTX, 0.3 μM), CPA and NECA relaxed myogenic precontraction induced by KCl (50 mM) (EC50=1.26×10−5 and 1.04×10−5 M, respectively), but CGS21680 (1 μM) did not cause any relaxation. DPCPX did not affect responses to CPA and NECA at a concentration of 10 nM, but a higher concentration (1 μM) of DPCPX and 10 μM of 8‐PT antagonized those responses.

These data lead us to the hypothesis that adenosine may mediate relaxation through two different inhibitory receptor subtypes; A1 receptors on the enteric neuron and A2B receptor on the smooth muscle in the guinea‐pig distal colon.

British Journal of Pharmacology (2000) 129, 871–876; doi:10.1038/sj.bjp.0703123
Possible mechanisms underlying the vasodilatation induced by olprinone, a phosphodiesterase III inhibitor, in rabbit coronary artery
Authors: Ohashi Masuo , Dohi Yasuaki , Itoh Takeo
Published: January 2005

The possible mechanisms underlying the vasodilatation induced by olprinone, a phosphodiesterase type III inhibitor, were investigated in smooth muscle of the rabbit coronary artery. Isometric force and membrane potential were measured simultaneously using endothelium‐denuded smooth muscle strips.

Acetylcholine (ACh, 3 μM) produced a contraction with a membrane depolarization (15.2±1.1 mV). In a solution containing 5.9 mM K+, olprinone (100 μM) hyperpolarized the resting membrane and (i) caused the absolute membrane potential level reached with ACh to be more negative (but did not reduce the delta membrane potential seen with ACh, 15.2±1.8 mV) and (ii) attenuated the ACh‐induced contraction. In a solution containing 30 mM K+, these effects were not seen with olprinone.

Glibenclamide (10 μM) blocked the olprinone‐induced membrane hyperpolarization. 4‐AP (0.1 mM) significantly attenuated the olprinone‐induced resting membrane hyperpolarization but TEA (1 mM) had no such effect.

Glibenclamide (10 μM), TEA (1 mM) and 4‐AP (0.1 mM), given separately, all failed to modify the inhibitory actions of olprinone on (i) the absolute membrane potential level seen with ACh and (ii) the ACh‐induced contraction.

It is suggested that olprinone inhibits the ACh‐induced contraction through an effect on the absolute level of membrane potential achieved with ACh in smooth muscle of the rabbit coronary artery. It is also suggested that glibenclamide‐sensitive, ATP‐sensitive K+ channels do not play an important role in the olprinone‐induced inhibition of the ACh‐induced contraction.

British Journal of Pharmacology (2000) 129, 1000–1006; doi:10.1038/sj.bjp.0703125
Pharmacological characterization of human recombinant melatonin mt1 and MT2 receptors
Authors: Browning Christopher , Beresford Isabel , Fraser Neil , Giles Heather
Published: January 2005

We have pharmacologically characterized recombinant human mt1 and MT2 receptors, stably expressed in Chinese hamster ovary cells (CHO‐mt1 and CHO‐MT2), by measurement of [3H]‐melatonin binding and forskolin‐stimulated cyclic AMP (cAMP) production.

[3H]‐melatonin bound to mt1 and MT2 receptors with pKD values of 9.89 and 9.56 and Bmax values of 1.20 and 0.82 pmol mg−1 protein, respectively. Whilst most melatonin receptor agonists had similar affinities for mt1 and MT2 receptors, a number of putative antagonists had substantially higher affinities for MT2 receptors, including luzindole (11 fold), GR128107 (23 fold) and 4‐P‐PDOT (61 fold).

In both CHO‐mt1 and CHO‐MT2 cells, melatonin inhibited forskolin‐stimulated accumulation of cyclic AMP in a concentration‐dependent manner (pIC50 9.53 and 9.74, respectively) causing 83 and 64% inhibition of cyclic AMP production at 100 nM, respectively. The potencies of a range of melatonin receptor agonists were determined. At MT2 receptors, melatonin, 2‐iodomelatonin and 6‐chloromelatonin were essentially equipotent, whilst at the mt1 receptor these agonists gave the rank order of potency of 2‐iodomelatonin>melatonin>6‐chloromelatonin.

In both CHO‐mt1 and CHO‐MT2 cells, melatonin‐induced inhibition of forskolin‐stimulated cyclic AMP production was antagonized in a concentration‐dependent manner by the melatonin receptor antagonist luzindole, with pA2 values of 5.75 and 7.64, respectively. Melatonin‐mediated responses were abolished by pre‐treatment of cells with pertussis toxin, consistent with activation of Gi/Go G‐proteins.

This is the first report of the use of [3H]‐melatonin for the characterization of recombinant mt1 and MT2 receptors. Our results demonstrate that these receptor subtypes have distinct pharmacological profiles.

British Journal of Pharmacology (2000) 129, 877–886; doi:10.1038/sj.bjp.0703130
The insulinotropic mechanism of the novel hypoglycaemic agent JTT‐608: direct enhancement of Ca2+ efficacy and increase of Ca2+ influx by phosphodiesterase inhibition
Authors: Mukai Eri , Ishida Hitoshi , Fujimoto Shimpei , Kajikawa Mariko , Okamoto Yoshimasa , Fujita Jun , Hamamoto Yoshiyuki , Tsuura Yoshiyuki , Yamada Yuichiro , Furukawa Noboru , Ohta Takeshi , Seino Yutaka
Published: January 2005

We examined the effects of the novel hypoglycaemic agent JTT‐608 [trans‐4‐(4‐methylcyclohexyl)‐4‐oxobutyric acid] on insulin secretion using rat pancreatic islets, and analysed the mechanism of its effect.

JTT‐608 augmented 8.3 mM glucose‐induced insulin secretion dose‐dependently, and there was a stimulatory effect of 100 μM JTT‐608 at both moderate and high concentrations (8.3, 11.1 and 16.7 mM) of glucose, but not at low concentrations (3.3 and 5.5 mM). In perifusion experiments, both phases of insulin release were enhanced, and the effect was eliminated 10 min after withdrawal of the agent.

In the presence of 200 μM diazoxide and a depolarizing concentration (30 mM) of K+, there was an augmentation of insulin secretion by 100 μM JTT‐608, not only under high levels of glucose but also under low levels, and the effects were abolished by 10 μM nitrendipine.

JTT‐608 augmented insulin secretion from electrically permeabilized islets in the presence of stimulatory concentrations (0.3 and 1.0 μM) of Ca2+, and the intracellular Ca2+ concentration ([Ca2+]i) response under 16.7 mM glucose, 200 μM diazoxide, and 30 mM K+ was also increased.

The cyclic AMP content in the islets was increased by 100 μM JTT‐608, and an additive effect to 1 μM forskolin was observed, but not to 50 μM 3‐isobutyl‐1‐methylxanthine (IBMX). JTT‐608 inhibited phosphodiesterase (PDE) activity dose‐dependently.

We conclude that JTT‐608 augments insulin secretion by enhancing Ca2+ efficacy and by increasing Ca2+ influx. This appears to be a result of the increased intracellular cyclic AMP concentration due to PDE inhibition.

British Journal of Pharmacology (2000) 129, 901–908; doi:10.1038/sj.bjp.0703133
Role of clot‐associated (‐derived) thrombin in cell proliferation induced by fibrin clots in vitro
Authors: Gandossi E , Lunven C , Berry C N
Published: January 2005

Thrombin is a potent mitogenic agent. Clot‐associated thrombin retains its amidolytic and pro‐aggregant activity. We therefore studied the ability of fibrin clots to induce proliferation in CCL39 cells (Chinese hamster lung fibroblasts), in the absence and presence of the thrombin inhibitors PPACK, recombinant hirudin (rHV2 Lys47) and heparin:antithrombin III.

Fibrin clots incubated for 48 h with CCL39 cells led to significant cell proliferation, which was dependent on the concentration of thrombin used to prepare the clots. Thus, clots prepared with 91 nmol l−1 thrombin produced a similar proliferation (231±21%) to that obtained with 50 nmol l−1 thrombin in solution (213±29%). Rabbit plasma clots led to a 499±41% increase in cell number under identical conditions.

Fibrin clot‐induced cell proliferation was inhibited by all three thrombin inhibitors with no difference in IC50 values compared to those obtained against thrombin in solution, suggesting that cell proliferation be due to thrombin leaching from the clots.

We found a time‐dependent increase in thrombin release from the clots attaining a plateau at 24 h (∼61% of the total thrombin used in clot formation). Clots separated from the cells using porous cell culture chamber inserts led to similar proliferation to that of clots in contact with the cells.

Thus fibrin‐clot induced CCL39 proliferation is due to thrombin released from the clots.

British Journal of Pharmacology (2000) 129, 1021–1027; doi:10.1038/sj.bjp.0703137
Endothelium‐dependent relaxation followed by contraction mediated by NK1 receptors in precontracted rabbit intrapulmonary arteries
Authors: Shirahase Hiroaki , Kanda Mamoru , Kurahashi Kazuyoshi , Nakamura Shohei
Published: January 2005

In the present study, we examined whether substance P (SP) and SP methyl ester (SPME), a selective NK1 agonist, cause biphasic responses consisting of endothelium‐dependent relaxation (EDR) and contraction (EDC) in precontracted rabbit intrapulmonary arteries.

In arteries contracted with PGF2α (2×10−6 M), SP as well as SPME caused only EDR at low concentration (10−9 M) and EDR followed by EDC at higher concentrations, indicating the involvement of NK1 receptors. The SP (10−8 M)‐induced EDR was abolished in arteries moderately contracted by PGF2α (5×10−7 M) and the EDC in arteries maximally contracted by PGF2α (10−5 M), indicating that EDR and EDC are inversely dependent on preexisting tone.

Indomethacin (10−8–10−6 M), a cyclo‐oxygenase inhibitor, and ozagrel (10−8–10−6 M), a TXA2 synthetase inhibitor attenuated the EDC in the SPME (10−7 M)‐induced biphasic response and markedly potentiated the EDR. AA‐861 (10−8–10−6 M), a 5‐lipoxygenase inhibitor, did not affect the EDR or EDC. L‐NG‐nitro‐arginine methyl ester (10−5–10−4 M), a nitric oxide synthase inhibitor, attenuated the EDR and slightly potentiated the EDC.

CP‐99994 (10−10–10−8 M), an NK1 antagonist, attenuated the EDC and potentiated the EDR in the SPME (10−7 M)‐induced biphasic response, while the NK2 antagonist SR‐48968 (10−9–10−7 M) had no effect. CP‐99994 attenuated the SPME (10−7 M)‐induced EDC under EDR‐blockade to a greater extent than the EDR under EDC‐blockade, indicating that CP‐99994 enhanced the EDR component by preferential inhibition of the EDC component.

In conclusion, NK1 agonists caused a biphasic endothelium‐dependent response (EDR and EDC) in submaximally precontracted intrapulmonary arteries. The EDC and EDR mediated by NK1 receptors may play physiological and/or pathophysiological roles in modulation of vascular tone.

British Journal of Pharmacology (2000) 129, 937–942; doi:10.1038/sj.bjp.0703140
Upregulation by glucocorticoids of responses to eosinopoietic cytokines in bone‐marrow from normal and allergic mice
Authors: Gaspar Elsas Maria Ignez C , Maximiano Elisabeth S , Joseph Danielle , Alves Letícia , Topilko Andrzej , Vargaftig B Boris , Xavier Elsas P
Published: January 2005

Since the production of eosinopoietic cytokines (GM‐CSF, IL‐3, IL‐5) is inhibited by glucocorticoids, while responsiveness to these cytokines is enhanced in bone‐marrow of allergic mice, we studied the ability of glucocorticoids to modulate murine bone‐marrow eosinopoiesis.

Progenitor (semi‐solid) and/or precursor (liquid) cultures were established from bone‐marrow of: (a) normal mice; (b) ovalbumin‐sensitized and challenged mice or (c) dexamethasone (1–5 mg kg−1) injected mice. Cultures were established with GM‐CSF (2 ng ml−1) or IL‐5 (1 ng ml−1), respectively, alone or associated with dexamethasone, hydrocortisone or corticosterone. Total myeloid colony numbers, frequency and size of eosinophil colonies, and numbers of eosinophil‐peroxidase‐positive cells were determined at day 7.

In BALB/c mice, dexamethasone (10−7 M) increased GM‐CSF‐stimulated myeloid colony formation (P=0.01), as well as the frequency (P=0.01) and size (P<0.01) of eosinophil colonies. Dexamethasone (10−7 M) alone had no effect. Dexamethasone (10−7–10−10 M) increased (P<0.002) eosinophil precursor responses to IL‐5. Potentiation by dexamethasone was still detectable: (a) on low density, immature, nonadherent BALB/c bone‐marrow cells, (b) on bone‐marrow from other strains, and (c) on cells from allergic mice. Hydrocortisone and corticosterone had similar effects. Dexamethasone administered in vivo, 24 h before bone‐marrow harvest, increased subsequent progenitor responses to GM‐CSF (P=0.001) and precursor responses to IL‐5 (P<0.001). These effects were blocked by RU 486 (20 mg kg−1, orally, 2 h before dexamethasone, or added in vitro at 10 μM, P<0.001).

Glucocorticoids, acting in vivo or in vitro, through glucocorticoid receptors, enhance bone‐marrow eosinopoiesis in naïve and allergic mice.

British Journal of Pharmacology (2000) 129, 1543–1552; doi:10.1038/sj.bjp.0703145
Active transport of the angiotensin‐II antagonist losartan and its main metabolite EXP 3174 across MDCK‐MDR1 and Caco‐2 cell monolayers
Authors: Soldner Andrea , Benet Leslie Z , Mutschler Ernst , Christians Uwe
Published: January 2005

We studied the functional interaction between transport and metabolism by comparing the transport of losartan and its active metabolite EXP 3174 (EXP) across cell monolayers.

Epithelial layers of Caco‐2 cells as well as MDR1, MRP‐1 and MRP‐2 overexpressing cells, in comparison to the respective wildtypes, were used to characterize the transcellular transport of losartan and EXP.

Losartan transport in MDCK‐MDR1 and Caco‐2 cells was saturable and energy‐dependent with a significantly greater basolateral‐to‐apical (B/A) than apical‐to‐basolateral (A/B) flux (ratio=31±1 in MDCK‐MDR1 and ratio 4±1 in Caco‐2 cells). The B/A flux of losartan was inhibited by cyclosporine and vinblastine, inhibitors of P‐glycoprotein and MRP. In contrast, no active losartan transport was observed in MRP‐1 or MRP‐2 overexpressing cells.

The metabolite was only transported in Caco‐2 cells with a B/A‐to‐A/B ratio of 5±1, while lacking active transport in the MDR1, MRP‐1 or MRP‐2 overexpressing cells. The B/A flux of EXP was significantly inhibited by cyclosporine and vinblastine.

In conclusion, losartan is transported by P‐glycoprotein and other intestinal transporters, that do not include MRP‐1 and MRP‐2. In contrast, the carboxylic acid metabolite is not a P‐glycoprotein substrate, but displays considerably higher affinity for other transporters than losartan, that again most probably do not include MRP‐1 and MRP‐2.

British Journal of Pharmacology (2000) 129, 1235–1243; doi:10.1038/sj.bjp.0703150
Endothelin‐1‐induced ETA receptor‐mediated nociception, hyperalgesia and oedema in the mouse hind‐paw: modulation by simultaneous ETB receptor activation
Authors: Piovezan Anna P , D'Orléans‐Juste Pedro , Souza Glória E P , Rae Giles A
Published: January 2005

Endothelin‐1 causes ETA receptor‐mediated enhancement of capsaicin‐induced nociception in mice. We have assessed if this hyperalgesic effect of endothelin‐1 is also accompanied by other pro‐inflammatory effects, namely nociception and oedema, and characterized the endothelin ET receptors involved.

Intraplantar (i.pl.) hind‐paw injection of endothelin‐1 (0.3–30 pmol) induced graded nociceptive responses (accumulated licking time: vehicle, 20.5±3.3 s; endothelin‐1 at 30 pmol, 78.1±9.8 s), largely confined to the first 15 min. Endothelin‐1 (1–10 pmol) potentiated ipsilateral capsaicin‐induced (0.1 μg, i.pl.; at 30 min) nociception (vehicle, 40.2±2.6 s; endothelin‐1 at 10 pmol, 98.4±5.8 s, but 30 pmol was inactive), and caused oedema (increase in paw weight 5 min after capsaicin: vehicle, 46.3±2.3 mg; endothelin‐1 at 30 pmol, 100.3±6.1 mg).

Selective ETB receptor agonists sarafotoxin S6c (up to 30 pmol) and IRL 1620 (up to 100 pmol) were inactive, whereas endothelin‐3 (up to 30 pmol) induced only modest oedema.

ETA receptor antagonists BQ‐123 (1 nmol, i.pl.) or A‐127722‐5 (6 μmol kg−1, i.v.) prevented all effects of endothelin‐1 (10 pmol), but the ETB receptor antagonist BQ‐788 (1 or 10 nmol, i.pl.) was ineffective.

BQ‐788 (10 nmol, i.pl.) unveiled hyperalgesic effects of 30 pmol endothelin‐1 and endothelin‐3. Sarafotoxin S6c (30 pmol, i.pl.) did not modify endothelin‐1‐induced (10 pmol) nociception or oedema, but abolished hyperalgesia.

Thus, endothelin‐1 triggers ETA receptor‐mediated nociception, hyperalgesia and oedema in the mouse hind‐paw. Simultaneous activation of ETB receptors by endothelin‐1 or selective agonists can limit the hyperalgesic, but not the nociceptive or oedematogenic, effects of the peptide.

British Journal of Pharmacology (2000) 129, 961–968, doi:10.1038/sj.bjp.0703154
Factors influencing the low‐frequency associated nicotinic ACh autoreceptor‐mediated depression of ACh release from rat motor nerve terminals
Authors: Prior Chris , Singh Shila
Published: January 2005

We have studied the inhibitory autoreceptor control of acetylcholine (ACh) release from rat motor nerve endings using an electrophysiological technique to quantify evoked ACh release in isolated hemidiaphragm muscles. Quantal ACh release (m) was estimated from the ratio of amplitudes of nerve evoked endplate currents and spontaneously occurring miniature endplate currents.

The nicotinic ACh receptor agonist cytisine (1 μM) decreased m at 0.5 Hz by around 20% but had no effect on m at 50 Hz. Changing the extracellular Ca2+ concentration from 1.8 mM to either 0.45 or 3.6 mM abolished the effect of cytisine on m at 0.5 Hz. The nicotinic ACh receptor antagonist hexamethonium (200 μM) increased m at 0.5 Hz by 15–20%.

The effects of cytisine and hexamethonium on m at 0.5 Hz were blocked by 10 μM verapamil, which itself significantly increased m. However, the effects of cytisine and hexamethonium on m at 0.5 Hz were not sensitive to 10 μM of the calmodulin antagonist, W‐7. This concentration of W‐7 attenuates effects on ACh release mediated by facilitatory prejunctional nicotinic ACh autoreceptors.

Our present observations are suggestive of actions of cytisine and hexamethonium to activate and inhibit respectively negative‐feedback prejunctional nicotinic ACh autoreceptors. Further, they strengthen the case for the existence of two separate and independent autoregulatory mechanisms for the control of ACh release from motor nerve terminals and give a preliminary insight into the cellular mechanism involved in the autoinhibition of ACh release.

British Journal of Pharmacology (2000) 129, 1067–1074; doi:10.1038/sj.bjp.0703161
The effect of P2 receptor antagonists and ATPase inhibition on sympathetic purinergic neurotransmission in the guinea‐pig isolated vas deferens
Authors: Sneddon P , Westfall T D , Todorov L D , Todorova S Mihaylova , Westfall D P , Nickel P , Kennedy C
Published: January 2005

Intracellular microelectrodes were used to record the transmembrane potential and excitatory junction potentials (e.j.p.s) produced by sympathetic nerve stimulation (1 Hz) in smooth muscle cells of the guinea‐pig isolated vas deferens.

The symmetrical 3′‐urea of 8‐(benzamido)naphthalene‐1,3,5‐trisulphonic acid (NF023) produced a concentration‐dependent inhibition of e.j.p. magnitude (IC50=4.8×10−6 M), but had no effect on the resting membrane potential of the smooth muscle cells.

Pyridoxal‐5‐phosphate (P‐5‐P) also depressed e.j.p. magnitude in a concentration‐dependent manner, but was less potent than NF023 (IC50=2.2×10−5 M). At 10−4 M and above P‐5‐P significantly depolarized the smooth muscle cells.

The nucleoside triphosphatase inhibitor 6‐N,N‐diethyl‐D‐β,γ‐dibromomethyleneATP (ARL 67156) (5×10−5 M) significantly increased e.j.p. amplitude. ARL 67156 (10−4 M) further increased e.j.p. amplitude such that they often reached threshold for initiation of action potentials, causing muscle contraction and expulsion of the recording electrode.

After reduction of e.j.p.s by NF023 or P‐5‐P (both 10−5 M), subsequent co‐addition of ARL 67156 (10−4 M) significantly increased their magnitude.

The overflow of endogenous ATP evoked by field stimulation of sympathetic nerves (8 Hz, 1 min) was measured by HPLC and flurometric detection. ARL 67156 (10−4 M) enhanced ATP overflow by almost 700% compared to control.

We conclude that for electrophysiological studies NF023 is preferable to other P2X receptor antagonists such as pyridoxalphosphate ‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS), suramin or P‐5‐P. Furthermore, breakdown of endogenous ATP by nucleoside triphosphatases is an important modulator of purinergic neurotransmission in the guinea‐pig vas deferens.

British Journal of Pharmacology (2000) 129, 1089–1094; doi:10.1038/sj.bjp.0703163
Effects of AMP derivatives on cyclic AMP levels in NG108‐15 cells
Authors: Ohkubo Satoko , Nakanishi Hironori , Kimura Junko , Matsuoka Isao
Published: January 2005

In NG108‐15 neuroblastoma×glioma hybrid cells, ATP stimulates intracellular cyclic AMP formation, which is inhibited by both adenosine (P1) and P2 receptor antagonists. In the present study, we examined the effects of several AMP derivatives in NG108‐15 cells and mouse neuroblastoma N18TG‐2 cells.

Adenosine 2′‐monophosphate (A2P), adenosine 3′‐monophosphate (A3P) and adenosine 5′‐phosphosulphate (A5PS) increased cyclic AMP levels with similar concentration‐dependencies in NG108‐15 cells.

Increases in cyclic AMP by AMP derivatives were inhibited by the P2 receptor antagonist PPADS, but not by suramin. Effects of AMP derivatives were also inhibited by P1 receptor antagonists ZM241385, XAC, DPCPX and partially by alloxazine. The ecto‐nucleotidase inhibitor α,β‐methyleneADP was without effect.

In contrast, AMP derivatives did not change cyclic AMP levels in N18TG‐2 cells. Accumulation of cyclic AMP in N18TG‐2 cells was stimulated by adenosine A2 receptor agonists CGS21680 and NECA, but not by ATP or β,γ‐methyleneATP, agonists for cyclic AMP production in NG108‐15 cells.

Reverse transcription‐coupled polymerase chain reaction (RT–PCR) analyses revealed that N18TG‐2 cells express both A2A and A2B receptors, while NG108‐15 cells express mainly A2A receptors.

AMP derivatives did not affect the P2X and P2Y receptors expressed in NG108‐15 cells.

These results suggest that A2P, A3P and A5PS act as agonists for cyclic AMP production and that these compounds are valuable tools for determinating the mechanism of ATP‐stimulated cyclic AMP response in NG108‐15 cells.

British Journal of Pharmacology (2000) 129, 1244–1250; doi:10.1038/sj.bjp.0703168
The stimulant effects of caffeine on locomotor behaviour in mice are mediated through its blockade of adenosine A2A receptors
Authors: Yacoubi Malika El , Ledent Catherine , Ménard Jean‐François , Parmentier Marc , Costentin Jean , Vaugeois Jean‐Marie
Published: January 2005

The locomotor stimulatory effects induced by caffeine (1,3,7‐trimethylxanthine) in rodents have been attributed to antagonism of adenosine A1 and A2A receptors. Little is known about its locomotor depressant effects seen when acutely administered at high doses. The roles of adenosine A1 and A2A receptors in these activities were investigated using a Digiscan actimeter in experiments carried out in mice. Besides caffeine, the A2A antagonist SCH 58261 (5‐amino‐7‐(β‐phenylethyl)‐2‐(8‐furyl)pyrazolo[4,3‐e]‐1,2,4‐triazolo[1,5‐c]pyrimidine), the A1 antagonist DPCPX (8‐cyclopentyl‐1,3‐dipropylxanthine), the A1 agonist CPA (N6‐cyclopentyladenosine) and A2A receptor knockout mice were used.

Caffeine had a biphasic effect on locomotion of wild‐type mice not habituated to the open field, stimulating locomotion at 6.25–25 mg kg−1 i.p. doses, while depressing it at 100 mg kg−1. In sharp contrast, caffeine dose‐dependently decreased locomotion in A2A receptor knockout mice over the whole range of tested doses.

The depressant effects induced by high doses of caffeine were lost in control CD1 mice habituated to the open field.

The A1 agonist CPA depressed locomotion at 0.3–1 mg kg−1 i.p. doses.

The A1 antagonist DPCPX decreased locomotion of A2A receptor knockouts and CD1 mice at 5 mg kg−1 i.p. and 25 mg kg−1 i.p. respectively.

DPCPX (0.2–1 mg kg−1 i.p.) left unaltered or even reduced the stimulant effect of SCH 58261 (1–3 mg kg−1 i.p.) on CD1 mice.

These results suggest therefore that the stimulant effect of low doses of caffeine is mediated by A2A receptor blockade while the depressant effect seen at higher doses under some conditions is explained by A1 receptor blockade.

British Journal of Pharmacology (2000) 129, 1465–1473; doi:10.1038/sj.bjp.0703170
Respiratory actions of tachykinins in the nucleus of the solitary tract: characterization of receptors using selective agonists and antagonists
Authors: Mazzone Stuart B , Geraghty Dominic P
Published: January 2005

The respiratory response to microinjection of tachykinins and analogues into the commissural nucleus of the solitary tract (cNTS) of urethane‐anaesthetized rats was investigated in the presence and absence of selective tachykinin NK1, NK2 and NK3 antagonists (RP 67580, SR 48968 and SR 142801, respectively).

All tachykinins, except for the selective NK2 agonist, [Nle10]‐NKA(4‐10), increased tidal volume (VT). The rank potency order of naturally‐occurring tachykinins was neurokinin A (NKA)substance P (SP)>>NKB, whereas the rank order for selective analogues was senktideseptide>> [Sar9,Met(O2)11]‐SP>>[Nle10]‐NKA(4‐10). Septide (NK1‐selective) and senktide (NK3‐selective) were 22 fold more potent (pD2∼12) at stimulating VT than SP (pD2∼10.5).

Tachykinin agonists produced varying degrees of respiratory slowing, independent of changes in VT. At doses producing maximum stimulation of VT, agonists induced either a mild (<10 breaths min−1 decrease; SP and septide), moderate (10–25 breaths min−1 decrease; NKA, NKB and [Sar9,Met(O2]‐SP) or severe (∼40 breaths min−1 decrease; senktide) bradypnoea. [Nle10]‐NKA(4‐10) produced a dose‐dependent bradypnoea without affecting VT.

RP 67580 significantly attenuated the VT response to SP (33 pmol) and NKA (10 pmol) but not NKB (100 pmol). In the presence of RP 67580, the mild bradypnoeic response to NKB was significantly enhanced whereas SP and NKA induced a bradypnoea which was not observed in the absence of RP 67580. SR 48968 had no effect on the VT response to SP or NKB, markedly enhanced the VT response to NKA and completely blocked the bradypnoeic response to [Nle10]‐NKA(4‐10). Only SR142801 attenuated the VT response to NKB.

The present data suggest that all three tachykinin receptors (NK1, NK2 and NK3) are present in the cNTS and are involved in the central control of respiration.

British Journal of Pharmacology (2000) 129, 1121–1131; doi:10.1038/sj.bjp.0703172
Nitrergic relaxation of the mouse gastric fundus is mediated by cyclic GMP‐dependent and ryanodine‐sensitive mechanisms
Authors: Selemidis Stavros , Cocks Thomas M
Published: January 2005

Ryanodine‐sensitive, Ca2+ release (‘Ca2+ sparks’) from the sarcoplasmic reticulum (SR) can activate plasmalemmal Ca2+‐activated K+ channels (KCa) to cause membrane hyperpolarization and smooth muscle relaxation. Since cyclic guanosine monophosphate (cyclic GMP) can modulate Ca2+ spark activity, the aim of the present study was to determine if Ca2+ spark‐like events are involved in NO‐dependent, NANC relaxations to electrical field stimulation (EFS) of mouse, longitudinal smooth muscle of the gastric fundus in isolated strips contracted to ∼40% of their maximum contraction.

NANC relaxations to EFS were almost abolished by both the NO synthase inhibitor, NG‐nitro‐L‐arginine (L‐NOARG; 100 μM) and the guanylate cyclase inhibitor, 1‐H‐oxodiazol‐[1,2,4]‐[4,3‐α] quinoxaline‐1‐one (ODQ; 10 μM). Also, ODQ abolished relaxations to the NO donor, sodium nitroprusside (SNP; 1 nM–30 μM). NANC relaxations and SNP‐evoked relaxations were both partly ryanodine (10 μM)‐ and nifedipine (0.3 μM)‐sensitive, but in each case, the inhibitory effects of ryanodine and nifedipine were additive.

Apamin (1 μM), charybdotoxin (0.1 μM), iberiotoxin (0.1 μM), tetraethylammonium (TEA; 1 mM), glibenclamide (10 μM) and 4‐aminopyridine (1 mM) had no effect on either NANC‐ or SNP‐evoked relaxations, the latter of which were also unaffected by high extracellular K+ (68 mM).

Caffeine (0.1–1 mM) caused concentration‐dependent relaxations of gastric fundus which were inhibited by ryanodine but unaffected by L‐NOARG.

Relaxation to ATP (30 μM) was abolished by nifedipine, partly inhibited by apamin and ryanodine, but was unaffected by L‐NOARG.

In conclusion, the results of the present study show that nitrergic relaxations in the mouse longitudinal gastric fundus occur via a cyclic GMP‐activated ryanodine‐sensitive mechanism, which does not appear to involve activation of K+ channels.

British Journal of Pharmacology (2000) 129, 1315–1322; doi:10.1038/sj.bjp.0703174
Active immunization with angiotensin I peptide analogue vaccines selectively reduces the pressor effects of exogenous angiotensin I in conscious rats
Authors: Gardiner S M , Auton T R , Downham M R , Sharp H L , Kemp P A , March J E , Martin H , Morgan P J , Rushton A , Bennett T , Glover J F
Published: January 2005

Male, Sprague‐Dawley rats were actively immunized with novel angiotensin vaccines, and their pressor responses to exogenous angiotensin I (AI) and angiotensin II (AII) were assessed in vivo. Serum antibody titres were also measured.

The most effective vaccine consisted of an AI analogue conjugated with a tetanus toxoid carrier protein and adjuvanted with aluminium hydroxide. When this vaccine was injected on days 0, 21 and 42, pressor responses to AI on day 63 were significantly inhibited (maximum, 8.9 fold shift), but responses to AII were unaffected. The anti‐angiotensin antibody titre was increased 32,100 fold, and, uniquely, these antibodies also cross‐reacted with angiotensinogen.

These findings indicate that active immunization against AI may be a useful approach for treating cardiovascular disorders involving the renin‐angiotensin system.

British Journal of Pharmacology (2000) 129, 1178–1182; doi:10.1038/sj.bjp.0703178
Both α1A‐ and α1B‐adrenergic receptor subtypes couple to the transient outward current (ITo) in rat ventricular myocytes
Authors: Homma Nobuo , Hirasawa Akira , Shibata Katsushi , Hashimito Keitaro , Tsujimoto Gozoh
Published: January 2005

Regulation of transient outward current (ITo) by α1‐adrenergic (α1AR) plays a key role in cardiac repolarization. α1ARs comprise a heterogeneous family; two natively expressed subtypes (α1A and α1B) and three cloned subtypes (α1a, α1b and α1d) can be distinguished. We have examined the electrophysiological role of each α1AR subtype in regulating ITo in isolated rat ventricular myocytes.

Reverse transcription‐PCR study revealed the presence of three subtype mRNAs (α1a, α1b and α1d) in rat myocytes.

Radioligand binding assay using [125I]‐HEAT showed that the inhibition curves for α1AAR‐selective antagonists (WB4101, 5‐methylurapidil, (+)‐niguldipine and KMD‐3213) in rat ventricles best fit a two‐site model, with 30% high and 70% low affinity binding sites. The high affinity sites were resistant to 100 μM chloroethylclonidine (CEC), while the low affinity sites were highly inactivated by CEC.

Whole cell voltage clamp study revealed that methoxamine reduced a 4‐aminopyridine(4‐AP)‐sensitive component of ITo in the isolated rat ventricle myocytes. Lower concentrations of KMD‐3213 (1 nM) or 5‐MU (10 nM) did not affect the methoxamine‐induced reduction of ITo. On the other hand, CEC treatment (100 μM) of isolated myocytes reduced the methoxamine‐induced reduction of ITo by 46%, and the remaining response was abolished by lower concentrations of KMD‐3213 or 5‐MU.

The results indicate that rat ventricular myocytes express transcripts of the three α1AR subtypes (α1a, α1b and α1d); however, two pharmacologically distinct α1AR subtypes (α1A and α1B) are predominating in receptor populations, with approximately 30% α1AAR and 70% α1BAR. Although both α1A and α1BAR subtypes are coupled to the cardiac ITo, α1BARs predominantly mediate α1AR‐induced effect.

British Journal of Pharmacology (2000) 129, 1113–1120; doi:10.1038/sj.bjp.0703179
Characterization of human recombinant α2A‐adrenoceptors expressed in Chinese hamster lung cells using intracellular Ca2+ changes: evidence for cross‐talk between recombinant α2A‐ and native α1‐adrenoceptors
Authors: Reynen P H , Martin G R , Eglen R M , MacLennan S J
Published: January 2005

Human α2A‐adrenoceptors expressed in Chinese hamster lung (CHL) fibroblasts have been pharmacologically characterized by measuring intracellular calcium (Ca2+i) changes using the Ca2+‐sensitive dye Fluo3‐AM, in conjunction with a fluorometric imaging plate reader (FLIPR).

Several α‐adrenoceptor agonists were examined including the α2‐adrenoceptor agonists UK‐14304, B‐HT 920, dexmedetomidine and A‐54741, the selective α1‐adrenoceptor agonist phenylephrine and the non‐selective adrenergic agonist noradrenaline. Of these only noradrenaline (mean pEC50=6.49) and A‐54741 (6.90) evoked changes in Ca2+i; A‐54741 was a partial agonist relative to noradrenaline, achieving only 33% of the noradrenaline maximum.

Ca2+i changes induced by noradrenaline and A‐54741 were antagonized by the α2‐selective antagonist rauwolscine (10 nM) and by the α1‐selective antagonists prazosin (0.1 nM) and doxazosin (1.0 nM).

Phenylephrine (100 μM) and UK‐14304 (10 μM) alone were ineffective in causing Ca2+i increase. In the presence of a fixed concentration of UK‐14304 (3.0 μM), phenylephrine induced concentration‐dependent increases in Ca2+i (mean pEC50=5.33). In the presence of phenylephrine (30.0 μM) UK‐14304 induced Ca2+i release (pEC50=6.92). The effects of phenylephrine were abolished by prazosin (1.0 nM) or rauwolscine (100 nM).

In saturation radioligand binding experiments using membranes of parental (non‐transfected) CHL cells there was a small, specific binding of [3H]‐prazosin (Bmax=24 fmol mg protein−1; pKD=10.24).

Collectively, these data suggest that α‐adrenoceptor agonist‐induced Ca2+i release in CHL fibroblasts transfected with the human α2A‐adrenoceptor is dependent upon co‐activation of the recombinant receptor and a native α1‐adrenoceptor.

British Journal of Pharmacology (2000) 129, 1339–1346; doi:10.1038/sj.bjp.0703184
Role of gap junctions and EETs in endothelium‐dependent hyperpolarization of porcine coronary artery
Authors: Edwards G , Thollon C , Gardener M J , Félétou M , Vilaine J ‐P , Vanhoutte P M , Weston A H
Published: January 2005

The effects of endothelium‐derived hyperpolarizing factor (EDHF: elicited using substance P or bradykinin) were compared with those of 11,12‐EET in pig coronary artery. Smooth muscle cells were usually impaled with microelectrodes through the adventitial surface.

Substance P (100 nM) and 11,12‐EET (11,12‐epoxyeicosatrienoic acid; 3 μM) hyperpolarized endothelial cells in intact arteries. These actions were unaffected by 100 nM iberiotoxin but were abolished by charybdotoxin plus apamin (each 100 nM).

Substance P (100 nM) and bradykinin (30 nM) hyperpolarized intact artery smooth muscle; Substance P had no effect after endothelium removal.

11,12‐EET hyperpolarized de‐endothelialized vessels by 12.6±0.3 mV, an effect abolished by 100 nM iberiotoxin. 11,12‐EET hyperpolarized intact arteries by 18.6±0.8 mV, an action reduced by iberiotoxin, which was ineffective against substance P. Hyperpolarizations to 11,12‐EET and substance P were partially inhibited by 100 nM charybdotoxin and abolished by further addition of 100 nM apamin.

30 μM barium plus 500 nM ouabain depolarized intact artery smooth muscle but responses to substance P and bradykinin were unchanged. 500 μM gap 27 markedly reduced hyperpolarizations to substance P and bradykinin which were abolished in the additional presence of barium plus ouabain.

Substance P‐induced hyperpolarizations of smooth muscle cells immediately below the internal elastic lamina were unaffected by gap 27, even in the presence of barium plus ouabain.

In pig coronary artery, 11,12‐EET is not EDHF. Smooth muscle hyperpolarizations attributed to ‘EDHF’ are initiated by endothelial cell hyperpolarization involving charybdotoxin‐ (but not iberiotoxin) and apamin‐sensitive K+ channels. This may spread electrotonically via myoendothelial gap junctions but the involvement of an unknown endothelial factor cannot be excluded.

British Journal of Pharmacology (2000) 129, 1145–1154; doi:10.1038/sj.bjp.0703188
The effects of central aromatic amino acid DOPA decarboxylase inhibition on the motor actions of L‐DOPA and dopamine agonists in MPTP‐treated primates
Authors: Treseder Sarah A , Jackson Michael , Jenner Peter
Published: January 2005

Endogenous L‐DOPA may act as a neuromodulator contributing to the production of motor activity. We now investigate the effects of the centrally acting aromatic amino acid dopa decarboxylase (AADC) inhibitor NSD‐1015 (3‐hydroxybenzyl hydrazine) on the motor actions of L‐DOPA and dopamine agonist drugs in MPTP treated common marmosets.

Pretreatment with NSD‐1015 (10–50 mg kg−1; i.p.) worsened baseline motor deficits in MPTP‐treated common marmosets. Similarly, it abolished L‐DOPA (5–18 mg kg−1 s.c.) induced locomotor activity and reversal of disability. NSD‐1015 pretreatment inhibited dopamine formation and elevated L‐DOPA levels in plasma.

The increase in locomotor activity and improvement in disability produced by the administration of the D‐1 agonist A‐86929 (0.03–0.04 mg kg−1 s.c.) or the D‐2 agonist quinpirole (0.05–0.3 mg kg−1 i.p.) was abolished by NSD‐1015 (25 mg kg−1 i.p.) pretreatment. While the effects of a low dose combination of A‐86929 (0.04 mg kg−1 s.c.) and quinpirole (0.05 mg kg−1 i.p.) were inhibited by NSD‐1015 (25 mg kg−1 i.p.), there was little effect on the action of a high dose combination of these drugs (0.08 mg kg−1 A‐86929 and 0.1 mg kg−1 quinpirole).

Following central AADC inhibition with NSD‐1015 (25 mg kg−1 i.p.), locomotor behaviour induced by administration of high dose combinations of A‐86929 (0.08 mg kg−1 s.c.) and quinpirole (0.1 mg kg−1 i.p.) was unaffected by L‐DOPA (5 mg kg−1 s.c.) pretreatment.

These results do not support a role for endogenous L‐DOPA in spontaneous or drug induced locomotor activity. Rather, they strengthen the argument for the importance of endogenous dopaminergic tone in the motor actions of dopamine agonists.

British Journal of Pharmacology (2000) 129, 1355–1364; doi:10.1038/sj.bjp.0703189
Distinct Ca2+ signalling mechanisms induced by ATP and sphingosylphosphorylcholine in porcine aortic smooth muscle cells
Authors: Chin Ting‐Yu , Chueh Sheau‐Huei
Published: January 2005

The increase in the cytosolic Ca2+ concentration ([Ca2+]i) following repetitive stimulation with ATP or sphingosylphosphorylcholine (SPC) in single porcine aortic smooth muscle cells was investigated using the Ca2+ indicator, fura‐2.

The ATP‐induced [Ca2+]i increase resulted from both Ca2+ release and Ca2+ influx. The former was stimulated by phospholipase C activation, while the latter occurred predominantly via the receptor‐operated Ca2+ channels (ROC), rather than the store‐operated Ca2+ channels (SOC) or the voltage‐operated Ca2+ channel (VOC). Furthermore, the P2X5 receptor was shown to be responsible for the ATP‐induced Ca2+ influx.

A reproducible [Ca2+]i increase was induced by repetitive ATP stimulation, but was abolished by removal of extracellular Ca2+ or inhibition of intracellular Ca2+ release using U‐73122 or thapsigargin, and was restored by Ca2+ readdition in the former case.

SPC only caused Ca2+ release, and the amplitude of the repetitive SPC‐induced [Ca2+]i increases declined gradually. However, a reproducible [Ca2+]i increase was seen in cells in which protein kinase C being inhibited, which increased the SPC‐induced Ca2+ influx, rather than IP3 generation.

In conclusion, although the amplitude of the ATP‐induced Ca2+ release, measured when Ca2+ influx was blocked, or of the Ca2+ influx when Ca2+ release was blocked, progressively decreased following repetitive stimulation, the overall [Ca2+]i increase for each stimulation under physiological conditions remained the same, suggesting that the Ca2+ stores were replenished by an influx of Ca2+ during stimulation. The SPC‐induced [Ca2+]i increase resulted solely from Ca2+ release and decreased gradually following repetitive stimulation, but the decrease could be prevented by stimulating Ca2+ influx, further supporting involvement of the intracellular Ca2+ stores in Ca2+ signalling.

British Journal of Pharmacology (2000) 129, 1365–1374; doi:10.1038/sj.bjp.0703190
Mechanisms of suppression of inducible nitric oxide synthase (iNOS) expression in RAW 264.7 cells by andrographolide
Authors: Chiou Wen‐Fei , Chen Chieh‐Fu , Lin Jin‐Jung
Published: January 2005

Andrographolide, an active component found in leaves of Andrographis paniculata, has been reported to exhibit nitric oxide (NO) inhibitory property in endotoxin‐stimulated macrophages, however, the detailed mechanisms remain unclear. In the present study we investigated the effect of andrographolide on the expression of inducible NO synthase (iNOS) mRNA, protein, and enzyme activity in RAW 264.7 macrophages stimulated with lipopolysaccharide (LPS) plus interferon‐γ (IFN‐γ).

RAW 264.7 cells stimulated with LPS/IFN‐γ activated NO production; in this condition andrographolide (1–100 μM) inhibited NO production in a dose‐dependent manner with an IC50 value of 17.4±1.1 μM. Andrographolide also reduces the expression of iNOS protein level but without a significant effect on iNOS mRNA. The reduction of iNOS activity is thought to be caused by decreased expression of iNOS protein.

In a protein stability assay, andrographolide moderately but significantly reduced the amount of iNOS protein as suggested by accelerating degradation. Furthermore, andrographolide also inhibited total protein de novo synthesis as demonstrated by [35S]‐methionine incorporation.

As a whole, these data suggest that andrographolide inhibits NO synthesis in RAW 264.7 cells by reducing the expression of iNOS protein and the reduction could occur through two additional mechanisms: prevention of the de novo protein synthesis and decreasing the protein stability via a post‐transcriptional mechanism. It is also possible that inhibition of iNOS protein expression and NO production under immune stimulation and/or bacteria infection may explain, in part, the beneficial effects of andrographolide as an anti‐inflammatory agent.

British Journal of Pharmacology (2000) 129, 1553–1560; doi:10.1038/sj.bjp.0703191
Modulation by dihydropyridine‐type calcium channel antagonists of cytokine‐inducible gene expression in vascular smooth muscle cells
Authors: Cattaruzza Marco , Wachter Rolf , Wagner Andreas H , Hecker Markus
Published: January 2005

The 1,4‐dihydropyridine nifedipine is frequently used in the therapy of hypertension and heart failure. In addition, nifedipine has been shown to exert distinct anti‐arteriosclerotic effects both in experimental animal models and in patients. In the present study we have investigated the hypothesis that the latter effect of this class of drugs is mediated by an interference with the expression of pro‐arteriosclerotic gene products in the vessel wall. Moreover, to elucidate as to whether nifedipine acts via L‐type calcium channel blockade, its effects were compared to those of another dihydropyridine, Bay w 9798, which has no calcium‐antagonistic properties in concentrations up to 10 μM, as verified by superfusion bioassay.

Both, nifedipine and Bay w 9798, in concentrations ranging from 0.01 to 1 μM, augmented the interleukin‐1β/tumour necrosis factor‐α (IL‐1β/TNF‐α)‐induced expression of the inducible isoform of nitric oxide synthase (iNOS) in rat aortic cultured smooth muscle cells (raSMC) 2–3 fold, as judged by RT–PCR and Western blot analyses.

In contrast, cytokine‐induced mRNA expression of monocyte chemoattractant protein 1 (MCP‐1) in these cells was down‐regulated by more than 60% in the presence of both dihydropyridines, as judged by RT–PCR and Northern blot analyses.

Nuclear run‐on assays and incubation with the transcription‐terminating drug actinomycin D revealed that both drugs acted at the level of mRNA synthesis rather than stability.

These findings suggest that 1,4‐dihydropyridines such as nifedipine affect the expression of both potentially pro‐arteriosclerotic (MCP‐1) and anti‐arteriosclerotic (iNOS) gene products in the vessel wall at the level of transcription, and that these effects are unrelated to their calcium channel‐blocking properties.

British Journal of Pharmacology (2000) 129, 1155–1162; doi:10.1038/sj.bjp.0703192
Histamine H3 receptors mediate inhibition of noradrenaline release from intestinal sympathetic nerves
Authors: Blandizzi Corrado , Tognetti Martina , Colucci Rocchina , Tacca Mario Del
Published: January 2005

The present study investigates whether presynaptic histamine receptors regulate noradrenaline release from intestinal sympathetic nerves. The experiments were performed on longitudinal muscle‐myenteric plexus preparations of guinea‐pig ileum, preincubated with [3H]‐noradrenaline.

In the presence of rauwolscine, electrically‐induced [3H]‐noradrenaline release was inhibited by histamine or R‐α‐methylhistamine, whereas it was unaffected by pyridylethylamine, impromidine, pyrilamine, cimetidine, thioperamide or clobenpropit. The inhibitory effects of histamine or R‐α‐methylhistamine were antagonized by thioperamide or clobenpropit, but not by pyrilamine or cimetidine. In the absence of rauwolscine, none of these drugs modified the release of [3H]‐noradrenaline.

The modulatory action of histamine was attenuated by pertussis toxin and abolished by N‐ethylmaleimide. Tetraethylammonium or 4‐aminopyridine enhanced the evoked tritium outflow and counteracted the inhibitory effect of histamine. However, the blocking effects of tetraethylammonium and 4‐aminopyridine were no longer evident when their enhancing actions were compensated by reduction of Ca2+ concentration in the superfusion medium.

Histamine‐induced inhibition of tritium output was enhanced by ω‐conotoxin or low Ca2+ concentration, whereas it was not modified by nifedipine, forskolin, rolipram, phorbol myristate acetate, H7 or lavendustin A.

The present results indicate that presynaptic H3 receptors, located on sympathetic nerve endings, mediate an inhibitory control on intestinal noradrenergic neurotransmission. It is suggested that these receptors are coupled to Gi/Go proteins which modulate the activity of N‐type Ca2+ channels through a direct link, thus reducing the availability of extracellular Ca2+ at the level of noradrenergic nerve terminals.

British Journal of Pharmacology (2000) 129, 1387–1396; doi:10.1038/sj.bjp.0703194