bluuebunny/crossref_metadata_2025 huggingface dataset:
Action of adenosine receptor antagonists on hypoxia‐induced effects in the rat hippocampus in vitro
Authors: Croning M.D.R. , Zetterstróm T.S.C. , Grahame‐Smith D.G. , Newberry N.R.
Published: July 2012

We have studied three hypoxia‐induced phenomena in the CA1 stratum pyramidale of the rat hippocampal slice: (a) the increase in extracellular potassium ion concentration ([K+]e) measured with ion‐sensitive microelectrodes, (b) the intracellularly‐recorded pyramidal cell hyperpolarization and (c) the extracellularly‐recorded depression of the synaptically‐evoked field potential recorded in stratum pyramidale.

The extracellular potassium ion concentration ([K+]e) rose from 3 mM to 4.1–4.4 mM at a time when the pyramidal cells hyperpolarized by about 6 mV and neurotransmission was virtually abolished.

Presumed glial cells depolarized in response to hypoxia. The shape and time course of this response was remarkably similar to the rise in [K+]e so induced. This is consistent with findings that glial cell membrane potential is dependent on transmembrane K+ gradient.

We investigated the effects of theophylline (100 μm) and 1,3‐dipropyl‐8‐cyclopentylxanthine (DPCPX, 0.1 μm) on these effects. We have found that these compounds attenuated by about half the hypoxia‐induced increase in [K+]e; however, they did not reduce the hypoxia‐induced hyperpolarization. We have confirmed that they dramatically reduced the suppression of excitatory transmission caused by the hypoxia. We conclude that adenosine A1 receptors may be involved in the alteration of K+ homeostasis in the hippocampal shoe during hypoxia.
Effects of the ETA/ETB receptor antagonist, bosentan on endothelin‐1‐induced myocardial ischaemia and oedema in the rat
Authors: Filep János G. , Fournier Alain , Földes‐Filep Éva
Published: July 2012

The purposes of this study were to assess the role of ETB receptors in mediating endothelin‐1 (ET‐1)‐induced myocardial ischaemia and oedema in rats and to study the inhibitory action of the novel non‐pep tide ETA/ETB receptor antagonist, bosentan on these actions of ET‐1

Intravenous bolus injection of ET‐1 (1 nmol kg−1) into anaesthetized rats produced marked ST segment elevation of the electrocardiogram without causing arrhythmias. ST segment elevation developed within 30–50 s and persisted for at least 30 min following injection of the peptide

Pretreatment of the animals with bosentan (10 mg kg−1, i.v.) inhibited on average by 96% the ST segment elevation elicited by ET‐1 (1 nmol kg−1) compared to the 82% inhibition observed with the ETA receptor‐selective antagonist, FR 139317 (2.5 mg kg−1, i.v.)

Bolus injection of ET‐1 (1 nmol kg−1, i.v.) to conscious chronically catheterized rats evoked a transient depressor response followed by a prolonged pressor effect. Corresponding to changes in blood pressure, a transient tachycardia and a sustained bradycardia were observed. ET‐1 (1 nmol kg−1) enhanced albumin extravasation by 119 and 93% in the left ventricle and right atrium, respectively, as measured by the local extravascular accumulation of Evans blue dye

Pretreatment of the animals with bosentan (10 mg kg−1) inhibited by 71 and 90% the depressor and pressor actions of ET‐1 (1 nmol kg−1) and the accompanying tachycardia and bradycardia, respectively. FR 139317 (2.5 mg kg−1) attenuated the pressor response to ET‐1 and accompanying bradycardia by 75%, without affecting the depressor action and accompanying tachycardia. ET‐1‐induced albumin extravasation was completely inhibited by bosentan (10 mg kg−1) both in the left ventricle and right atrium, compared to the 86% inhibition observed with FR 139317 (2.5 mg kg−1)

Like ET‐1, the ETB receptor‐selective agonist, IRL 1620 (0.3 and 1 nmol kg−1, i.v.) also produced dose‐dependent ST segment elevation in anaesthetized rats and enhanced albumin extravasation (up to 141% of control) in the left ventricle and right atrium, respectively, in conscious rats. These effects of IRL 1620 were completely prevented by bosentan (10 mg kg−1)

These results indicate that ETB receptors, albeit to a lesser extent than ETA receptors, are also involved in mediating ET‐1‐induced myocardial ischaemia and oedema in the rat, and suggest the therapeutic potential for bosentan in the treatment of ischaemic myocardial diseases.
Effects of 5‐HT receptor agonists on depolarization‐induced [3H]‐noradrenaline release in rabbit hippocampus and human neocortex
Authors: Allgaier C. , Warnke P. , Stangl A.P. , Feuerstein T.J.
Published: July 2012

The present study attempted to determine whether noradrenaline (NA) release in rabbit hippocampus and human neocortex is modulated by presynaptic 5‐hydroxytryptamine (5‐HT) receptors

Slices of rabbit hippocampus and human neocortex, loaded with [3H]‐noradrenaline ([3H]‐NA) were superfused and the effects of 5‐hydroxytryptamine (5‐HT) receptor ligands on electrically evoked [3H]‐NA release were investigated

In rabbit hippocampus, 5‐HT, 5‐carboxamidotryptamine (5‐CT; 32 μm) and 2‐CH3‐5‐HT (32 μm) increased [3H]‐NA release elicited with 360 pulses/3 Hz. Facilitation of transmitter release was not influenced by the 5‐HT3 receptor antagonist, tropisetron but was prevented by the α2‐adrenoceptor antagonist, rauwolscine. When autoinhibition was avoided by stimulating the tissue with 4 pulses/100 Hz (pseudo‐one pulse‐(POP) stimulation), 2‐CH3‐5‐HT decreased evoked transmitter release, whereas 5‐HT and 5‐CT had no effect. Inhibition caused by 2‐CH3‐5‐HT was not affected by tropisetron but counteracted by the α2‐adrenoceptor ligands, clonidine and rauwolscine. Inhibition caused by clonidine was diminished in the presence of 5‐CT or 2‐CH3‐5‐HT

In human neocortex, [3H]‐NA release elicited with 360 pulses/3 Hz was increased by 10 μm 5‐HT and 32 μm 5‐CT, whereas 2‐CH3‐5‐HT was ineffective. [3H]‐NA release evoked with a modified POP stimulation (2 bursts of 4 pulses/100 Hz, 3.5 min apart) was not affected by 2‐CH3‐5‐HT or 5‐CT

The present results indicate that 5‐HT, 2‐CH3‐5‐HT and 5‐CT can act on presynaptic a2‐autoreceptors as partial agonists (2‐CH3‐5‐HT; in rabbit hippocampal tissue) or antagonists (5‐HT and 5‐CT; in tissue of rabbit hippocampus and human neocortex). Furthermore the existence of autoinhibition dictates whether these drugs cause facilitation of release, inhibition or have no effect.
Involvement of superoxide and xanthine oxidase in neutrophil‐independent rat gastric damage induced by NO donors
Authors: Lamarque D. , Whittle B.J.R.
Published: July 2012

Nitric oxide (NO) and the superoxide anion can interact to form the cytotoxic moiety, peroxynitrite. The involvement and potential source of superoxide in the gastric mucosal damage induced by local infusion of NO donors, has now been investigated in the pentobarbitone‐anaesthetized rat

Local intra‐arterial infusion of the NO donor, sodium nitroprusside (40 μg kg−1 min−1) for 10 min induced macroscopically apparent gastric mucosal injury

This mucosal damage was dose‐dependently reduced by prior administration of a systemically acting form of superoxide dismutase conjugated with polyethylene glycol (500–2000 iu kg−1, i.v.)

Likewise, the mucosal damage induced by nitroprusside was dose‐dependently reduced by prior administration of the xanthine oxidase inhibitor, allopurinol (20–100 mg kg−1, i.p. or 100 mg kg−1, p.o.)

Pretreatment with allopurinol (100 mg kg−1, i.p.) also reduced the mucosal injury induced by local intra‐arterial infusion of the nitrosothiol, S‐nitroso‐N‐acetyl‐penicillamine (40 μg kg−1 min−1), but not that induced by local infusion of endothelin‐1 (5 pmol kg−1 min−1), indicating specificity of action

Prior administration (4h) of rabbit anti‐rat neutrophil serum (0.4 ml kg−1, i.p.), which reduced circulating neutrophils by 90%, did not significantly protect against mucosal injury induced by nitroprusside

Intravenous administration of the platelet‐activating factor receptor antagonists, WEB 2086 (1 mg kg−1) or BN 52021 (10 mg kg−1), or the thromboxane synthase inhibitor, OKY 15181 (25 mg kg;−1), did not modify mucosal damage induced by nitroprusside, showing lack of involvement of these neutrophil‐derived mediators

These findings indicate the involvement of superoxide in the injurious actions of the NO donors, implicating a cytotoxic role of peroxynitrite. Xanthine oxidase, but not neutrophils, appears to be a source of the superoxide.
Differential modulation of voltage‐activated conductances by intracellular and extracellular cyclic nucleotides in leech salivary glands
Authors: Everill Brian , Berry Michael S.
Published: July 2012

Two‐electrode voltage clamp was used to study the effects of adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) and guanosine 3′:5′‐cyclic monophosphate (cyclic GMP) on voltage‐dependent ion channels in salivary gland cells of the leech, Haementeria ghilianii

Intracellular cyclic AMP specifically blocked delayed rectifier K+ channels. This was shown by use of 3‐isobutyl‐l‐methylxanthine (IBMX, a phosphodiesterase inhibitor), forskolin (an activator of adenylyl cyclase) and intracellular injection of cyclic AMP and its dibutyryl and 8‐bromo analogues. Cyclic AMP appeared to be the second messenger for the putative neuroglandular transmitter, 5‐hydroxytryptamine

Intracellular injection of cyclic GMP specifically potentiated high‐voltage‐activated (HVA) Ca2+ current and the effect was mimicked by zaprinast, an inhibitor of cyclic GMP‐dependent phosphodiesterase

Extracellularly, cyclic GMP and cyclic AMP specifically decreased the amplitude and increased the rate of inactivation of HVA Ca2+ current. These effects of the cyclic nucleotides are identical to those known for extracellular ATP, which activates a presumed purinoceptor. The pyrimidine nucleotide, UTP, was almost equipotent to ATP (threshold dose < 10−6 M), indicative of a vertebrate‐type nucleotide receptor. However, suramin (5 × 10−5 M), a non‐specific P2‐receptor antagonist, failed to block the effects of 5 × 10−6 m ATP (higher suramin doses could not be reliably tested because of the depolarization and increase in membrane conductance produced by the drug)

Activation of the putative purinoceptor by ATP did not affect inward rectifier Na+/K+ current which is known to be potentiated by intracellular cyclic AMP and reduced by intracellular cyclic GMP

The preparation may provide a useful model for study of nucleotide actions, and interactions, in channel modulation. It has technical advantages such as large cells (1200 μm in diameter) which lack intercellular coupling and may be individually dissected for biochemical studies.
Restoration of blood pressure by choline treatment in rats made hypotensive by haemorrhage
Authors: Ulus Ismail. H. , Arslan B. Yener , Savci Vahide , Kiran Burhan K.
Published: July 2012

Intracerebroventricular (i.c.v.) injection of choline (25–150 μg) increased blood pressure in rats made acutely hypotensive by haemorrhage. Intraperitoneal administration of choline (60 mg kg−1) also increased blood pressure, but to a lesser extent. Following i.c.v. injection of 25 μg or 50 μg of choline, heart rate did not change, while 100 μg or 150 μg i.c.v. choline produced a slight and short lasting bradycardia. Choline (150 μg) failed to alter the circulating residual volume of blood in haemorrhaged rats

The pressor response to i.c.v. choline (50 μg) in haemorrhaged rats was abolished by pretreatment with mecamylamine (50 μg, i.c.v.) but not atropine (10 μg, i.c.v.). The pressor response to choline was blocked by pretreatment with hemicholinium‐3 (20 μg, i.c.v.)

The pressor response to i.c.v. choline (150 μg) was associated with a several fold increase in plasma levels of vasopressin and adrenaline but not of noradrenaline and plasma renin

The pressor response to i.c.v. choline (150 μg) was not altered by bilateral adrenalectomy, but was attenuated by systemic administration of either phentolamine (10 mg kg−1) or the vasopressin antagonist [β‐mercapto‐β,β‐cyclopenta‐methylenepropionyl1, O‐Me‐Tyr2,Arg8‐vasopressin (10 μg kg−1)

It is concluded that the precursor of acetylcholine, choline, can increase and restore blood pressure in acutely haemorrhaged rats by increasing central cholinergic neurotransmission. Nicotinic receptor activation and an increase in plasma vasopressin and adrenaline level appear to be involved in this effect of choline.
Differential effects of acute and chronic fluoxetine administration on the spontaneous activity of dopaminergic neurones in the ventral tegmental area
Authors: Prisco Simona , Esposito Ennio
Published: July 2012

Electrophysiological techniques were used to study the effects of fluoxetine and citalopram on the basal activity of dopaminergic neurones in the ventral tegmental area (VTA) and substantia nigra, pars compacta (SNc) of rats

Acute i.v. injection of fluoxetine (20–1280 μg kg−1) caused a dose‐dependent inhibition of the firing rate of VTA dopaminergic neurones, but did not affect the activity of dopaminergic cells in the SNc. Citalopram (20–1280 μg kg−1, i.v.) inhibited the firing rate of dopaminergic neurones in the VTA, but its effect (maximal inhibition: 14 ±7%) was less pronounced than that of fluoxetine (maximal inhibition: 34 ±7%)

Pretreatment with mesulergine (80 μg kg−1, i.v.), a 5‐hydroxytryptamine2C/2B (5‐HT2C/2B) receptor antagonist, blocked the inhibitory effect of fluoxetine on VTA dopaminergic cells. Selective lesions of 5‐hydroxytryptaminergic neurones by the neurotoxin, 5,7‐dihydroxytryptamine (5,7‐DHT), abolished the fluoxetine‐induced reduction of VTA dopaminergic activity

In a series of experiments, fluoxetine (10 mg kg−1, i.p.) was administered once daily for 21 consecutive days. Acute i.v. administration of fluoxetine (20–1280 μg kg−1, 72 h after the last i.p. injection) did not cause any change in the basal firing rate of VTA dopaminergic neurones in treated rats, whereas it induced the typical inhibitory effect in control animals. A group of rats chronically treated with fluoxetine, received i.v. w‐chlorophenylpiperazine (mCPP; 10–320 μg kg−1), a 5‐HT2C/2B receptor agonist. This drug significantly inhibited VTA dopaminergic function in control rats, but did not modify the basal activity of dopaminergic cells in animals given chronic fluoxetine

It is concluded that fluoxetine inhibits dopaminergic function in the VTA by enhancing the synaptic levels of 5‐HT, which possibly acts through the 5‐HT2C/2B receptor subtype. Repeated treatment with fluoxetine induces tolerance to its inhibitory effect on dopaminergic activity, possibly as a consequence of down‐regulation of 5‐HT2C/2B receptors. The effects of fluoxetine on VTA dopaminergic cell activity might be relevant for its therapeutic actions and may explain the origin of the reported cases of akathisia.
Vasoconstrictor responses to the P2X‐purinoceptor agonist (β, γ‐methylene‐L‐ATP in human cutaneous and renal blood vessels
Authors: von Kiigelgen Ivar , Krumme Bernd , Schaible Ulrike , Schollmeyer Peter J. , Rump Lars Christian
Published: July 2012

Strips of human saphenous veins and of human renal arteries and veins were superfused with Krebs‐Henseleit solution at 37°C. Constrictor responses were elicited by exogenous noradrenaline and the P2X‐purinoceptor‐selective agonist, β,γ‐methylene‐L‐ATP

In human saphenous veins, β,γ‐methylene‐L‐ATP (0.3–30 μm; EC50 2.2 μm) induced marked constrictor responses. The maximal response to β,γ‐methylene‐L‐ATP was similar to the maximal response to noradrenaline. The P2‐purinoceptor antagonist suramin (30 μm) shifted the concentration‐response curve of β,γ‐methylene‐L‐ATP to the right (apparent pKB value 4.8); suramin (100 μm) markedly inhibited the responses to P,y‐methylene‐L‐ATP. The preferential P2X‐purinoceptor antagonist, pyridoxal‐phosphate‐6‐azophenyl‐2,4,‐disulphonic acid (PPADS; 3 μm) slightly reduced the response to β,γ‐methylene‐L‐ATP. At a ten times higher concentration (30 μm), PPADS almost abolished the responses to p,y‐methylene‐L‐ATP. PPADS (30 μm), in contrast, caused no significant change in the concentration‐response curve of noradrenaline

In extrarenal and intrarenal arteries, EC50 values and maximal responses to noradrenaline were similar when compared with responses to noradrenaline in saphenous veins. Noradrenaline also constricted extrarenal veins. However, in contrast to the results obtained on saphenous veins, p,y‐methylene‐L‐ATP caused almost no constrictor responses in extrarenal veins and arteries and only moderate responses in intrarenal arteries

The results demonstrate marked differences in responsiveness of human blood vessels to the selective P2X‐purinoceptor agonist, β,γ‐methylene‐L‐ATP, suggesting tissue differences in the occurrence or operation of P2X‐purinoceptors in human vascular tissues. Moreover, the results indicate that PPADS blocks P2X‐purinoceptors in human isolated blood vessels as previously demonstrated in animal blood vessels.
Participation of 5‐HT1‐like and 5‐HT2A receptors in the contraction of human temporal artery by 5‐hydroxytryptamine and related drugs
Authors: Verheggen Raphaela , Freudenthaler Stefan , Meyer‐Dulheuer Frauke , Kaumann Alberto J.
Published: July 2012

We investigated the hypothesis that, as in some other large human arteries, 5‐HT‐induced contraction of the temporal artery is mediated through two co‐existing receptor populations, 5‐HT1‐ like‐and 5‐HT2A. Temporal arterial segments were obtained from patients undergoing brain surgery and rings prepared set up to contract with 5‐HT and related agents. Fractions of maximal 5‐HT responses mediated through 5‐HT1‐like and 5‐HT2A receptors, f1, and f2 = 1 ‐ f1, were estimated by use of the 5‐HT2A‐selective antagonist ketanserin.

In rings with intact endothelium 5‐HT evoked contractions with a ‐log EC50, m of 7.0. Ketanserin (10–1000 nM) antagonized part of the 5‐HT‐induced contractions. Ketanserin‐resistant components of 5‐HT‐induced contractions were found with ‐log EC50, m of 6.9 and f1 of 0.17 (100 nM ketanserin) and ‐ log EC50, m of 6.4 and f1 of 0.20 (1000 nM ketanserin).

In rings with endothelial function attenuated by enzymatic treatment, 5‐HT caused contractions with a ‐log EC50, m of 7.2 that were partially blocked by ketanserin. Ketanserin‐resistant components of 5‐HT‐induced contractions were found with ‐log EC50, m 7.4 and f1 of 0.16 (100 nM ketanserin) and ‐log EC50, m of 7.5 and f1 of 0.14 (1000 nM ketanserin).

The ketanserin‐resistant component of 5‐HT‐evoked contraction was blocked by methiothepin (100–1000 nM) consistent with mediation through 5‐HT1‐like receptors.

In rings with intact endothelium the 5‐HT1‐like‐selective agonist, sumatriptan, caused small contractions with a ‐log EC50, m of 6.5 and intrinsic activity of 0.21 with respect to 5‐HT that were resistant to blockade by 1000 nM ketanserin but antagonized by 100 nM methiothepin.

In rings with intact endothelium the 5‐HT2A receptor partial agonist SK&F 103829 (2,3,4,5‐ tetrahydro‐8[methyl sulphonyl]‐1H3‐benzazepin‐7‐01 methensulphonate) contracted rings with a ‐log EC50, m of 5.0 and an intrinsic activity of 0.49 with respect to 5‐HT; the effects were antagonized by ketanserin 1000 nM.

We conclude that 80–86% of the maximum 5‐HT‐evoked contraction of human temporal artery is mediated through 5‐HT2A receptors, the remainder through 5‐HT1‐like‐receptors, regardless of whether or not endothelium is functional. The 5‐HT1‐like‐receptors are more likely to be 5‐HT1Dβ receptors than 5‐HT1Dα receptors and sumatriptan is a full agonist for these receptors. As found in arteries of other species, SK&F 103829 is a partial agonist for 5‐HT2A receptors of human temporal artery.
Release of vasopressin and oxytocin by excitatory amino acid agonists and the effect of antagonists on release by muscarine and hypertonic saline, in the rat in vivo
Authors: Bisset G.W. , Fairhall K.M.
Published: July 2012

It has been claimed that glutamate is the dominant excitatory neurotransmitter in neuroendocrine regulation. The evidence is derived mainly from in vitro experiments.

We have investigated in vivo a possible role of excitatory amino acids (EAAs) in the neural control of release of vasopressin (AVP) and oxytocin from the neurohypophysis.

In rats under ethanol anaesthesia in which a diuresis was maintained by a constant fluid load, the i.c.v. injection of glutamate and the synthetic agonists α‐amino, 3‐hydroxy‐5‐methyl‐isoxazole‐4‐propionate (AMPA) and N‐methyl‐D‐aspartate (NMDA) produced an antidiuretic response (ADR) which was abolished by an AVP antagonist. For AMPA and NMDA it was shown that this ADR was accompanied by increased urinary excretion of AVP and oxytocin.

The selectivity of antagonists was tested in this system. D‐2‐Amino‐5‐phosphonopentanoate (D‐AP5) blocked the responses to NMDA but not to AMPA; 6‐cyano‐7‐nitroquinoxaline‐2, 3‐dione (CNQX) blocked the responses to both agonists.

The ADR to muscarine and hypertonic saline i.c.v., and the increase in excretion of AVP and oxytocin in response to muscarine, were blocked by CNQX but not by D‐AP5.

The results suggest that hypertonic saline releases AVP and muscarine releases both AVP and oxytocin, at least in part, by activating a glutaminergic input to the SON and PVN involving an AMPA receptor. This input could function as a terminal interneurone in afferent neural pathways to these nuclei.
Increase in insulin release from rat pancreatic islets by quinolone antibiotics
Authors: Maeda N. , Tamagawa T. , Niki I. , Miura H. , Ozawa K. , Watanabe G. , Nonogaki K. , Uemura K. , Iguchi A.
Published: July 2012

The present study was undertaken to elucidate the mechanism(s) of hypoglycaemia caused by quinolone antibiotics. We investigated the effects of various quinolone antibiotics on insulin release in rat pancreatic islets.

At a non‐stimulatory concentration of 3 mM glucose, lomefloxacin (LFLX) or sparfloxacin at 1 mM and pipemidic acid (0.1‐1 mM) induced slight insulin release but tosufloxacin or enoxacin up to 100 μm did not.

At the stimulatory concentration of 10 mM glucose, all quinolones augmented insulin release in a dose‐dependent manner. LFLX (100 μm) shifted the dose‐response curve of glucose‐induced insulin release to the left without altering the maximal response.

At 10 mM glucose, LFLX (100 μm) increased insulin release augmented by forskolin (5 μm) or 12‐0‐tetradecanoyl phorbol‐13‐acetate (100 nM) but not by raising the K+ concentration from 6 to 25 mM.

Verapamil (50 μm) or diazoxide (50–400 μm) antagonized the insulinotropic effect of LFLX.

These data suggest that quinolone antibiotics may cause hypoglycaemia by increasing insulin release via blockade of ATP‐sensitive K+ channels.
Rise in cytosolic Ca2+ concentration induced by P2‐purinoceptor activation in isolated myocytes from the rat gastrointestinal tract
Authors: Blottière Hervé M. , Loirand Gervaise , Pacaud Pierre
Published: July 2012

The changes in the free cytosolic Ca2+ concentration ([Ca2+]i) in response to agonists of P2‐purinoceptors were studied in myocytes isolated from the longitudinal muscle layer of different regions of the rat gastrointestinal tract (stomach, jejunum, ileum, caecum and colon). [Ca2+]i was estimated by emission from the fluorescent dye, indo‐1.

ATP and the P2Y‐purinoceptor agonist, 2‐methylthio‐ATP (2‐MeSATP), transiently increased [Ca2+]i in single myocytes from all segments of the gastrointestinal tract, whereas α,β‐methylene‐ATP, a P2X‐purinoceptor agonist, had no effect.

The rise in [Ca2+]i induced by ATP and 2‐MeSATP was maintained in Ca2+‐free solution but was abolished by depletion of the intracellular store with thapsigargin (1 μm).

Single myocytes from stomach, caecum and colon also responded to UTP by a transient increase in [Ca2+]i.

Individual myocytes responded to ATP, 2‐MeSATP and UTP in a nearly all‐or‐nothing manner. The increasing of agonist concentration enhanced the number of responding cells but did not increase the amplitude of the [Ca2+]i rise.

These results suggest that myocytes from the longitudinal layer of gastrointestinal muscle do not possess functional P2X‐purinoceptors and that agonists of P2Y and P2U‐purinoceptors induced a rise in [Ca2+]i, probably via an all‐or‐nothing mobilization of Ca2+ from intracellular stores.
Evidence for functional α2D‐adrenoceptors in the rat intestine
Authors: Liu Lu , Coupar Ian M.
Published: July 2012

Peristaltic contractions were induced in segments of rat ileum by raising the intramural pressure. A mean pressure of 4.3±0.2 cmH20 (n = 112) was required to initiate rhythmic contractions of the longitudinal muscle (preparatory phase) and associated volume expulsions caused by circular muscle contraction. The frequency of peristalsis remained constant over two 15 min periods of stimulation.

The α2‐adreoceptor agonists, clonidine, UK 14,304, B‐HT 920, B‐HT 933 and the selective α2A‐ agonists, oxymetazoline and guanfacine, caused concentration‐dependent inhibition of peristaltic contractions. The potency order and IC50 values (nM) were: clonidine (2.81)≥oxymetazoline (4.23) ≥ UK 14,304 (4.48) ≥ guanfacine (5.51) > B‐HT 920 (78.72) > B‐HT 933 (442.48).

High concentrations of clonidine, amounting to more than 10 and 100 times the IC50 value in the peristaltic reflex (30 and 300 nM respectively), failed to inhibit the cholinergic contractile response to transmural electrical stimulation over the range of 2.5 to 40 Hz.

α2‐Adrenoceptor antagonists were used to determine the subtype of presynaptic α2‐adrenoceptor in rat ileum. All the antagonists tested caused parallel shifts to the right of the concentration‐response regression line to clonidine and none, except ARC 239, influenced the rate of peristalsis. However, ARC 239 significantly decreased the frequency of control period peristaltic contractions. The order of affinity of the antagonists against clonidine (pKB values in parentheses) was RX 821002 (8.99) > phentolamine (8.07) > BRL 44408 (7.43) ≥ rauwolscine (7.41) ≥ yohimbine (7.28) > prazosin (5.86) ≥ ARC 239 (5.74).

These results, when compared with binding and functional data from various other tissues and cell lines, are consistent with the presence of presynaptic α2D‐adrenoceptors in rat ileum. Further evidence is provided that this subtype of α2‐adrenoceptor is probably located proximal to the final cholinergic neurones in the reflex arc.
Oxytocin receptors expressed and coupled to Ca2+ signalling in a human vascular smooth muscle cell line
Authors: Yazawa Hidenori , Hirasawa Akira , Horie Kuniko , Saita Yuji , Iida Emi , Honda Kazuo , Tsujimoto Gozoh
Published: July 2012

In a human vascular smooth muscle cell line (HVSMC), binding experiments with [3H]‐arginine8‐vasopressin (AVP) have shown the existence of a homogeneous population of binding sites with affinity (Kd value) of 0.65 nM and a maximum number of binding sites (Bmax) of 122 fmol mg−1 protein.

Nonlabelled compounds compete for [3H]‐AVP binding in the HVSMC membrane with an order of potency of oxytocin > lyspressin ≥ AVP > Thr4, Gly7‐oxytocin > (β‐mercapto‐β‐β‐cyclopentamethylenepropionyl‐O‐Me Tyr2, Arg8) vasopressin > desmopressin > OPC21268 > OPC31260. This order was markedly different from that observed in rat vascular smooth muscle cells (A10), a well‐established V1A receptor system.

In HVSMC both oxytocin and AVP increased inositol 1,4,5‐trisphosphate (IP3) production and [Ca2+]i response, but the efficacy of the responses was greater for oxytocin than AVP.

Reverse transcription‐polymerase chain reaction (RT‐PCR) assay detected only oxytocin receptor but not V1A or V2 receptors in HVSMC, whereas only V1A receptors were found in A10 cells.

In conclusion, in HVSMC only oxytocin receptors are expressed among the vasopressin receptor family, and they coupled to phosphatidyl inositol (PI) turnover/Ca2+ signalling. This unexpected observation should provide new insight into the functional role of the oxytocin receptor in a human vascular smooth muscle cell line.
Suppression of reperfusion‐induced arrhythmias with combined administration of 5‐HT2 and thromboxane A2 antagonists
Authors: Shaw Linda A. , Coker Susan J.
Published: July 2012

The effects of the 5‐HT2 antagonist, ICI 170,809 and the thromboxane A2 antagonist, ICI 192,605, given alone and in combination (n = 12 per group), were examined in anaesthetized rats. Haemodynamics and arrhythmias induced by permanent coronary artery occlusion or by reperfusion after 5 min of ischaemia were monitored.

In a study on reperfusion‐induced arrhythmias, the only significant effect of ICI 170,809 (1 mg kg−1, i.v.) was a reduction in the number of ventricular premature beats (VPBs). ICI 192,605 (1 mg kg−1 min−1, i.v.) did not alter reperfusion‐induced arrhythmias. However, in combination, when compared with controls, these drugs caused significant reductions in the incidence of ventricular tachycardia (VT), 100% to 58%; ventricular fibrillation (VF), 92% to 33%; and the mortality due to sustained VF, 67% to 17%. There was also a significant reduction in the number of VPBs following reperfusion.

In a second study with lower doses of drugs, ICI 170,809 (0.3 mg kg−1) and ICI 192,605 (0.3 mg kg−1 min−1) had no significant effects on reperfusion‐induced arrhythmias either alone or in combination.

A third study examined the effects of the higher doses of the drugs on ischaemia‐induced arrhythmias. Neither drug alone, nor in combination, altered the incidence of ischaemia‐induced VT, VF, the mortality, or the number of VPBs.

These results indicate that, in contrast to the administration of either drug alone, combined administration of a 5‐HT2 antagonist and a thromboxane A2 antagonist caused marked suppression of reperfusion‐induced but not ischaemia‐induced arrhythmias.
Different pharmacological characteristics in L6 and C2C12 muscle cells and intact rat skeletal muscle for amylin, CGRP and calcitonin
Authors: Pittner Richard A. , Wolfe‐Lopez Deborah , Young Andrew A. , Beaumont Kevin
Published: July 2012

We compared the ability of rat amylin, rat calcitonin gene‐related peptide (CGRP) and rat and salmon calcitonins to elevate cyclic AMP levels and to inhibit [U‐14C]‐glucose incorporation into glycogen in insulin‐stimulated intact rat soleus muscle and in two cell lines derived from rodent skeletal muscle, L6 and C2C12

In intact soleus muscle, both amylin (EC50s of 0.7‐6.1 nM) and salmon calcitonin (EC50s of 0.5‐1.4 nM) were more potent than CGRP (EC50s of 5.6–15.8 nM) and were much more potent than rat calcitonin (EC50s of 50–137 nM) at stimulating cyclic AMP production, activating glycogen phosphorylase and inhibiting insulin‐stimulated [14C]‐glycogen formation.

In contrast, in both L6 and C2C12 cells, CGRP (EC50s of 0.042‐0.12 nM) stimulated cyclic AMP formation and inhibited insulin‐stimulated [U‐14C]‐glucose incorporation into glycogen approximately 1000 times more potently than amylin (EC50s 34–240 nM), while salmon calcitonin was without measurable effect.

There was a correlation between elevation of cyclic AMP and inhibition of insulin‐stimulated [U‐14C]‐glucose incorporation into glycogen evoked by these peptides in both intact muscle (r2 = 0.69, P <0.0004) and muslce cell lines (r2 = 0.96, P <0.0001).

In conclusion, the effects of amylin, CGRP, and calcitonin on soleus muscle glycogen metabolism appear to be mediated by adenylyl cyclase‐coupled receptors which show a pharmacological profile similar to high affinity amylin binding sites that have been previously reported in rat brain. In contrast, the effects of amylin and CGRP in L6 and C2C12 rodent muscle cell lines appear to be mediated by adenylyl cyclase‐coupled receptors that behave like CGRP receptors.
Modulation of vasorelaxant responses to potassium channel openers by basal nitric oxide in the rat isolated superior mesenteric arterial bed
Authors: McCulloch Audrey I. , Randall Michael D.
Published: July 2012

We have used the isolated buffer‐perfused mesenteric arterial bed of the rat to assess the modulation of vasorelaxation to potassium channel openers (KCOs) by basal nitric oxide.

The dose‐response curves to the KCOs, levcromakalim and pinacidil, in preconstricted preparations were significantly shifted to the left in the presence of the nitric oxide synthase inhibitor (100 μm) NG‐nitro‐L‐arginine methyl ester (levcromakalim, ED50=4.47±0.70 nmol vs. 1.73±0.26 nmol, P<0.001; pinacidil, ED50=16.1±4.8 nmol vs. 5.43±1.10 nmol, P<0.001). The vasorelaxant responses to papaverine, a vasodilator which acts independently of potassium channels was unaffected by NG‐nitro‐L‐arginine methyl ester (L‐NAME).

Removal of the endothelium, by perfusion with the detergent CHAPS (0.3%), significantly (P<0.001) increased the potency of levcromakalim as a vasodilator (ED50 4.47±0.70 nmol vs. 2.59±0.31 nmol). The subsequent administration of L‐NAME following perfusion with CHAPS did not lead to any additional enhancement of responses to levcromakalim.

The presence of the non‐selective adenosine antagonist, 8‐phenyltheophylline (8‐PT, 10 μm) significantly (P<0.001) shifted the dose‐response curve to levcromakalim to the left (ED50 4.47±0.70 nmol vs. 1.11±0.32 nmol). In the presence of both L‐NAME and 8‐PT, the dose‐response curve to levcromakalim was also significantly (P<0.01) shifted to the left compared with control (ED50 in the presence of both L‐NAME and 8‐PT was 0.42±0.08 nmol).

The presence of 8‐bromo cyclic GMP (10 μm) reversed the increase potency of levcromakalim, observed following inhibition of nitric oxide synthase (ED50 in the presence of L‐NAME was 0.59±0.01 nmol and in the presence of 8‐bromo cyclic GMP plus L‐NAME the ED50 was 3.17±0.80 nmol). However in the absence of L‐NAME, the cell permeable analogue of cyclic GMP, 8‐bromo cyclic GMP, did not affect the dose‐response curve to levcromakalim compared with control (control ED50 value was 4.16±0.52 nmol vs. 3.85±1.13 nmol in the presence of 8‐bromo cyclic GMP).

The present investigation demonstrates that both basal nitric oxide and adenosine modulate vasorelaxation to the KCOs levcromakalim and pinacidil. The modulatory effect of nitric oxide may be mediated via cyclic GMP.
Enhancement of sympathetic purinergic neurotransmission in the guinea‐pig isolated vas deferens by the novel ecto‐ATPase inhibitor ARL 67156
Authors: Westfall T.D. , Kennedy C. , Sneddon P.
Published: July 2012

Field stimulation of the sympathetic nerves of the guinea‐pig isolated vas deferens with trains of pulses for 20 s at 1–8 Hz produced characteristic biphasic contractions. The effect of the novel ecto‐ATPase inhibitor, 6‐N,N‐diethyl‐D‐β,γ‐dibromomethyleneATP (ARL 67156, formerly known as FPL 67156), on the magnitude of the initial, predominantly purinergic peak of this response was studied in order to determine the influence of enzymatic degradation of adenosine 5′‐triphosphate (ATP) on its action as a neurotransmitter.

The peak magnitude of the response to nerve stimulation was significantly increased in a concentration‐dependent manner by ARL 67156 (5–100 μm) and the size of the neurogenic response at 4 Hz was approximately doubled in the presence of ARL 67156 (100 μm).

ARL 67156 (100 μm) has a rapid onset of action. The enhancing effect on neurogenic contractions was maximal after 10 min, was well maintained for at least 30 min and was rapidly reversed, with responses returning to control levels 10 min after washout.

The neurogenic contraction in the presence of prazosin (0.1 μm) was purely purinergic, as it was abolished by the P2‐purinoceptor antagonist, PPADS (100 μm). ARL 67156 (100 μm) produced a similar degree of enhancement of neurogenic responses in the absence and presence of prazosin, supporting the view that the enhancing effects of ARL 67156 on neurogenic contractions result from potentiation of the action of ATP.

Exogenous ATP and α,β‐methyleneATP produced rapid transient contractions. Responses to ATP were increased in magnitude and duration in the presence of ARL 67156 (100 μm), whereas those to the stable analogue, α,β‐methyleneATP were not significantly affected.

Contractions to exogenous noradrenaline (10 μm) and KCl (40 mM) were significantly enhanced by ARL 67156 (100 μm), but this potentiation was abolished by PPADS (100 μm). Therefore, this effect of the ecto‐ATPase inhibitor may be due to a build up of endogenous ATP, increasing the sensitivity of the smooth muscle to other agonists.

It is concluded that ARL 67156 potentiates the action of ATP, and that when ATP acts as a neurotransmitter its postjunctional actions are greatly attenuated by enzymatic degradation.
Modulation by nitric oxide and prostaglandin of the renal vascular response to angiotensin II (3–8)
Authors: Yoshida Makoto , Kikukawa Masanobu , Hisa Hiroaki , Satoh Susumu
Published: July 2012

The aim of this study was to investigate the renal vascular response to angiotensin II (3–8) (AIV). The effect of the nitric oxide synthase inhibitor, NG‐nitro‐L‐arginine methyl ester (L‐NAME) or the cyclo‐oxygenase inhibitor, indomethacin on the AIV‐induced response was examined in anaesthetized spontaneously hypertensive rats (SHR) and normotensive Wistar‐Kyoto rats (WKY).

Intrarenal infusion of AIV produced a biphasic vasoconstrictor response. The vasoconstriction developed rapidly to reach a maximum followed by a partial recovery to a sustained lesser level of vasoconstriction. The initial maximum response was enhanced by L‐NAME but not affected by indomethacin treatment. The simultaneous administration of L‐NAME and indomethacin prevented the partial recovery resulting in a greater sustained level of constriction.

A stable vasoconstriction of relatively constant magnitude was observed with angiotensin II (AII) infusion. The AII vasoconstriction was enhanced by L‐NAME but not changed by indomethacin. The combination of these inhibitors further enhanced the AII‐induced vasoconstriction in WKY, but not in SHR.

Pretreatment with the AII receptor antagonist, losartan, inhibited the vasoconstriction induced by AIV and AII.

These results suggest that nitric oxide and prostaglandins may modulate the renal vascular response to AIV.
In situ assessment of the role of the β1, β2‐ and β3‐adrenoceptors in the control of lipolysis and nutritive blood flow in human subcutaneous adipose tissue
Authors: Barbe Pierre , Millet Laurence , Galitzky Jean , Lafontan Max , Berlan Michel
Published: July 2012

The involvment of β1, β2‐ and β3‐adrenoceptors in the control of lipolysis and nutritive blood flow was investigated in abdominal subcutaneous adipose tissue of healthy young adults by use of an in situ microdialysis technique.

Dialysis probes were infused either with isoprenaline (non‐selective β‐adrenoceptor agonist), CGP 12,177 (selective β3‐adrenoceptor agonist having β1‐/β2‐antagonist properties), dobutamine (selective β1‐adrenoceptor agonist) or terbutaline (selective β2‐adrenoceptor agonist). The recovery of each probe used for perfusion was calculated by an in vivo calibration method. The local blood flow was estimated through the measurement of the escape of ethanol infused simultaneously with the drugs included in the probe.

Isoprenaline infusion at 0.01 μm had a weak effect while higher concentrations of isoprenaline (0.1 and 1 μm) caused a rapid, sustained and concentration‐dependent increase of glycerol outflow; the maximum increase was 306±34% with 1 μm. Isoprenaline also increased the nutritive blood flow in adipose tissue; a significant effect appeared at 0.1 μm isoprenaline and was greater at 1 μm.

CGP 12,177 (10 and 100 μm) increased the glycerol concentration in the dialysate (128±8 and 149±12%, respectively) and nutritive blood flow. Terbutaline and dobutamine (100 μm) both provoked rapid and similar increases in glycerol outflow (252±18 and 249±18%, respectively). Both, terbutaline and dobutamine increased nutritive blood flow.

It is concluded that β1 and β2‐adrenoceptor subtypes are both mainly involved in the mobilization of lipids and in the control of nutritive blood flow. β3‐Adrenoceptors play a weaker role in the control of lipolysis and nutritive blood flow in human subcutaneous abdominal adipose tissue.
Cisapride‐induced prolongation of cardiac action potential and early afterdepolarizations in rabbit Purkinje fibres
Authors: Puisieux François L. , Adamantidis Monique M. , Dumotier Bérengére M. , Dupuis Bernard A.
Published: July 2012

Cisapride, a gastrointestinal prokinetic agent, has been associated with cases of Torsades de Pointes but its effects on the cardiac action potential have not been described. We investigated its electrophysiological effects on rabbit isolated Purkinje fibres. The results demonstrated that cisapride (0.01 ×10 μm) lengthened concentration‐dependently the action potential duration without modifying other parameters and induced early after depolarizations and subsequent triggered activity. This typical class III antiarrhythmic effect, that showed ‘reverse’ rate‐dependence and was reduced by increasing external K concentration, can account for clinical arrhythmogenesis.
Angiotensin II‐elicited signal transduction via AT1 receptors in endothelial cells
Authors: Pueyo Maria E. , N'Diaye Nina , Michel Jean‐Baptiste
Published: July 2012

. Angiotensin II (AII) actions are mediated by two distinct types of receptors: AT1, which includes two subtypes, AT1A and AT1B, and AT2. AII produces vasoconstriction on the vascular wall acting directly on smooth muscle cells via AT1 receptors. AII receptors have recently been demonstrated on endothelial cells. But the pharmacological characteristics of these receptors and the intracellular signal pathways coupled to them remain unclear.

. The aim of this work was to characterize the AII receptor subtypes in rat aortic endothelial cells (RAEC) in primary culture and to evaluate the signal pathways coupled to these receptors by measuring the activation of phospholipase C (PLC) and phospholipase A2 (PLA2).

. Labelled AII bound to RAEC in a specific, saturable manner. Scatchard analysis showed a Kd of 1.87±0.49 nM and a Bmax of 50.2±10.9 × 103 sites per cell. AII was displaced by the AT1‐specific antagonist, DuP753 with a Ki of 17.37 ± 1.49 nM, but not by the AT2 receptor analogues CGP42771B or PD123177. These data were confirmed by the finding of AT1 mRNA in endothelial cells. Analysis of RNA expression by RT‐PCR showed the presence of both subtypes, AT1A and AT1B, in endothelial cells, whereas smooth muscle cells express only AT1A.

. The activation of PLC and PLA2 in response to AII was evaluated by measuring inositol phosphate production and arachidonic acid release, respectively. Both were enhanced by AII in a dose‐dependent manner, and inhibited by DuP753, but not by PD123177.

. We conclude that AT1 receptors are expressed by endothelial cells in primary culture and that phospholipase C and phospholipase A2 are activated via this receptor.
Protein kinase C isoforms in bovine aortic endothelial cells: role in regulation of P2Y‐ and P2U‐purinoceptor‐stimulated prostacyclin release
Authors: Patel Viral , Brown Colin , Boarder Michael R.
Published: July 2012

. Enhanced synthesis of prostacyclin (PGI2) and inositol polyphosphates in bovine aortic endothelial cells in response to ATP and ADP is mediated by co‐existing P2Y‐ and P2U‐purinoceptors. Here we examine the regulation of these responses by isoforms of protein kinase C (PKC).

. Immunoblots with antisera specific for 8 different PKC isoforms revealed the presence of α, ∍ and ζ, while no immunoreactivity was found for β, γ, δ, η and θ isoforms. PKC‐α was largely cytosolic in unstimulated cells and almost all translocated to the membrane (Triton X‐100 soluble) after a 1 min treatment with the PKC activating phorbol myristate acetate (PMA); PKC‐∍ was always in a Triton X‐100 insoluble membrane fraction, while PKC‐ζ was found in both soluble and membrane bound (Triton X‐100 soluble) forms in the unstimulated cells and was unaffected by PM A.

. Treatment with PM A for 6 h led to a 90% downregulation of PKC‐α, while the immunoreactivity to the e and ζ isoforms remained largely unchanged.

. After either 10 min or 6 h exposure to PM A the PGI2 response to activation of both receptors was enhanced, while the inositol 1,4,5‐trisphosphate response to P2Y‐purinoceptor activation was substantially attenuated and the P2U‐purinoceptor response was unchanged. Thus the PGI2 response to PM A under conditions when 90% of the PKC‐α was lost resembles that seen on acute stimulation of PKC by PMA, and the PGI2 response does not correlate with the phospholipase C response.

. Inhibition of PKC with the isoform non‐selective inhibitors, Ro 31–8220 and Go 6850 abolished the PGI2 response to both P2U‐ and P2Y‐purinoceptor stimulation. However, Go 6976, which preferentially inhibits Ca2+ sensitive isoforms (such as PKC‐α) and not Ca2+ insensitive isoforms (such as PKC‐∍), had no effect on the PGI2 response.

. The results show that there is a requirement for PKC in the stimulation of PGI2 production by endothelial P2Y‐ and P2U‐purinoceptors. Both downregulation and inhibition studies show that PKC‐α is not responsible for the regulation of the response to P2‐purinergic stimulation, and imply that the response is mediated by PKC‐∍ (PKC‐ζ is unresponsive to PMA), or an as yet uncharacterized PKC isoform.
Effects of phosphorothioated neuropeptide Y Y1‐receptor antisense oligodeoxynucleotide in conscious rats and in human vessels
Authors: Sun Xiang Ying , Zhao Xiao He , Erlinge David , Edvinsson Lars , Fallgren Bo , Wahlestedt Claes , Hedner Thomas
Published: July 2012

. Metabolically stabilized (phosphorothioate) human and rat NPY Yx receptor oligodeoxynucleotides (ODNs) complimentary to the rat or human Y1 mRNA were synthesized; [sense (rY1‐SODN, 5′‐AATTCAACTCTGTTCTCC‐3′), antisense (hY1‐ASODN, 5′‐CCTGGGAAAATAATGTTG‐3′ and rY1‐ASODN, 5′‐GGAGAACAGAGTTGAATT‐3′) and mismatches (hY1‐MMODN, 5′‐CCTGAGATAA‐TAAGGTTG‐3′ and rY1‐MM 5′‐GTAGATCAGAGATGAAGT‐3′)] and used to modulate cardiovascular function in vitro in human vessels as well as in vivo in the rat.

. The objectives of the experiments were to assess the influence of the NPY Y1 receptor on vasomotor function human resistance arteries in vitro and to investigate the contribution of the NPY receptor system to cardiovascular haemodynamics in vivo.

. Human subcutaneous resistance arteries removed from patients who underwent surgery for non‐vascular diseases were incubated in vitro with the stabilized phosphorothioated hY1‐receptor ASODN or MMODN (10−7 to 10−5 m).

. In human resistance vessels preincubated with hY1‐AS (10−7 to 10−5 m), the contractile response to NPY was significantly reduced in a dose‐dependent fashion. No effects were observed in the hY1‐MMODN‐incubated vessels at lower concentrations (10−7 m to 10−6 m).

. The haemodynamic effects of the phosphorothioated rY1‐ASODN, SODN or MMODN were investigated in conscious rats during 48 h of continuous infusions. The continuous infusion with the rY1‐ASODN did not change MAP while the rY1‐SODN unexpectedly induced an early (10–20 h) increase in ambulatory MAP and the rY1‐MMODN a late (24–44 h) increase.

. Contractile responses to NPY (2, 4, 8, 16 and 32 μg kg−1) were significantly reduced in the rats treated with long‐term infusion of rY1‐ASODN (2.1 mg kg−1 h−1, i.v. infusion for 48 h) compared with animals treated with rY1‐SODN and MMODN, as well as animals treated with saline and glucose. Notably, the group infused with the rY1‐SODN showed an exaggerated response to tested doses of NPY.

. We conclude that the incubation of human subcutaneous arteries with a metabolically stabilized 18 base pair hY1‐ASODN and long‐term infusion with a corresponding rY1‐ASODN attenuate NPY‐induced vasoconstriction.