bluuebunny/crossref_metadata_2025 huggingface dataset:
Poly‐l‐arginine‐mediated release of acetylcholine from parasympathetic nerves in rat and guinea‐pig airways
Authors: Spina D. , Goldie R.G.
Published: July 2012

The synthetic cationic polypeptide, poly‐l‐arginine (0.03–1 mg ml−1) induced concentration‐dependent contraction of guinea‐pig and rat isolated trachea. In guinea‐pig isolated trachea, this response was attenuated in the presence of the muscarinic cholinoceptor antagonist, atropine (0.1 μm) and augmented by the acetylcholinesterase inhibitor, ecothiophate (0.1 μm). The neuronal sodium channel blocker, tetrodotoxin (3 μm) failed to alter the contractile response to poly‐l‐arginine and acetylcholine.

The contractile response to poly‐l‐arginine in rat isolated trachea was inhibited in the presence of atropine (0.1 μm) and the 5‐hydroxytryptamine (5‐HT) receptor antagonist, methysergide (1 μm). Treatment of rat tracheal preparations with capsaicin (100 μm) or tetrodotoxin (3 μm) failed to alter the contractile response to poly‐l‐arginine. In contrast, ecothiophate (0.1 μm) augmented the contractile response to poly‐l‐arginine in rat isolated trachea.

Electrical field stimulation (5 Hz, 2 min) of epithelium‐denuded guinea‐pig tracheal preparations preloaded with [3H]‐choline resulted in a contractile response and the simultaneous efflux of radioactivity into the superfusate. Both these responses were abolished in the presence of tetrodotoxin (1.5 μm). Poly‐l‐arginine (1 mg ml−1) also increased the efflux of total radioactivity from epithelium‐denuded guinea‐pig isolated tracheal preparations preloaded with [3H]‐choline, but this response was tetrodotoxin‐insensitive. The negatively charged polyanion, heparin (1 mg ml−1) failed to increase significantly the efflux of radioactivity from epithelium‐denuded preparations.

In conclusion, the synthetic cationic polypeptide, poly‐l‐arginine, caused contraction of guinea‐pig isolated tracheal preparations via the release of acetylcholine from parasympathetic nerves. Similarly, poly‐l‐arginine‐induced contraction of rat isolated trachea is secondary to the release of acetylcholine from parasympathetic nerves and/or the release of mast cell‐derived 5‐HT.
Stereoselective and non‐stereoselective actions of isoflurane on the GABAA receptor
Authors: Hall A.C. , Lieb W.R. , Franks N.P.
Published: July 2012

Acutely dissociated cerebellar Purkinje neurones from 8–14 day old rats were studied under voltage clamp in the whole‐cell patch‐clamp configuration. Cl− currents induced by bath application of γ‐aminobutyric acid (GABA) were measured (using symmetrical Cl− solutions) at both low (2 μm) non‐desensitizing and high (300 μm) desensitizing concentrations of GABA.

At 2 μm GABA, the bicuculline‐sensitive Cl− currents were potentiated by racemic isoflurane and both of its optical isomers. Isoflurane had no effect on membrane current in the absence of GABA. The dose‐response data for potentiation by racemic isoflurane could be fitted with a Hill equation with an EC50 = 320 ± 20 μm isoflurane and a Hill coefficient of h = 2.7 ± 0.4 (means ± s.e.mean).

The potentiations produced by the optical isomers of isoflurane at 2 μm GABA were stereoselective at moderate and high anaesthetic concentrations. The maximum stereoselectivity, about two fold, occurred at the EC50 concentration for general anaesthesia (310 μm isoflurane), with S(+)‐isoflurane being more effective than R(−)‐isoflurane. At sub‐anaesthetic concentrations, the stereoselectivity was less marked and vanished at the lowest concentration used (77 μm isoflurane).

The sustained residual current remaining after exposure of neurones to a desensitizing concentration of GABA (300 μm) was inhibited non‐stereoselectively, but only at high concentrations of isoflurane. The ratio of inhibitions by S(+)‐ and R(−)‐isoflurane (mean ± s.e.mean) was 1.14 ± 0.21 at 770 μm isoflurane. At the EC50 concentration for general anaesthesia, however, the inhibition was barely significant.

The above results are discussed in relation to the possible role of the GABAA receptor channel in general anaesthesia.
Involvement of apamin‐sensitive K+ channels in antigen‐induced spasm of guinea‐pig isolated trachea
Authors: Yamauchi Hideyuki , Miura Motohiko , Ichinose Masakazu , Ishikawa Jun , Nakajima Natsuko , Tomaki Masafumi , Inoue Hiroshi , Maeyama Kazutaka , Watanabe Takehiko , Shirato Kunio
Published: July 2012

In order to examine whether K+ channels play a role in antigen‐induced airway responses, the effect of K+ channel blockers on antigen‐induced airway smooth muscle contraction and mediator release was examined in vitro in guinea‐pigs actively sensitized with ovalbumin (OA).

Tracheal strips from sensitized animals were suspended in organ baths under a resting tension of 1 g and isometric tension was continuously measured. Cumulative concentration‐response curves to OA (0.1–1000 ng ml−1) or histamine (10 nm‐1 mm) were obtained in the presence and absence of K+ channel blockers.

OA (10, 100 or 1000 ng ml−1) was incubated with minced lung tissues from the same animals for 15 min in the presence and absence of K+ channel blockers, and released histamine and leukotriene C4 (LTC4) in the incubating medium were measured.

Apamin, a small conductance Ca2+‐activated K+ channel (PK,Ca) blocker, (0.1, 0.3 and 1 μm) significantly inhibited OA‐induced smooth muscle contraction, while charybdotoxin (ChTX, 10 nm), an intermediate and large conductance PK,Ca blocker, and iberiotoxin (IbTX, 3 nm), a large conductance PK,Ca blocker, were without effect. Apamin (0.3 μm) had no effect on exogenously administered histamine‐induced airway smooth muscle contraction, suggesting that the inhibition of OA‐induced contraction by apamin did not occur at the smooth muscle level.

The inhibition of OA‐induced contraction by apamin (0.3 μm) was not significantly affected by pretreatment with a leukotriene antagonist, ONO‐1078 (10 μm), but was abolished by pretreatment with a histamine H1‐receptor blocker, pyrilamine (1 μm).

Apamin by itself (up to 0.1 μm) had no effect on spontaneous histamine release from minced lung tissues. Histamine release induced by low and intermediate concentrations of OA (10 and 100 ng ml−1) was significantly suppressed by apamin pretreatment (P < 0.05 and P < 0.001), whereas LTC4 release was not affected. ChTX (0.1 μm) and IbTX (10 nm) had no significant effect on either spontaneous or OA (100 ng ml−1)‐induced histamine release.

These results suggest that apamin partially but substantially inhibits antigen‐induced smooth muscle contraction, presumably by inhibiting antigen‐induced histamine release from airway mast cells through small conductance PK,Ca. closure.
Pharmacological properties of the cloned α1A/D‐adrenoceptor subtype are consistent with the α1A‐adrenoceptor characterized in rat cerebral cortex and vas deferens
Authors: Kenny B.A. , Naylor A.M. , Greengrass P.M. , Russell M.J. , Friend S.J. , Read A.M. , Wyllie M.G.
Published: July 2012

The pharmacological characteristics of cloned mammalian α1A/D‐, α1B‐ and α1C‐adrenoceptor subtypes expressed in rat 1 fibroblasts were determined in comparison to the binding and functional properties of these subtypes in rat tissues.

Analysis of [3H]‐prazosin binding to membrane homogenates from rat 1 fibroblast cells expressing each of the α1‐subtypes indicated high affinity binding to a single population of binding sites. Binding affinities were similar for α1A/D‐, α1B‐ and α1C‐subtypes (Kds: 0.13, 0.10 and 0.15 nm respectively) although a higher density of α1B‐ and α1C‐receptors (Bmax: 4068 and 10,323 fmol mg−1 protein respectively) were expressed in comparison to α1A/D (838 fmol mg−1).

Displacement of [3H]‐prazosin from membranes expressing cloned α1‐adrenoceptor subtypes revealed that 5‐methyl‐urapidil, WB 4101, benoxathian and phentolamine displayed high affinity and selectivity for α1A/D‐ over α1B‐subtypes. These compounds also had high affinity and selectivity for α1C‐ over α1B‐subtypes. 5‐Methyl‐urapidil showed selectivity for α1C (Ki 0.60 ± 0.16 nm) over both α1A/D (Ki, 9.8 ± 2.8 nm) and α1B (Ki 57.2 ± 12 nm) subtypes. Prazosin and doxazosin were not subtype selective.

In comparison to [3H]‐prazosin a similar pharmacological profile was obtained with [125I]‐HEAT using cloned α1A/D‐, α1B‐ and α1C‐adrenoceptors expressed in rat 1 fibroblasts.

The affinities of prazosin, WB 4101, 5‐methyl‐urapidil, phentolamine and benoxathian at cloned α1A/D‐receptors were consistent with α1A affinities determined with chlorethylclonidine‐treated rat cortical membranes. Affinities at cloned α1B‐receptors were consistent with α1B affinities determined with rat liver membranes.

Using the epididymal rat vas deferens as a functional measure of α1A affinity, prazosin (pA2 9.23 ± 0.28), WB 4101 (pA2 9.58 ± 0.12), phentolamine (pKB 7.90 ± 0.16), benoxathian (pKB 9.21 ± 0.21) and 5‐methyl‐urapadil (pKB 8.51 ± 0.16) were potent antagonists of noradrenaline‐induced contractions.

At present, evidence from cloning studies suggests the existence of at least three α1‐adrenoceptor subtypes. In contrast to the recent proposal for α1‐adrenoceptor classification, the pharmacology of the cloned α1A/D (or α1D)‐adrenoceptor is more consistent with that of an α1A‐adrenoceptor characterized in rat cerebral cortex and vas deferens.
Replacement of salt by a novel potassium‐ and magnesium‐enriched salt alternative improves the cardiovascular effects of ramipril
Authors: Mervaala Eero M.A. , Paakkari Ilari , Laakso Juha , Nevala Riikka , Teräväinen Tertt M‐Lüsa , Fyhrquist Frej , Vapaatalo Heikki , Karppanen Heikki
Published: July 2012

The influence of salt (sodium chloride; NaCl) (an additional 6% in the diet) and that of a novel sodium‐reduced, potassium‐, magnesium‐, and l‐lysine‐enriched salt alternative on the cardiovascular effects of ramipril was studied in stroke‐prone spontaneously hypertensive rats in a 6‐week study. The intake of sodium chloride was adjusted to the same level by adding the salt alternative at a 1.75 times higher amount than regular salt.

Salt produced a marked rise in blood pressure and induced cardiac hypertrophy and significant mortality, while the salt alternative neither increased blood pressure nor caused any mortality and produced less cardiac hypertrophy than salt.

Ramipril treatment at a daily dose of 3 mg kg−1normalized blood pressure and prevented the development of cardiac hypertrophy of rats on control diet. These effects of ramipril were blocked by the addition of salt but were only slightly attenuated by the addition of the salt alternative. The mortality in the salt group was prevented by ramipril.

Responses of mesenteric arterial rings in vitro were examined at the end of the study. Salt, but not the salt alternative, increased vascular contractile responses to noradrenaline. Ramipril treatment improved the arterial relaxation responses to acetylcholine and to sodium nitroprusside. The vascular relaxation enhancing effect of ramipril was blocked by salt but only slightly attenuated by the salt alternative.

Ramipril treatment did not significantly increase plasma renin activity in the presence or in the absence of salt supplementation. The salt alternative did not cause hyperkalaemia, either alone or in combination with ramipril treatment.

Both salt supplementations, irrespective of ramipril treatment, induced a six to eight fold increase in the urinary excretion of calcium. There was an expected 90 to 140% rise in the urinary excretion of magnesium and 200% rise in the urinary excretion of potassium in the salt alternative group. Salt also produced an approximately 50% increase in magnesuria.

Our findings suggest that replacement of salt by the potassium‐, magnesium‐ and l‐lysine‐enriched salt alternative improves the cardiovascular effects of ramipril. In the present study the beneficial effect was related to the increased intakes of potassium and/or magnesium and l‐lysine from the salt alternative because the amount of sodium chloride was the same.
Modulation of antagonist binding to histamine H1‐receptors by sodium ions and by 2‐amino‐2‐hydroxymethyl‐propan‐1,3‐diol HCl
Authors: Gibson W.J. , Roques T.W. , Young J.M.
Published: July 2012

NaCl (100 mm) reduced the potency of (+)‐N‐methyl‐4‐methyldiphenhydramine ((+)‐QMDP) as an inhibitor of the binding of [3H]‐mepyramine to histamine H1‐receptors on guinea‐pig cerebellar membranes to a greater extent than that of mepyramine, consistent with the greater inhibitory effect of Na+on the binding of [3H]‐QMDP than on the binding of [3H]‐mepyramine.

The concentration of 2‐amino‐2‐hydroxymethyl‐propan‐1,3‐diol HCl (Tris, HCl) buffer, pH 7.5, present had little effect on the temelastine‐insensitive binding of [3H]‐mepyramine, but caused a concentration‐dependent inhibition of the binding of [3H]‐mepyramine sensitive to 1 μm temelastine (H1‐receptor binding), with an approximate IC50 of 75 mm, assuming that complete inhibition would have been achieved.

Inhibition of [3H]‐mepyramine binding by Na+was more marked in 10 mm than in 50 mm Tris HCl and was not evident in 200 mm Tris HCl.

The Kd for the temelastine‐sensitive binding of [3H]‐mepyramine measured in 10 mm Tris HCl, 0.24 ± 0.01 nm, was increased by 2.2 ± 0.2 fold by 100 mm NaCl, without any significant change in the maximum binding (Bmax). The Bmax for [3H]‐mepyramine was similarly unchanged in 50 mm Tris HCl, but the Kd was increased 2.5 ± 0.2 fold.

The Kd for the temelastine‐sensitive binding of [3H]‐mepyramine was also increased in 50 mm, compared with 10 mm, N‐[2‐hydroxyethyl]piperazine‐N′‐[2‐ethanesulphonic acid] KOH (HEPES.KOH) buffer (Kd 0.25 ± 0.02 nm in 10 mm HEPES), but the evidence for an interaction between HEPES and Na+was less clear.

The effect of 100 mm NaCl on the inhibition of [3H]‐mepyramine binding in 10 mm Tris HCl was examined for a range of antagonists. The decrease in potency caused by Na+was greatest for triprolidine, (+)‐chlorpheniramine and benzilylcholine (9.6–10.3 fold increase in Kd values) but the binding of mepyramine and promethazine was much less affected (1.8 and 1.9 fold increase in Kd respectively). The Kd for temelastine was not significantly changed. In contrast to the general decrease in antagonist affinity in the presence of Na+, the Kd for MDL 16,455A (4‐[1‐hydroxy‐4‐[4‐(hydroxydiphenylmethyl)‐1‐piperidinyl]butyl]‐α,α‐dimethylbenzene acetic acid) was increased, but only by 1.5 fold.

It is concluded that Na+can act as an allosteric effector of the binding of antagonists at the histamine H1‐receptor. Tris HCl also appears to have an allosteric action at the H1‐receptor.
The identification of apparently novel cyclic AMP and cyclic GMP phosphodiesterase activities in guinea‐pig tracheal smooth muscle
Authors: Burns Fiona , Stevens Patricia A. , Pyne Nigel J.
Published: July 2012

Phosphodiesterase (PDE) activities that were capable of hydrolysing cyclic AMP (Km = 6.8 ± 2 μm) and cyclic GMP (Km = 6.7 ± 1.6 μm) were isolated from tracheal smooth muscle. These enzyme(s) activities were insensitive to stimulation by calcium/calmodulin and to inhibition by cyclic GMP, rolipram (type IV inhibitor) and siguazodan (type III inhibitor). Zaprinast was a relatively poor inhibitor of both cyclic AMP and cyclic GMP hydrolysis (IC50 = 46 ± 9 μm and 45 ± 14 μm respectively). These results suggest that tracheal smooth muscle may contain an apparently novel PDE. However, KCl (30 mm) which facilitates calcium entry in cells, depressed bradykinin‐stimulated intracellular cyclic AMP formation, suggesting that the type I PDE may be functionally present. We suggest that considerable caution be exercised in identifying apparently novel PDE isoforms.
Close correlation of the cardioprotective effect of FK409, a spontaneous NO releaser, with an increase in plasma cyclic GMP level
Authors: Kita Yasuhiro , Sugimoto Toshiko , Hirasawa Yoshimi , Yoshida Keizo , Maeda Kazuhiro
Published: July 2012

FK409 ((±)‐(E)‐ethyl‐2‐[(E)‐hydroxyimino]‐5‐nitro‐3‐hexeneamide), which has been reported by us to be a new spontaneous nitric oxide (NO) releaser, prevented myocardial infarction following occlusion and reperfusion in rat coronary artery and increased plasma cyclic GMP level in rats, dose‐dependently and significantly at 32 mg kg−1. Isosorbide dinitrate (ISDN), which is the most popular orally active NO donor used in the treatment of ischaemic cardiovascular diseases, did not show significant effects at 32 mg kg−1 in either experiment. Therefore, it is suggested that FK409 can attenuate myocardial injury during ischaemia and reperfusion in contrast to ISDN and a change in plasma cyclic GMP level may serve as an indicator of the cardioprotective effect of NO‐releasing drugs.
Loss of contractile activity of endothelin‐1 induced by electrical field stimulation‐generated free radicals
Authors: Yasuda Nobuyuki , Kasuya Yoshitoshi , Yamada Goro , Hama Hiroshi , Masaki Tomoh , Goto Katsutoshi
Published: July 2012

Electrical field stimulation (EFS; 10 V, 10 Hz, 2 ms) of porcine coronary artery strips precontracted with 10 nm endothelin‐1 (ET‐1) for 5 min caused a biphasic response, consisting of a slight contraction during EFS and a marked and irreversible relaxation just after EFS. This irreversible relaxation after EFS has never been investigated. In the present study, we have investigated the mechanism of the relaxation after EFS.

The EFS‐induced response was not affected by the presence or absence of endothelium and was insensitive to 10 μm tetrodotoxin (TTX).

In the presence of free radical scavengers (40 u ml−1 superoxide dismutase (SOD), 1200 u ml−1 catalase or 80 mm d‐mannitol), the relaxation after EFS was significantly inhibited. Moreover, relaxation after EFS was not observed in porcine coronary artery strips precontracted with 20 mm KCl.

In a cascade experiment, EFS of Krebs‐Ringer solution containing 10 nm ET‐1 induced marked suppression of the contractile activity of ET‐1 in porcine coronary artery strips, which was in accord with the observed decrease in release of immunoreactive ET‐1 (ir‐ET‐1). This effect of EFS was significantly inhibited by each of the free radical scavengers, 3 mm vitamin C, 40 u ml−1 SOD, 1200 u ml−1 catalase and 80 mm d‐mannitol.

The exchange of 95% O2/5% CO2 gas for 95% N2/5% CO2 gas significantly inhibited the EFS‐induced decrease in release of ir‐ET‐1.

Neither superoxide anions generated by xanthine (10 μm) plus xanthine oxidase (0.1 u ml−1) nor hydrogen peroxide (10 μm) exogenously added to Krebs‐Ringer solution containing 10 nm ET‐1 affected the level of ir‐ET‐1.

Generation of hydroxyl radicals was detected in the EFS‐applied Krebs‐Ringer solution. The EFS‐induced generation of hydroxyl radicals was dependent on the period of stimulation and O2‐bubbling, and significant generation of hydroxyl radicals was detectable with stimulation of over 5 min. Moreover, hydroxyl radicals generated in 50 mm NaCl solution containing 10 nm ET‐1 by H2O2 plus Fe2+, i.e. the Fenton reaction, significantly decreased the level of ir‐ET‐1.

These findings suggest that oxygen‐derived hydroxyl radicals generated by EFS of porcine coronary artery strips inactivate ET‐1, probably by structural modification. Thus, porcine coronary artery strips precontracted with ET‐1 are potently relaxed by EFS.
The involvement of bradykinin B1 and B2 receptor mechanisms in cytokine‐induced mechanical hyperalgesia in the rat
Authors: Davis A.J. , Perkins M.N.
Published: July 2012

Interleukin‐1β (IL‐1β), IL‐2 and IL‐8 induced a mechanical hyperalgesia following intra‐articular (i.artic.) injection into rat knee joints, whereas IL‐6 and tumour necrosis factor α (TNF‐α) were without effect.

Co‐administration of IL‐1 receptor antagonist (0.1 μg) with IL‐1β (1 u), IL‐2 (10 u) or IL‐8 (0.1 u) prevented the subsequent development of the hyperalgesia.

Co‐administration of desArg9Leu8BK (0.5–5 nmol) with IL‐1β (1 u), IL‐2 (10 u) or IL‐8 (0.1 u) reduced the level of hyperalgesia at 1, 4 and 6h post administration, whereas Hoe 140 (5 pmol) antagonized the hyperalgesia only at the 1 h time point.

Intravenous administration of desArg9Leu8BK (10 nmol kg−1) or Hoe 140 (100 pmol kg−1) following IL‐1β (1 u), IL‐2 (10 u), or IL‐8 (0.1 u) reversed the subsequent hyperalgesia.

Administration of desArg9BK into joints 24 h after pre‐treatment with IL‐1β (1 u) produced analgesia at low doses (50 pmol) and hyperalgesia at a higher dose (0.5 nmol). Both these effects were blocked by desArg9Leu8BK (0.5 nmol).

Administration of desArg9BK (0.5 nmol i.artic.) to animals 24 h after pre‐treatment with IL‐2 (1–100 u) or IL‐8 (0.1–10 u) had no effect on the load tolerated by the treated joint.

Administration of indomethacin (1 mg kg−1, s.c.) prior to IL‐1β (1 u i.artic.) prevented the development of hyperalgesia. Administration of desArg9BK (5 pmol‐0.5 nmol, i.artic.) to animals 24 h after indomethacin and IL‐1β pretreatment had no effect on the load tolerated by the treated joint.

These data suggest that both bradykinin B1 and B2 receptors are involved in the induction and maintenance of cytokine‐induced hyperalgesia. They also show that the induction of B1 receptor‐mediated hyperalgesia requires both cyclo‐oxygenase products and IL‐1 in vivo.
Effect of in vivo desensitization to leukotriene B4 on eosinophil infiltration in response to C5a in guinea‐pig skin
Authors: Pettipher E.R. , Salter E.D. , Showell H.J.
Published: July 2012

The effect of in vivo desensitization to leukotriene B4 (LTB4) on eosinophil infiltration in response to recombinant C5a was examined in guinea‐pig skin.

LTB4 (10–300 ng) and C5a (1–10 μg) caused a dose‐dependent increase in the levels of eosinophil peroxidase activity (a measure of eosinophil infiltration) 4h after injection into guinea‐pig skin. Leukotriene B4 and C5a were approximately equipotent on a molar basis. Platelet activating factor (0.01–10 μg) also caused eosinophil accumulation but was much less active than LTB4 or C5a.

20‐Hydroxy‐LTB4 caused a dose‐dependent desensitization of eosinophil responses to LTB4 (ED50 = 1.6 μg kg−1, s.c.) and partially reduced responses to C5a. At a dose of 20‐hydroxy‐LTB4 (10 μg) which inhibited responses to LTB4 completely, responses to C5a were reduced by 56.5 ± 1.8% (n = 5). The structurally related metabolite of 20‐hydroxy‐LTB4, 20‐carboxy‐LTB4, which does not cause desensitization to the effects of LTB4, did not inhibit eosinophil infiltration in response to C5a.

The LTB4 receptor antagonist, SC‐41,930 (10 mg kg−1, p.o.), also inhibited eosinophil accumulation in response to C5a by 63.0 ± 3.9% (n = 5) at a dose which inhibited responses to LTB4 by 86.5 ± 1.9% (n = 5).

These data indicate that eosinophil infiltration in response to C5a may, in part, be mediated by the generation of secondary chemotactic factors such as LTB4.
Effects of chronic infusions of α‐trinositol on regional and cardiac haemodynamics in conscious rats
Authors: Gardiner S.M. , Kemp P.A. , Fallgren B. , Bennett T.
Published: July 2012

Male, Long Evans rats (350–450 g) were chronically instrumented for the measurement of renal, mesenteric and hindquarters haemodynamics, and were given three consecutive, 24 h infusions of vehicle (sterile saline at 0.3 ml h−1; n = 8) or α‐trinositol (d‐mjyo‐inositol‐1,2,6‐triphosphate) at 5, 20 and 80 mg kg−1 h−1 (0.3 ml h−1; n = 9). During infusion of α‐trinositol at 5 or 20 mg kg−1 h−1, cardiovascular changes were little different from those seen during saline infusion. However, during infusion of α‐trinositol at 80 mg kg−1 h−1 there were increases in hindquarters vascular conductance, renal flow and vascular conductance, that were all significantly different from the changes seen in the saline group. Infusion of α‐trinositol at the high dose in naive rats (n = 8) had even more marked vasodilator effects.

Two groups of rats (n = 8 in each), chronically instrumented for the measurement of cardiac haemodynamics, were given 48 h infusions of saline (0.3 ml h−1) or α‐trinositol (2 mg kg−1 bolus, 80 mg kg−1 h−1 infusion at 0.3 ml h−1). During the infusion of saline, there were slight reductions in heart rate, cardiac index, peak aortic flow, dF/dtmax and central venous pressure. In the animals receiving α‐trinositol, with the exception of central venous pressure, all the above variables, together with total peripheral conductance, increased.

These results, collectively, indicate that incremental infusions of α‐trinositol do not reveal its full vasodilator potential, possibly due to concurrent activation of counter‐regulatory vasoconstrictor mechanisms. However, infusion of α‐trinositol at a high dose causes substantial increases in renal, mesenteric and hindquarters flows and vascular conductances, supported by significant increases in indices of cardiac inotropism. Such effects, in the absence of significant hypotension, tachycardia or signs of desensitization, give α‐trinositol a unique cardiovascular profile.
Factors underlying the increased sensitivity to field stimulation of urinary bladder strips from streptozotocin‐induced diabetic rats
Authors: Tammela Teuvo L.J. , Briscoe Janice A.K. , Levin Robert M. , Longhurst Penelope A.
Published: July 2012

The responses of bladder strips from control, streptozotocin‐diabetic, and sucrose‐drinking rats to electrical field stimulation were investigated. Sucrose‐drinking rats were included as additional controls because they have enlarged bladders as a result of non‐diabetic diuresis.

Bladder strips from diabetic rats developed more spontaneous activity than those from the two control groups. Indomethacin reduced the amplitude and frequency of spontaneous contractions suggesting that they resulted from endogenous prostaglandin formation. Tetrodotoxin (TTX) had little effect, while α,β‐methylene ATP caused increases in spontaneous activity.

Bladder strips from diabetic rats responded to field stimulation with greater contractions than controls in the absence of antagonists as well as in the presence of atropine and α,β‐methylene ATP. Increasing TTX concentrations caused a step‐wise depression of the contractile response to electrical stimulation which was not affected by preincubation with either atropine or α,β‐methylene ATP.

Atropine and indomethacin had no effect on stength‐duration curves constructed to measure threshold contractile responses to five pulses stimulation. The curves were shifted to the right by both TTX and α,β‐methylene ATP, indicating that the responses were neurogenic in nature and at least partially, the result of stimulation of P2‐purinoceptors. In the absence of drugs, bladder strips from diabetics responded at lower voltages and pulse widths than those of control and sucrose‐drinking rats, suggesting that they were more excitable.

The response curve of bladder strips from diabetics to field stimulation at increasing voltage was shifted upwards and to the left compared to strips from control or sucrose‐drinking rats.

Bladder strips from diabetics responded to stimulation at increasing pulse width with greater responses than those from control or sucrose‐drinking rats. At 1.0 ms pulse width, the TTX‐resistant response of strips from diabetic rats was still greater than that of the other groups, indicating that a myogenic component was also involved.

The data suggest that bladder strips from diabetic rats are more excitable than those of control or sucrose‐drinking rats. This may result from diabetes‐induced decreases in bladder lipid or other membrane changes, and/or be a result of partial depolarization, perhaps related to diabetic neuropathy.
Benzodiazepine‐induced intestinal motor disturbances in rats: mediation by ω2 (BZ2) sites on capsaicin‐sensitive afferent neurones
Authors: Bonnafous Cécile , Scatton Bernard , Buéno Lionel
Published: July 2012

The central and peripheral effects of the ω (benzodiazepine) site ligands, clonazepam, alpidem, zolpidem, triazolam, flumazenil, ethyl β carboline‐3‐carboxylate (β‐CCE) and N‐methyl β carboline‐3‐carboxylate (β‐CCM) on intestinal myoelectrical activity were evaluated in conscious rats, chronically fitted with Nichrome electrodes implanted on the duodenum and jejunum. The localization of the ω (benzodiazepine) receptors involved in these effects was evaluated by use of systemic and perivagal capsaicin treatments.

When administered intraperitoneally (i.p.) the ω site inverse agonists β‐CCE and β‐CCM, and the ω site antagonist flumazenil, did not affect the duodeno‐jejunal motility. Alpidem and zolpidem, two selective ω1 site agonists induced an inhibition of migrating myoelectric complexes (MMCs) only at a high dose (5 mg kg−1). In contrast, clonazepam (a mixed ω1/ω2 agonist) and triazolam (a preferential ω2 site agonist) disrupted the MMC‐pattern at doses as low as 0.05 mg kg−1, the effect of triazolam being of much longer duration than that of clonazepam. None of these drugs altered MMC‐pattern when administered centrally (i.c.v.).

Administered i.p. or i.c.v. prior to triazolam, alpidem blocked the effect of triazolam on duodeno‐jejunal spike activity. Administered i.p. prior to triazolam, flumazenil suppressed the triazolam‐induced MMC‐disruption. Previous systemic but not perivagal capsaicin treatment suppressed the effects of clonazepam on MMCs.

It is concluded that ω‐site agonists but not antagonist or inverse agonists, administered systemically induce intestinal motor disturbances which may be linked to activation of ω2 (BZ2) sites located on nonvagal capsaicin‐sensitive afferent neurones.
Cyclothiazide unmasks AMPA‐evoked stimulation of [3H]‐L‐glutamate release from rat hippocampal synaptosomes
Authors: Barnes Janine M. , Dev Kumlesh K. , Henley Jeremy M.
Published: July 2012

The effect of α‐amino‐3‐hydroxy‐5‐methylisoxazolepropionate (AMPA) on Ca2+‐sensitive, tetrodotoxin (TTX)‐insensitive K+‐stimulated [3H]‐L‐glutamate release from rat hippocampal synaptosomes was determined. AMPA in the presence, but not in the absence of cyclothiazide, a drug which blocks AMPA receptor desensitization, elicited a dose‐dependent increase in K+‐stimulated [3H]‐L‐glutamate release but had no effect on basal release. The AMPA/cyclothiazide stimulation was blocked by CNQX and by GYKI 52466, an antagonist at the cyclothiazide site. These results indicate that AMPA receptors are present on presynaptic terminals and suggest that they may play a role in the regulation of neurotransmitter release.
Suppression of VEGF‐induced angiogenesis by the protein tyrosine kinase inhibitor, lavendustin A
Authors: Hu D.E. , Fan T.‐P.D.
Published: July 2012

Vascular endothelial growth factor (VEGF) is a heparin‐binding angiogenic factor which specifically acts on endothelial cells via distinct membrane‐spanning tyrosine kinase receptors. Here we used the rat sponge implant model to test the hypothesis that the angiogenic activity of VEGF can be suppressed by protein tyrosine kinase (PTK) inhibitors.

Neovascular responses in subcutaneous sponge implants were determined by measurements of relative sponge blood flow by use of a 133Xe clearance technique, and confirmed by histological studies and morphometric analysis.

Daily local administration of 250 ng VEGF165 accelerated the rate of 133Xe clearance from the sponges and induced an intense neovascularisation. This VEGF165‐induced angiogenesis was inhibited by daily co‐administration of the selective PTK inhibitor, lavendustin A (10 μg), but not its negative control, lavendustin B (10 μg). Blood flow measurements and morphometric analysis of 8‐day‐old sponges showed that lavendustin A reduced the 133Xe clearance of VEGF 165‐treated sponges from 32.9 ± 1.5% to 20.9 ± 1.6% and the total fibrovascular growth area from 62.4 ± 6.1% to 21.6 ± 6.8% (n = 12, P <0.05).

Co‐injection of suramin (3 mg), an inhibitor of heparin‐binding growth factors, also suppressed the VEGF165‐elicited neovascular response. In contrast, neither lavendustin A nor suramin produced any effect on the basal sponge‐induced angiogenesis.

When given alone, low doses of VEGF165 (25 ng) or basic fibroblast growth factor (bFGF; 10 ng) did not modify the basal sponge‐induced neovascularisation. However, co‐administration of these two peptides to a single sponge together caused a significant increase in the rate of 133Xe clearance and angiogenesis similar to that seen with the high dose of VEGF 165 (250 ng) acting alone. This VEGF/bFGF neovascular response was also blocked by daily co‐administration of lavendustin A (10 μg), suramin (3 mg) or a monoclonal anti‐bFGF antibody (DG2, 1 μg), but not lavendustin B (10 μg).

These results suggest that selective inhibition of PTK could have therapeutic potential in angiogenic diseases where VEGF plays a dominant role. Furthermore, blockade of the angiogenic activity of VEGF and VEGF bFGF by suramin reveals an alternative strategy in angiosuppression.
Failure of nitric oxide donors to alter arrhythmias induced by acute myocardial ischaemia or reperfusion in anaesthetized rats
Authors: Barnes Christopher S. , Coker Susan J.
Published: July 2012

The aim of the present studies was to examine the effects of nitric oxide donors on arrhythmias induced by coronary artery occlusion and reperfusion, and on cardiac cyclic nucleotides. Experiments were performed in pentobarbitone‐anaesthetized rats prepared for occlusion of the left coronary artery.

Sodium nitroprusside (0.1, 0.3 and 1 μg kg−1 min−1) had no significant effects on the incidence of ventricular tachycardia, total ventricular fibrillation or the mortality resulting from 25 min of acute myocardial ischaemia when compared with values in controls. In addition, there was no alteration in the number of ventricular premature beats that occurred in survivors.

3‐Morpholinosydnonimine‐N‐ethylcarbamide (SIN‐1, 10, 20 and 40 μg kg−1 min−1) caused marked hypotension but did not alter the incidence or severity of ischaemia‐induced arrhythmias. In rats subject to abrupt reperfusion after 5 min of myocardial ischaemia, lower doses of SIN‐1 (1, 3 and 10 μg kg−1 min−1) still caused significant reductions in systolic and diastolic blood pressure but were devoid of antiarrhythmic activity.

In separate experiments in sham‐operated rats, sodium nitroprusside (1 μg kg−1 min−1), isosorbide dinitrate (30 and 60 μg kg−1 min−1) and SIN‐1 (20 and 40 μg kg−1 min−1) had no significant effects on cardiac cyclic GMP content.

These results indicate that nitric oxide donors do not alter arrhythmias induced by acute coronary artery occlusion or reperfusion in anaesthetized rats. Although increases in total cardiac cyclic GMP could not be detected, the results suggest that, at least in the rat, cyclic GMP does not influence these arrhythmias.
Inhibitory action of SR33557 on L‐type calcium current in single ventricular myocytes of rat
Authors: Yasui Kenji , Palade Philip
Published: July 2012

The effect of SR33557 on L‐type Ca2+ currents in rat ventricular myocytes was investigated by use of the whole‐cell patch‐clamp technique.

SR33557 inhibited Ca2+ current (ICa) in a concentration‐dependent manner without change in the current‐voltage relationship.

The inhibitory effect of SR33557 on ICa was dependent on the holding potential (Vh). The IC50 values were estimated to be 2.2 times 10−8 m at Vh = −50 mV and 9.0 times 10−6 m at Vh = −80 mV. SR33557 (10−7 m) shifted the steady state inactivation curve of ICa toward more negative potentials. Thus, the affinity of the drug for inactivated channels was considerably higher than for resting channels.

Blockade of ICa by SR33557 was both tonic and use‐dependent.

The time constant of onset of block was 36.4 s at − 50 mV and 41.9 ± 11.1 s at − 40 mV.

The time course of unblock was voltage‐dependent. The time constant declined from 400.7 ± 68.1 s at − 50 mV to 5.2 ± 1.2 s at − 80 mV.

The rate of block of ICa was related to the number of openings per unit time and to the amount of time spent depolarized. The affinity of drug for open channels was considered to be similar to that for inactivated channels.

These results suggest that SR33557 inhibits L‐type Ca2+ current through binding to both open and inactivated channels in rat ventricular myocytes.
Interacting roles of nitric oxide and ATP in the pulmonary circulation of the rat
Authors: Hasséssian Haroutioun , Burnstock Geoffrey
Published: July 2012

The potentiating effect of NG‐nitro‐l‐arginine methyl ester, (l‐NAME) a nitric oxide synthesis inhibitor, on responses of the rat pulmonary vascular pressure (PVP) to purinoceptor agonists was examined.

At a constant flow of 23 ml min−1 the PVP was 22.4 ± 2.5 mmHg (n = 15), and treatment with 100 μm l‐NAME for 15 min was without effect on the PVP. After the tone was raised with 28 nmol 9,11‐dideoxy‐11α, 9α‐epoxymethano‐prostaglandin F2α (U‐46619), the PVP was 29.4 ± 3.3 mmHg and treatment with 100 μm l‐NAME was still without effect on the PVP. It appears that there is a graded release of nitric oxide in response to different levels of steady shear stress and in our experimental model the threshold for detection was not reached under basal conditions.

In contrast, when the circulation was challenged with 30 s step, additive increases in flow between 11 and 50 ml min−1 (n = 8), treatment with 100 μm l‐NAME produced a significant (P <0.05) increase in PVP suggesting that changes in flow‐derived forces evoke the release of nitric oxide. This was evident for flow rates above 30 ml min−1.

In preparations in which tone was raised with U‐46619, a dose of 1 times 10−8 mol ATP or 2‐meSATP evoked a drop in PVP while α,β‐meATP produced an increase in PVP under constant flow of 23 ml min−1. After treatment with 100 μm l‐NAME, all three purinoceptor agonists evoked an increase in PVP. The increase in PVP evoked by α,β‐meATP was not affected by l‐NAME. These results suggest that P2Y‐purinoceptor stimulation evokes the release of nitric oxide to produce vasodilatation.

Under conditions of constant flow and basal pressure, 100 μm l‐NAME significantly (P <0.05) potentiated the increase in PVP evoked by 1 times 10−6 mol ATP, although the increase evoked by 1 times 10−8 mol α,β‐meATP, which was of similar magnitude, was not affected. These results indicate that a blockade of evoked nitric oxide release is responsible for the potentiation of the increase in PVP evoked by ATP.

This study shows that, while nitric oxide does not appear to be released in the pulmonary circulation of the rat under constant flow conditions, nitric oxide release evoked by purinoceptor agonists attenuates increases in pulmonary vascular pressure.
Acute effects of the β3‐adrenoceptor agonist, BRL 35135, on tissue glucose utilisation
Authors: Liu Yong‐Ling , Stock Michael J.
Published: July 2012

The acute effects of BRL 35135 (BRL) on tissue glucose utilisation index (GUI) in vivo were investigated in anaesthetized rats by use of 2‐deoxy‐[3H]‐glucose.

Intravenous injection of BRL caused a dose‐dependent increase in GUI in skeletal muscle, and white and brown adipose tissue; plasma insulin and fatty acid concentrations were also increased. Chronic treatment with BRL added to the diet caused a 34 fold increase in basal GUI of brown adipose tissue (BAT), but had no effect on GUI in other tissues. After chronic treatment, the acute tissue response to an intravenous maximal dose of BRL had disappeared completely in all tissues apart from the soleus muscle.

A high dose (20 mg kg−1) of the non‐selective β‐antagonist, propranolol, inhibited the acute effect of BRL on GUI in BAT, but failed to affect GUI in muscle. A lower dose (1 mg kg−1) of the antagonist also inhibited the BAT response, but had little or no effect on the response in Type I (working) muscles such as soleus and adductor longus (ADL), and potentiated the response in Type II (non‐working) muscles such as tibialis and extensor digitorium longus (EDL).

A low dose (1 mg kg−1) of the selective β1‐antagonist, atenolol, had no effect on the BRL response but the same dose of the selective β2‐antagonist, ICI 118551, potentiated significantly the effect of BRL on GUI in most muscles without altering plasma insulin levels.

It is concluded that: (i) the heterogeneous tissue responses of different muscle fibre types in the presence of β‐antagonists indicates that BRL affects muscle GUI directly, in addition to effects mediated by increases in plasma insulin concentration; (ii) the resistance of the BRL response to conventional β‐adrenoceptor antagonists implicates an atypical adrenoceptor mediating the GUI response in skeletal muscle, but this may not be identical to the adipose tissue β3‐adrenoceptor; (iii) the potentiation of BRL responses by ICI 118551 indicates an inhibitory β2‐adrenoceptor‐mediated component in the muscle GUI response to BRL.
Anaesthetic modulation of nicotinic ion channel kinetics in bovine chromaffin cells
Authors: Charlesworth P. , Richards C.D.
Published: July 2012

We have investigated the action of the anaesthetics methoxyflurane, methohexitone and etomidate on the nicotinic acetylcholine receptor channel of bovine adrenal chromaffin cells using the whole cell patch clamp technique.

Spectral analysis of macroscopic currents evoked by 25 μm carbachol revealed that each of the agents tested reduced the lifetime of the channel open state in a dose‐dependent manner. The whole cell current was inhibited in a concentration‐dependent fashion by each agent.

Channel gating parameters were calculated from single channel studies and the results used to test models explaining the modulation of nicotinic acetylcholine receptor channels by anaesthetics.

Each of the agents studied reduced the mean channel open time in a concentration‐dependent manner. Anaesthetic concentrations reducing mean open time by 50% were: 370 μm methoxyflurane, 30 μm methohexitone or 23 μm etomidate.

Methohexitone and etomidate produced an increase in the number of brief closures within bursts, while no such increase was observed with methoxyflurane. Despite these inter‐burst gaps, mean burst length was reduced by each of the agents tested.

It is concluded that a simple sequential blocking model fails to account for the action of these anaesthetics. An extended model, in which blocked channels can close, may be applicable.
Activation by ATP of a P2U‘nucleotide’ receptor in an exocrine cell
Authors: Martin Shaun C. , Shuttleworth Trevor J.
Published: July 2012

We employed the perforated patch whole‐cell technique to investigate the effects of ATP and other related nucleotides on membrane conductances in avian exocrine salt gland cells.

ATP (10μm‐1 mM) evoked an increase in maxi‐K+ and Cl− conductances with a reversal potential of −35mV. At lower concentrations of ATP (≤100 μm) responses were generally oscillatory with a sustained response observed at higher concentrations (≥200 μm).

Both oscillatory and sustained responses were abolished by the removal of bath Ca2+. In cells preincubated in extracellular saline containing reduced Ca2+, the application of ATP resulted in a transient increase in current.

As increasing concentrations of ATP (and related nucleotides) evoked a graded sequence of events with little run‐down we were able to establish a rank order of potency in single cells. The order of potency of ATP analogues and agonists of the various P2‐receptor subtypes was UTP>ATP = 2‐methylthio‐ATP>ADP. Adenosine (1μm‐1mm), AMP (1μm‐1μm), α,β‐methylene‐ATP (1 μM−1 mm) and α,β‐methylene‐ATP (1μm‐ mm) were without effect.

In conclusion, although unable to preclude a role for a P2Y‐receptor, our results suggest that ATP binds to a P2U‐receptor increasing [Ca2+]i and subsequently activating Ca2+‐sensitive K+ and Cl−currents.
Prevention by the cannabinoid antagonist, SR141716A, of cannabinoid‐mediated blockade of long‐term potentiation in the rat hippocampal slice
Authors: Collins Dawn R. , Pertwee Roger G. , Davies Stephen N.
Published: July 2012

Incubation of rat hippocampal slices in the presence of the synthetic cannabinoid (−)−11‐OH‐Δ8–dimethylheptyl tetrahydrocannabinol (HU‐210) (100 nM) prevented the induction of long‐term potentiation (LTP). Slices co‐incubated with both3HU‐210 (100 nM) and the cannabinoid antagonist, SR141716A (100 nM), exhibited tetanically induced LTP, comparable to control slices. Intriguingly, co‐incubation with3HU‐210 and SR141716A prevented the induction of the early, short‐term phase of LTP.
Blockade by sigma site ligands of high voltage‐activated Ca2+ channels in rat and mouse cultured hippocampal pyramidal neurones
Authors: Church John , Fletcher Elizabeth J.
Published: July 2012

The effects of a series of structurally‐dissimilar σ site ligands were examined on high voltage‐activated Ca2+ channel activity in two preparations of cultured hippocampal pyramidal neurones.

In mouse hippocampal neurones under whole‐cell voltage‐clamp, voltage‐activated Ca2+ channel currents carried by barium ions (IBa) were reduced with the rank order (IC50 values in μm): 1S,2R‐(–)‐cis‐N‐methyl‐N‐[2‐(3,4‐dichlorophenyl)ethyl]‐2‐(1‐pyrrolidinyl)cyclohexylamine (7.8) > rimcazole (13) > haloperidol (16)>ifenprodil (18)>opipramol (32)>carbetapentane (40)=1‐benzylspiro[1,2,3,4‐tetrahy‐dronaphthalene‐1,4‐piperidine] (42) >caramiphen (47) >dextromethorphan (73). At the highest concentrations tested, the compounds almost abolished IBa in the absence of any other pharmacological agent.

The current‐voltage characteristics of the whole‐cell IBa were unaffected by the test compounds. The drug‐induced block was rapid in onset and offset, with the exceptions of carbetapentane and caramiphen where full block was achieved only after two to three voltage‐activated currents and was associated with an apparent increase in the rate of inactivation of IBa

In rat hippocampal neurones loaded with the Ca2+‐sensitive dye Fura‐2, rises in intracellular free Ca2+ concentration evoked by transient exposure to 50 mM K+‐containing medium, either in the absence or in the presence of 10 μm nifedipine (to block L‐type high voltage‐activated Ca2+ channels), were also reversibly attenuated by the σ ligands. The rank order potencies for the compounds in these experimental paradigms were similar to that observed for blockade of IBa in the electrophysiological studies.

These results indicate that, at micromolar concentrations, the compounds tested block multiple subtypes of high voltage‐activated Ca2+ channels. These actions, which do not appear to be mediated by high‐affinity σ binding sites, may play a role in some of the functional effects previously described for the compounds.
The promotion of patent airways and inhibition of antigen‐induced bronchial obstruction by endogenous nitric oxide
Authors: Persson Magnus G. , Friberg Sten G. , Gustafsson Lars E. , Hedqvist Per
Published: July 2012

The aim of the present study was to investigate the role of nitric oxide (NO), histamine and leukotrienes in bronchial obstruction. For this, guinea‐pigs immunised against ovalbumin were studied under anaesthesia during challenge with antigen or agonists.

Challenge with nebulised antigen (0.1‐1 mg) elicited dose‐dependent increases in insufflation pressure which were abolished by combined administration of histamine and leukotriene antagonists.

Challenge with nebulised antigen (0.1‐1 mg) also elicited dose‐dependent increases in the concentration of endogenous nitric oxide in the exhaled air. After an initial peak, exhaled NO concentrations returned to pre‐challenge levels.

The increase in insufflation pressure and in exhaled NO caused by ovalbumin challenge was inhibited by combined administration of histamine and leukotriene antagonists.

In non‐immunised guinea‐pigs, challenge of the airways with nebulised histamine (10–1000 nmol) or leukotriene C4 (LTC4, 30–300 pmol) elicited dose‐dependent increases in insufflation pressure and in concentrations of endogenous NO in exhaled air.

The increase in exhaled NO correlated with the increase in insufflation pressure in response to ovalbumin, histamine and LTC4. An inhibitor of endogenous NO synthesis, Nω‐nitro‐l‐arginine methylester (l‐NAME, 30 mg kg−1 i.v.) abolished NO exhalation, and markedly augmented the airway responses to ovalbumin, histamine, or LTC4.

The potentiation by l‐NAME of the increase in insufflation pressure in response to ovalbumin or histamine was prevented by exogenous NO (20 p.p.m.) in the inhaled air.

The results indicate that endogenous NO has an inhibitory effect on bronchial obstruction. Increased NO release during allergen challenge is likely to be due to actions of histamine and leukotrienes.