bluuebunny/crossref_metadata_2025 huggingface dataset:
In vitro denervation of the rat vas deferens through hypothermic storage
Authors: Jurkiewicz N.H. , Garcia A.G. , Jurkiewicz A.
Published: July 2012

The rat vas deferens was excised, stored at 4–6°C and tested after 24, 48, 72 or 96 h for its contractile activity and for the presence of innervation.

The maximal contractile capacity of the vas, tested through cumulative concentrations of barium chloride (3 × 10−2 m) was progressively reduced from about 110 mm to about 63 mm after 72 h, without further decay after 96 h. Spontaneous rhythmic contractions were practically absent.

A loss of endogenous pools of catecholamines was indicated by four parameters: (a) a decline of about 80% after 24 h and of more than 95% after 48 h of the contractile effect of the indirect sympathomimetic agonist tyramine; (b) a fall of about 20%, 50% and 85% on the concentration of noradrenaline, respectively after 24, 48 and 72 h; (c) a fall of about 25% and 90% after respectively 24 and 48 h, of the activity of dopamine‐β‐hydroxylase (DBH); (d) a decline of noradrenaline‐induced histofluorescence on cross sections of the vas.

A loss of neuronal uptake capacity was indicated by: (a) a progressive variation of the apparent affinity for adrenaline, expressed as pD2 values, that increased by about 1.5 log units (corresponding to a 30 fold potentiation) after 72 h, and (b) a reduction of the ability of cocaine to potentiate the contractile effects of adrenaline.

The pD2 values for barium chloride, 5‐hydroxytryptamine (5‐HT) and histamine were not significantly changed, while the corresponding value for acetylcholine was slightly but significantly reduced by about 0.8 log units.

6 The maximal heights of concentration‐response curves for noradrenaline, acetylcholine, histamine and 5‐HT were reduced by 42–66% in relation to controls. However, when this reduction was measured in relation to the corresponding barium effect, by means of the relative responsiveness ratio (p), a small though significant increase was observed for noradrenaline, and a fall for the other drugs.

7 It is concluded that: (1) the values for the various biochemical and pharmacological parameters decline at different rates, though revealing altogether that denervation is completed by at least 85% after 72 h of hypothermic storage; (2) two of the results, i.e., the lack of spontaneous rhythmic contractions and the lack of increased contractile effects for acetylcholine, 5‐HT and histamine, indicate that in these conditions the vas is devoid of the so‐called nonspecific signs of denervation.
Histamine H3 receptors modulate the release of [3H]‐acetylcholine from slices of rat entorhinal cortex: evidence for the possible existence of H3 receptor subtypes
Authors: Clapham J. , Kilpatrick G.J.
Published: July 2012

The effect of agents which interact with the histamine H3 receptor on potassium‐stimulated tritium release from slices of rat entorhinal cortex preloaded with [3H]‐choline is described. We have examined the effects of the selective H3 receptor agonist, (R)‐α‐methylhistamine (RAMH), and a number of H3 receptor antagonists, including the selective compound thioperamide, on the potassium‐stimulated release of tritium.

In the presence of mepyramine and ranitidine, RAMH (0.01–10 μm) inhibited potassium‐stimulated tritium release in a concentration‐dependent manner, EC50 = 0.11 μm. The maximum inhibition was approximately 50%.

Thioperamide displaced the RAMH concentration‐response curve to the right yielding a pKB value of 8.4. There was no change in the maximum response to RAMH.

Other H3 receptor antagonists, including impromidine and burimamide, also caused rightwards displacement of the linear portion of the RAMH concentration‐response curve. However, phenylbutanoylhistamine and betahistine, which are reported to be relatively potent H3 receptor antagonists, showed very low affinity.

Thioperamide (0.001–1 μm) alone enhanced the potassium‐stimulated release of tritium in a concentration‐dependent manner. Maximum effects were observed at 0.1–1 μm thioperamide, enhancing release by approximately 20%.

Results are discussed in terms of the regulatory role of H3 receptors on acetylcholine release and the possible existence of H3 receptor subtypes.
Bradykinin‐induced activation of nociceptors: receptor and mechanistic studies on the neonatal rat spinal cord‐tail preparation in vitro
Authors: Dray A. , Patel I.A. , Perkins M.N. , Rueff A.
Published: July 2012

The effects of bradykinin on nociceptors have been characterized on a preparation of the neonatal rat spinal cord with functionally connected tail maintained in vitro. Administration of bradykinin to the tail activated capsaicin‐sensitive peripheral fibres and evoked a concentration‐dependent (EC50 = 130 nm) depolarization recorded from a spinal ventral root (L3‐L5).

The response to bradykinin was unaffected by the peptidase inhibitors, bestatin (0.4 mm), thiorphan (1 μm), phosphoramidon (1 μm) and MERGETPA (10 μm) or by the presence of calcium blocking agents, cadmium (200 μm) and nifedipine (10 μm).

Inhibition of cyclo‐oxygenase with indomethacin (1–5 μm), aspirin (1–10 μm) and paracetamol (10–50 μm) consistently attenuated responses to bradykinin.

The effect of bradykinin was mimicked by the phorbol ester PDBu, an activator of protein kinase C. The response to bradykinin was attenuated following desensitization to PDBu but desensitization to bradykinin did not induce a cross‐desensitization to PDBu. The protein kinase C inhibitor staurosporine (10–500 nm) consistently attenuated the effects of PDBu and bradykinin.

Bradykinin responses were reversibly enhanced by dibutyryl cyclic AMP (100 μm). However dibutyryl cyclic GMP (0.5 mm) and nitroprusside (10 μm) produced prolonged block of responsiveness to bradykinin. Prolonged superfusion with pertussis toxin did not affect responses to bradykinin.

The B1‐receptor agonist des Arg9‐bradykinin (10–100 μm) was ineffective alone or after prolonged exposure of the tail to lipopolysaccharide (100 ng ml−1) or epidermal growth factor (100 ng ml−1) to induce B1 receptors. The B1‐receptor antagonist, des Arg9 Leu8‐bradykinin (10 μm) did not attenuate the response to bradykinin. A number of bradykinin B2 antagonists selectively and reversibly attenuated the response to bradykinin. The rank order potency was Hoe 140 > LysLys [Hyp3,Thi5,8,d‐Phe7]‐bradykinin > d‐Arg[Hyp3, Thi5,8, d‐Phe7]‐bradykinin = d‐Arg[Hyp2,Thi5,8, d‐Phe7]‐bradykinin.

These data show that bradykinin produces concentration‐dependent activation of peripheral nociceptors in the neonatal rat tail. The responses were unaffected by calcium channel block and were partially dependent on the production of prostanoids. Bradykinin‐evoked responses were consistent with the activation of protein kinase C‐dependent mechanisms. Cyclic GMP‐dependent mechanisms may be involved in bradykinin‐receptor desensitization whereas cyclic‐AMP dependent mechanisms increase fibre excitability and facilitate bradykinin‐induced responses. The effects of bradykinin were mediated by a B2 receptor.
Endothelium‐dependent relaxation of rabbit middle cerebral artery to a histamine H3‐agonist is reduced by inhibitors of nitric oxide and prostacyclin synthesis
Authors: Kim L. Ea , Javellaud J. , Oudart N.
Published: July 2012

The possible involvement of prostanoids and endothelium‐derived relaxing factor (EDRF) in the vasodilatation induced by a histamine H3‐agonist was examined in the rabbit perfused middle cerebral artery preconstricted with K+ (50 mm).

The endothelium‐dependent relaxation to (R)‐α‐methylhistamine [(R)‐α‐MeHA] was competitively antagonized by thioperamide (an H3‐antagonist) with a pA2 of 9.05, but unaffected by propranolol, atropine, l‐ and d‐sulpiride. This effect was stereoselective since the (S)‐isomer was 100 times less potent than the (R)‐isomer.

Two inhibitors of nitric oxide synthesis, NG‐nitro‐l‐arginine methyl ester (l‐NAME) and NG‐monomethyl‐l‐arginine (l‐NMMA), inhibited the relaxation induced by (R)‐α‐methylhistamine. The inhibitory effects of 10−5 m NG‐nitro‐l‐arginine methyl ester and 10−5 m NG‐monomethyl‐l‐arginine were reversed by equimolar concentrations of l‐arginine, but strongly enhanced by 10−4 m tranylcypromine. Tranylcypromine alone (10−5 m−10−4 m) partially reduced the (R)‐α‐methylhistamine‐induced relaxation. Both dexamethasone and indomethacin also inhibited this relaxation.

The results suggest that the H3‐mediated relaxation of the rabbit middle cerebral artery may involve release of both a prostanoid, probably prostacyclin, and endothelium‐derived relaxing factor. The relaxant effects of these two endogenous compounds appear to be synergistic.
Interaction of the central analgesic, tramadol, with the uptake and release of 5‐hydroxytryptamine in the rat brain in vitro
Authors: Driessen B. , Reimann W.
Published: July 2012

Tramadol is a centrally acting analgesic with low opioid receptor affinity and therefore presumably other mechanisms of analgesic action. Tramadol inhibits noradrenaline uptake but since 5‐hydroxytryptamine (5‐HT) is also involved in the modulation of pain perception, we tested the effects of tramadol on 5‐HT uptake and release in vitro.

Tramadol inhibited the uptake of [3H]‐5‐HT into purified rat frontal cortex synaptosomes with an IC50 of 3.1 μm. The (+)‐enantiomer was about four times more potent than the (−)‐enantiomer; the main metabolite of tramadol, O‐desmethyltramadol, was about ten times less potent.

Rat frontal cortex slices were preincubated with [3H]‐5‐HT, then superfused and stimulated electrically. Tramadol facilitated the basal outflow of [3H]‐5‐HT, at concentrations greater than 1 μm, while the uptake inhibitor 5‐nitroquipazine enhanced both basal and stimulation‐evoked overflow. Effects of the (+)‐enantiomer were more potent than either the racemate, the (−)‐enantiomer or the principal metabolite.

The effects of tramadol on the basal outflow of [3H]‐5‐HT were almost completely abolished when the superfusion medium contained a high concentration of the selective 5‐HT uptake blocker, 6‐nitroquipazine.

The results provide evidence for an interaction of tramadol with the neuronal 5‐HT transporter. An intact uptake system is necessary for the enhancement of extraneuronal 5‐HT concentrations by tramadol indicating an intraneuronal site of action.
The effects of dietary treatment with essential fatty acids on sciatic nerve conduction and activity of the Na+/K+ pump in streptozotocin‐diabetic rats
Authors: Lockett M.J. , Tomlinson D.R.
Published: July 2012

This study examined the effects of dietary essential fatty acid supplementation (5% (w/w) evening primrose oil) upon sciatic motor nerve conduction velocity and 86Rb+ pumping in sciatic nerve endoneurial preparations in rats with 4 to 5 weeks of streptozotocin‐induced diabetes.

Control diabetic rats (dietary supplementation with 5% (w/w) hydrogenated coconut oil) exhibited a reduction in motor nerve conduction velocity (16%; P < 0.05) compared to similarly‐fed non‐diabetic controls, but there was no significant alteration in ouabain‐sensitive 86Rb+ pumping, a parameter reflecting activity of the Na+/K+ pump.

Treatment of diabetic rats with evening primrose oil prevented completely the development of the motor nerve conduction velocity deficit without affecting the severity of diabetes. Evening primrose oil treatment did not significantly affect motor nerve conduction velocity of non‐diabetic animals.

Evening primrose oil treatment caused a significant reduction in activity of the Na+/K+ pump in sciatic nerves of diabetic animals (45%; P < 0.05).

These results suggest that the acute conduction velocity defect arising in streptozotocin‐diabetic rats, and the actions of evening primrose oil upon this, are independent of any effect on activity of the Na+/K+ pump. Other putative mechanisms are discussed.
The electrical and mechanical responses of the rabbit saphenous artery to nerve stimulation and drugs
Authors: Nally Jane E. , Muir T.C.
Published: July 2012

Electrical and mechanical responses to field stimulation (1–64 Hz, 0.5 ms supramaximal voltage) were recorded simultaneously in the rabbit saphenous artery. The electrical response consisted entirely of excitatory junction potentials (e.j.ps) which were abolished by α, β methylene ATP (α,β MeATP, 10−6 m) and by tetrodotoxin (TTX, 10−6 m) but were unaffected by the α1‐adrenoceptor antagonist, prazosin (10−6 m). No additional electrical response was evoked by field stimulation, even in the presence of normetanephrine (NMN) and desmethylimipramine (DMI, each 10−6 m), which block neuronal and extraneuronal uptake of noradrenaline (NA) respectively. Action potentials to field stimulation were produced only in the presence of tetraethylammonium (10−3 m) which also enhanced the contraction.

Contractions to field stimulation were reduced (by some 50%) by prazosin (10−6 m) and abolished by the additional presence of α,β MeATP (10−6 m), which blocks purinoceptors by desensitization, suggesting the involvement of both NA and an ATP‐like substance in the contractile response.

Idazoxan (10−6 m) which blocks prejunctional α2‐adrenoceptors, significantly increased the amplitude of both e.j.ps and the contraction to field stimulation (10 pulses, 1–4 Hz, 0.5 ms, supramaximal voltage).

NA (10−2 m by pressure ejection) did not alter membrane potential even in the presence of NMN and DMI (each 10−6 m). ATP (10−2 m by pressure ejection) produced a concentration‐dependent, α,β MeATP‐sensitive depolarization.

In tissues desensitized by constant infusion of α,β MeATP (10−6 m) contractions to NA (10−7−3 × 10−5 m), histamine (10−7 − 3 × 10−5 m) and KCl (1–1.6 × 10−2 m) were unaffected.

Following restoration of the membrane potential, after an initial depolarization, α,β MeATP (4 × 10−6 m) did not change the amplitude of electrotonic hyperpolarizing current pulses significantly but abolished evoked e.j.ps. The rates of recovery of evoked e.j.ps and the depolarization to ATP (10−2 m by pressure ejection) following desensitization to α,β MeATP (10−6 m) were comparable. These results suggest that the effects of α,β MeATP are mediated selectively via receptors (purinoceptors).

Suramin (10−3 m) abolished e.j.ps and the prazosin (10−6 m), insenstive component of the contractile response and antagonized contractions to α,β MeATP (10−7−10−5 m), ATP (10−5−10−3 m), histamine (10−7−3 × 10−5 m) and 5‐hydroxytryptamine (10−7−10−5 m) but those to NA (10−7−10−5 m) and KCl (1−1.6 × 10−2 m) were unaffected. Suramin is insufficiently selective, under these conditions, as a purinoceptor antagonist.
Adenosine receptors in rat basophilic leukaemia cells: transductional mechanisms and effects on 5‐hydroxytryptamine release
Authors: Abbracchio Maria P. , Paoletti Anna M. , Luini Alberto , Cattabeni Flaminio , DE Matteis M. Antonietta
Published: July 2012

The presence of adenosine receptors linked to adenylate cyclase activity and their functional role in calcium‐evoked 5‐hydroxytryptamine (5‐HT) release was investigated in rat basophilic leukaemia (RBL) cells, a widely used model for studying the molecular mechanisms responsible for stimulus‐secretion coupling.

In [3H]‐5‐HT‐loaded cells triggered to release by the calcium ionophore A23187, a biphasic modulation of 5‐HT secretion was induced by adenosine analogues, with inhibition of stimulated release at nm and potentiation at μm concentrations, suggesting the presence of adenosine receptor subtypes mediating opposite effects on calcium‐dependent release. This was also confirmed by results obtained with other agents interfering with adenosine pharmacology, such as adenosine deaminase and the non‐selective A1/A2 antagonist 8‐phenyl‐theophylline.

Similar biphasic dose‐response curves were obtained with a variety of adenosine analogues on basal adenylate cyclase activity in RBL cells, with inhibition and stimulation of adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) production at nm and μm concentrations, respectively. The rank order of potency of adenosine analogues for inhibition and stimulation of adenylate cyclase activity and the involvement of G‐proteins in modulation of cyclic AMP levels suggested the presence of cyclase‐linked A1 high‐affinity and A2‐like low‐affinity adenosine receptor subtypes. However, the atypical antagonism profile displayed by adenosine receptor xanthine antagonists on cyclase stimulation suggested that the A2‐like receptor expressed by RBL cells might represent a novel cyclase‐coupled A2 receptor subtype.

Micromolar concentrations of adenosine analogues could also increase inositol phospholipid hydrolysis and inositol tris‐phosphate formation in both unstimulated cells and in cells triggered to release by the calcium ionophore. The stimulation was constant, small and additive to that exerted by the calcium ionophore.

It is concluded that RBL cells express both A1 and A2‐like adenosine receptors which exert opposite effects on 5‐HT release and intracellular cyclic AMP levels. However, besides modulation of cyclic AMP levels, additional transduction pathways, such as modulation of phospholipase C activity, may contribute to the release response evoked by adenosine analogues in this cell‐line.
Nedocromil sodium prevents in vivo generation of the eosinophilotactic substance induced by PAF but fails to antagonize its effects
Authors: e Silva Patrícia M.R. , Martins Marco A. , Castro‐Faria‐Neto Hugo C. , Bozza Patricia T. , Pires Ana L.A. , Renato  , Cordeiro S.B. , Vargaftig B. Boris
Published: July 2012

The intrathoracic injection of platelet activating factor (PAF) into rats induced a decrease in the pleural leucocyte numbers within 15 min, accompanied by a marked exudation, maximal 1 h later. After 6 h, concomitantly with the reduction of exudation, a marked increase in the number of mononuclear cells, neutrophils and eosinophils was observed. Within 24 h, the pleural eosinophil accumulation peaked and persisted up to 96 h.

Topical treatment with nedocromil sodium affected pleural exudation by PAF under conditions where systemic meclizine was ineffective. Nedocromil sodium blocked, dose‐dependently, the increase in the pleural content of mononuclear cells, neutrophils and eosinophils, observed 6 h after PAF administration, as well as the eosinophilia 24 h later. Moreover, the co‐incubation of peritoneal eosinophils with nedocromil sodium did not interfere with the migration triggered by PAF.

The transfer of the 6 h‐PAF pleural washings from donor to recipient rats caused a selective pleural eosinophilia, which was clearly inhibited when nedocromil sodium was administered to donor, but not to recipient animals, showing that this drug interferes with the generation rather than with the expression of the eosinophilotactic activity(ies).

These findings indicate that the nedocromil sodium interferes with PAF‐induced exudation and leucocyte accumulation, by a mechanism other than its ability to reduce the local effects of histamine and which may relate to suppression of the eosinophilotactic principle generation.
Reduced α1‐ and β2‐adrenoceptor‐mediated positive inotropic effects in human end‐stage heart failure
Authors: Steinfath Markus , Danielsen Wiebke , von der Leyen Heiko , Mende Ulrike , Meyer Wilfried , Neumann Joachim , Nose Monika , Reich Torsten , Schmitz Wilhelm , Scholz Hasso , Starbatty Jutta , Stein Birgitt , Döring Volker , Kalmar Peter , Haverich Axel
Published: July 2012

α1‐Adrenoceptor (phenylephrine in the presence of propranolol) and β2‐adrenoceptor (fenoterol)‐mediated positive inotropic effects were investigated in human ventricular preparations isolated from five nonfailing (prospective organ donors) and from eight explanted failing hearts with end‐stage idiopathic dilative cardiomyopathy (NYHA IV).

For comparison, the nonselective β‐adrenoceptor agonist isoprenaline, the phosphodiesterase (PDE) inhibitor 3‐isobutyl‐1‐methylxanthine (IBMX), the cardiac glycoside dihydroouabain, and calcium were studied.

Furthermore, the influence of IBMX on adenosine 3′: 5′‐cyclic monophosphate (cyclic AMP) PDE activity as well as total β‐adrenoceptor density, β1‐ and β2‐adrenoceptor subtype distribution, and α1‐adrenoceptor density were compared in nonfailing and failing human heart preparations. The radioligands (−)‐[125I]‐iodocyanopindolol for β‐adrenoceptor binding and [3H]‐prazosin for α1‐adrenoceptor binding were used.

The inotropic responses to calcium and dihydroouabain in failing human hearts were unchanged, whereas the maximal α1‐ and β2‐adrenoceptor‐mediated positive inotropic effects were greatly reduced. The inotropic effects of the other cyclic AMP increasing compounds, i.e. isoprenaline and IBMX, were also reduced to about 60% of the effects observed in nonfailing controls. The potency of these compounds was decreased by factors 4–10.

The basal PDE activity and the PDE inhibition by IBMX were similar in nonfailing and failing preparations.

The total β‐adrenoceptor density in nonfailing hearts was about 70 fmol mg−1 protein. In failing hearts the total number of β‐adrenoceptors was markedly reduced by about 60%. The β1/β2‐adrenoceptor ratio was shifted from about 80/20% in nonfailing to approximately 60/40% in failing hearts which was due to a selective reduction of β1‐adrenoceptors. The β2‐adrenoceptor population remaining unchanged. α1‐Adrenoceptor density was increased from about 4 fmol mg−1 protein in nonfailing to 10 fmol mg−1 protein in failing hearts.

Changes in PDE activity and adrenoceptor downregulation cannot completely explain the reduced positive inotropic effects of α1‐ and β2‐adrenoceptor agonists in failing human hearts. This supports the hypothesis that impairment of other processes such as the coupling between receptor and effector system, i.e. the respective G‐proteins, are equally important in end‐stage heart failure.
Penicillin‐induced potentiation of glycine receptor‐operated choride current in rat ventro‐medial hypothalamic neurones
Authors: Tokutomi Naofumi , Agopyan Nadia , Akaike Norio
Published: July 2012

Effects of penicillin G (PCN) on glycine (Gly)‐evoked Cl− current (IGly) were investigated in acutely dissociated rat ventro‐medial hypothalamic (VMH) neurones by the whole cell mode of patch clamp technique.

When PCN was applied simultaneously with Gly, PCN depressed IGly like a Cl− channel blocker.

The PCN‐induced blocking action was clearly observed at a low PCN concentration (30 u), while the maximal blockade was achieved by 600 u (units per 10 ml) PCN.

When tested solution containing both PCN and Gly was quickly substituted with one containing Gly only, a new rebound‐like transient current (IT) which also passed through Cl− channel, was elicited.

The peak amplitude of IT induced by PCN at concentrations higher than 100 u was greater than that induced by glycine alone. We termed this phenomenon PCN‐induced potentiation of IGly. In all cells tested, PCN potentiated IGly.

At a lower PCN concentration below 30 u, IT generation was not clear in the presence of 10−5 m γ‐aminobutyric acid. With PCN a higher concentration than 300 u, IT amplitude was treater than that of the original peak IGly. This was observed in 18 neurones out of 21. The maximal amplitude of the IT was achieved with 600 u PCN.
Intervascular and stimulus selectivity of nitrendipine and related derivatives in KCl and prostaglandin F2α precontracted porcine arteries
Authors: Kojda Georg , Klaus Wolfgang , Werner Günther , Fricke Uwe
Published: July 2012

Dihydropyridine‐type calcium entry blockers exhibit a different vasodilator potency depending on the arterial tissue (intervascular selectivity) as well as on the precontracting stimulus used (stimulus selectivity). In addition, the structure of their ester side chains seems to influence their activity.

Vascular activity of nitrendipine and six related 3‐ester side chain derivatives was investigated in isolated coronary, ulnar and basilar arteries of the pig following precontraction with KCl or prostaglandin F2α (PGF2α).

After depolarization, all dihydropyridines exhibited a weak preferential action on coronary arteries. Bay E 6927 produced the strongest effect in all vessel types. By contrast, precontraction with PGF2α resulted in a marked preferential action in basilar arteries, although higher concentrations of the dihydropyridines were required for half maximal vasorelaxation. In each case, ulnar arteries were less sensitive.

Except with Bay O 5572, the most bulky substituted and least active derivative, only moderate differences were observed within the dihydropyridines studied. On the other hand, there was a pronounced increase in the ratios of the half maximal active concentrations required after precontraction of the vessels with PGF2α compared to KCl (stimulus selectivity) following a limited prolongation of the 3‐ester side chain up to an isopropyl‐group.

It is suggested that the observed shift in the intervascular selectivity after precontraction with PGF2α is a consequence of different contractile mechanisms in the three vessel types studied. The degree of the stimulus selectivity may also depend on the structure of the dihydropyridines.
Comparison of the contractile effects of endothelin‐1 and sarafotoxin S6b in goat isolated cerebral arteries
Authors: Salom Juan B. , Torregrosa Germán , Miranda Francisco J. , Alabadí José A. , Alborch Enrique
Published: July 2012

The effects of endothelium‐derived endothelin‐1 and snake venom‐derived sarafotoxin S6b, peptides with striking structural and functional similarities, were examined and compared in isolated middle cerebral arteries of goats.

Endothelin‐1 and sarafotoxin S6b contracted cerebral arteries in a concentration‐dependent manner. The potency of endothelin‐1 (EC50 = 4.9 (3.9–6.2) × 10−10 m) was about ten times higher than that of sarafotoxin S6b (EC50 = 5.5 (4.4–6.9) × 10−9 m). The tension returned to basal values after repeated washings and contraction with endothelin‐1 could be reproduced. Endothelin‐1 and sarafotoxin S6b induced further contraction in arteries precontracted with prostaglandin F2α (10−5 m).

Mechanical removal of the endothelium or incubation with indomethacin (10−5 m) displaced the concentration‐response curves to endothelin‐1 and, more pronouncedly, to sarafotoxin S6b to the left. The maximum response to sarafotoxin S6b was also increased by either of these two treatments.

Incubation in ‘nominally’ Ca2+‐free medium attenuated the vasoconstrictor response to endothelin‐1 but not to sarafotoxin S6b, which was inhibited after incubation in Ca2+‐free medium to which EGTA (10−4 m) had been added. Pretreatment with caffeine (2 × 10−2 m) in Ca2+‐free medium abolished responses to endothelin‐1 and sarafotoxin S6b.

Bay K 8644 (10−10 m, 10−8 m) enhanced and nicardipine (10−10 m, 10−8 m) inhibited in a concentration‐dependent manner vasoconstrictor response to endothelin‐1. Response to sarafotoxin S6b was only affected by 10−8 m Bay K 8644 or nicardipine.

It is concluded that endothelin‐1 and sarafotoxin S6b are potent vasoconstrictors of goat cerebral arteries, having direct effects on smooth muscle which are counteracted by the endothelium through the release of a vasodilatator substance, probably prostacyclin. Both endothelin‐1 and sarafotoxin S6b depend on extracellular Ca2+ and on intracellular, caffeine‐sensitive Ca2+ stores to develop vasoconstriction. However, endothelin‐1 depends to a larger extent than sarafotoxin S6b on free extracellular Ca2+.
Kinetic modulation of guinea‐pig cardiac L‐type calcium channels by fendiline and reversal of the effects of Bay K 8644
Authors: Schreibmayer Wolfgang , Tripathi Onkar , Tritthart Helmut A.
Published: July 2012

The modulation of L‐type calcium channel current (ICa) by fendiline, a diphenylalkylamine type of calcium channel blocker was investigated on guinea‐pig ventricular myocytes by use of the whole‐cell patch‐clamp technique.

Fendiline‐induced block of ICa is accompanied by modulation of the channel kinetics in a complex manner. The time course of ICa inactivation is significantly faster and the channel availability (f∞) curve is shifted considerably to more negative potentials by fendiline. These findings can be interpreted qualitatively in terms of a modulated receptor.

When the 1,4‐dihydropyridine agonist (4R, 4S)‐Bay K 8644 was added in presence of 30 μm fendiline a further reduction of ICa instead of the expected stimulatory effect was observed.

A similar ‘paradoxical’ inhibition of ICa was produced by the pure agonist enantiomer (4S)‐Bay K 8644. Thus this novel effect of Bay K 8644 cannot be attributed to changes in affinity of the 1,4‐dihydropyridine receptor site for (4R)‐Bay K 8644 during fendiline action.

The IC50 for fendiline was reduced to 3.0 ± 0.1 μm (control value: 17.0 ± 2.4 μm) and the Hill slope in its presence was increased to 1.90 ± 0.1 (control value: 1.39 ±0.23) by 1 μm (4R, 4S)‐Bay K 8644.

(4R,4S)‐Bay K 8644 caused the expected stimulation of ICa in the presence of verapamil, diltiazem and nifedipine, overcoming the inhibitory effect of these calcium channel blockers.

The ‘paradoxical’ inhibitory effect of the agonist Bay K 8644 can be explained in terms of an allosteric interaction between fendiline and the dihydropyridine agonist.
Effects of the neutral endopeptidase inhibitor, SQ 28,603, on regional haemodynamic responses to atrial natriuretic peptide or proendothelin‐1 [1–38] in conscious rats
Authors: Gardiner S.M. , Kemp P.A. , Bennett T.
Published: July 2012

Regional haemodynamic responses to atrial natriuretic peptide (ANP, 0.5 nmol kg−1) or proendothelin‐1 [1–38] (1.0 nmol kg−1) were assessed in the same conscious Long Evans rats before and 20 min after administration of the novel neutral endopeptidase (NEP) inhibitor, SQ 28,603 (50 mg kg−1, i.v.). In a separate experiment, responses to endothelin‐1 (0.5 nmol kg−1), angiotensin I (0.25 nmol kg−1) and angiotensin II (0.12 nmol kg−1) were measured before and after administration of SQ 28,603.

SQ 28,603 alone had no significant cardiovascular effects but caused significant prolongation of the hypotensive, tachycardic and renal and mesenteric vasoconstrictor effects of ANP. However, the early vasodilator and late vasoconstrictor responses in the hindquarters were not affected significantly.

SQ 28,603 caused significant attenuation of the pressor effects of proendothelin‐1 [1–38] but the associated bradycardia was unchanged. SQ 28,603 caused a significant inhibition of the renal and mesenteric vasoconstrictor effects of proendothelin‐1 and also inhibited the initial vasodilator and subsequent vasoconstrictor responses in the hindquarters vascular bed.

SQ 28,603 had no significant effects on the haemodynamic responses to endothelin‐1, angiotensin I, or angiotensin II.

The results are consistent with SQ 28,603 not only inhibiting NEP that is involved in the degradation of ANP, but also suppressing activity of the enzyme involved in the conversion of proendothelin‐1 [1–38] to endothelin‐1.
Effect of the hypoglycaemic drug (−)‐AZ‐DF‐265 on ATP‐sensitive potassium channels in rat pancreatic β‐cells
Authors: Ronner Peter , Le Hang Tu , Kraebber Markus J. , Higgins Timothy J.
Published: July 2012

It has previously been shown that the hypoglycaemic drug (−)‐AZ‐DF‐265 stimulates insulin release from mouse islets; in the presence of 3 mm glucose it also inhibits 86Rb‐efflux from 86Rb‐loaded islets. Based on these data we tested the hypothesis that (−)‐AZ‐DF‐265 inhibits ATP‐sensitive potassium channels.

We voltage‐clamped the plasma membrane of single rat pancreatic β‐cells in the whole‐cell configuration and measured the current flowing through ATP‐sensitive potassium channels. (−)‐AZ‐DF‐265 was applied to the outside of the cell; it inhibited the current half‐maximally at a concentration of 1.2 ± 0.2 nm with a Hill coefficient of 0.7 ± 0.1. The inhibition was reversible on washing.

In intact RINm5F cells, the sulphonylurea [3H]‐glibenclamide bound with an affinity of 0.24 ± 0.01 nm and a Hill coefficient of 2.1 ± 0.4. Treatment with (−)‐AZ‐DF‐265 led to the displacement of [3H]‐glibenclamide; this effect was half‐maximal at 8.6 ± 1.7 nm and displayed a Hill coefficient of 0.53 ± 0.01. Meglitinide and tolbutamide, which represent, respectively, the benzamido and sulphonylurea moieties of glibenclamide, showed Hill coefficients of 0.8 ± 0.1 and 0.9 ± 0.1, respectively.

At pH 7.4 and with phosphate/borate buffer, (−)‐AZ‐DF‐265 had an apparent octanol/water partition coefficient of about 53, which is intermediate between the partition coefficients of glibenclamide and glipizide. Nevertheless, (−)‐AZ‐DF‐265 bound only to a minor degree to glass and plastic.

In conclusion, (−)‐AZ‐DF‐265 inhibits ATP‐sensitive potassium channels and displaces [3H]‐glibenclamide from the sulphonylurea receptor.
Impaired cyclic nucleotide‐mediated vasorelaxation may contribute to closure of the human umbilical artery after birth
Authors: Renowden S. , Edwards D.H. , Griffith T.M.
Published: July 2012

The mechanical and biochemical effects of agents that relax vascular smooth muscle either through elevation of guanosine 3′:5′‐cyclic monophosphate (cyclic GMP) or adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) levels were compared in isolated ring preparations of human umbilical artery and rat aorta. Tone was established by preconstriction with 5‐hydroxytryptamine.

The endothelium‐dependent vasodilator calcium ionophore (A23187) (which stimulates endothelium‐derived relaxing factor [EDRF] release and thus acts through soluble guanylyl cyclase), sodium nitroprusside (which stimulates soluble guanylyl cyclase directly), and atrial natriuretic peptide (which stimulates particulate guanylyl cyclase) relaxed rat aorta but not human umbilical artery.

Sodium nitroprusside, 10 μm, increased cyclic GMP levels from 10 to 390 pmol mg−1 protein at 2 min in rat aorta, as compared with a slower, relatively attenuated rise from 5 to 116 pmol mg−1 protein after 15 min in human umbilical artery. The rise in cyclic GMP in the umbilical artery was not significantly augmented by the cyclic GMP phosphodiesterase inhibitor, MB22948. Atrial natriuretic peptide increased cyclic GMP levels in rat aorta but not in human umbilical artery.

Forskolin, 10 μm, which stimulates both soluble and particulate adenylyl cyclase, maximally relaxed rat aorta and increased cyclic AMP levels from 15 to 379 pmol mg−1 protein at 15 min, but did not significantly relax or increase cyclic AMP levels in human umbilical artery. After preincubation with the cyclic nucleotide phosphodiesterase inhibitor, IBMX, 10 μm forskolin increased cyclic AMP levels to 1365 pmol mg−1 protein at 30 min in human umbilical arteries, but these high levels were not accompanied by mechanical relaxation.

8‐Bromo‐cyclic GMP and 8‐bromo‐cyclic AMP which are lipophilic analogues of cyclic GMP and cyclic AMP, both maximally relaxed the rat aorta at a concentration of 10 μm, but did not significantly relax the human umbilical artery.

The findings indicate that elevated cyclic nucleotide levels are not associated with mechanical relaxation of the post‐partum human umbilical artery, as in other vessels such as rat aorta. This impaired response to cyclic nucleotides may contribute to closure of the umbilical artery after birth.
Cibacron blue stimulation of surfactant secretion in rat type II pneumocytes
Authors: Griese Matthias , Gobran Laurice I. , Rooney Seamus A.
Published: July 2012

The effect of cibacron blue, a selective P2y‐purinoceptor antagonist in some systems, on the stimulatory effect of adenosine 5′‐triphosphate (ATP) on [3H]‐phosphatidylchloine secretion was examined in primary cultures of rat type II pneumocytes prelabelled by overnight culture with [3H]‐choline.

Cibacron blue alone stimulated phosphatidylcholine secretion in a concentration‐dependent manner in the range 10−4−10−3 m. At a concentration of 10−4 or lower, cibacron blue had no effect on ATP‐induced phosphatidylcholine secretion but at 10−4−10−3 m it increased the effect of ATP. Enhancement of the ATP effect was apparent whether cibacron blue was added before or together with ATP. Cibacron blue also increased ATP‐induced secretion in the presence of the P1‐purinoceptor antagonist, xanthine amine congener (10−5 m).

The stimulatory effect of cibacron blue on phosphatidylcholine secretion was additive to those of 5′ (N‐ethylcarboxyamido) adenosine (NECA) and terbutaline but less than additive to that of ATP.

Cibacron blue alone had no effect on formation of cyclic AMP or inositol phosphate and when added stimultaneously with ATP it did not affect the ATP‐induced increase in these second messengers. Preincubation of the cells with cibacron blue before addition of ATP, however, resulted in antagonism of the ATP‐induced increase in cyclic AMP and inositol phosphates. Preincubation with ATP had the same effect. The stimulatory effects of NECA and terbutaline on cyclic AMP formation were enhanced by preincubation with cibacron blue.

Thus, ATP‐induced phosphatidylcholine secretion in type II cells is not diminished by the P2y‐antagonist, cibacron blue. On the contrary, cibacron blue stimulates phosphatidylcholine secretion. Cibacron blue may act as a P2‐agonist in type II pneumocytes.
Inhibitory effect of methysergide on calcitonin gene‐related peptide‐induced vasodilatation and ocular irritative changes in the rabbit
Authors: Krootila Kari , Oksala Olli , Zschauer Aino , Palkama Arto , Uusitalo Hannu
Published: July 2012

Calcitonin gene‐related peptide (CGRP) is involved in ocular neurogenic inflammation in the rabbit, causing vasodilatation in the anterior uvea, breakdown of the blood‐aqueous barrier, increase in the intraocular pressure (IOP) and rise in the adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) content in the aqueous humour. So far there is no means of preventing these CGRP‐induced ocular effects.

In the present study, the effect of intravenous methysergide (1–10 mg kg−1, b.w.) on CGRP‐induced changes in the IOP, blood‐aqueous barrier and cyclic AMP content in the aqueous humour was studied in vivo. The effect of methysergide on CGRP‐induced vasodilatation both in vivo and in vitro was also investigated.

Methysergide decreased intraocular pressure but had only a transient effect on blood pressure. Methysergide decreased the regional blood flow in ocular tissues by 53–65%, but did not have such a vasoconstrictor effect in most extra‐ocular tissues studied.

Methysergide inhibited CGRP‐induced vasodilatation, increase in the IOP, breakdown of the blood‐aqueous barrier and increase in the cyclic AMP content in the aqueous humour in vivo.

In vitro, methysergide alone did not have effects on the vascular tone in isolated ophthalmic artery of rabbit. However, it potentiated noradrenaline (NA)‐induced contraction. There were no differences in the IC50 values for CGRP on the NA‐induced contraction in the presence and absence of methysergide, indicating that methysergide has no direct effect on the vasorelaxant effect of CGRP in vitro.

The present study demonstrates that in the rabbit eye methysergide inhibits CGRP‐induced changes. One inhibitory mechanism of methysergide may be to enhance the effect of a vasoconstrictor (NA) to antagonize the vasodilator effect of CGRP. The present findings suggest that a methysergide‐sensitive mechanism may be used to limit some pathophysiological conditions in the eye that involve neurogenic inflammation and the release of CGRP.
Study of mechanisms of glucocorticoid hypertension in rats: endothelial related changes and their amelioration by dietary fish oils
Authors: Yin K. , Chu Z.M. , Beilin L.J.
Published: July 2012

To investigate possible mechanisms of increased systolic blood pressure after 1 weeks treatment with dexamethasone and its amelioration by fish oil feeding, we have examined the reactivity of aortic rings and perfused mesenteric resistance vessels.

Thirty six Sprague‐Dawley rats were initially divided into two groups and fed a semisynthetic diet containing either (10% by weight) hydrogenated coconut oil and safflower oil mixture (HCO/S) (24 rats) or fish oil (12 rats) for 5 weeks. From the end of the fourth week, dexamethasone (1.25 mg ml−1) in drinking water, was given to half the rats on hydrogenated coconut oil (HCO/S + Dex) and to the fish oil‐fed group (fish oil + Dex).

One week of dexamethasone treatment raised systolic blood pressure in the HCO/S + Dex rats but not in the fish oil + Dex group.

Endothelium‐dependent relaxation to acetylcholine (ACh) was decreased in aortic rings taken from HCO/S + Dex rats compared to rats on HCO/S alone. Relaxant responses to ACh of aortic rings from rats given fish oil + Dex were intermediate between the three groups. Aortic endotlielium‐independent responses to sodium nitroprusside (SNP) were unchanged between the groups, while aortic contractile responses to noradrenaline were similar in all the groups.

In the perfused mesenteric resistance artery, sensitivity to noradrenaline was decreased in rats given fish oil and dexamethasone compared to the other two groups. There were no differences in resistance vessel relaxation to ACh or SNP between groups.

Serum corticosterone levels, used as a marker of dexamethasone absorption, were substantially suppressed in dexamethasone‐treated rats but levels were higher in rats on fish oil than on HCO/S diets.

We suggest that the glucocorticoid‐induced rise in systolic blood pressure may be due in part to decreased aortic compliance as a consequence of impaired endothelium‐dependent relaxation and perhaps reduced nitric oxide synthesis. Fish oil feeding may ameliorate this rise in blood pressure through (i) changes in dexamethasone absorption, (ii) decrease in reactivity to noradrenaline of perfused mesenteric resistance arteries, (iii) an increase in endothelium‐dependent relaxation to ACh or a combination of these three factors.
Effects of imipramine on the transient outward current in rabbit atrial single cells
Authors: Delpón Eva , Tamargo Juan , Sánchez‐Chapula José
Published: July 2012

. The effects of imipramine on action potential characteristics and transient outward potassium current (It) of rabbit isolated atrial myocytes were studied using the whole‐cell configuration of the patch‐clamp technique.

Imipramine, 3 μm, decreased action potential amplitude and lengthened the action potential duration measured at 50% of repolarization, whereas it did not modify the final phase of repolarization or the resting membrane potential. These results are similar to those reported in multicellular rabbit atrial preparations.

Imipramine, 0.1–100 μm, induced a concentration‐dependent inhibition of the peak amplitude of It, a shortening of the time to peak current and an increase in the inactivation rate. The acceleration of the current inactivation is to a major extent responsible for the decrease in the integral of the outward current measured at 50 ms after the start of the pulse.

The drug‐induced block of It was not associated with changes in the voltage‐dependence of the steady‐state inactivation curve or in the process of recovery from inactivation of the current. Extrapolation to zero block shows that imipramine did not block It before its activation at the onset of the depolarization. These results suggested that imipramine does not affect the inactivated or the resting state of It channels.

It is concluded that in rabbit isolated atrial cells, imipramine inhibits It and that this effect is responsible for the lengthening of the action potential duration produced by this drug.
Coeliac haemodynamic effects of endothelin‐1, endothelin‐3, proendothelin‐1 [1–38] and proendothelin‐3 [1–41] in conscious rats
Authors: Gardiner S.M. , Kemp P.A. , Compton A.M. , Bennett T.
Published: July 2012

Conscious, chronically‐instrumented, Long Evans rats were given bolus doses of endothelin‐1, endothelin‐3 (both at 0.01 and 0.1 nmol kg−1), proendothelin‐1 [1–38] and proendothelin‐3 [1–41] (both 0.1 and 1 nmol kg−1) in order to compare their effects on coeliac haemodynamics, because it has been reported that, in conscious dogs, endothelin‐1 has paradoxical, prolonged hyperaemic vasodilator effects in this vascular bed. Measurements were made also of mesenteric and hindquarters haemodynamics for comparison. In a separate experiment, endothelin‐1 (0.1 nmol kg−1) was given before and 20 min after the onset of an infusion of mecamylamine (50 μmol kg h−1) to ensure that the responses measured were not confounded by rapid reflex changes in autonomic activity.

None of the peptides caused any increases in coeliac flow or any sustained rises in coeliac vascular conductance, although such changes were clear‐cut in the hindquarters vascular bed following the higher dose of endothelin‐1 and endothelin‐3. In animals treated with mecamylamine the regional haemodynamic effects of the higher dose endothelin‐1 were not different from those in animals with intact baroreflexes.

Although the lower dose of both endothelin‐1 and endothelin‐3 caused less marked coeliac than mesenteric vasoconstriction, this difference was not apparent with the higher dose of the peptides, or with proendothelin‐1 [1–38]. However, proendothelin‐3 [1–41] had less marked coeliac and hindquarters vasoconstrictor effects than proendothelin‐1 [1–38], in spite of both peptides causing similar changes in mesenteric haemodynamics. These differences could have been due to regional differences in the conversion of proendothelin‐1 [1–38] and proendothelin‐3 [1–41] to endothelin‐1 and endothelin‐3, respectively. Our findings contradict the proposition that exogenous proendothelin‐3[1–41] is not converted into endothelin‐3 in vivo.

Since, in contrast to endothelin‐1 and endothelin‐3, proendothelin‐1 [1–38] and proendothelin‐3 [1–41] caused only small hindquarters vasodilatations, but large vasoconstrictions, it is feasible that the vasodilator responses to exogenous endothelin‐1 and endothelin‐3 are pharmacological phenomena, and that, in vivo, the production of endothelins from proendothelins exerts widespread vasoconstrictor effects.
Characterization of 8‐OH‐DPAT‐induced hypothermia in mice as a 5‐HT1A autoreceptor response and its evaluation as a model to selectively identify antidepressants
Authors: Martin Keith F. , Phillips Ian , Hearson Mitchell , Prow Michael R. , Heal David J.
Published: July 2012

8‐Hydroxy‐2‐(di‐n‐propylamino)tetralin (8‐OH‐DPAT) dose‐dependently induced hypothermia in mice.

The 5‐HT1A receptor partial agonists, buspirone, gepirone and ipsapirone, also dose‐dependently induced hypothermia.

The 8‐OH‐DPAT temperature response was antagonized by the 5‐HT1 receptor antagonists quipazine (2 mg kg−1, i.p.), (±)‐propranolol (10 mg kg−1, i.p.). (±)‐pindolol (5 mg kg−1, i.p.), spiroxatrine (0.5 mg kg−1, i.p.) and metitepine (0.05 mg kg−1, i.p.), but not by 5‐HT2 (ketanserin) or 5‐HT3 (MDL 72222, GR 38032F) receptor antagonists.

The response was also antagonized by the dopamine D2 receptor antagonists, haloperidol and BRL 34778. No other catecholamine or muscarinic receptors were involved in mediating the response.

Destruction of 5‐hydroxytryptamine (5‐HT)‐containing neurones with the neurotoxin, 5,7‐dihydroxytryptamine (75 μg, i.c.v.), abolished the response to 8‐OH‐DPAT indicating that the 5‐HT1A receptors involved were located on 5‐HT neurones.

6 Chronic antidepressant treatment down‐regulated this 8‐OH‐DPAT response. In addition, chronic administration of anxiolytics and neuroleptics was also effective in this respect. Down‐regulation was also observed following repeated administration of 8‐OH‐DPAT (0.5 mg kg−1, s.c.), (±)‐pindolol (10 mg kg−1, i.p.) and ketanserin (0.5 mg kg−1, i.p.).

7 In conclusion, these data confirm that 8‐OH‐DPAT‐induced hypothermia is mediated by 5‐HT1A autoreceptors. They also indicate that the response involves D2 receptors. The present study also shows that a wide range of antidepressant drugs down‐regulate this response although this property is not restricted to antidepressant treatments. Therefore, care should be exercised when interpreting data from this paradigm.