bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparison of haemoconcentration induced by big endothelin‐1 and endothelin‐1 in mice
Authors: Okumura Hiroshi , Ashizawa Naoki , Kobayashi Fujio , Arai Koshi , Asakura Rieko , Ashikawa Nami , Matsuura Akihiro
Published: July 2012

The profile of haemoconcentration induced by big endothelin‐1 (big ET‐1), a precursor of endothelin‐1 (ET‐1), was compared with that induced by endothelin‐1 in mice.

ET‐1(1.5 nmol kg−1, i.v.) increased haematocrit in mice, which reached a maximum at 5 min and then returned to the control value within 30 min after the administration, this occurred at the same time as changes in the plasma immunoreactive endothelin‐1 and rat atrial natriuretic peptide (rANP)‐like activities (IR‐ET‐1 and IR‐rANP, respectively).

Big ET‐1(2.5–15 nmol kg−1, i.v.) also caused a significant and dose‐dependent increase in haematocrit, that lasted over 3 h although elevated plasma IR‐ET‐1 and IR‐rANP had almost been restored to the initial levels within 10 min after big ET‐1 injection.

A metalloproteinase inhibitor, phosphoramidon (10 mg kg−1, i.v.), which inhibits the activity of endothelin converting enzyme (ECE), delayed the onset of big ET‐1‐induced haemoconcentration, but failed to alter the maximal value and the duration of the haemoconcentration.

Pretreatment with phosphoramidon (10 mg kg−1, i.v.) did not affect the big ET‐1‐induced change in plasma IR‐ET‐1, while significant delay of the disappearance of plasma IR‐rANP and significant suppression of a sustained increase in tissue IR‐ET‐1 were observed.

These results suggest that ET‐1, not in plasma but in tissue, plays an important role in the pathogenesis of big ET‐1‐induced long‐lasting haemoconcentration, in which unknown factors besides rANP are involved.
Pharmacological characterization of the metabotropic glutamate receptor inhibiting d‐[3H]‐aspartate output in rat striatum
Authors: Lombardi Grazia , Alesiani Marina , Leonardi Patrizia , Cherici Giovanna , Pellicciari Roberto , Moroni Flavio
Published: July 2012

The effects of several agonists of the metabotropic glutamate receptor (mGluR) were studied in adult rat striatal slices by measuring (i) KCl (30 mm)‐induced output of previously taken up d‐[3H]‐aspartate (Asp), (ii) forskolin (30 μm)‐induced adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) accumulation and (iii) phophoinositide (PI) hydrolysis.

K+‐induced efflux of d‐[3H]‐Asp was inhibited by the following mGluR agonists: (1S,3S,4S)‐(carboxycyclopropyl)glycine (l‐CCG‐I), (1S,3R)‐1‐aminocyclopentane‐1,3‐dicarboxylic acid (1S,3R‐ACPD) and quisqualic acid (Quis). 2‐Amino‐4‐phosphonobutyrate (l‐AP4) was inactive up to 300 μm. The maximal inhibition of d‐[3H]‐Asp output was 60 ± 8%. The EC50s of mGluR agonists were: 0.5 μm for l‐CCG‐I, 100 μm for 1S,3R‐ACPD and 100 μm for Quis.

Forskolin‐induced cyclic AMP accumulation was also inhibited by mGluR agonists. The maximal inhibition was 50 ± 4% and was obtained at a concentration of 10 μm for l‐CCG‐I and 100 μm for 1S,3R‐ACPD. The EC50s for this inhibition were: 0.9 μm for l‐CCG‐I and 20 μm for 1S,3R‐ACPD. Quis (300 μm) inhibited cyclic AMP accumulation by approximately 20%. l‐AP4 slightly potentiated cyclic AMP accumulation.

PI hydrolysis was stimulated by mGluR agonists. The most potent compound was Quis (100 μm), which increased inositol phosphate formation up to 2.2 fold over control values. Its EC50 was 15 μm.l‐CCG‐I and 1S,3R‐ACPD increased inositol phosphate formation by approximately 1.8 fold and their EC50 values were 30 and 25 μm, respectively. l‐AP4 did not affect PI hydrolysis.

In conclusion, mGluR agonists that reduce d‐[3H]‐Asp output have a pharmacological profile similar to that of mGluR agonists inhibiting cyclic AMP accumulation. l‐CCG‐I appears to be a relatively selective agonist for the mGluR receptor which inhibits d‐[3H]‐Asp efflux and cyclic AMP accumulation, while Quis appears to act preferentially on the mGluR receptor linked to the metabolism of PIs.
Prejunctional modulation of noradrenaline release in mouse and rat vas deferens: contribution of P1‐ and P2‐purinoceptors
Authors: Kurz Katharina , von Kügelgen Ivar , Starke Klaus
Published: July 2012

Prejunctional purinoceptors modulating the release of noradrenaline were compared in mouse and rat vas deferens. Tissue slices were preincubated with [3H]‐noradrenaline and then superfused and stimulated electrically, in most experiments by trains of 60 pulses, 1 Hz.

In mouse vas deferens, 2‐chloroadenosine (IC50 0.24 μm), β,γ‐methylene‐ATP (IC50 3.8 μm), α,β‐methylene‐ATP (IC50 2.9 μm) and 2‐methylthio‐ATP (only 30 μm tested) reduced the evoked overflow of tritium. 8‐Cyclopentyl‐1,3‐dipropylxanthine (DPCPX), 10 nm, antagonized the effect of 2‐chloroadenosine (apparent pKB 10.2) as well as of β,γ‐methylene‐ATP (apparent pKB 9.6) and α,β‐methylene‐ATP. Suramin, 300 μm, attenuated the effect of 2‐chloroadenosine at best very slightly, antagonized the effect of β,γ‐methylene‐ATP (apparent pKB 4.5) and, when combined with DPCPX 10 nm, caused a further marked shift to the right of the concentration‐response curve of β,γ‐methylene‐ATP beyond the shift produced by DPCPX alone.

In rat vas deferens, 2‐chloroadenosine (IC50 0.20 μm), β,γ‐methylene‐ATP (IC50 4.8 μm); α,β‐methylene‐ATP (IC50 3.0 μm) and 2‐methylthio‐ATP (only 30 μm tested) also reduced the evoked overflow of tritium. DPCPX, 10 nm, antagonized the effect of 2‐chloroadenosine (apparent pKB 9.7) as well as of β,γ‐methylene‐ATP (apparent pKB 9.6) and α,β‐methylene‐ATP. Suramin, 300 μm, did not change the effect of 2‐chloroadenosine, attenuated the effect of β,γ‐methylene‐ATP at best very slightly and, when combined with DPCPX, caused at best a very small shift to the right of the concentration‐response curve of β,γ‐methylene‐ATP beyond the shift produced by DPCPX alone.

It is concluded that prejunctional purinoceptor mechanisms in mouse and rat vas deferens are similar. In either species, both nucleosides such as adenosine and nucleotides such as β,γ‐methylene‐ATP activate a common release‐inhibiting receptor which is a P1‐ or, more specifically, A1‐purinoceptor. There seems to be no need to postulate the existence of a novel prejunctional P3‐purinoceptor. Moreover, the sympathetic terminal axons possess an additional P2‐purinoceptor in both species which is activated by some nucleotides such as β,γ‐methylene‐ATP and 2‐methylthio‐ATP, although the activation of the P2‐purinoceptor by β,γ‐methylene‐ATP is difficult to demonstrate in the rat.
PPADS selectively antagonizes P2X‐purinoceptor‐mediated responses in the rabbit urinary bladder
Authors: Ziganshin Airat U. , Hoyle Charles H.V. , Bo Xuenong , Lambrecht Günter , Mutschler Ernst , Bäumert Hans G. , Burnstock Geoffrey
Published: July 2012

Pyridoxalphosphate‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS), an inhibitor of P2X‐purinoceptor‐mediated responses in rabbit vas deferens, was investigated for its ability to antagonize contractions evoked by α,β‐methylene ATP (α,β‐MeATP), carbachol and electrical field stimulation in the rabbit urinary bladder detrusor muscle.

PPADS (1–30 μm) caused concentration‐dependent inhibition of contractions to the stable P2X‐purinoceptor agonist, α,β‐MeATP, decreasing the maximum response to α,β‐MeATP (30 μm) at concentrations of 3–30 μm. The pD2 value for α,β‐MeATP in the absence of PPADS was 6.52 ± 0.10 (8). In the presence of PPADS at concentrations of 1, 3, 10 and 30 μm the negative log concentrations of α,β‐MeATP that cause the same contractile response as the pD2 value were significantly different from control, being respectively 6.17 ± 0.09 (8), 5.64 ± 0.12 (7), 5.15 ± 0.23 (7) and 4.78 ± 0.22 (5).

PPADS (1–30 μm) caused concentration‐dependent inhibition of contractions to stimulation of intramural purinergic nerves (1–32 Hz). There was a greater inhibition at lower frequencies (1–8 Hz) than at higher frequencies (16–32 Hz). PPADS, 30 μm, did not produce significantly greater antagonism than 10 μm.

PPADS (30 μm) had no significant influence on the contractile potency of carbachol: the pD2 values of carbachol in the absence and presence of PPADS were not significantly different being 6.42 ± 0.16 (5) and 6.33 ± 0.18 (5), respectively. However, PPADS caused a small, but significant, suppression of the maximal response of carbachol, reducing it by approximately 9%.

Radioligand binding studies carried out on rabbit bladder membranes with [3H]‐α,β‐methylene ATP ([3H]‐α,β‐MeATP) showed that PPADS concentration‐dependently inhibited the binding of [3H]‐α,β‐MeATP to P2X‐purinoceptors, while the binding of [3H]‐quinuclidinyl benzilate to muscarinic cholinoceptors was not affected.

Thus, PPADS (1–30 μm) antagonized responses mediated via P2X‐purinoceptors in the rabbit urinary bladder. It was selective for P2‐purinoceptor‐mediated contractions rather than those mediated via muscarinic receptors. Binding studies demonstrated that the antagonistic effect of PPADS is via a direct interaction with P2X‐purinoceptors.
Suppression by intradermal administration of heparin of eosinophil accumulation but not oedema formation in inflammatory reactions in guinea‐pig skin
Authors: Teixeira M.M. , Hellewell P.G.
Published: July 2012

Heparin is widely used in the treatment of thrombotic disorders and as an aid in surgery. Antiinflammatory effects of heparin have also been described. In this study, we have investigated the effects of locally‐injected heparin on the oedema formation and eosinophil accumulation induced by various inflammatory stimuli in guinea‐pig skin.

Heparin dose‐dependently suppressed the accumulation of 111In‐labelled eosinosphils induced by i.d. injection of zymosan‐activated plasma (ZAP). The 111In‐eosinophil accumulation induced by other inflammatory stimuli (compound 48/80, platelet activating factor, interleukin‐8 and in a passive cutaneous anaphylaxis reaction) was also suppressed by locally‐injected heparin.

Oedema formation in response to these same stimuli was not altered by the local injection of heparin.

Fucoidin, a negatively‐charged sulphated algal polymer, had no effect on the 111In‐eosinophil accumulation or oedema formation induced by ZAP. Nevertheless, fucoidin significantly suppressed the oedema formation induced by i.d. injection of cationic protein‐containing extracts of Schistosoma mansoni larvae. Heparin also inhibited oedema induced by the extracts, suggesting that both fucoidin and heparin were effectively neutralizing the cationic protein of the extracts to inhibit their oedema‐inducing activity.

Thus, heparin significantly inhibited the accumulation of 111In‐eosinophils, but not oedema formation, induced by various inflammatory stimuli. This, taken together with the lack of effect of fucoidin, suggests that heparin interferes with the process of eosinophil trafficking by a mechanism that does not depend on neutralisation of the charge of the stimulatory molecules.
Role of prostaglandins and nitric oxide in acute inflammatory reactions in guinea‐pig skin
Authors: Teixeira M.M. , Williams T.J. , Hellewell P.G.
Published: July 2012

Oedema formation in skin is dependent on a synergism between mediators that increase vascular permeability and mediators that enhance local blood flow. Leukocyte accumulation is also enhanced by mediators that increase local blood flow. In this study, we have investigated whether nitric oxide (NO), an important endogenous vasodilator, could modulate oedema formation and leukocyte accumulation in guinea‐pig skin.

Local administration of the NO synthesis inhibitor, NG‐nitro‐l‐arginine methyl ester (l‐NAME), dose‐dependently inhibited the oedema formation induced in response to intradermal injection of bradykinin or histamine. l‐NAME, but not NG‐nitro‐d‐arginine methyl ester (d‐NAME), also inhibited oedema formation in response to i.d. injection of platelet‐activating factor (PAF), zymosan‐activated plasma (ZAP) and in a passive cutaneous anaphylactic (PCA) reaction.

N‐iminoethyl‐l‐ornithine (l‐NIO) was less effective and about 100 times less potent than l‐NAME in inhibiting bradykinin‐induced oedema formation. The cyclo‐oxygenase inhibitor, ibuprofen, had little effect on oedema responses induced by bradykinin, PAF and in a PCA reaction. On the other hand, histamine‐induced oedema formation was significantly suppressed by ibuprofen.

The inhibition by l‐NAME of bradykinin‐induced oedema formation was reversed by co‐injection of sodium nitroprusside (SNP) or prostaglandin E1 (PGE1).

L‐NAME inhibited 111In‐eosinophil and 111In‐neutrophil accumulation induced by i.d. injection of ZAP. 111In‐eosinophil accumulation induced by PAF and in the PCA reaction was also inhibited by l‐NAME but not by d‐NAME.

Co‐injection of SNP or PGE1, reversed the inhibition by l‐NAME of ZAP‐induced oedema formation and 111In‐neutrophil accumulation. SNP, but not PGE1, also reversed the effects of l‐NAME on ZAP‐induced 111In‐eosinophil accumulation.

l‐NAME caused a significant decrease in basal cutaneous blood flow when injected alone or with bradykinin. Again, SNP or PGE1 reversed the effects of l‐NAME suggesting that the inhibitory action of l‐NAME on oedema formation and cell accumulation was due to an inhibition of vasodilator tone in the microcirculation.

Thus, it appears that in guinea‐pig skin the inhibition of the production of endogenous NO inhibits both leukocyte accumulation and oedema formation induced by different mediators of inflammation. Since administration of l‐NAME also causes a local decrease in basal blood flow, we suggest that this is the mechanism by which it exerts anti‐inflammatory effects in this model.
Antagonism of levcromakalim by imidazoline‐ and guanidine‐derivatives in rat portal vein: involvement of the delayed rectifier
Authors: Ibbotson Timothy , Edwards Gillian , Weston Arthur H.
Published: July 2012

In rat whole portal veins, guanabenz (100 nm to 10 μm) and antazoline (100 nm to 100 μm) each increased the amplitude, frequency and duration of spontaneous contractions. In addition, guanabenz (30 μm) and antazoline (30 μm) each antagonized the ability of levcromakalim (3 nm to 10 μm) to inhibit the spontaneous contractions of this tissue.

Whole‐cell voltage‐clamp recordings were made from freshly‐isolated rat portal vein cells dispersed by a collagenase/pronase enzyme treatment. The ability of several agents (antazoline, cirazoline, clonidine, guanabenz and phentolamine, each containing an imidazoline or guanidine moiety), to modulate potassium (K) currents and to inhibit the actions of levcromakalim was investigated.

Antazoline, cirazoline, clonidine, guanabenz and phentolamine (each at a concentration of 30 μm) had little effect on control non‐inactivating currents but inhibited the delayed‐rectifier current, IK(V).

Levcromakalim (1 μm) induced a non‐inactivating current, IK(ATP), and also inhibited the delayed rectifier current, IK(V).

Glibenclamide (1 μm) had no effect on control delayed rectifier or non‐inactivating currents, but it inhibited the simultaneous induction of IK(ATP) and reduction of IK(V) produced by levcromakalim (1 μm).

Antazoline, cirazoline, clonidine and guanabenz (each at a concentration of 30 μm) prevented the induction of IK(ATP) by levcromakalim (1 μm). Phentolamine (30 μm) and clonidine (30 μm) each inhibited the IK(ATP) generated by levcromakalim (1 μm).

It is concluded that a variety of agents which possess either an imidazoline (antazoline, cirazoline, clonidine and phentolamine) or a guanidine (guanabenz) moiety within their structure inhibit the delayed rectifier current, IK(V). This action may thus be mediated via a so‐called non‐adrenoceptor imidazoline binding site. Furthermore, the ability of these ligands to inhibit IK(V) and to antagonize both the induction of IK(ATP) and the vasorelaxation produced by levcromakalim is consistent with the view that the channel (KATP) which underlies IK(ATP) is a voltage‐insensitive state of the delayed rectifier K‐channel (KV).
The guinea‐pig distal colon ‐ a sensitive preparation for the investigation of 5‐HT4 receptor‐mediated contractions
Authors: Wardle Kay A. , Sanger Gareth J.
Published: July 2012

Experiments were designed to characterize pharmacologically the contractile responses to 5‐hydroxytryptamine (5‐HT) in the guinea‐pig isolated distal colon longitudinal muscle‐myenteric plexus preparation (LMMP).

In the presence of methiothepin (100 nm) and granisetron (1 μm), 5‐HT (10 pm −‐ 10 nm) produced concentration‐dependent contractile responses of the guinea‐pig distal colon LMMP, with a pEC50 of 9.2 ± 0.08.

Responses to 5‐HT were mimicked by a series of tryptamine analogues, with the following rank order of potency; 5‐HT > 5‐MeOT >> 5‐CT > tryptamine > 2‐Me‐5‐HT. All were found to be full agonists.

Responses to 5‐HT were also mimicked by a series of substituted benzamide analogues. Their rank order of potency was 5‐HT > renzapride > cisapride > (S)‐zacopride > (R)‐zacopride > metoclopramide. All were full agonists relative to 5‐HT.

The benzimidazolone derivatives, BIMU 1 and BIMU 8 were approximately equipotent partial agonists (intrinsic activities of 0.8 ± 0.07 and 0.5 ± 0.08 respectively) in the guinea‐pig distal colon.

Tropisetron produced a rightward displacement of the 5‐HT concentration‐effect curve, yielding an apparent pA2 of 6.4 ± 0.1. The slope of the Schild plot (1.3 ± 0.1) was significantly greater than unity.

SDZ 205 557 produced a concentration‐dependent shift to the right of the 5‐HT concentration‐response curve, yielding an estimated pA2 of 7.8 ± 0.1 and a slope which did not significantly deviate from unity. SDZ 205 557 produced similar pKB estimates (7.3–7.9) when tested against 5‐MeOT, renzapride and 5‐CT, indicating a common site of action.

The pharmacological profile of the 5‐HT‐evoked contractions of the guinea‐pig distal colon LMMP are consistent with activity at the 5‐HT4 receptor. Furthermore, of the models of this receptor described in the literature, the guinea‐pig distal colon appears to be the most sensitive model to date, making it a useful tool in the investigation of 5‐HT4 receptor‐mediated events.
Profile of capsaicin‐induced mouse ear oedema as neurogenic inflammatory model: comparison with arachidonic acid‐induced ear oedema
Authors: Inoue Hideo , Nagata Nobuyuki , Koshihara Yasuko
Published: July 2012

We have investigated the mechanism of capsaicin‐induced mouse ear oedema compared with that of arachidonic acid (AA)‐induced ear oedema, and evaluated the possible involvement of neuropeptides in the development of capsaicin‐induced oedema.

Topical application of capsaicin (0.1–1.0 mg per ear) to the ear of mice produced immediate vasodilatation and erythema followed by the development of oedema which was maximal at 30 min after the treatment. This oedema was of shorter duration with less swelling than AA‐induced oedema (2.0 mg per ear).

Capsaicin‐induced ear oedema was unaffected when inhibitors of arachidonate metabolites including platelet activating factor (PAF) were administered before capsaicin (250 μg per ear) application, while these agents significantly prevented AA‐induced oedema. Dexamethasone, histamine H1 and/or 5‐hydroxytryptamine (5‐HT) antagonists, and substance P (SP) antagonists were effective in inhibiting both models. Furthermore, a Ca2+‐channel blocker and the capsaicin inhibitor, ruthenium red, were effective inhibitors of capsaicin oedema but had no effect on AA‐induced oedema.

Phosphoramidon (50 μg kg−1, i.v.), an endopeptidase inhibitor, markedly (P < 0.001) enhanced only capsaicin‐induced ear oedema, but bestatin (0.5 mg kg−1, i.v.), an aminopeptidase, failed to enhance oedema formation.

Neuropeptides (1–100 pmol per site) such as rat calcitonin gene‐related peptide (CGRP), SP, neurokinin A (NKA), and vasoactive intestinal peptide (VIP), which are released from capsaicin‐sensitive neurones, caused ear oedema by intradermal injection. Furthermore, a synergistic effect of CGRP (10 fmol per site) and SP (10 pmol per site) on oedema formation was observed.

The oedema induced by neuropeptides was significantly (P < 0.05 or P < 0.001) inhibited when cyproheptadine (20 mg kg−1, p.o.), a histamine H1 and 5‐HT antagonist, was administered before injection. In contrast, nifedipine (50 mg kg−1, p.o.), a Ca2+‐channel blocker, and indomethacin (10 mg kg−1, p.o., except for NKA), a cyclo‐oxygenase inhibitor, had little effect on neuropeptide‐induced oedema.

These results suggest that the mechanism of capsaicin‐induced ear oedema is different from that of AA‐induced oedema and suggest that the development of capsaicin‐induced ear oedema is primarily mediated by neuropeptides. The neuropeptides released after activation of sensory nerves cause an increase of vascular permeability by interactions with endothelial cells and by histamine (and 5‐HT) release from mast cells.
Effect of activation on adhesion of flowing neutrophils to cultured endothelium: time course and inhibition by a calcium channel blocker (nitrendipine)
Authors: Perry I. , Buttrum S.M. , Nash G.B.
Published: July 2012

Adhesion of neutrophils to vascular endothelium plays an important role in inflammation and thrombosis. Modulation of adhesion may be therapeutic in these conditions.

A flow model was used to quantify adhesion of neutrophils to human cultured umbilical vein endothelial cells. The time course of the neutrophil response to activation by N‐formyl‐methionyl‐leucyl‐phenylalanine (fMLP, 10−7 m) was studied and the inhibitory effects of the calcium‐channel blockers, nitrendipine and nifedipine, were investigated.

Neutrophils adhered firmly to the endothelial cells without rolling, but initial attachment was highly dependent on shear stress; doubling the stress from 0.05 to 0.1Pa decreased the number of neutrophils adhering by over 80%.

Adhesion rapidly increased after activation of neutrophils by fMLP, peaking at 1–3 min post‐treatment, and then decreased over the next 10–12 min. A monoclonal antibody to the β2‐integrin component CD18 inhibited adhesion by over 80% for activated or unactivated cells.

The Ca‐channel blocker, nitrendipine, but not nifedipine, significantly inhibited the fMLP‐induced increase of adhesion in a dose‐dependent manner (10−8 to 10−6 m). Dihydropyridines may be useful agents for modifying neutrophil function.
Mediation of the hypotensive action of systemic clonidine in the rat by α2‐adrenoceptors
Authors: Hieble J. Paul , Kolpak David C.
Published: July 2012

During the past few years it has been shown that the sympatholytic effect resulting from localized microinjection of clonidine and other imidazolines into the rostral ventrolateral medulla (RVL) results from activation of ‘imidazoline’ receptors (I1 receptors) rather than from an α2‐adrenoceptor‐mediated effect.

The relative contributions of these two receptor systems to the hypotensive action of systemically administered clonidine have not been studied. Clonidine has afffinity for both I1 and α2‐adrenoceptors; guanabenz represents a useful pharmacological tool, since it activates only the α2‐adrenoceptor.

Antagonists acting at both I1 and α2‐adrenoceptors (idazoxan) and at only α2‐adrenoceptors (SK&F 86466; 6‐chloro‐3‐methyl‐2,3,4,5‐tetrahydro‐3‐benzazepine) are available. Idazoxan (1 mg kg−1, i.v.) and SK&F 86466 (3 mg kg−1, i.v.) produced an equivalent degree of blockade of the pressor response to guanabenz or clonidine in the pithed rat, a response mediated by the α2‐adrenoceptor.

Guanabenz (30 μg kg−1, i.v.) and clonidine (10 μg kg−1, i.v.) lowered blood pressure in the chloralose‐anaesthetized spontaneously hypertensive rat by 48 ± 4.6 mmHg and 44 ± 5.4 mmHg, respectively; this response, for either agonist, was blocked by both idazoxan and SK&F 86466.

These data show that the hypotensive effect of intravenously administered clonidine results from activation of central α2‐adrenoceptors, with no significant contribution from an I1‐mediated effect. Thus clonidine can lower blood pressure by different receptor mechanisms, dependent on the route of administration.
Insulin‐mediated sensitization of adenylyl cyclase activation
Authors: Feldman Ross D.
Published: July 2012

Insulin may be an important regulator of vascular function. We have previously studied lymphocyte β‐adrenoceptors as a model for the human vascular β‐adrenoceptor. To examine the effects of insulin on human β‐adrenoceptor responsiveness, adenylyl cyclase activity, cyclic AMP‐dependent protein kinase activity and β‐adrenoceptor radioligand binding assays were performed on permeabilized mononuclear leukocytes.

With acute exposure to insulin in vitro, followed by washing and permeabilization there was a dose‐dependent increase in both lymphocyte NaF‐stimulated activity and β‐adrenoceptor‐stimulated adenylyl cyclase activity paralleling an increase in β‐adrenoceptor‐stimulated protein kinase A activity. Manganese‐, forskolin‐ and forskolin plus guanylimidodiphosphate‐stimulated adenylyl cyclase activities were not altered by insulin pretreatment. Additionally, mononulcear leukocyte β‐adrenoceptor density, proportion of externalized receptors and receptor affinity for agonist were not altered.

The data indicate that acute exposure to insulin sensitizes G‐protein‐stimulated adenylyl cyclase activity. These findings suggest a potential role for insulin in the regulation of β‐adrenoceptor responsiveness in man.
Tacrine‐induced increase in the release of spontaneous high quantal content events in Torpedo electric organ
Authors: Cantí Carles , Martí Eulàlia , Marsal Jordi , Solsona Carles
Published: July 2012

The anticholinesterases, tacrine (100 μm) and physostigmine (60 μm) had different effects on the amplitude distribution and kinetics of miniature endplate currents (m.e.p.cs) recorded extracellularly from the electric organ of Torpedo marmorata.

Tacrine increased the ratio of giant miniatures (larger than 4 mV of amplitude) to more than 20% of recorded spontaneous events. In the presence of physostigmine such events represented only 4%.

Both tacrine and physostigmine increased the rise time and the decay phase of normal‐sized m.e.p.cs when compared to control conditions. Both effects were significantly greater for tacrine.

We have tested the specificity of the tacrine effect on ectoenzyme activities associated with plasma membranes of these pure cholinergic nerve endings. Tacrine does not act unspecifically on every ectoenzyme, because it is not able to block the ectoapyrase activity even at a concentration 100 fold greater than that required to inhibit 94% of AChE.

We conclude that the differential effects of tacrine and physostigmine can be explained in terms of undetermined presynaptic actions of tacrine, while comparable effects of the two compounds can be explained through a shared anticholinesterase activity.
Chloride secretion in response to guanylin in colonic epithelia from normal and transgenic cystic fibrosis mice
Authors: Cuthbert A.W. , Hickman Margaret E. , MacVinish Lesley J. , Evans M.J. , Colledge W.H. , Ratcliff Rosemary , Seale P.W. , Humphrey P.P.A.
Published: July 2012

Guanylin, a 15 amino acid endogenous gut peptide, increased the short circuit current (SCC) in the epithelium of the mouse colon, but only when applied to the apical and not the basolateral surface.

By use of selective blockers of epithelial ion transport and modification of the bathing solution, it was concluded that guanylin increased electrogenic chloride secretion but also had a minor effect on electrogenic sodium absorption. In addition there were small residual currents which remained unresolved.

The threshold concentration of guanylin causing a SCC increase was less than 50 nm, but at concentrations 40 times greater no indication of a maximally effective concentration was found.

Two guanylin isomers with the same amino acid sequence but with the disulphide bridges joined in an alternate fashion showed no activity. Thus only guanylin with the greatest structural homology to heat stable enterotoxin (STa) showed biological activity.

The action of guanylin was virtually eliminated in colonic epithelia from transgenic cystic fibrosis (CF) mice. As these animals lack the chloride channel coded by the CF gene sequence, it is likely that the final effector process in murine colonic epithelia involves the CFTR (cystic fibrosis transmembrane conductance regulator) chloride channel.

Opportunistic infections of the gut generating STa lead to diarrhoeal conditions via an action of the toxin on apical guanylin receptors. Thus, as discussed, the CF heterozygote may have a genetic advantage in this circumstance.
β1‐ and β2‐adrenoceptor antagonist activities of ICI‐215001, a putative β3‐adrenoceptor agonist
Authors: Tesfamariam Belay , Allen Guy T.
Published: July 2012

The present study was undertaken to characterize the β3‐adrenoceptor agonist activity of ICI‐215001 and to determine whether it exhibits additional activities on β1‐ and β2‐adrenoceptors in isolated spontaneously beating atrium, trachea and ileum of guinea‐pig.

In guinea‐pig atrium, isoprenaline, a non‐selective β‐adrenoceptor agonist, caused concentration‐dependent, positive chronotropic effects that were inhibited by atenolol, a selective β1‐antagonist. ICI‐215001 also competitively antagonized the increase in heart rate caused by isoprenaline.

ICI‐215001 exhibited low intrinsic activity at increasing the beating rate of atrium and no activity on resting or induced tone of tracheal strips.

In strips of guinea‐pig trachea, contracted submaximally with carbachol, isoprenaline, caused concentration‐dependent relaxations. Both ICI‐118551, a selective β2‐adrenoceptor antagonist, and ICI‐215001 competitively inhibited the relaxations caused by isoprenaline.

In isolated strips of guinea‐pig ileum longitudinal smooth muscle contracted with histamine, isoprenaline and ICI‐215001 caused relaxations which were inhibited by alprenolol, a β‐adrenoceptor antagonist with modest affinity for β3‐adrenoceptors, but were resistant to ICI‐118551 and atenolol.

These results indicate that ICI‐215001 exhibits β3‐adrenoceptor agonist activity as demonstrated by relaxations mediated via atypical β‐adrenoceptors in the longitudinal smooth muscle of guinea‐pig ileum. Further, the studies demonstrate that ICI‐215001 can act as an antagonist at β1‐ and β2‐adrenoceptors in situations where its intrinsic agonist activity is low.
Ontogenic increase in PGE2 and PGF2α receptor density in brain microvessels of pigs
Authors: Li D.Y. , Varma D.R. , Chemtob S.
Published: July 2012

The hypothesis that the relative vasoconstrictor ineffectiveness of prostaglandin E2 (PGE2) and PGF2α on cerebral vessels of newborn pigs might be due to fewer receptors for these prostanoids was tested by comparing receptors for PGE2 (EP) and PGF2α (FP) in cerebral microvessels from newborn and adult pigs.

Specific binding of [3H]‐PGE2 and [3H]‐PGF2α to membranes prepared from brain microvessels showed that EP and FP receptor density (Bmax) in tissues from newborn animals was less than 50% of that determined in tissues from adults. By contrast, estimates of affinity (KD) were unchanged.

Specifically bound [3H]‐PGE2 to brain microvessels from both the newborn and adult was displaced by AH 6809 (EP1‐selective antagonist) by 80–90%, and only by approximately 30–35% by both 11‐deoxy PGE1 (EP2/EP3 agonist) and M&B 28,767 (EP3 agonist); butaprost (EP2 agonist) was completely ineffective.

PGE2, 17‐phenyl trinor PGE2 (EP1 agonist), PGF2α and fenprostalene (PGF2α analogue) caused significantly less increase in inositol 1,4,5‐triphosphate (IP3) in brain microvessels from the newborn than in those from adult pigs. The stimulation of IP3 by PGE2 and 17‐phenyl trinor PGE2 was almost completely inhibited by the EP1 antagonist, AH 6809.

PGE2, 11‐deoxy PGE1 and M&B 28,767 produced small reduction of adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) production in adult vessels but no effect in newborn tissues.

The lower density of EP and FP receptors in microvessels of newborn pigs compared to adults may explain the reduced ability of PGE2 and PGF2α to stimulate production of IP3 in tissues from newborn animals. This in turn, may provide an explanation for previous observations demonstrating that these prostanoids elicit contraction of adult cerebral microvessels, but exert minimal effects on these vessels in newborn animals.
Comparative analysis of the vagal stimulation of gastric acid secretion in rodent isolated stomach preparations
Authors: Welsh Nicola J. , Shankley Nigel P. , Black James W.
Published: July 2012

Electrical field stimulation produced a tetrodotoxin‐sensitive, frequency‐dependent, release of acid from isolated, lumen‐perfused, stomach preparations from mouse, immature rat and guinea‐pig.

In the guinea‐pig and mouse preparations, the frequency‐dependent response was abolished by hexamethonium, acetylcholine (ACh) muscarinic (M) and histamine H2‐receptor blockade, consistent with the hypothesis that the vagal ACh acts indirectly by stimulating the release of endogenous histamine.

In contrast, in the rat preparation the frequency‐dependent response was partially refractory to all of these inhibitors. However, a combination of H2‐ and ACh M‐receptor blockade did abolish the effect.

We conclude that vagal‐stimulated acid secretion in the rat, unlike the other two species, behaves as though there is a direct innervation of the oxyntic cells by either cholinergic or noncholinergic neurones.
Evoked noradrenaline release in the rabbit ear artery: enhancement by purines, attenuation by neuropeptide Y and lack of effect of calcitonin gene‐related peptide
Authors: Maynard K.I. , Burnstock G.
Published: July 2012

Adenosine (30 μm) and its analogues 5′‐N‐ethylcarboxaminoadenosine (5 and 30 μm) and l‐phenylisopropyladenosine (5 and 30 μm), potentiated the evoked but not spontaneous release of tritiated noradrenaline in the rabbit central ear artery.

Prejunctional inhibition of the evoked but not spontaneous release of tritiated noradrenaline by 100 nm neuropeptide Y is greater at 2 min than at 10 min after superfusion of the peptide.

Calcitonin gene‐related peptide (2.63 to 263 nm) did not affect the evoked or spontaneous release of tritiated noradrenaline in this preparation.

These results are discussed in terms of prejunctional modulation of sympathetic transmission in the rabbit central ear artery.
Role of endopeptidase 3.4.24.16 in the catabolism of neurotensin, in vivo, in the vascularly perfused dog ileum
Authors: Barelli H. , Fox‐Threlkeld J.E.T. , Dive V. , Daniel E.E. , Vincent J.P. , Checler F.
Published: July 2012

The degradation of tritiated and unlabelled neurotensin (NT) following close intra‐arterial infusion of the peptides in ileal segments of anaesthetized dogs was examined.

Intact NT and its catabolites recovered in the venous effluents were purified by chromatography on Sep‐Pak columns followed by reverse‐phase h.p.l.c. and identified by their retention times or by radioimmunoassay.

The half‐life of neurotensin was estimated to be between 2 and 6 min. Four labelled catabolites, corresponding to free tyrosine, neurotensin (1–8), neurotensin (1–10) and neurotensin (1–11), were detected.

Neurotensin (1–11) was mainly generated by a phosphoramidon‐sensitive cleavage, probably elicited by endopeptidase 24–11.

Two endopeptidase 3.4.24.16 inhibitors, phosphodiepryl 03 and the dipeptide Pro‐Ile, dose‐dependently potentiated the recovery of intact neurotensin. Furthermore, both agents inhibited the formation of neurotensin (1–10), the product that results from the hydrolysis of neurotensin by purified endopeptidase 3.4.24.16. In contrast, the endopeptidase 3.4.24.15 inhibitor Cpp‐AAY‐pAB neither protected neurotensin from degradation nor modifed the production of neurotensin (1–10).

Our study is the first evidence to indicate that endopeptidase 3.4.24.16 contributes to the catabolism of neurotensin, in vivo, in the dog intestine.
Studies of the nucleoside transporter inhibitor, draflazine, in the human myocardium
Authors: Böhm Michael , Weinhold Christian , Schwinger Robert H.G. , Müller‐Ehmsen Jochen , Böhm Dieter , Reichenspurner Hermann , Reichart Bruno , Erdmann Erland
Published: July 2012

The aim of the present study was to determine the effect of the nucleoside transporter inhibitor, draflazine, on the force of contraction in human myocardium and the affinity of the compound for the nucleoside transporter. Nucleoside transport inhibitors, like draflazine, are of potential importance for cardiopreservation of donor hearts for heart transplantation.

Functional experiments were performed in isolated electrically driven (1 Hz, 1.8 mmol l−1 Ca2+) human atrial trabeculae and ventricular papillary muscle strips. The affinity of draflazine for the myocardial nucleoside transporter was studied in isolated membranes from human ventricular myocardium and human erythrocytes in radioligand binding experiments using [3H]‐nitrobenzylthioinosine ([3H]‐NBTI). Dipyridamole was studied for comparison.

In membranes from human myocardium and erythrocytes, [3H]‐NTBI labelled 1.18 pmol mg−1 protein and 23.0 pmol mg−1 protein, respectively, nucleoside transporter molecules with a KD value of 0.8 nmol l−1. Draflazine concentration‐dependently inhibited binding of [3H]‐NBTI to myocardial and erythrocyte membranes with a Ki‐value of 4.5 nmol l−1. The potency as judged from the Ki values was ten times greater than that of dipyridamole in both myocardial and erythrocyte membranes.

Draflazine, at concentrations up to 100 μmol l−1, did not produce negative inotropic effects in atrial and ventricular myocardium.(−)‐N6‐phenylisopropyladenosine (R‐PIA) and carbachol did not reduce force of contraction in ventricular myocardium, but exerted concentration‐dependent direct negative inotropic effects in atrial myocardium.

The data provide evidence that draflazine specifically binds to the nucleoside transporter of the human heart and erythrocytes with high affinity. The compound does not produce negative inotropic effects at concentrations as high as 100 μmol l−1.

Draflazine could be a useful agent for cardiopreservation because it does not produce cardiodepressant effects. Thus, it may be possible to perfuse explanted hearts directly with this agent without the hazard of cardiodepression.
The renal functional responses to 5‐HT1A receptor agonist, flesinoxan, in anaesthetized, normotensive rat
Authors: Chamienia Andrzej L. , Johns Edward J.
Published: July 2012

The present study was designed to examine the effects of a centrally acting 5‐HT1A receptor agonist, flesinoxan, on the cardiovascular system and renal haemodynamics and excretory function.

In chloralose‐urethane anaesthetized Wistar rats, i.v. administration of bolus doses of flesinoxan, at 30, 100, 300 and 1000 μg kg−1, caused significant, dose‐dependent decreases in mean arterial pressure, of 33 ± 2 mmHg (P < 0.001) and heart rate of 57 ± 9 beats min−1 (P < 0.001) at the highest dose used. Despite this substantial fall in perfusion pressure there were no meaningful changes in the renal excretion of water and sodium. In a second group of rats, reduction of renal perfusion pressure mechanically to the same values as observed in rats given flesinoxan (i.e. 100, 92, 84 and 76 mmHg) produced reductions in urine flow, absolute and fractional sodium excretions reaching a maximum of 74, 86 and 84% respectively (all P < 0.001) at the lowest pressure. These reductions were significantly larger than those seen in the previous group of animals.

In the group of rats subjected to renal denervation, flesinoxan produced changes in blood pressure and heart rate which were not different from those observed in intact animals. However, the reduction in pressure was accompanied by significant decreases in urine flow of 71%, absolute sodium excretion of 68% and fractional sodium excretion of 67% (all P < 0.001) at the highest dose, which were all significantly greater than the changes seen in the innervated animals but were not different from those observed when renal perfusion pressure was reduced mechanically.

The findings of this investigation showed that flesinoxan was effective in lowering blood pressure and heart rate in the anaesthetized rat, which was probably due to decreased sympathetic nerve activity. Renal excretion of water and sodium was well preserved in the face of the flesinoxan‐induced hypotension. The maintenance of fluid excretion with flesinoxan appeared to be mediated via changes in renal nerve activity, since it did not occur when the kidney was denervated.
Modulatory effects of NMDA on phosphoinositide responses evoked by the metabotropic glutamate receptor agonist 1S,3R‐ACPD in neonatal rat cerebral cortex
Authors: Challiss R.A. John , Mistry Rajendra , Gray David W. , Nahorski Stefan R.
Published: July 2012

The effect of NMDA‐receptor stimulation on phosphoinositide signalling in response to the metabotropic glutamate receptor agonist 1‐aminocyclopentane‐1S,3R‐dicarboxylic acid (1S,3R‐ACPD) has been examined in neonatal rat cerebral cortex slices.

Total [3H]‐inositol phosphate ([3H]‐InsPx) accumulation, in the presence of 5 mm LiCl, in [3H]‐inositol pre‐labelled slices was concentration‐dependently increased by 1S,3R‐ACPD (EC50 16.6 μm) and, at a maximally effective concentration, 1S,3R‐ACPD (300 μm) increased [3H]‐InsPx accumulation by 12.8 fold over basal values.

[3H]‐InsPx accumulation stimulated by 1S,1R‐ACPD was enhanced by low concentrations of NMDA (3–30 μm), but not by higher concentrations (> 30 μm). [3H]‐InsPx accumulations stimulated by 1S,3R‐ACPD in the absence or presence of 10 μm NMDA were linear with time, at least over the 15 min period examined; however, in the presence of 100 μm NMDA the initial enhancement of 1S,3R‐ACPD‐stimulated phosphoinositide hydrolysis progressively decreased with time.

In the presence of a maximal enhancing concentration of NMDA (10 μm), the response to 1S,3R‐ACPD (300 μm) was increased 1.9 fold and the EC50 for agonist‐stimulated [3H]‐InsPx accumulation decreased about 4 fold. The enhanced response to the metabotropic agonist was concentration‐dependently inhibited by competitive and uncompetitive antagonists of NMDA‐receptor activation.

1S,3R‐ACPD also stimulated inositol 1,4,5‐trisphosphate (Ins(1,4,5)P3) mass accumulation with an initial peak response (5–6 fold over basal) at 15 s decaying to a smaller (2 fold), but persistent elevated accumulation (1–10 min).

Co‐addition of 10 or 100 μm NMDA enhanced the initial peak Ins(1,4,5)P3 response to 1S,3R‐ACPD. However, the enhancing effect was only maintained over 10 min in the presence of 10 μm NMDA, whilst in contrast, 100 μm NMDA ceased to cause a significant enhancement of the metabotropic response by 5 min and completely suppressed 1S,3R‐ACPD‐stimulated Ins(1,4,5)P3 accumulation at 10 min.

Both basal and 1S,3R‐ACPD‐stimulated Ins(1,4,5)P3 accumulations were reduced when slices were incubated in nominally Ca2+‐free medium. Under these conditions only a concentration‐dependent enhancement of the response was observed (EC50 for NMDA facilitation of 1S,3R‐ACPD‐stimulated Ins(1,4,5)P3 accumulation of 32 μm).

These experiments have revealed that at low concentrations, NMDA can dramatically potentiate 1S,3R‐ACPD‐stimulated phosphoinositide hydrolysis, probably by a Ca2+‐dependent facilitation of agonist‐stimulated phosphoinositide‐specific phospholipase C activity. Higher concentrations of NMDA result in time‐dependent inhibition of the metabotropic agonist‐stimulated response. We believe the former effect could be fundamental in glutamate receptor ‘cross‐talk’, whereas the latter may reflect a Ca2+‐dependent neurotoxic effect of NMDA on the neonatal cerebral cortex slices.
Cardiovascular and behavioural effects of centrally administered tachykinins in the rat: characterization of receptors with selective antagonists
Authors: Picard Pierre , Regoli Domenico , Couture Réjean
Published: July 2012

The effects of intracerebroventricular (i.c.v.) injection of selective and potent NK1 (RP 67580), NK2 (SR 48968) and NK3 (R 486, [Trp7, β‐Ala8]NKA(4–10)) receptor antagonists were assessed on the cardiovascular and behavioural responses elicited by the i.c.v. injection of substance P (SP), neurokinin A (NKA) or [MePhe7]neurokinin B ([MePhe7]NKB) in the conscious freely moving rat.

SP, NKA and [MePhe7]NKB (5–650 pmol) evoked dose‐dependent increases in mean arterial blood pressure (MAP) and heart rate (HR) with the rank order of potency SP > NKA > [MePhe7]NKB. The cardiovascular responses were accompanied by excessive face washing, grooming and wet dog shakes.

The cardiovascular effects and face washing behaviour induced by SP (25 pmol) were significantly reduced by the pre‐injection (i.c.v., 5 min earlier) of RP 67580 (6.5 nmol). However, this antagonist failed to affect the central effects of 25 pmol NKA or [MePhe7]NKB.

The cardiovascular and behavioural responses (except for wet dog shakes) elicited by NKA (25 pmol) were significantly reduced by 6.5 nmol SR 48968. However, the latter antagonist had no effect on the SP or [MePhe7]NKB‐mediated responses.

Both cardiovascular and behavioural effects produced by either SP or NKA (25 pmol) were completely abolished when rats were pretreated with a combination of RP 67580 (6.5 nmol) and SR 48968 (6.5 nmol), yet this combination of antagonists failed to modify the central effects of [MePhe7]NKB.

R 486 (6.5 nmol) inhibited the cardiovascular effects as well as wet dog shakes produced by [MePhe7]NKB, but it was inactive against the responses induced by either SP or NKA.

None of the tachykinin receptor antagonists or agonists caused motor impairment or respiratory distress. All antagonists blocked in a reversible manner and were devoid of intrinsic activity except R 486 (6.5 nmol) which produced a transient increase of MAP and HR.

These results suggest that the central effects of SP, NKA and [MePhe7]NKB are primarily mediated by central NK1, NK2 and NK3 receptors, respectively. However, a minor activation of NK2 receptors by SP and NK1 receptors by NKA was seen during blockade of both receptors. This study therefore supports the existence of functional NK1, NK2 and NK3 receptors in the adult rat brain.
Pharmacological relevance of peripheral type benzodiazepine receptors on motor nerve and skeletal muscle
Authors: Chiou L.C. , Chang C.C.
Published: July 2012

Effects of agonists and antagonists of peripheral and central benzodiazepine receptors (pBZR and cBZR) on neuromuscular transmission were studied in mouse isolated phrenic nerve‐diaphragm preparations.

Ro5–4864, a pBZR agonist, had no effect on the neuromuscular transmission but increased muscle contractility and antagonized the tetanic fade induced by neostigmine.

Ro5–4864 inhibited the regenerative tonic endplate depolarization caused by repetitive stimulation in the presence of neostigmine without affecting the amplitude and decay time of miniature and evoked single endplate potentials.

All the effects of Ro5–4864 were shared by PK11195, a pBZR antagonist, but not by clonazepam and flumazenil, a cBZR agonist and antagonist, respectively.

It is suggested that peripheral type benzodiazepine receptors modulate presynaptic function and muscle contraction.