bluuebunny/crossref_metadata_2025 huggingface dataset:
Inflammatory mechanisms in the passive cutaneous anaphylactic reaction in the rabbit: evidence that novel mediators are involved
Authors: Hellewell Paul G. , Jose Peter J. , Williams Timothy J.
Published: July 2012

We have examined the mechanisms of local oedema formation in the passive cutaneous anaphylactic (PCA) reaction in the rabbit.

IgE‐containing antiserum was injected i.d. and allowed to sensitize skin sites for periods up to 240 h. Antigen (bovine gamma globulin) was injected i.d. or i.v. and local oedema formation assessed by the accumulation of i.v. injected 125I‐labelled rabbit serum albumin. Potential inhibitors were mixed with antigen prior to i.d. injection or were administered i.v.

Maximum oedema formation was observed when a sensitization period of 48–72 h was used. Oedema formation in the PCA reaction was of short duration with a t1/2 of approximately 15 min. No evidence of late oedema formation (up to 6 h) was found.

Local oedema formation in the PCA was reduced by indomethacin suggesting that vasodilator, oedema‐potentiating prostaglandins were released. However, it was likely that other vasodilators were also generated.

Antihistamines were poor inhibitors of oedema formation as were PAF antagonists, a 5‐lipoxygenase inhibitor, a kallikrein inhibitor, a bradykinin antagonist and anti‐C5a antibody.

Local oedema formation in the PCA was partially reduced by neutrophil depletion and colchicine suggesting that neutrophil‐dependent mediators were involved.

Exudate fluid from anaphylactic reactions in the rabbit peritoneal cavity contained permeability‐increasing activity when injected into rabbit skin. This activity is now being characterized.

A vasodilator prostaglandin appears to be released in the rabbit PCA reaction but none of the established permeability‐increasing mediators appears to be involved. Thus, there may be novel inflammatory mediators generated in this reaction which may have relevance for human allergic skin diseases.
Effect of murine recombinant interleukin‐5 on the cell population in guinea‐pig airways
Authors: Iwama T. , Nagai H. , Suda H. , Tsuruoka N. , Koda A.
Published: July 2012

An intratracheal injection of murine recombinant interleukin 5 (mrIL‐5, 2–15 μg/0.25 ml/animal) induced a dose‐dependent increase in the number of macrophages, eosinophils, neutrophils and epithelial cells in the bronchoalveolar lavage fluid (BALF) of guinea‐pigs 24 h after administration. Bovine serum albumin (15 μg/0.25 ml/animal), used as a reference material, did not cause any change of this type.

The intratracheal administration of mrIL‐5 at a dose of 15 μg showed a tendency to increase the number of these pulmonary inflammatory cells and epithelial cells in the BALF at 12 h with a significant increase observed at 24 h.

Prednisolone (20 mg kg−1, i.p.) inhibited the mrIL‐5‐induced increase in macrophages, eosinophils, neutrophils and epithelial cells. Ketotifen (2 mg kg−1, i.p.) reduced the mrIL‐5‐induced increase in the eosinophil, neutrophil and epithelial cell populations. The simultaneous injection of 2% disodium cromoglycate (DSCG) into the trachea prevented the mrIL‐5‐induced increase in the number of airway epithelial cells, without affecting changes in the other inflammatory leukocytes.

These results suggest that mrIL‐5 is a potent inducer of lung inflammation, in terms of increased inflammatory leukocytes and epithelial cells in guinea‐pig BALF. Prednisolone, DSCG and ketotifen are effective against mrIL‐5‐induced pulmonary inflammation, especially the desquamation of bronchial epithelial cells.
Blockade by local anaesthetics of the single Ca2+‐activated K+ channel in rat hippocampal neurones
Authors: Oda Masao , Yoshida Atsuya , Ikemoto Yoshimi
Published: July 2012

Effects of local anaesthetics on single Ca2+‐activated K+ channels were investigated using the inside‐out configuration of the patch‐clamp technique in single pyramidal neurones, which were freshly dissociated from rat hippocampus by use of proteolytic enzymes.

No significant effect was observed when 2 mm benzocaine was applied on either side of the membrane patch, or when 3 mm lignocaine or QX‐314 was applied to the external surface of the membrane.

Lignocaine 1 mm, applied to the internal surface, slightly reduced the amplitude of the single K+ channel current. When applied to the internal surface, QX‐314 reduced the amplitude of the K+ channel current, accompanied by an increase in noise in the open channel current, suggesting a fast flickering block. The blocking effect of QX‐314 on the outward current increased with depolarization, suggesting a binding site for the drug at an electrical distance of about 0.5 across the membrane field.

The open time histogram showed one exponential component and the closed time histogram showed at least two components. The mean open time of the outward current was increased when the amplitude was reduced by the drugs.

The ionized form of the local anaesthetics had a similar action on the Ca2+‐activated K+ channels to that on Na+ channels, that is, they enter into the channel from the cytoplasmic side to induce open channel block. The blocking kinetics, however, might be so fast that they were beyond the frequency response of our recording apparatus, thus the recorded current amplitude was decreased. In contrast the K+ channel was not accessible via hydrophobic pathways for the neutral form, which is also known to block the sodium channel.
Role of muscarinic receptor subtypes in central antinociception
Authors: Bartolini A. , Ghelardini C. , Fantetti L. , Malcangio M. , Malmberg‐Aiello P. , Giotti A.
Published: July 2012

The ability to modify the pain threshold by the two M1‐muscarinic agonists: McN‐A‐343 and AF‐102B and by the specific M2‐agonist arecaidine was examined in mice and rats by using three different noxious stimuli: chemical (writhing test), thermic (hot‐plate test) and mechanical (paw pressure test).

In the mouse hot‐plate test McN‐A‐343 (20–50 μg per mouse i.c.v.) and AF‐102B (1–10 mg kg−1 i.p.) produced significant antinociception which was prevented by atropine (1 μg per mouse i.c.v.) and by the two selective M1 antagonists: pirenzepine (0.01 μg per mouse i.c.v.) and dicyclomine (0.08 μg per mouse i.c.v. or 10 mg kg−1 i.p.) but not by the specific M2‐antagonist AFDX‐116 (0.1 μg per mouse i.c.v.), naloxone (1 mg kg−1 i.p.) or by the acetylcholine (ACh) depletor hemicholinium‐3 (HC‐3) (1 μg per mouse i.c.v.). McN‐A‐343 and AF‐102B were able to increase the pain threshold also in the mouse acetic acid writhing test and in rat paw pressure test. These antinociceptive effects were completely prevented by dicyclomine (0.08 μg per mouse i.c.v. or 10 mg kg−1 i.p.) but not by AFDX‐116 (0.1 μg per mouse or rat i.c.v.).

In contrast with the M1‐agonists, the M2‐agonist arecaidine (0.1–2 μg per mouse or rat i.c.v.) did not induce antinociception in all three analgesic tests. However, arecaidine, at the same i.c.v. doses, was able to reduce the pain threshold in the hot‐plate and paw pressure tests.

The site of muscarinic control of the pain threshold is localized in the CNS since drugs which do not cross the blood‐brain barrier such as McN‐A‐343, pirenzepine and arecaidine exerted their effects only if injected i.c.v.

On the basis of the above findings and existing literature we suggest that the postsynaptic muscarinic receptors involved in antinociception belong to the M1 subtype. Nevertheless, presynaptic autoreceptors (M2 subtype) may play a role in pain regulation since they are involved in modulation of endogenous ACh release.
Role of the adrenal medulla in the metabolic and pressor actions of 8‐OH‐DPAT
Authors: Bouhelal Rochdi , Mir Anis K.
Published: July 2012

Intravenous administration of 8‐hydroxy‐2‐(di‐n‐propylamino)tetralin (8‐OH‐DPAT, 150 μg kg−1) into conscious sham‐operated rats caused significant increases in basal glycaemia with minor changes in basal insulinaemia. Glucose‐stimulated (intravenous glucose tolerance test) plasma insulin levels were significantly inhibited in 8‐OH‐DPAT‐treated sham‐operated animals. These metabolic changes were associated with significant and sustained falls in blood pressure (BP) and heart rate (HR) preceded by transient (< 5 min) increases only in BP.

In adrenodemedullated animals, 8‐OH‐DPAT failed to cause an initial vasoconstriction, hyperglycaemia, or inhibition of glucose‐stimulated plasma insulin despite eliciting falls in BP and HR that were comparable to those observed in sham‐operated animals.

Noradrenaline, adrenaline and dopamine levels in the adrenal tissue were reduced by about 95% in adrenodemedullated rats as compared to sham‐operated rats. A functionally intact adrenal cortex was indicated by the presence of corticosterone in the plasma of both adrenodemedullated and sham‐operated rats.

The present data demonstrate that 8‐OH‐DPAT mediates an initial increase in BP and changes in metabolic parameters via intact adrenal medulla and may thus be consequential to the release of adrenaline, whereas the sustained cardiovascular effects of 8‐OH‐DPAT are not.
Oxidative modification of low‐density lipoproteins and the inhibition of relaxations mediated by endothelium‐derived nitric oxide in rabbit aorta
Authors: Plane Frances , Bruckdorfer K. Richard , Kerr Paul , Steuer Alan , Jacobs Michael
Published: July 2012

The mechanism by which Cu2+‐oxidized low‐density lipoproteins (oxLDL) inhibit acetylcholine (ACh)‐evoked relaxations mediated by endothelium‐derived nitric oxide (EDRF) in rabbit aortic rings was investigated. The proposed role of lysophosphatidylcholine (LPC) in the inhibition was also studied.

The kinetics of lipid peroxidation of native low‐density lipoproteins (LDL) from individual donors, as measured by changes in conjugated diene concentration, were related to the inhibitory effects of the resultant oxLDL. It was found that the more susceptible LDL was to oxidation, the greater the inhibition.

No correlation was found between the inhibitory effects of oxLDL and LPC content.

Synthetic 1‐palmitoyl LPC produced an inhibition of ACh‐induced relaxations and when added to precontracted rings evoked nitric oxide‐mediated endothelium‐dependent relaxation. This latter effect was not elicited by oxLDL.

Synthetic 1‐palmitoyl (10 μm) had no effect on relaxations evoked by glyceryl trinitrate in endothelium‐denuded aortic rings in contrast to the inhibition found previously for oxLDL.

Concentrations of oxLDL and phospholipase A2‐treated LDL which inhibited relaxation contained very different LPC concentrations. Unlike oxLDL, the inhibitory effects of phospholipase A2‐treated LDL preparations were independent of the donors and showed no lag period.

We sugggest that there are differences in the mechanisms by which oxLDL and 1‐palmitoyl LPC exert their inhibitory effects on relaxation.

The inhibition of relaxation by oxLDL (1–2 mg protein ml−1) was prevented by the presence of high‐density lipoproteins (HDL; 1–2 mg protein ml−1).

It is proposed that prevention of the inhibition of relaxation by HDL is consistent with the inhibitory factor(s) being lipophilic constituents of oxLDL. However, variations in the inhibitory effects of oxLDL preparations are not due to differences in their LPC content and factors other than LPC must contribute to the inhibition.
Response to atrial natriuretic peptide, endopeptidase 24.11 inhibitor and C‐ANP receptor ligand in the rat
Authors: Wilkins M.R. , Settle S.L. , Kirk J.E. , Taylor S.A. , Moore K.P. , Unwin R.J.
Published: July 2012

The present studies compared the renal and hypotensive response to (a) exogenous atrial natriuretic peptide (ANP) (99–126), (b) an endopeptidase‐24.11 inhibitor (candoxatrilat) and (c) an antagonist of ANP clearance receptors (SC 46542) in conscious rats.

Infusion of low‐dose‐ANP (100 ng kg−1 min−1) produced a gradual increase in urinary sodium and guanosine 3′:5′‐cyclic monophosphate (cyclic GMP) excretion without significant change in glomerular filtration rate (GFR) or fractional lithium clearance (FeLi). There was a significant fall in blood pressure.

Infusion of high‐dose ANP (300 ng kg−1 min−1) produced a brisk, 3 fold increase in urinary sodium and cyclic GMP excretion along with a rise in GFR, but had no significant effect on FeLi compared to the control group. The renal response was accompanied by a pronounced fall in blood pressure.

Candoxatrilat or SC 46542, alone, had no significant effect on sodium excretion compared to control animals. Both compounds enhanced the natriuretic and cyclic GMP responses to a low‐dose ANP infusion (100 ng kg−1 min−1) to levels similar to, or greater than, those observed with the high‐dose ANP (300 ng kg−1 min−1). However, unlike high‐dose ANP, these renal effects were not accompanied by a significant change in GFR and neither compound potentiated the hypotensive effect of the low‐dose ANP infusion. Only candoxatrilat when given with ANP produced a marked rise in FeLi.

Similarly, combined administration of candoxatrilat and SC 46542 (without exogenous ANP) induced an increase in sodium and cyclic GMP excretion comparable to high‐dose ANP but did so without a significant increase in GFR and with a significantly smaller fall in blood pressure. Interestingly, there was no increase in FeLi with the combination of the two compounds, suggesting that the major contribution to sodium excretion came from SC 46542.

6 Both candoxatrilat and SC 46542 increased sodium and cyclic GMP excretion in the rat A‐V fistula model of heart failure, a model hyporesponsive to infusions of ANP, without significant change in blood pressure.

7 These data show that candoxatrilat and SC 46542 do not simply reproduce the effects of an ANP infusion but preferentially enhance the natriuretic response to ANP. Inhibition of E‐24.11 may potentiate a tubule action of ANP while the renal mechanism of action of the C‐ANP receptor ligand needs further study. Both manipulations are of potential value in the management of heart failure.
MDL 26,479: a potential cognition enhancer with benzodiazepine inverse agonist‐like properties
Authors: Miller Jerry A. , Dudley Mark W. , Kehne John H. , Sorensen Stephen M. , Kane John M.
Published: July 2012

The present study investigated biochemical, electrophysiological and behavioural properties of the novel cognition enhancer, MDL 26,479 (5‐(3‐fluorophenyl)‐2,4,‐dimethyl‐3H‐1,2,4‐triazole‐3‐thione).

The 5‐aryl‐1,2,4‐triazole, MDL 26,479, potently (0.22 ± 0.05 mg kg−1) inhibited [3H]‐flumazenil (Ro15–1788) binding in mouse cortex but was ineffective in vitro at displacing radioligand binding to the GABAA receptor complex.

Parenteral administration of MDL 26,479 (1 mg kg−1) or the benzodiazepine (BZD) inverse agonist methyl 6,7‐dimethoxy‐4‐ethyl‐β‐carboline‐3‐carboxylate (DMCM) (0.3 mg kg−1) increased cortical ex vivo binding of [3H]‐hemicholinium‐3 ([3H]‐HC‐3), a marker for cholinergic activation. This effect of MDL 26,479 was blocked by pretreatment with the antagonist flumazenil (1 mg kg−1).

MDL 26,479 (20 μm) and DMCM (1 μm) increased excitation in the hippocampal long‐term potentiation (LTP) slice preparation; however, unlike DMCM, the effect of MDL 26,479 was not blocked by flumazenil.

In behavioural studies, MDL 26,479 did not exhibit adverse properties characteristic of drugs associated with the GABAA receptor complex. It lacked convulsant, anxiogenic, anxiolytic, or depressant effects. Since MDL 26,479 lacks activity with the BZD receptor in vitro we suggest that it acts via the GABAA receptor complex at another site on this receptor or in an as yet undefined manner or an active metabolite is formed in vivo.

6 Previous work showed that MDL 26,479 enhances learning acquisition in animal models. The present study suggests that at least some of the cognition enhancing properties are due to the enhancement of cortical and hippocampal cholinergic function and LTP.
Suramin inhibits excitatory junction potentials in guinea‐pig isolated vas deferens
Authors: Sneddon Peter
Published: July 2012

. Intracellular microelectrode recording techniques were used to investigate the action of the putative P2‐purinoceptor antagonist, suramin, on sympathetic neurotransmission in the guinea‐pig isolated vas deferens.

. The resting membrane potential of the control cells was 67.4 ± 0.7 mV (n = 48). Field stimulation of the sympathetic nerves innervating the vas deferens produced excitatory junction potentials (e.j.ps) which reached a mean magnitude of 8.5 ± 0.8 mV (n = 23) when fully facilitated at a stimulation frequency of 0.5 Hz.

. Introduction of suramin 1–100 μm produced no change in the resting membrane potential of the smooth muscle cells, but gradually reduced e.j.p. magnitude. Suramin, 20 μm, reduced the mean magnitude of the fully facilitated e.j.ps to 1.4 ± 0.3mV (n = 18).

. After suramin‐induced inhibition of e.j.ps, nerve stimulation at 1–8 Hz resulted in summation of e.j.ps to a subthreshold level. Subsequent introduction of the α‐adrenoceptor antagonists, prazosin or phentolamine (1 μm) did not reduce the magnitude of the summated e.j.ps.

. The results support the proposal that e.j.ps in vas deferens are mediated by adenosine 5′‐triphosphate, and not by noradrenaline, and confirm that suramin can antagonize responses mediated via P2‐purinoceptors.
H3 receptor antagonist, thioperamide, inhibits adrenal steroidogenesis and histamine binding to adrenocortical microsomes and binds to cytochrome P450
Authors: LaBella F.S. , Queen G. , Glavin G. , Durant G. , Stein D. , Brandes L.J.
Published: July 2012

Thioperamide (TP), an imidazole and a highly potent, specific antagonist of the histamine H3 receptor, inhibited the secretion of cortisol from bovine isolated adrenocortical cells (IC50 0.20 μm) and, in the rat (5 mg kg−1) prevented both basal and stress‐induced secretion of corticosterone.

In adrenocortical microsomes, low affinity binding of [3H]‐histamine (KD 27.7 μm) was potently inhibited by TP (Ki 0.33 μm).

In adrenocortical microsomal membranes, both histamine and TP yielded type II difference absorption spectra, characteristic of the interaction between imidazole and cytochrome P450 enzymes. Dissociation constants for binding to P450, calculated from spectral data, were 15.9 μm and 1.5 mm for histamine, and 0.3 μm and 3.7 μm for TP.

In view of previously reported evidence for an intracellular mediator role of histamine in platelets, the present findings suggest a physiological role for histamine in the modulation of adrenal P450 monooxygenases that generate adrenocortical steroids.

The results suggest that direct adrenocortical inhibition by thioperamide at a non‐H3 intracellular site must be taken into account in studies designed to elucidate functional roles of H3 receptors.
Cardiac α1‐adrenoceptor densities in different mammalian species
Authors: Steinfath Markus , Chen Yi‐Yue , Lavický Jan , Magnussen Olaf , Nose Monika , Rosswag Sebastian , Schmitz Wilhelm , Scholz Hasso
Published: July 2012

α1‐Adrenoceptor densities were studied in cardiac membrane preparations from several mammalian species including human failing hearts under identical experiment conditions; the α1‐adrenoceptor antagonist, [3H]‐prazosin, was used as radioligand. End‐stage heart failure (NYHA IV) in human hearts was due to idiopathic dilated cardiomyopathy.

The ventricular α1‐adrenoceptor densities were not significantly different in guinea‐pig, mouse, pig, calf, and man (11 to 18 fmol mg−1 protein) but about 5 to 8 fold smaller than in rat (about 90 fmol mg−1 protein). Right and left ventricular receptor densities were similar in these species.

A sufficient amount of right and left atrial tissue was obtained from rabbit, pig, calf, and man only. The α1‐adrenoceptor densities in both atria of these species were found to be at the detection limit of the method used (less than 8 fmol mg−1 protein).

The equilibrium dissociation constant (KD) was similar in all species studied ranging from 0.047 ± 0.006 to 0.063 ± 0.007 nmol l−1.

It is concluded that differences in α1‐adrenoceptor density between atria and ventricles may exist in mammalian species. The exceptionally high density of these receptors in rat ventricles seem to be a particular feature in these animals.
The α2‐adrenoceptors of the human retinoblastoma cell line (Y79) may represent an additional example of the α2C‐adrenoceptor
Authors: Gleason Marie M. , Hieble J. Paul
Published: July 2012

In agreement with the literature, correlation of the ability of a series of agonists and antagonists to displace [3H]‐rauwolscine binding shows the α2‐adrenoceptors of HT29 cells, NG108–15 cells, OK cells and homogenates of rat sublingual gland to represent four distinct subtypes.

[3H]‐rauwolscine also bound with high affinity (KD = 0.30 ± 0.10 mm) to a human retinoblastoma cell line (Y79). Specific binding represents 73% of total binding, and a Bmax of 38 ± 1 fmol mg−1 protein was determined.

Correlation of antagonist affinities against [3H]‐rauwolscine with corresponding values in the other four tissue sources showed the Y79 cells to resemble most closely the OK cells, the prototype example of an α2C‐adrenoceptor, with a correlation coefficient of 0.90 and a regression slope of 1.01 being obtained for 10 antagonists in these two systems.

Comparison of KD values for [3H]‐rauwolscine also showed a similarity between the OK cells (0.19 ± 0.07 nm) and Y79 cells.

These data suggest that the human retinoblastoma cell line may represent an additional example of the α2C‐adrenoceptor subtype.
Attenuation of tachykinin‐induced airflow obstruction and microvascular leakage in immature airways
Authors: Tokuyama Kenichi , Yokoyama Tatsuya , Morikawa Akihiro , Mochizuki Hiroyuki , Kuroume Takayoshi , Barnes Peter J.
Published: July 2012

To study the effect of maturation on substance P (SP)‐ and neurokinin A (NKA)‐induced airflow obstruction and airway microvascular leakage (MVL), we have measured changes in both lung resistance (RL) and extravasation of Evans blue dye in anaesthetized immature (aged 14 ± 1 days) and adult guinea‐pigs (aged 80 ± 3 days).

RL and its recovery after hyperinflation at 5 min were measured for 6 min after i.v. SP (0.2, 1 and 30 nmol kg−1), NKA (1 and 10 nmol kg−1) or vehicle (0.9% NaCl). After measurement of RL, MVL in trachea, main bronchi and intrapulmonary airways was also examined.

The order of potency in inducing airflow obstruction did not change with age (NKA> SP) but immature animals required a larger dose of SP or NKA than adults to cause a significant increase in RL.

The order of potency in inducing airway microvascular leakage was sp> NKA in both immature and adult animals. The amount of extravasated dye after SP was significantly less in immature airways, especially in central airways.

Phosphoramidon (2.5 mg kg−1), a neutral endopeptidase (NEP) inhibitor, significantly increased RL after 0.2 nmol kg−1 SP only in adult airways. Phosphoramidon enhanced the dye extravasation after 0.2 nmol kg−1 SP in both immature and adult airways with a significantly greater amount of dye in adult animals, suggesting that mechanisms other than changes in NEP activity may be responsible for this age‐related difference.

RL after hyperinflation following SP was not correlated with the degree of extravasation of Evans blue dye in immature animals, whereas it was closely correlated in adult animals.

SP and NKA may be less potent in causing both bronchoconstriction and microvascular leakage in immature airways.

Airway oedema caused by microvascular leakage may contribute less in immature airways to airflow obstruction after SP or NKA.
Dependence of endotoxin‐induced vascular hyporeactivity on extracellular l‐arginine
Authors: Schott Cathy A. , Gray Gillian A. , Stoclet Jean‐Claude
Published: July 2012

The dependence on extracellular l‐arginine of vascular hyporeactivity induced by bacterial lipopolysaccharide (LPS) was studied in vivo in rats infused with LPS and in vitro in endothelium‐denuded rat thoracic aortic rings exposed to LPS.

Infusion of LPS during 50 min at a dose of 10 mg kg−1 h−1 produced a significant impairment of the pressor effect of noradrenaline, while in tissues collected 60 min after the start of LPS infusion, no significant alteration in either plasma arginine concentration or aortic arginine content was found compared to saline‐infused controls (where plasma arginine was 78.5 ± 7 μm and aortic arginine 394 ± 124 nmol g−1 tissue).

Incubation of isolated, endothelium‐denuded aortic rings with LPS (10 μg ml−1) in the absence of l‐arginine for 4 h at 37°C produced a 6 fold (P < 0.01) rightward shift in the noradrenaline concentration‐effect curve compared to polymyxin B (1 μg ml−1, a LPS neutralizing agent) and reduced by 15% the maximum observed tension.

The presence of l‐arginine (100 μm) during the incubation with LPS and throughout the following contraction experiments caused a 15 fold (P < 0.01) increase in the EC50 of noradrenaline and greater depression (45%) of the maximum observed tension compared to polymyxin B‐treated controls. Responses in control, non LPS‐treated rings were unaffected by the presence of l‐arginine.

The addition of l‐arginine to rings incubated with LPS in the absence of l‐arginine and maximally precontracted with noradrenaline (10 μm) induced a dose‐dependent relaxation. The EC50 of l‐arginine was 8.0 ± 0.3 μm.

The reactivity of LPS‐treated rings to noradrenaline both in the absence and presence of l‐arginine was restored to control levels by NG‐nitro‐l‐arginine methyl ester (l‐NAME, 300 μm), an inhibitor of NO production and by methylene blue (3 μm), an inhibitor of guanylate cyclase.

Incubation of isolated aortae in the absence of l‐arginine did not significantly decrease the tissue arginine content, whether LPS (10 μg ml−1) was present or not. Similarly, the presence of l‐arginine (100 μm) in the incubation medium did not modify the tissue arginine content.

These results show that the LPS‐induced impairment of vasoconstriction elicited by noradrenaline is dependent on extracellular l‐arginine, although the tissue arginine content is not depleted after LPS pretreatment, and that circulating l‐arginine is sufficient to activate maximally the vascular l‐arginine/NO pathway in endotoxaemic rats.
Comparison of the effects of selective inhibitors of phosphodiesterase types III and IV in airway smooth muscle with differing β‐adrenoceptor subtypes
Authors: Tomkinson Adrian , Karlsson Jan‐Anders , Raeburn David
Published: July 2012

The relaxant properties of the type IV adenosine 3′,5′‐cyclic monophosphate phosphodiesterase (cyclic AMP PDE) inhibitor, rolipram and the β2‐selective and non‐selective β‐adrenoceptor agonists salbutamol and isoprenaline, were compared on the guinea‐pig, bovine, and mouse trachea and porcine bronchus all precontracted with methacholine (EC30).

Rolipram and both β‐agonists produced concentration‐dependent reversal of the methacholine‐induced tone in the four airway preparations.

Isoprenaline and salbutamol were similar in potency on the guinea‐pig (−log10IC50:8.43, 8.06) and bovine (−log10 IC50:8.52, 8.40) airways. In contrast, salbutamol was much less potent than isoprenaline on the mouse trachea (> 1000 fold) and the porcine bronchus (> 100 000 fold).

The potency of rolipram approached that of isoprenaline on the guinea‐pig and bovine trachea (β2‐adrenoceptors predominate). However, rolipram was significantly less active than isoprenaline on the porcine bronchus (1000 fold) and mouse trachea (> 2000 fold) where β2‐adrenoceptors predominate.

Siguazodan, the type III cyclic AMP PDE inhibitor, produced concentration‐dependent relaxations of the porcine bronchus and guinea‐pig trachea contracted with methacholine. Siquazodan was 100 fold more active than rolipram in pig tissues indicating the type III isoenzyme may be of greater functional significance in this tissue. In contrast, siguazodan was 15 times less potent that rolipram in guinea‐pig airways suggesting a greater role for the type IV PDE.

These findings may reflect a possible relationship between the β2‐adrenoceptor subtype and the functional importance of the type IV PDE isoenzyme. A similar relationship may exist between β1‐adrenoceptors and the PDE type III isoenzyme.
The local anti‐inflammatory action of dexamethasone in the rat carrageenin oedema model is reversed by an antiserum to lipocortin 1
Authors: Duncan G.S. , Peers S.H. , Carey F. , Forder R. , Flower R.J.
Published: July 2012

A local pre‐injection of 1 μg dexamethasone sodium phosphate strongly inhibited (> 60% inhibition at 3 h; P < 0.001 at all time points) the development of carrageenin‐induced paw oedema in the rat induced by a subplantar injection of 0.1 ml, 2% carrageenin.

Coinjection of a polyclonal rabbit antiserum raised against human 1–188 recombinant lipocortin 1, which also recognised the rat protein, reversed the inhibitory action of dexamethasone (P < 0.05 at 4 h and 5 h). At the highest volume used (40 μl) control antisera were without any effect.

These data further support the concept that lipocortin 1 is involved in the anti‐inflammatory mechanism of action of the glucocorticoids.
Sympathetic innervation and α‐adrenoceptor profile of blood vessels in the posterior region of the rabbit knee joint
Authors: Najafipour Hamid , Ferrell William R.
Published: July 2012

Experiments were performed to determine the nature of adrenoceptors mediating neurally‐induced vasoconstriction of blood vessels in the posterior region of the rabbit knee joint capsule.

Electrical stimulation of the posterior articular nerve resulted in frequency‐dependent vasoconstriction which was maximal at 10 Hz. This response was mediated predominantly by α2‐adrenoceptors as it was only slightly reduced by prazosin administration and was not only abolished but converted into a dilator response by the α2‐adrenoceptor antagonist rauwolscine. Further experiments with another specific α1‐adrenoceptor antagonist YM‐12617 showed that the frequency‐response curve in the presence of this antagonist did not differ significantly from control.

Neurally‐induced vasoconstriction did not appear to have a purinergic component as it was unaffected by the P2x‐purinoceptor desensitiser α,β methylene ATP.

The rank‐order of potency of α‐adrenoceptor agonists given as a bolus by close intra‐arterial injection was: adrenaline = UK‐14304 > clonidine > phenylephrine, suggesting that the vasoconstrictor effects were mediated predominantly by postjunctional α2‐adrenoceptors.

The α2‐adrenoceptor antagonist rauwolscine converted the constrictor response to close intra‐arterial injection of adrenaline into a dilator response. The vasoconstrictor responses to UK‐14304, clonidine and phenylephrine were substantially inhibited by rauwolscine. The α1‐adrenoceptor antagonist prazosin failed to inhibit the vasoconstrictor responses to adrenaline, clonidine and UK‐14304 and resulted in enhancement of their constrictor effects.

The enhancement of the responses to the α1 and α2 agonists by prazosin appeared to be specifically related to this agent as administration of YM‐12617 did not show such enhancement. The dose‐response curves to both clonidine and UK‐14304 in the presence of YM‐12617 did not differ significantly from control responses. Responses to phenylephrine were significantly reduced by YM‐12617, indicating the presence of post‐junctional α1‐adrenoceptors.

These results show almost complete reversal of the adrenoceptor profile compared to results obtained in an earlier in vitro study, where responses were mediated predominantly by α1‐adrenoceptors with a small population of postjunctional α2‐adrenoceptors (Ferrell & Khoshbaten, 1989). This suggests that the differing environment in vitro may not completely reflect the conditions prevailing in vivo.
Identification of a novel calcium antagonist binding site in rat brain by SR 33557
Authors: Kenny Barry A. , Fraser Stuart , Speeding Michael
Published: July 2012

In K+‐depolarized taenia preparations from guinea‐pig caecum SR 33557 was a potent antagonist of Ca2+‐induced contractions and antagonized the effect of the calcium channel activator Bay K 8644.

SR 33557 displayed high affinity (pKi 9.54 ± 0.04, nH 1.01) for the [3H]‐(±)‐PN 200‐110 binding site in rat cerebral cortex membranes. In the presence of 5 mm Ca2+ this affinity was reduced (pKi 8.82 ± 0.01, nH 1.05) whilst the affinity of nitrendipine was unaffected by this concentration of Ca2+.

Saturation binding experiments in rat cerebral cortex carried out in the absence and presence of SR 33557 (0.1–1.0 nm) indicated an apparently competitive interaction at the dihydropyridine site, in that SR 33557 reduced the KD of [3H]‐(±)‐PN 200‐110 binding without any effect on Bmax. In kinetic experiments, the rate of dissociation of [3H]‐(±)‐PN 200‐110 from rat cerebral cortex was unchanged in the presence of SR 33557 (5 nm).

d‐cis‐diltiazem fully reversed the inhibition [3H]‐nitrendipine binding to rat cerebral cortex produced by SR 33557 indicating the site of action of SR 33557 to be distinct from the dihydropyridine (DHP) binding site.

Saturation analysis indicated that [3H]‐SR 33557 (0.01–0.8 nm) labelled a single class of binding sites in rat cerebral cortex membranes with high affinity (KD 0.12 ± 0.01, Bmax 222 ± 20 fmol mg−1 protein), although kinetic data indicated the existence of negative cooperativity between the binding sites.

In competition studies, a variety of different calcium antagonists displayed similar affinity for [3H]‐SR 33557 and [3H]‐(±)‐PN 200‐110 sites. The [3H]‐SR 33557 site was sensitive to the inhibitory effect of divalent cations. The affinity of Cd2+ was 0.026 ± 0.015 mm and the rank order of affinity was Cd2+ > Ca2+ > Mn2+ > Mg2+ > Na+.

We propose that SR 33557 labels a distinct site in rat cerebral cortex. The coupling between the SR 33557 and DHP site appears to be very close, resulting in apparently competitive interactions in some experimental protocols but can be revealed as negatively allosteric in other circumstances.
Neurokinin‐induced changes in pial artery diameter in the anaesthetized guinea‐pig
Authors: Beattie D.T. , Stubbs C.M. , Connor H.E. , Feniuk W.
Published: July 2012

The effects of selective neurokinin agents on pial artery diameter, measured with an on‐line image analyser, have been studied in anaesthetized guinea‐pigs in order to characterize the neurokinin receptors present on pial arteries.

Perivascular injection of either substance P (0.01–1 μm) or the selective NK1 receptor agonists, substance P methyl ester (SPOMe, 0.01–1 μm) and GR73632 (0.1 μm), increased pial artery diameter.

In contrast, the selective NK2 receptor agonist, GR64349 (1 μm), produced a small vasoconstriction while the NK3 receptor‐selective agonist, senktide (1 μm) was inactive.

Co‐administration of GR82334 (1 μm), a selective NK1 receptor antagonist, inhibited the vasodilatation produced by SPOMe (0.1 μm) but not that caused by calcitonin gene‐related peptide (CGRP, 0.01 μm).

The results are consistent with an involvement of NK1 receptors in the neurokinin‐induced increase in guinea‐pig pial artery diameter.
Influences of the endothelium and hypoxia on neurogenic transmission in the isolated pulmonary artery of the rabbit
Authors: MacLean Margaret R. , McCulloch K.M. , MacMillan J.B. , McGrath J.C.
Published: July 2012

The effects of nitric oxide (10−6 m), Nω‐nitro‐l‐arginine methylester (l‐NAME, 10−4 m, an inhibitor of nitric oxide synthase), endothelium removal, hypoxia and selective α‐adrenoceptor antagonists on responses to nerve electrical field‐stimulation (EFS) were studied in the rabbit isolated pulmonary artery.

EFS induced frequency‐dependent contractions which were abolished by prazosin (α1‐adrenoceptor antagonist) and unaffected by rauwolscine (α2‐adrenoceptor antagonist). EFS‐induced responses were potentiated by l‐NAME and inhibited by nitric oxide. The effect of l‐NAME was reversed by the presence of l‐arginine (2 × 10−4 m), which had no effect on its own. In the presence of l‐NAME, the EFS‐induced responses were reduced by rauwolscine and the residual responses were abolished by prazosin.

Removal of the vascular endothelium increased the maximum contractile response to EFS but did not inhibit the ability of l‐NAME to potentiate contractile responses to EFS.

Hypoxia inhibited the contractile response to EFS. This effect of hypoxia was also seen in the presence of l‐NAME and in endothelium rubbed preparations.

In conclusion, the endothelium modulates EFS‐induced contractions in the rabbit pulmonary artery. The contraction induced by EFS was inhibited by nitric oxide, but potentiated by the nitric oxide‐synthase inhibitor, l‐NAME. The effect of l‐NAME was not mediated solely through the endothelium and revealed involvement of α2‐adrenoceptors in EFS‐induced contraction. Hypoxia inhibited neurogenic responses in rabbit isolated pulmonary arteries.
Conversion of glyceryl trinitrate to nitric oxide in tolerant and non‐tolerant smooth muscle and endothelial cells
Authors: Salvemini Daniela , Pistelli Alessandra , Vane John
Published: July 2012

Exposure of smooth muscle cells (SMC) to glyceryl trinitrate (GTN, 75–600 μm) for 30 min led to a concentration‐dependent increase in nitrite (NO2−), one of the breakdown products of nitric oxide (NO). This was not affected by 30 min pretreatment of the cells with 0.5 mm of sulphobromophthalein (SBP) an inhibitor of glutathione‐S‐transferase (GST), by metyrapone or SKF‐525A inhibitors of cytochrome P450. These experiments were confirmed by organ bath studies using rabbit aortic strips denuded of endothelium and contracted with phenylephrine. Thus, a 30 min incubation of the strips with 0.5 mm SPB, metyrapone or SKF‐525A did not affect the relaxations in response to GTN (10−10−10−6 m).

Potentiation of the anti‐platelet effect of GTN (44 μm) by endothelial cells (EC, 40 × 103 cells) was not affected by prior incubation of EC with SBP, metyrapone or SKF‐525A (all at 0.5 mm).

Potentiation of the antiplatelet activity of GTN (11–352 μm) by small numbers of SMC (24 × 103 cells) or EC (40 × 103 cells) treated with indomethacin (10 μm) was attenuated when the SMC or EC were treated in culture with a high concentration of GTN (600 μm) for 18 h beforehand (referred to as ‘tolerant’ cells). In addition, tolerant SMC produced far less NO2− than non‐tolerant SMC.

Exposure of non‐tolerant SMC or EC (105 cells) to GTN (200 μm) for 3 min increased (3–4 fold) the levels of guanosine 3′:5′‐cyclic monophosphate (cyclic GMP). This increase was much less (≤ 1 fold) in the tolerant SMC or EC (105 cells). The basal levels of cyclic GMP were similar in normal or tolerant SMC or EC. Sodium nitroprusside (80 μm) or atrial natriuretic factor (ANF, 10−7 m) increased the levels of cyclic GMP in normal or tolerant SMC or EC to the same extent.

The anti‐platelet effects of GTN (44 μm) were potentiated by the sulphydryl donor N‐acetylcysteine (NAC, 0.5 mm). Incubation of GTN (150–1200 μm) for 30 min with NAC (0.1–1 mm) led to a concentration‐dependent increase in NO2− formation. The reduced ability of tolerant SMC or EC to potentiate the anti‐platelet activity of GTN was restored by NAC (0.5 mm). These anti‐aggregatory effects were abolished by concurrent co‐incubation with oxyhaemoglobin (10 μm) indicating that they were due to NO release.

Thus, in SMC or EC, metabolism of GTN to NO does not depend on glutathione‐S‐transferase or the cytochrome P450 system. Furthermore, when compared to normal cells, tolerant SMC or EC metabolize GTN to NO less effectively as assessed by the reduced capacity to potentiate the antiplatelet effects of GTN, to release NO2− and to increase the level of cyclic GMP. This decrease in NO formation shows that tolerance to GTN is mainly due to impaired biotransformation of GTN to NO. NAC, by directly forming NO from GTN, compensates for this failing mechanism.
Histamine H1‐receptor‐mediated inositol phospholipid hydrolysis in DDT1MF‐2 cells: agonist and antagonist properties
Authors: White Teresa E. , Dickenson John M. , Hill Stephen J.
Published: July 2012

The effect of histamine H1‐receptor stimulation on inositol phospholipid hydrolysis has been investigated in the hamster vas deferens smooth muscle cell line, DDT1MF‐2.

Histamine (EC50 = 27 μm) stimulated the accumulation of [3H]‐inositol phosphates in DDT1MF‐2 cells prelabelled with [3H]‐myo‐inositol. 2‐Thiazolylethylamine (EC50 42 μm) produced a maximal response of similar magnitude to histamine while the maximal response obtained with Nα‐methylhistamine (EC50 = 72 μm) and 2‐pyridylethylamine (EC50 = 85 μm) were much lower (circa 65%, histamine = 100%).

The H1‐selective agonists 2‐(3‐fluorophenyl)‐histamine (2‐FPH) and 2‐(3‐chlorophenyl)‐histamine (2‐CPH) both appeared to act as partial H1‐agonists in this system. Both compounds produced maximal responses of only 30% (with respect to histamine = 100) and were able to antagonize the inositol phosphate response to histamine (estimated Kp = 10.4 and 18.9 μm for 2‐FPH and 2‐CPH respectively).

The response to histamine was antagonized by the H1‐antagonists, mepyramine (KD 0.4 nm), (+)‐chlorpheniramine (KD 1.2 nm) and promethazine (KD 0.3 nm). Furthermore, the (−)‐isomer of chlorpheniramine was approx. three orders of magnitude less potent than the corresponding (+)‐isomer.

The response to histamine (0.1 mm) was not altered by prior treatment of cells with pertussis toxin (100 ng ml−1; 24 h) whereas the inositol phosphate response to adenosine A1‐receptor stimulation in this cell line was significantly attenuated under these conditions.

These data indicate that histamine‐stimulated inositol phospholipid hydrolysis in DDT1MF‐2 cells is mediated via a classical H1‐receptor. Furthermore, the results also suggest that histamine H1‐ and adenosine A1‐receptors activate phospholipase C in DDT1MF‐2 cells via two different G‐protein‐coupled pathways.
Charybdotoxin‐sensitive K+ channels regulate the myogenic tone in the resting state of arteries from spontaneously hypertensive rats
Authors: Asano Masahisa , Masuzawa‐Ito Kaoru , Matsuda Tomohiro
Published: July 2012

To determine the possible role of Ca2+‐activated K+ (KCa) channels in the regulation of resting tone of arteries from spontaneously hypertensive rats (SHR), the effects of agents which interact with these channels on tension and 86Rb efflux were compared in endothelium‐denuded strips of carotid, femoral and mesenteric arteries from SHR and normotensive Wistar‐Kyoto rats (WKY).

Strips of carotid, femoral and mesenteric arteries from SHR exhibited a myogenic tone; that is, the resting tone decreased when either the Krebs solution was changed to a 0‐Ca2+ solution or 10−7 m nifedipine was added.

The addition of charybdotoxin (ChTX, 10−9−10−7 m), a blocker of large conductance KCa channels, to the resting strips of these arteries produced a concentration‐dependent contraction, which was significantly greater in SHR than in WKY. Relatively low concentrations of tetraethylammonium (0.05–5 mm) produced a concentration‐dependent contraction which was similar to the ChTX‐induced contraction in these strips.

The ChTX‐induced contractions in SHR were greatly attenuated by 10−7 m nifedipine and by 3 × 10−6 m cromakalim, a K+ channel opener. Cromakalim alone abolished the myogenic tone in SHR.

The addition of apamin (a blocker of small conductance KCa channels, up to 10−6 m), or of glibenclamide (a blocker of ATP‐sensitive K+ channels, up to 5 × 10−6 m), to the resting strips failed to produce a contraction.

In resting strips of carotid, femoral and mesenteric arteries preloaded with 86Rb, the basal 86Rb efflux rate constants were significantly greater in SHR than in WKY. The addition of 10−7 m nifedipine to the resting strips decreased the basal 86Rb efflux rate constants only in SHR.

The cellular Ca2+ uptake in the resting state of carotid and femoral arteries from SHR was significantly increased when compared to WKY, and this increase in SHR was significantly reduced by 10−7 m nifedipine.

These results suggest that the ChTX‐sensitive KCa channels were highly activated to regulate the myogenic tone in the resting state of carotid, femoral and mesenteric arteries from SHR. The increased KCa channel functions in SHR arteries appeared to be secondary to the increased Ca2+ influx via L‐type voltage‐dependent Ca2+ channels in the resting state of these arteries.
Nitric oxide is involved in the regulation of angiogenesis
Authors: Pipili‐Synetos Eva , Sakkoula Eleni , Maragoudakis M.E.
Published: July 2012

The in vivo model of the chick embryo chorioallantoic membrane (CAM) was used to study the involvement of nitric oxide (NO) in angiogenesis. The nitrovasodilator sodium nitroprusside (NaNP) and the amino acid, l‐arginine, inhibited angiogenesis, assessed as both collagenous protein biosynthesis and vascular density. NG‐monomethyl‐l‐arginine (l‐NMMA), an NO synthase inhibitor, increased both collagenous protein biosynthesis and vascular density, indicating that this agent promotes angiogenesis. These results suggest that NO may participate in the regulation of angiogenesis. Manipulation of NO synthesis therefore, may prove to be another approach for controlling angioproliferative diseases.
Cholecystokinin as a factor in the enhanced potency of spinal morphine following carrageenin inflammation
Authors: Stanfa Louise C. , Dickenson Anthony H.
Published: July 2012

Cholecystokinin (CCK) has been shown to diminish opioid analgesia. Here we investigate whether changes in the physiological levels of spinal CCK are responsible for the enhanced potency of spinal morphine in animals following carrageenin inflammation, as compared with normal animals.

Single dorsal horn nociceptive neurones were recorded in intact halothane‐anaesthetized rats in the presence and absence of carrageenin‐induced inflammation and comparisons were made between the two groups of animals. Inflammation was induced by the injection of 100 μl of 2% λ‐carrageenin into the hind paw.

The inhibitory effect of intrathecal morphine on the C‐fibre‐evoked responses of the neurones was enhanced in the carrageenin‐treated animals such that the effects of 0.25 μg and 10 μg of morphine in normal animals were comparable to those of 0.01 μg and 2.5 μg in the carrageenin animals. The effect of 0.2 mg kg−1 of the CCKB antagonist, L‐365,260, on the antinociceptive potency of intrathecal morphine was examined in both groups of animals. In normal animals, L‐365,260 produced a significant enhancement in the effect of morphine indicating a tonic CCK modulation in these animals, but it had no effect on the inhibitions produced by either dose of morphine in the carrageenin animals.

The inhibition of the C‐fibre‐evoked response produced by intrathecal morphine in the presence of 1 μg of CCK was examined in both groups of animals. CCK attenuated the effects of morphine only in animals with carrageenin inflammation, having no effect on the action of morphine in normal animals.

The effects of both CCK and L‐365,260 were therefore dependent on the inflammatory state of the animal, with each drug being active in opposite situations.

We propose that in normal animals, morphine may produce a maximal stimulation of the release of CCK such that exogenous CCK is unable to reduce further the analgesic effects under these conditions. However, the differential effects of the agonist and antagonist in the normal and inflamed rats points to a role of CCK in the enhanced opiate actions. This enhancement of the potency of spinal morphine in inflammation is best explained by a reduction in spinal CCK release by morphine in this state.