bluuebunny/crossref_metadata_2025 huggingface dataset:
4, 6‐Dibromo‐3‐hydroxycarbazole (an analogue of caffeine‐like Ca2+ releaser), a novel type of inhibitor of Ca2+‐induced Ca2+ release in skeletal muscle sarcoplasmic reticulum
Authors: Takahashi Yukiko , Furukawa Ken‐Ichi , Kozutsumi Daiske , Ishibashi Masami , Kobayashi Jun'ichi , Ohizumi Yasushi
Published: July 2012

4,6‐Dibromo‐3‐hydroxycarbazole (DBHC) was synthesized as an analogue of bromoeudistomin D (BED), a powerful Ca2+ releaser, and its pharmacological properties were examined.

In Ca2+ electrode experiments, DBHC (100 μm) markedly inhibited Ca2+ release from the heavy fraction of sarcoplasmic reticulum (HSR) induced by caffeine (1 mm) and BED (10 μm).

DBHC (0.1 to 100 μm) inhibited 45Ca2+ release induced by Ca2+ from HSR in a concentration‐dependent manner.

DBHC (100 μm) abolished 45Ca2+ release induced by caffeine (1 mm) and BED (10 μm) in HSR.

Inhibitory effects of calcium‐induced calcium release (CICR) blockers such as procaine, ruthenium red and Mg2+ on 45Ca2+ release were clearly observed at Ca2+ concentrations from pCa 7 to pCa 5.5, and were decreased at Ca2+ concentrations higher than pCa 5.5 or lower than pCa 7. However, DBHC decreased Ca2+ release induced by Ca2+ over the wide range of extravesicular Ca2+ concentrations.

[3H]‐ryanodine binding to HSR was suppressed by ruthenium red, Mg2+ and procaine, but was not affected by DBHC up to 100 μm.

[3H]‐ryanodine binding to HSR was enhanced by caffeine and BED. DBHC antagonized the enhancement in a concentration‐dependent manner.

9‐[3H]‐Methyl‐7‐bromo‐eudistomin D, an 3H‐labelled analogue of BED, specifically bound to HSR. Both DBHC and caffeine increased the KD value without affecting the Bmax value, indicating a competitive mode of inhibition.

These results suggest that DBHC binds to the caffeine binding site to block Ca2+ release from HSR. This drug is a novel type of inhibitor for the CICR channels in SR and may provide a useful tool for clarifying the Ca2+ releasing mechanisms in SR.
The role of nitric oxide in the altered vascular reactivity of pregnancy in the rat
Authors: Nathan Lauren , Cuevas Janis , Chaudhuri Gautam
Published: July 2012

Pregnancy is characterized by a decrease in systemic vascular resistance and a blunting of the angiotensin II (AII) pressor response. We studied the role of nitric oxide (NO) and prostanoids in these vascular changes of pregnancy in anaesthesized, ganglion blocked non‐pregnant and pregnant rats.

Inhibition of NO synthesis with NG‐nitro‐l‐arginine methyl ester (l‐NAME) led to an increase in mean arterial pressure (MAP) which was of a significantly greater magnitude in pregnant rats in late gestation than in non‐pregnant rats, or rats in mid‐gestation.

The pressor response to varying doses of AII was attenuated during late pregnancy, and this attenuation was partially reversed by l‐NAME.

The pressor response to varying doses of a vasocontrictor, phenylephrine (PE), was also attenuated in late pregnancy. However, this attenuation was not reversed by l‐NAME.

Inhibition of prostanoid biosynthesis with meclofenamate did not alter basal MAP, nor the pressor response to varying doses of AII or PE in pregnant and non‐pregnant animals.

It is concluded that (a) increased NO synthesis occurs during late gestation and contributes both to the decrease in systemic vascular resistance, as well as the blunting of the pressor response to AII during pregnancy, and (b) prostaglandins are not important in the maintenance of basal vascular tone, or the blunting of the pressor response to AII during pregnancy.
Suppression by the sumatriptan analogue, CP‐122,288 of c‐fos immunoreactivity in trigeminal nucleus caudalis induced by intracisternal capsaicin
Authors: Cutrer F. Michael , Schoenfeld David , Limmroth Volker , Panahian Nariman , Moskowitz Michael A.
Published: July 2012

The effects of an intravenously administered sumatriptan analogue were examined on c‐fos‐like immunoreactivity (c‐fos‐LI), a marker of neuronal activation, evoked within trigeminal nucleus caudalis (TNC) and other brain stem regions 2 h after intracisternal injection of the irritant, capsaicin (0.1 ml, 0.1 mm), in pentobarbitone‐anaesthetized Hartley guinea‐pigs.

C‐fos‐LI was assessed in eighteen serial sections (50 μm) using a polyclonal antiserum. A weighted average, reflecting total expression within lamina I, IIo of TNC was obtained from three representative levels (i.e., at −0.225 mm, −2.475 mm and −6.975 mm.)

Capsaicin caused significant labelling within lamina I, IIo, a region containing axonal terminations of small unmyelinated C‐fibres, as well as within the nucleus of the solitary tract, area postrema and medial reticular nucleus. A similar distribution of positive cells was reported previously after intracisternal injection of other chemical irritants such as autologous blood or carrageenin.

Pretreatment with a conformationally restricted sumatriptan analogue (with some selectivity for 5‐HT1B and 5‐HT1D receptor subtypes) CP‐122,288, reduced the weighted average by approximately 50–60% (P<0.05) in lamina I, IIo at ≥ 100 pmol kg−1, i.v., but did not decrease cell number within area postrema, nucleus of the solitary tract or medial reticular nucleus. A similar pattern was reported previously following sumatriptan, dihydroergotamine or CP‐93,129 administration after noxious meningeal stimulation.

We conclude that modifications at the amino‐ethyl side chain of sumatriptan dramatically enhance the suppression of c‐fos expression within TNC, a finding consistent with its remarkable potency against neurogenic plasma protein extravasation within dura mater. CP‐122,288 and related analogues may serve as an important prototype for drug development in migraine and related headaches.
Further investigations into adenosine A1 receptor‐mediated contraction in rat colonic muscularis mucosae and its augmentation by certain alkylxanthine antagonists
Authors: Reeves J.J. , Jarvis J.E. , Sheehan M.J. , Strong P.
Published: July 2012

The alkylxanthine antagonists, 8‐phenyltheophylline (8‐PT), 8‐p‐sulphophenyltheophylline (8‐SPT) and 1,3,7‐trimethylxanthine (caffeine) produced rightward displacements of contractile concentration‐effect curves to 5′‐N‐ethylcarboxamidoadenosine (NECA) in rat isolated colonic muscularis mucosae (RCMM) with concentration‐ratios consistent with adenosine receptor blockade. The non‐xanthine antagonist, 9 fluro‐2‐(2‐furyl)‐5,6‐dihydro [1,2,4] triazo to [1,5‐c]‐quinazin‐imine (CGS15943A) also antagonized contractions to NECA with an affinity (pKB 8.1–8.5) consistent with adenosine A1 receptor blockade.

In addition to producing rightward shifts of the concentration‐response curves, the maximum contractions to 5′‐N‐ethylcarboxamidoadenosine (NECA) were also markedly increased in the presence of 8‐PT (by 83 ± 16% at 1 μm), 8–SPT (by 37 ± 7% at 10 μm) and caffeine (by 45 ± 5% at 100 μm) but were unaffected by CGS15943A (at 0.01 and 0.03 μm).

As with NECA, the maximum contractions to the adenosine A1 receptor agonists R‐phenylisopropyladenosine (R‐PIA) and N‐[(1S, trans)‐2‐hydroxyclopentyl] adenosine (GR79236) were both antagonized and augmented by 8‐PT. In addition, the contractions to NECA in the presence of 8–PT (1 μm) were inhibited by nanomolar concentrations of 8‐cyclopentyl‐1,3‐dipropylxanthine (DPCPX).

The non‐selective phosphodiesterase (PDE) inhibitor, 3‐isobutyl‐1‐methylxanthine (1 μm) produced a marked increase in the NECA maximum without producing a rightward shift in the NECA curve, whereas a higher concentration (10 μm) virtually abolished responses. The PDE type III inhibitor, milrinone (1 μm), the type IV inhibitor, rolipram (10 μm), and the type V PDE inhibitor, zaprinast (3 μm), were all without effect on NECA responses in RCMM.

Partial inhibitions of contractions to NECA were produced by indomethacin (at 3 or 10 μm) or piroxicam (at 3 μm). Responses to GR79236 were also partially inhibited by indomethacin. In the presence of indomethacin, 8‐PT was still able to enhance markedly the maximum contractions obtained to NECA in RCMM.

The present study has shown that certain alkylxanthine antagonists (but not the non‐xanthine CGS15943A) produced a marked augmentation of adenosine A1 receptor‐mediated contractions in RCMM. The mechanism of this augmentation is, as yet, not known but is unlikely to result from inhibition of PDE. This study has also shown that adenosine A1 receptor‐induced contractions in RCMM are mediated, in part, via products of the cyclo‐oxygenase pathway.
Pharmacological characterization of α1‐adrenoceptor subtypes in rat heart: a binding study
Authors: Noguchi Hideki , Muraoka Ryusuke , Kigoshi Shigeru , Muramatsu Ikunobu
Published: July 2012

The α1‐adrenoceptor subtypes of rat heart were characterized in binding experiments performed with [3H]‐prazosin as the radiolabel. The specific binding to the α1‐adrenoceptors was determined with 0.3 μm prazosin, because phentolamine (10 μm) was insufficient to inhibit completely the specific binding of high concentrations of [3H]‐prazosin.

In saturation experiments, [3H]‐prazosin bound to two distinct affinity sites (pKD = 10.39 and 8.19). The proportion of the low affinity sites was approximately 84% of total specific binding. Membranes pretreated with chloroethylclonidine (CEC, 10 μm) also showed two distinct affinity sites for [3H]‐prazosin, although the maximum numbers of high and low affinity sites were reduced by 86 and 64%, respectively.

In competition experiments, [3H]‐prazosin (100 pm) binding was inhibited by WB4101 (2‐(2,6‐dimethoxy‐phenoxyethyl)aminomethyl‐1,4‐benzodioxane) and 5‐methylurapidil. The inhibition curves displayed shallow slopes which could be subdivided into high and low affinity components (pK1 = 10.43 and 8.36 for WB4101, 8.62 and 6.61 for 5‐methylurapidil). However, unlabelled prazosin or HV723 (α‐ethyl‐3,4,5‐trimethoxy‐α‐(3‐((2‐(2‐methoxyphenoxy)‐ethyl)amino)propyl)benzeneacetonitrile fumarate) competed for [3H]‐prazosin binding monophasically (pK1 = 10.34 and 8.28, respectively). In CEC‐pretreated membranes, prazosin, WB4101, 5‐methylurapidil and HV723 antagonized the [3H]‐prazosin (100 pm) binding monophasically (pKI = 9.70, 9.56, 8.60 and 8.82, for each antagonist).

On the other hand, 1000 pm [3H]‐prazosin binding was inhibited by unlabelled prazosin biphasically (pKI = 10.49 and 8.49). HV723 did not discriminate both prazosin‐high and low affinity sites (pKI = 8.18).

These results suggest the presence of at least three distinct α1‐adrenoceptor subtypes in rat hearts (two prazosin‐high affinity sites and one prazosin‐low affinity site). According to the recent α1‐adrenoceptor subclassifications, one of the former two sites corresponds to the α1B subtype with low affinities for WB4101 and 5‐methylurapidil and sensitive to CEC, while another site with relatively high affinities for WB4101 and 5‐methylurapidil may be classical α1A, cloned α1C, α1D subtypes or their mixture. The prazosin‐low affinity site corresponds to putative α1L subtype with low affinity for HV723, which may be predominantly involved in the positive inotropic response to phenylephrine.
Inhibition of morphine withdrawal by the association of RB 101, an inhibitor of enkephalin catabolism, and the CCKB antagonist PD‐134,308
Authors: Maldonado Rafael , Valverde Olga , Ducos Bertrand , Blommaert Armand G , Fournie‐Zaluski Marie‐Claude , Roques Bernard P.
Published: July 2012

The effects induced in rats on naloxone‐precipitated morphine withdrawal syndrome by the new mixed inhibitor of enkephalin catabolism able to cross the blood‐brain barrier RB 101 (N‐{(R,S)‐2‐benzyl‐3[(S)(2‐amino‐4‐methylthio)butyl dithio]‐1‐ox‐opropyl‐l‐phenylalanine benzyl ester) given alone or associated with the selective CCKB antagonist, PD‐134,308, were investigated.

The systemic administration of RB 101 (5, 10 and 20 mg kg−1, i.v.) elicited a significant decrease in 8 of the 14 withdrawal signs evaluated. PD‐134,308 (3 mg kg−1, i.p.) did not modify the expression of morphine abstinence when given alone, but induced a strong facilitation of RB 101 responses (12 of 14 withdrawal signs were decreased). This potentiation was particularly intense in peripherally mediated withdrawal signs.

In order to clarify the biochemical mechanisms implicated in these responses, the effects induced by the association of RB 101 and PD‐134,308 on the occupation of brain opioid receptors by endogenous enkephalins were also investigated in mice. PD‐134,308, as well as RB 101, inhibited [3H]‐diprenorphine binding to opioid receptors. These results suggest that an increase in endogenous enkephalin levels induced by PD‐134,308 could participate in the facilitation of RB 101 behavioural responses.

RB 101 has a promising potential role in the management of the opiate withdrawal syndrome. CCKB antagonists, such as PD‐134,308 may be useful in potentiating this anti‐withdrawal effect.
Functional effects of long‐term activation on human β2‐ and β3‐adrenoceptor signalling
Authors: Nantel François , Bouvier Michel , Strosberg A. Donny , Marullo Stefano
Published: July 2012

The functional effects of long‐term activation of β‐adrenoceptors were investigated by measuring adenylyl cyclase activity, cyclic AMP accumulation and cyclic AMP‐dependent protein kinase activity in CHW and L cells expressing either human β2‐ or β3‐adrenoceptors.

Pre‐incubation of CHW and L cells expressing β2‐adrenoceptors with 10 μm isoprenaline for 24 h produced a marked reduction in the total receptor number and dramatically reduced the capacity of the receptor to stimulate adenylyl cyclase maximally.

In contrast, the ability of β3‐adrenoceptors to stimulate adenylyl cyclase maximally was not affected by pre‐incubation with the agonist in either cell type. However, a significant reduction of isoprenaline potency and a sustained down‐regulation of β3‐adrenoceptor number was observed in L but not in CHW cells.

Maximal levels of intracellular cyclic AMP concentrations were reached during the first hour of receptor activation with isoprenaline in all four cell lines. In the absence of phosphodiesterase inhibitors, cyclic AMP decreased to basal levels within 24 h of continuous stimulation. This phenomenon occurred more rapidly in cells expressing the β2‐ than the β3‐adrenoceptors.

These results confirm that, at the level of adenylyl cyclase stimulation and cyclic AMP accumulation, the β3‐adrenoceptor is more resistant than the β3‐adrenoceptor to long‐term desensitization. However, when cyclic AMP‐dependent protein kinase activity was considered, a 24 h stimulation of β2‐ and β3‐adrenoceptor expressing cells led to the desensitization of the kinase in L but not in CHW cells.

In conclusion, long‐term desensitization may have distinct functional effects on cell signalling depending on the receptor subtype and the cell type considered. These findings might have practical implications for future strategies involving long‐term therapies with receptor agonists.
Inhibition by sodium nitroprusside of the expression of inducible nitric oxide synthase in rat neutrophils
Authors: Mariotto S. , Cuzzolin L. , Adami A. , Del Soldato P. , Suzuki H. , Benoni G.
Published: July 2012

A well‐known nitric oxide (NO)‐releasing compound, sodium nitroprusside (SNP), decreases in a dose‐dependent manner NO synthase (NOS) activity induced in rat neutrophils by treatment with lipopolysaccharide (LPS). This inhibitory action of SNP seems not to be due to its direct effect on the enzyme activity. The strong nitrosonium ion (NO+) character of SNP could be responsible for its inhibition of NOS induction in neutrophils.
Comparison of the pharmacological profile of S‐nitrosothiols, nitric oxide and the nitrergic neurotransmitter in the canine ileocolonic junction
Authors: De Man Joris G. , Boeckxstaens Guy E. , De Winter Benedicte Y. , Moreels Tom G. , Misset Menno E. , Herman Arnold G. , Pelckmans Paul A.
Published: July 2012

1 In organ bath experiments, hydroquinone (30–100 μm) and hydroxocobalamin (30–100 μm) concentration‐dependently inhibited the relaxations induced by NO (0.3–30 μm) but not those by nitroglycerin (GTN, 1 μm) in the canine ileocolonic junction (ICJ). Hydroxocobalamin reduced the relaxation to low frequency (2 Hz) stimulation of the non‐adrenergic, non‐cholinergic (NANC) nerves, whereas hydroquinone only reduced the NANC nerve‐mediated relaxations to electrical stimulation at 16 Hz, 0.5 ms.

2 Relaxations to S‐nitroso‐l‐cysteine (CysNO, 1–30 μm), or S‐nitroso‐N‐acetyl‐d,l‐penicillamine (SNAP, 1–30 μm) were not inhibited by hydroquinone (30–100 μm), hydroxocobalamin (30–100 μm), pyrogallol (30–100 μm) or l‐cysteine (1–3 μm). Hydroquinone (100 μm) only reduced the relaxation to 10 μm CysNO. Hydroxocobalamin, but not hydroquinone, pyrogallol or m‐cysteine, potentiated the relaxations to the lowest concentration (1 μm) of S‐nitrosoglutathione (GSNO, 1–30 μm).

3 In the superfusion bioassay, hydroquinone (100 μm) and hydroxocobalamin (1 μm) concentration‐dependently inhibited the biological activity of authentic NO (1–4pmol) to the same extent as that of the transferable nitrergic factor, released from the canine ICJ in response to NANC nerve stimulation (8–16 Hz, 2 ms). Responses to GTN (10 pmol) or adenosine 5′‐triphosphate (10 nmol) were not affected.

4 In conclusion, the nitrosothiols CysNO, SNAP and GSNO relax the canine ileocolonic junction, but these relaxations, pharmacologically, behave differently from the NANC nerve‐mediated relaxations. From the bioassay experiments, we conclude that the nitrergic factor, released in response to NANC nerve stimulation of the canine ICJ, behaves pharmacologically like NO but not like a nitrosothiol. Therefore, we suggest NO, and not CysNO, SNAP or GSNO as the inhibitory NANC neurotransmitter in the canine ICJ.
Effects of tyrosine kinase inhibitors on the contractility of rat mesenteric resistance arteries
Authors: Toma Catalin , Jensen Peter E. , Prieto Dolores , Hughes Alun , Mulvany Michael J. , Aalkjær Christian
Published: July 2012

1 A pharmacological characterization of tyrosine kinase inhibitors (TKI) belonging to two distinct groups (competitors at the ATP‐binding site and the substrate‐binding site, respectively) was performed, based on their effects on the contractility of rat mesenteric arteries.

2 Both the ATP‐site competitors (genistein and its inactive analogue, daidzein) and the substrate‐site competitors (tyrphostins A‐23, A‐47 and the inactive analogue, A‐1) reversibly inhibited noradrenaline (NA, (10 μm)) and KCl (125 mm) induced contractions, concentration‐dependently. Genistein was slightly but significantly more potent than daidzein; the tyrphostins were all less potent than genistein, and there were no significant differences between the individual potencies. The tyrosine kinase substrate‐site inhibitor bis‐tyrphostin had no inhibitory effect.

3 Genistein, daidzein, A‐23 and A‐47 each suppressed the contraction induced by Ca2+ (1 μm) in α‐toxin permeabilized arteries. A‐1 and bis‐tyrphostin had little or no effect on contraction of the permeabilized arteries.

4 Genistein was significantly more potent than daidzein with respect to inhibition of the contraction induced by 200 nm Ca2+ in the presence of NA (100 μm) and GTP (3 μm). The effect of A‐23, A‐47, A‐1 and bis‐tyrphostin was similar in permeabilized arteries activated with Ca2+ (200 nm) + NA (100 μm) + GTP (3 μm) and permeabilized arteries activated with 1 μm Ca2+.

5 Genistein (30 μm) reduced the fura‐2 measured intracellular calcium activity ([Ca2+]i) in arteries stimulated with NA but had no effect on [Ca2+]i in arteries stimulated with KCl (125 mm).

6 The potent effect of the TKIs in this study is consistent with a role for tyrosine kinases in the mechanisms which regulate both cytoplasmic Ca2+ levels and the effect of Ca2+ on the contractile apparatus in smooth muscle cells in resistance arteries. However, the results must be interpreted cautiously because the enzyme inhibitors may have a poor specificity in intact tissues and because the presumed inactive analogues had potent effects.
Effects of semotiadil fumarate, a novel Ca2+ antagonist, on cytosolic Ca2+ level and force of contraction in porcine coronary arteries
Authors: Kageyama Masaaki , Yanagisawa Teruyuki , Taira Norio
Published: July 2012

1 The mechanisms of action of semotiadil fumarate, a novel Ca2+ antagonist, were examined by measuring the cytosolic Ca2+ level ([Ca2+]i) and force of contraction in porcine coronary arteries, and by determining [3H]‐pyrilamine binding to bovine cerebellar membranes.

2 Semotiadil or verapamil (0.1 and 1 μm) inhibited both the high KCl‐induced increases in [Ca2+]i and force in a concentration‐dependent manner.

3 Histamine (30 μm) produced transient increases followed by sustained increases in [Ca2+]i and force, which were inhibited by semotiadil and verapamil (1 and 10 μm).& The agents were different in that semotiadil reduced the maximum [Ca2+]i and force responses to histamine, but not pD2 values, whereas verapamil did reduce the pD2 values for histamine, but not the maximum responses.

4 Verapamil (10 μm), but not semotiadil, inhibited histamine‐induced increases in [Ca2+]i and force in Ca2+‐free solution. Neither semotiadil nor verapamil affected the increases in [Ca2+]i and force induced by caffeine. Semotiadil even at the higher concentration (10 μm) did not displace specific binding of [3H]‐pyrilamine to bovine cerebellar membranes.

5 These results suggest that semotiadil inhibits both KCl‐ and histamine‐induced contractions mainly by blocking voltage‐dependent L‐type Ca2+ channels.
The effect of nitric oxide donors on haemodynamics and blood flow distribution in the porcine carotid circulation
Authors: van Gelderen E. Marcel , De Bruijne Emile L.E. , Agteresch Hendrik J. , Saxena Pramod R.
Published: July 2012

1 The role of nitric (NO) in the regulation of capillary and arteriovenous anastomotic blood flow was evaluated in the carotid circulation of the pig. For this purpose, the effect of intracarotid (i.c.) infusions of saline and two NO donors, nitroprusside sodium (NPR) and S‐nitroso‐N‐acetylpenicillamine (SNAP) in concentrations of 3–100 μg min−1 was studied on systemic haemodynamics and carotid blood flow and its distribution in anaesthetized pigs with low arteriovenous anastomotic blood flow, by use of the radioactive microsphere method.

2 Apart from heart rate, which increased after both NPR and SNAP, no major changes in systemic haemodynamic variables were observed. In contrast to saline, both NPR and SNAP increased common carotid blood flow, vascular conductance and vascular pulsations dose‐dependently.

3 The distribution of the carotid artery blood flow over capillary and arteriovenous anastomotic fraction remained stable after saline infusions. Both NPR and SNAP enhanced total capillary blood flow and conductance. In contrast to NPR, arteriovenous anastomotic blood flow and conductance were increased by SNAP.

4 At the tissue level, capillary blood flow increases following NPR or SNAP were reflected by an increase in both extracerebral and dural blood flow without changes in total brain blood flow.

5 These results indicate that both NO donors cause arteriolar dilatation together with enhanced vascular pulsations in the carotid circulation of the pig. Probably by way of a ‘steal’ phenomenon, this pronounced arteriolar dilatation limits the effect of NO donors on arteriovenous anastomoses.

6 The results of the present investigation support the contention that dilatation of intra‐ and extracranial arteries and arteriovenous anastomoses leads to increased vascular pulsations, which (rather than increased blood flow) could, at least in part, be responsible for the headache caused by nitrovasodilators.
Delayed‐type hypersensitivity‐induced increase in vascular permeability in the mouse small intestine: inhibition by depletion of sensory neuropeptides and NK1 receptor blockade
Authors: Kraneveld A.D. , Buckley T.L. , van Heuven‐Nolsen D. , van Schaik Y. , Koster A. Sj. , Nijkamp F.P.
Published: July 2012

1 This study investigates the effects of capsaicin‐induced depletion of sensory neuropeptides and of neurokinin! (NK1) receptor blockade on delayed‐type hypersensitivity (DTH)‐induced changes of vascular permeability in the small intestine of the mouse.

2 The DTH reaction in the small intestine was elicited by dinitrofluorobenzene (DNFB)‐contact sensitization followed by oral dinitrobenzene sulphonic acid (DNBS) challenge. To assess vascular leakage the accumulation of the plasma marker, Evans blue (EB), was measured 2, 24 and 48 h after the challenge.

3 The small intestinal DTH reaction was characterized by a significant increase in vascular permeability 24 h after the challenge of previously sensitized mice when compared to vehicle‐sensitized mice (P<0.05, ANOVA). Capsaicin‐induced depletion of sensory neuropeptides, two weeks before the sensitization, completely inhibited the DTH‐induced increase in small intestinal vascular permeability at 24 h (P<0.05, ANOVA). Vehicle/control: 108.2 ± 8.6 ngEB mg−1 dry weight; vehicle/DTH 207.8 ±25.1 ngEB mg−l dry weight; capsaicin/control: 65.8 ± 11.9 ng EB mg−1 dry weight; capsaicin/ DTH: 84.3 ± 7.6 ng EB mg−1 dry weight.

4 The tachykinins, substance P and neurokinin A (1.5 to 50 × 10−11 mol per mouse, i.v.), induced an increase in vascular leakage in the small intestine of naive mice. The specific NK1 receptor antagonist, RP67580 (10−9 mol per mouse, i.v.) was the most effective in reducing the substance P‐induced plasma extravasation when compared with other NK receptor antagonists, FK224 and FK888.

5 Treatment of DNFB‐sensitized mice with RP67580 (10−9 mol per mouse, i.v.) immediately before and 1 h after the DNBS challenge resulted in a significant reduction of the DTH‐induced increase in vascular permeability at 24h (vehicle/control: 107.5 ±8.8 ng EB mg−1 dry weight; RP67580/control: 95.4±5.4ng EB mg−1 dry weight; vehicle/DTH: 206.6± 22.6ng EB mg−1 dry weight; RP67580/DTH: 132.6±13.6 ng EB mg−1 dry weight, P<0.05, ANOVA).

6 These results suggest that sensory nerves are involved in the development of small intestinal DTH reactions in the mouse. NK1 receptors could play an important role in the initiation of the DTH‐induced changes in vascular leakage.
An enhancing effect of 5‐hydroxytryptamine on electrically evoked atropine‐resistant contraction of guinea‐pig proximal colon
Authors: Kojima Shu‐Ichi , Shimo Yasuo
Published: July 2012

In the presence of atropine (0.2 μm) and indomethacin (2 μm), the effects of 5‐hydroxytryptamine (5‐HT) have been studied on electrically‐evoked, neurogenic contractions of the guinea‐pig proximal colon in vitro.

5‐HT, at higher concentrations than 1 nm, caused an increase in electrically (1 Hz, 0.3 ms, 160 mA)‐evoked, atropine‐resistant contractions in a concentration‐dependent manner and at 30 nm produced a maximal effect (pEC50 value of 8.20 ± 0.11, n = 6). The enhancing effects of 5‐HT on the electrically evoked contractions were mimicked by α‐methyl‐5‐HT (pEC50 value of 6.59 ± 0.05, n = 6).

Both hexamethonium (100 μm) and spantide (10 μm), selective antagonists for nicotinic and tachykinin receptors respectively, significantly reduced the enhancement of the electrically evoked contractions by 5‐HT (30 nm).

DAU 6285 (3 μm), a 5‐HT4 receptor antagonist, abolished the enhancing action of 5‐HT (30 nm), but metitepine (0.03 μm), a 5‐HT1/5‐HT2 receptor antagonist, ketanserin (0.01 μm), a 5‐HT2 receptor antagonist, and ondansetron (1 μm), a 5‐HT3 receptor antagonist, had no effect on the enhancement. The enhancing effects of a‐methyl‐5‐HT (1 μm) were also abolished by DAU 6285 (3 μm).

Both 5‐HT (30 nm) and α‐methyl‐5‐HT (1 μm) had no effect on contractions to exogenous substance P (0.15–0.3 nm).

These results indicate that in the guinea‐pig proximal colon, 5‐HT produced an enhancement of atropine‐resistant neurogenic contraction induced by electrical field stimulation through pre‐junctional mechanisms and that the enhancement is mediated by the stimulation of 5‐HT4 receptors located on intramural preganglionic cholinergic neurones and tachykininergic neurones.
Acute versus chronic administration of phosphodiesterase inhibitors on allergen‐induced pulmonary cell influx in sensitized guinea‐pigs
Authors: Banner Katharine H. , Page Clive P.
Published: July 2012

The aims of this study were to determine which phosphodiesterase (PDE) isoenzymes are involved in the control of eosinophil accumulation in the airways of ovalbumin (OVA)‐immunized guinea‐pigs by the use of isoenzyme selective inhibitors and to compare the effects of acute versus chronic administration of PDE isozyme inhibitors on pulmonary cell influx in ovalbumin‐immunized guinea‐pigs.

Guinea‐pigs were sensitized and subsequently challenged with aerosolized OVA. Twenty four hours later bronchoalveolar lavage (BAL) was performed to permit assessment of inflammatory cell accumulation. A significant increase in the number of eosinophils was observed in the lavage fluid from OVA‐immunized (13.6 ± 1.4 × 104ml−1 in acute experiments and 10.1 ± 1.4 × 104 ml−1 in chronic experiments) animals compared with sham‐treated controls (5.6 ± 0.6 × 104 ml−1 in acute experiments and 5.1 ± 0.6 × 104 ml−1 in chronic experiments). There was no difference in neutrophil, mononuclear cell or total cell numbers between the two groups.

Acute administration of a high dose of selective and non‐selective PDE inhibitors by the i.p. route had no significant effect on eosinophil accumulation in the airways.

Chronic administration of a low dose (3 mg kg−1, i.p., twice daily for 7 days) of the type IV PDE inhibitor, RO 20–1724, and the PDE III/IV inhibitor, zardaverine, produced a significant inhibition of eosinophil accumulation (46% and 59% respectively).

These results suggest that the type IV PDE isoenzyme plays a role in the control of allergen‐induced eosinophil infiltration into the airways, but indicate that a period of low dose chronic treatment with a type IV or mixed type III/IV PDE inhibitor is necessary for eosinophil accumulation in the airways to be reduced.
Inhibition of calcium release from the sarcoplasmic reticulum of rabbit aorta by hydralazine
Authors: Gurney Alison M. , Allam Marina
Published: July 2012

The mechanism of hydralazine‐induced vasorelaxation was investigated in rabbit isolated aorta, by determining its ability to interfere with force development under a variety of conditions.

Hydralazine relaxed phenylephrine‐contracted aorta with half maximal relaxation at 17 μm and maximal relaxation above 100 μm. At 200 μm, hydralazine had little effect on contractions induced by 25 mm or 50 mm K+.

Hydralazine was equally effective at inhibiting contractile responses to phenylephrine in the absence or presence of extracellular Ca2+. Responses to phenylephrine in Ca2+‐free solution were blocked to the same degree whether hydralazine was applied during filling of the sarcoplasmic reticulum (SR) Ca2+ stores or after filling was complete. Caffeine‐induced contractions were less sensitive to block by hydralazine.

Thapsigargin, cyclopiazonic acid, ryanodine, nifedipine and diltiazem all failed to block the inhibitory effect of hydralazine on tonic contractions to phenylephrine in the presence of extracellular Ca2+. However, when cyclopiazonic acid was applied either with diltiazem or ryanodine, substantial inhibition of the hydralazine response was observed.

We propose that tonic contractions to phenylephrine are largely maintained by Ca2+ cycling through the SR, with Ca2+ entering the smooth muscle cell being sequestered by the SR eventually to leak out through IP3‐activated channels close to the contractile proteins. Sequestration of Ca2+ would employ two pathways, one sensitive to inhibitors of the SR Ca2+‐ATPase and the other to Ca antagonists. We further suggest that, in the presence of extracellular Ca2+ and phenylephrine, the leakage of Ca2+ through IP3‐activated channels is significantly reduced only if both routes for SR Ca2+ accumulation are blocked or the Ca2+‐ATPase is blocked while the SR is made leaky with ryanodine.

We conclude that the main action of hydralazine is to block the IP3‐dependent release of Ca2+ from the sarcoplasmic reticulum. Thus conditions that diminish the contribution of IP3‐induced Ca2+ release to tension can inhibit the hydralazine‐induced vasorelaxation.
Interaction of clozapine with the histamine H3 receptor in rat brain
Authors: Rodrigues A. Alves , Jansen F.P. , Leurs R. , Timmerman H. , Prell G.D.
Published: July 2012

We examined possible interactions between neuroleptics and the histamine H3 receptor and found an interaction of clozapine with this receptor. In competition binding experiments, using the H3 antagonist, [125I]‐iodophenpropit, we observed a Ki of 236 ± 87 nM. Functionally, clozapine was studied on the H3‐mediated inhibition of [3H]‐5‐hydroxytryptamine ([3H]‐5‐HT) release from rat brain cortex slices. Clozapine acts as an antagonist with an apparent KB value of 79.5 nM.
Stimulation of human umbilical vein endothelial cell proliferation by A2‐adenosine and β2‐adrenoceptors
Authors: Sexl Veronika , Mancusi Gudrun , Baumgartner‐Parzer Sabina , Schütz Wolfgang , Freissmuth Michael
Published: July 2012

Adenosine is known to stimulate capillary outgrowth and endothelial cell proliferation, but the underlying mechanism has not been identified. In order to identify the receptor subtype involved, the effects of adenosine receptor agonists and antagonists on human umbilical vein endothelial cell (HUVEC) proliferation were investigated.

Raising intracellular adenosine levels by use of the adenosine transport inhibitor, 4‐nitro‐benzylthioinosine (NBMPR) did not affect cell growth. This observation suggests that stimulation of an extracellular adenosine receptor generates the mitogenic signal.

In the presence of adenosine deaminase (ADA), which was used to remove adenosine present in the culture medium, the adenosine receptor agonists N‐ethylcarboxamidoadenosine (NECA, non‐selective) and CGS21680 (A2A‐receptor‐selective) stimulated [3H]‐thymidine incorporation with a half‐maximum effect at about 10 nM, while N6‐cyclopentyladenosine (CPA, A1selective) was about 100 fold less potent. The adenosine receptor antagonist, xanthine amine congener (XAC) produced a concentration‐dependent decrease in endothelial cell proliferation with a half‐maximum effect at about 10 nM. Hence, stimulation of an endothelial A2A‐adenosine receptor seems responsible for the mitogenic signal.

In the presence of ADA, isoprenaline is also able to stimulate [3H]‐thymidine incorporation with a half maximal effect of about 3 nM, an effect, which is reversed by the highly β2‐selective antagonist, ICI 118,551. In the absence of ADA, isoprenaline exerts only a minor stimulatory effect. Combination of A2A adenosine and β2‐adrenoceptor agonists did not further enhance [3H]‐thymidine incorporation when compared to the sole addition of each agonist. We therefore conclude that both receptors stimulate endothelial cell proliferation via a common signal transduction pathway.

Both receptors are coupled to stimulation of adenylyl cyclase via the stimulatory G protein Gs. However, direct activation of downstream effectors in the cyclic AMP‐signalling cascade (Gs with cholera toxin, adenylyl cyclase with forskolin, protein kinase A with 8Br‐cyclic AMP) not only failed to mimic the action of receptor‐activation, but even reduced cell proliferation.

Similarly, pertussis toxin‐treatment which inactivated the Gi2 protein present in HUVEC and thus inhibited cell proliferation per se, did not impair the ability of A2A‐receptor agonists to stimulate cell proliferation. This suggests that the A2A‐adenosine and β2‐adrenoceptor‐mediated stimulation of endothelial cell proliferation occurs via a mechanism that is independent of Gs and Gi.
Hypertension and enhanced β‐adrenoceptor‐mediated facilitation of noradrenaline release produced by chronic blockade of adenosine receptors
Authors: Guimarães S. , Albino‐Teixeira A. , Paiva M.Q.
Published: July 2012

The study was undertaken to compare the β‐adrenoceptor‐mediated facilitation of noradrenaline release in the tail artery of vehicle‐treated rats and of rats rendered hypertensive by chronic administration of 1,3‐dipropyl‐8‐sulphophenylxanthine (DPSPX). Artery rings were loaded with [3H]‐noradrenaline, and five periods of electrical stimulation (1 Hz for 2 min) were applied. To eliminate the influence of prejunctional α2‐adrenoceptors, the tissues were pre‐exposed to 1 μm phenoxybenzamine.

Isoprenaline caused a concentration‐dependent increase of tritium overflow elicited by electrical stimulation. It was more effective in arteries from DPSPX‐treated than in those from vehicle‐treated rats; isoprenaline (27.8 nM) increased by 30% tritium overflow in vessels from vehicle‐treated rats whereas isoprenaline (7.0 nM) produced a 30% increase in vessels from DPSPX‐treated animals. Furthermore, the maximal effect of isoprenaline was a 32.6% increase in control rats but a 48.6% increase in DPSPX‐treated rats.

These results show that the sympathetic nerve endings of the rat tail artery are endowed with prejunctional β‐adrenoceptors which mediate facilitation of noradrenaline release elicited by electrical stimulation. They also suggest that adenosine receptors and β‐adrenoceptors interact at the prejunctional level and that impairment of this ‘talk’ may lead to the development of a hypertensive state.
Contrasting effects of the imidazol(in)e α2‐adrenoceptor agonists, medetomidine, clonidine and UK 14,304 on extraneuronal levels of noradrenaline in the rat frontalcortex: evaluation using in vivo microdialysis and synaptosomal uptake studies
Authors: Dalley J.W. , Stanford S.C.
Published: July 2012

In vivo microdialysis in halothane‐anaesthetized rats and synaptosomal [3H]‐noradrenaline uptake studies in vitro were used to evaluate the effects of imidazole (medetomidine) and imidazoline (clonidine and UK 14,304) α2‐adrenoceptor agonists on extraneuronal levels of noradrenaline in the frontal cortex.

Levels of noradrenaline in the dialysate were increased by a depolarizing concentration of K+ (60 mM for 20 min) and substantially attenuated by reducing Ca2+ supply in the perfusate. These results suggest that spontaneous efflux of noradrenaline in the cortex is regulated predominantly by cation‐dependent exocytotic mechanisms.

At a low perfusion concentration (0.5 μm), medetomidine, clonidine and UK 14,304 all reduced the level of noradrenaline in cortical dialysates. Continuous perfusion of the selective α2‐adrenoceptor antagonist, atipamezole (0.5 μm) caused a sustained increase in noradrenaline efflux and reversed the inhibitory effects of medetomidine. All these changes are consistent with drug actions at presynaptic α2‐adrenoceptors.

Higher concentrations of medetomidine (5–50 μm), but not clonidine or UK 14,304, evoked a non‐desensitizing increase in noradrenaline efflux. This effect was not antagonized by 0.5 μm atipamezole.

The tricyclic noradrenaline reuptake inhibitor, desmethylimipramine (0.5–50 μm), increased noradrenaline efflux in a concentration‐dependent manner.

The specific uptake of [3H]‐noradrenaline into cortical synaptosomes was inhibited by medetomidine and desmethylimipramine with IC50 values of approximately 7μm and 8mUm respectively. Neither clonidine nor UK 14,304 inhibited [3H]‐noradrenaline uptake.

These results indicate that micromolar concentrations of the selective α2‐adrenoceptor agonist, medetomidine, can augment extraneuronal levels of noradrenaline in the rat frontal cortex; this effect seems to involve an inhibition of noradrenaline reuptake rather than an action at α2‐adrenoceptors.
Actions of the novel neuroprotective agent, lifarizine (RS‐87476), on voltage‐dependent sodium currents in the neuroblastoma cell line, N1E‐115
Authors: McGivern Joseph G. , Patmore Leslie , Sheridan Robert D.
Published: July 2012

The actions of the neuroprotective agent, lifarizine (RS‐87476‐190), on voltage‐dependent Na+ currents have been examined in the neuroblastoma cell line, N1E‐115, using the whole‐cell variant of the patch clamp technique.

At a holding potential of −80 mV, lifarizine reduced the peak Na+ current evoked by a 10 ms depolarizing step with an IC50 of 1.3μm. At holding potentials of −100 and −60mV the IC50 concentrations of lifarizine were 7.3 μm and 0.3μm, respectively.

At a holding potential of −100 mV, most channels were in the resting state and the IC50 value for inhibition of Na+ current should correspond to the dissociation constant of lifarizine for resting channels (KR). Kr was therefore estimated to be 7.3 μm.

In the absence of lifarizine, recovery from inactivation following a 20 s depolarization from −100 mV to 0 mV was complete within 2 s. However, in the presence of 3 μm lifarizine recovery took place in a biexponential fashion with time constants of 7 s and 79 s.

Lifarizine (1 μm) had no effect on steady‐state inactivation curves when conditioning pre‐pulses of 1 s duration were used. However, when pre‐pulse durations of 1 min were used the curves were shifted to the left by lifarizine by about 10 mV. Analysis of the shifts induced by a range of lifarizine concentrations revealed that the apparent affinity of lifarizine for the inactivated state of the channel (KI) was 0.19 μm.

Lifarizine (1 μm) had no effect on chloramine‐T‐modified Na+ currents, suggesting no significant open channel interaction.

Taken together, these data show that lifarizine is a potent voltage‐dependent inhibitor of Na+ currents in N1E‐115 cells and that the voltage‐dependence arises from an interaction of the compound with the inactivated state of the channel. The possible contribution of Na+ current inhibition to the neuroprotective actions of lifarizine is discussed.
Electrocardiographic interactions between pinacidil, a potassium channel opener and class I antiarrhythmic agents in guinea‐pig isolated perfused heart
Authors: Yang Quan , Padrini Roberto , Bova Sergio , Piovan Donatella , Magnolfi Gianna
Published: July 2012

Drugs that shorten action potential duration could decrease the Na‐channel blocking effect of class I antiarrhythmic agents by reducing the availability of Na channel in the inactivated state.

This hypothesis was tested in guinea‐pig perfused heart, measuring the surface ECG effects of three class I drugs endowed with different binding kinetics (15 μm mexiletine, 10 μm quinidine and 3 μm flecainide) in the presence of increasing concentrations of pinacidil (10 μm, 30 μm, 50 μm), a potassium channel opener that shortens action potential duration.

The ECG parameters measured were: the QRS interval, i.e. the intraventricular conduction time; the JT interval, which reflects the duration of ventricular repolarization; the ratio between JTpeak (the time from the end of QRS and the peak of T wave) and JT interval, which quantifies changes in the morphology of the T wave.

At the concentrations tested all the antiarrhythmic drugs widened the QRS complex by 55–60%. Flecainide did not significantly change JT interval, but quinidine prolonged and mexiletine shortened it. Mexiletine also decreased the JTpeak/JT ratio. Pinacidil by itself decreased the JT interval and the JT peak/JT ratio in a dose‐dependent way, but did not affect QRS duration.

In the presence of fixed antiarrhythmic drug concentrations, however, pinacidil decreased the QRS prolongation induced by mexiletine (−17%) and quinidine (− 8%), but not that induced by flecainide: this effect was already maximal at the lower concentration tested (10 μm) and there was no relationship between pinacidil‐induced JT shortening and QRS changes. To explain this unexpected result it has been supposed that, at the driving frequency used (4 Hz), myocardial cells were partially depolarized and that pinacidil could repolarize them, thus decreasing the number of inactivated Na channels and the effects of drugs that (mainly or partly) block the channels in the inactivated state. In agreement with this hypothesis, an additional series of experiments carried out with 15 μm mexiletine at a lower stimulation rate (2 Hz) showed only a negligible loss of QRS effect (−2.3%) at any pinacidil concentration.

Flecainide, but not quinidine and mexiletine, antagonized the JT shortening induced by pinacidil; furthermore, no drug modified the JTp/JT decrease induced by pinacidil.

These results indicate that: (a) an antagonism between class I antiarrhythmic drugs and pinacidil is possible; (b) mexiletine is the most involved among the drugs tested; (c) the interaction is not related to pinacidil‐induced repolarization shortening, but probably to changes in membrane resting potential. The possible clinical implications need to be defined.
Additive effect of ADP and CGRP in modulation of the acetylcholine receptor channel in Xenopus embryonic myocytes
Authors: Liou Jau‐Cheng , Fu Wen‐Mei
Published: July 2012

We have previously shown that the activation of either protein kinase A (PKA) or protein kinase C (PKC) enhanced the responses of muscle membrane to acetylcholine (ACh) by increasing the mean open time of embryonic‐type ACh channels in Xenopus cultured myocytes. In the present study, we further investigated the interaction between these two kinases in the modulation of ACh channels by using the receptor ligands, adenosine diphosphate (ADP) and calcitonin gene‐related peptide (CGRP) which selectively activate PKC and PKA, respectively.

ADP concentration‐dependently increased the mean open time of embryonic‐type ACh channels and 0.3 mM ADP is sufficient to achieve the maximal potentiating effect, α, β‐Methylene ATP and PMA (phorbol 12‐myristate 13‐acetate) but not adenosine, AMP, dibutyryl cyclic GMP have similar potentiating action.

Suramin (0.3 mM) pretreatment abolished the potentiating effect of ADP but left that of PMA unchanged.

CGRP increased the mean open time of embryonic‐type ACh channels in a concentration‐dependent manner and 1 μm CGRP produced the maximal effect.

The maximal effects of both ADP (0.3 mM) and CGRP (1 μm) in the prolongation of mean open time of ACh channels were additive.

These results suggest that the modulation of embryonic‐type ACh channels by the endogenously released ligands via the activation of PKA and PKC is additive and possibly different sites of ACh channels may be involved in the potentiation effect of either PKC or PKA.