bluuebunny/crossref_metadata_2025_split huggingface dataset:
Evidence that spontaneous contractile activity in the rat myometrium is not inhibited by NO‐mediated increases in tissue levels of cyclic GMP
Authors: Hennan James K. , Diamond Jack 
Published: January 2005

There is conflicting evidence in the literature concerning the role of cyclic GMP in the regulation of myometrial contractility and the importance of hormonal status on the uterine response to cyclic GMP‐elevating agents. The objective of the present study was to investigate further the importance of cyclic GMP in the control of uterine contractility, by monitoring the effects of cyclic GMP‐elevating agents on spontaneous contractions and cyclic GMP levels in myometrial strips from pregnant rats and from ovariectomized rats under the influence of oestrogen and/or progesterone.
Sodium nitroprusside (SNP) 5 mM, atrial natriuretic peptide (ANP) 100 nM, L‐arginine 1 mM and 8‐bromo‐cyclic GMP 100 mM had no relaxant effect on the spontaneous contractions of myometria from pregnant rats or from ovariectomized rats under the influence of oestrogen or progesterone.
Tissue levels of cyclic GMP were significantly elevated by SNP in all treatment groups, including pregnant animals. For example, in ovariectomized, progesterone‐treated rats, SNP raised cyclic GMP levels approximately 8 fold from a basal level of 2.9±0.4 pmol mg−1 protein to 24.8±4.0 pmol mg−1 protein. ANP increased cyclic GMP levels approximately 2 fold in all treatment groups, except in the pregnant animals. L‐Arginine elevated cyclic GMP significantly only in ovariectomized, vehicle‐treated myometria.
The activity of cyclic GMP‐dependent protein kinase (PKG) was significantly increased (3 fold) in myometria exposed to SNP (5 mM). Thus, the inability of SNP to relax uterine preparations was not due to a failure of SNP‐elevated cyclic GMP to activate PKG.
The more potent NO donor, S‐nitroso‐N‐acetylpenicillamine (SNAP), at a concentration of 100 μM was able to inhibit spontaneous contractions significantly in myometrial preparations from both non‐ovariectomized and ovariectomized rats treated with oestrogen or progesterone.
Tissue levels of cyclic GMP were markedly increased by SNAP at concentrations of 10, 30 and 100 μM. At 100 μM, cyclic GMP levels increased from 1.9±0.2 pmol mg−1 protein to 74.0±18.0 pmol mg−1 protein. However, complete or partial blockade of SNAP‐induced increases in cyclic GMP levels by the soluble guanylyl cyclase inhibitor, ODQ (25 μM), had no effect on the relaxant response to SNAP. Thus, the relaxant effect of SNAP in this tissue appears to be mediated via a mechanism independent of cyclic GMP.
Taken as a whole, the results of the present study indicate that cyclic GMP does not play an important role in the control of contractility in the rat uterus.

British Journal of Pharmacology (1998) 123, 959–967; doi:10.1038/sj.bjp.0701678
Opposite effects of nitric oxide donors on DNA single strand breakage and cytotoxicity caused by tert‐butylhydroperoxide
Authors: Guidarelli Andrea , Sestili Piero , Cantoni Orazio 
Published: January 2005

The effects of three different NO donors on tert‐butylhydroperoxide (tB‐OOH)‐induced DNA cleavage and toxicity were investigated in U937 cells.
Treatment with S‐nitroso‐N‐acetyl‐penicillamine (SNAP, 1–30 μm), while not in itself DNA‐damaging, potentiated the DNA strand scission induced by 200 μm tB‐OOH in a concentration‐dependent fashion. The enhancing effects of SNAP were observed with two different techniques for the assessment of DNA damage. Decomposed SNAP was inactive. S‐nitrosoglutathione (GSNO, 300 μm) and (Z)‐1‐[(2‐aminoethyl)‐N‐(2‐ammonioethyl) amino]diazen‐1‐ium‐1,2‐diolate (DETA‐NO, 1 mm) also increased DNA cleavage generated by tB‐OOH and these responses, as well as that mediated by SNAP, were prevented by the NO scavenger 2‐phenyl‐4,4,5,5‐tetramethylimidazolin‐1‐oxyl‐3‐oxide (PTIO).
SNAP neither inhibited catalase activity nor increased the formation of DNA lesions in cells exposed to H2O2. Furthermore, SNAP did not affect the rate of rejoining of the DNA single strand breaks generated by tB‐OOH.
Under the conditions utilized in the DNA damage experiments, treatment with tB‐OOH alone or associated with SNAP did not cause cell death. However, SNAP as well as GSNO markedly reduced the lethal response promoted by millimolar concentrations of tB‐OOH and these effects were abolished by PTIO. Decomposed SNAP was inactive.
It is concluded that low levels of NO donors, which probably release physiological concentrations of NO, enhance the accumulation of DNA single strand breaks in U937 cells exposed to tB‐OOH. This NO‐mediated effect appears to (a) not depend on inhibition of either DNA repair (which would increase the net accumulation of DNA lesions by preventing DNA single strand break removal) or catalase activity (which would also enhance the net accumulation of DNA lesions since H2O2 is one of the species mediating the tB‐OOH‐induced DNA cleavage) and (b) be caused by enforced formation of tB‐OOH‐derived DNA‐damaging species. In contrast to these results, similar concentrations of NO prevented cell death caused by millimolar concentrations of tB‐OOH. Hence, DNA single strand breakage generated by tB‐OOH in the absence or presence of NO does not represent a lethal event.

British Journal of Pharmacology (1998) 123, 1311–1316; doi:10.1038/sj.bjp.0701683
Mechanisms of relaxations of bovine isolated bronchioles by the nitric oxide donor, GEA 3175
Authors: Hernández Medardo , Elmedal Britt , Mulvany Michael J. , Simonsen Ulf 
Published: January 2005

The present study was designed to investigate the effects and mechanisms of relaxation induced by the nitric oxide (NO) donor, GEA 3175 (a 3‐aryl‐substituted oxatriazole derivative) on bovine bronchioles (effective lumen diameter 200–800 μm) suspended in microvascular myographs for isometric tension recording.
In segments of bovine bronchioles contracted to 5‐hydroxytryptamine, GEA 3175 (10−8–10−4 M) induced concentration‐dependent reproducible relaxations. These relaxations were slow in onset compared to other NO‐donors such as 3‐morpholinosydonimine‐hydrochloride (SIN‐1) and S‐nitroso‐N‐acetylpenicillamine (SNAP).
In 5‐hydroxytryptamine‐contracted preparations the order of relaxant potency (pD2) was: salbutamol (7.80)>GEA 3175 (6.18)>SIN‐1 (4.90)>SNAP (3.55). In segments contracted to acetylcholine, the relaxant responses were reduced and GEA 3175 relaxed the bronchioles with pD2=4.41±0.12 and relaxations of 66±10% (n=4), while SNAP and salbutamol caused relaxations of 19±6% (n=4) and 27±6% (n=8) at the highest concentration used, respectively.
Oxyhaemoglobin (10−5 M), the scavenger of nitric oxide, caused rightward shifts of the concentration‐relaxation curves to GEA 3175 and NO. 1H‐[1,2,4]oxadiazolo[4,3,‐a]quinoxalin‐1‐one (ODQ, 3×10−6 M) and LY 83583 (10−6 M), the inhibitors of soluble guanylate cyclase, also reduced the relaxations induced by GEA 3175 and nitric oxide. However, ODQ did not affect salbutamol‐evoked relaxation in the bovine small bronchioles.
GEA 3175‐induced relaxations were reduced in potassium‐rich (60 mmol l−1 K+) solution. Glibenclamide (10−6 M) markedly inhibited the relaxations induced by the opener of ATP‐sensitive K+ channels, levcromakalim (3×10−8–10−5 M), but it did not modify the relaxations induced by GEA 3175 or salbutamol. Apamin (5×10−7 M), a blocker of the small Ca2+‐activated K+‐channels did not affect the relaxations to GEA 3175. In contrast, blockers of large Ca2+‐activated K+‐channels, charybdotoxin (3×10−8–10−7 M) and iberiotoxin (10−8 M), did inhibit the relaxations to GEA 3175. The combination of apamin and charybdotoxin did not induce an additional inhibitory effect on the relaxations to GEA 3175 compared to charybdotoxin alone.
In preparations where a concentration‐response curve to GEA 3175 or NO was first obtained in the presence of LY 83583, incubation with charybdotoxin (10−7 M) did produce an additional inhibitory effect of the relaxations. However, in the presence of ODQ (3×10−6 M), iberiotoxin (10−8 M) did not produce additional reduction of the NO‐ or GEA 3175‐induced relaxations.
The present results suggest that the slow‐releasing NO‐donor GEA 3175 is more potent than the traditional NO donors in inducing relaxations of bovine bronchioles. GEA 3175, as for exogenously added NO, elicits relaxations through a cyclic GMP‐dependent mechanism followed by opening of large conductance Ca2+‐activated K+‐channels.

British Journal of Pharmacology (1998) 123, 895–905; doi:10.1038/sj.bjp.0701684
Interaction of N‐benzoyl‐D‐phenylalanine and related compounds with the sulphonylurea receptor of β‐cells
Authors: Schwanstecher Christina , Meyer Miriam , Schwanstecher Mathias , Panten Uwe 
Published: January 2005

The structure activity relationships for the insulin secretagogues N‐benzoyl‐D‐phenylalanine (NBDP) and related compounds were examined at the sulphonylurea receptor level by use of cultured HIT‐T15 and mouse pancreatic β‐cells. The affinities of these compounds for the sulphonylurea receptor were compared with their potencies for KATP‐channel inhibition. In addition, the effects of cytosolic nucleotides on KATP‐channel inhibition by NBDP were investigated.
NBDP displayed a dissociation constant for binding to the sulphonylurea receptor (KD value) of 11 μM and half‐maximally effective concentrations of KATP‐channel inhibition (EC50 values) between 2 and 4 μM (in the absence of cytosolic nucleotides or presence of 0.1 mM GDP or 1 mM ADP).
In the absence of cytosolic nucleotides or presence of GDP (0.1 mM) maximally effective concentrations of NBDP (0.1–1 mM) reduced KATP‐channel activity to 47% and 44% of control, respectively. In the presence of ADP (1 mM), KATP‐channel activity was completely suppressed by 0.1 mM NBDP.
The L‐isomer of N‐benzoyl‐phenylalanine displayed a 20 fold lower affinity and an 80 fold lower potency than the D‐isomer.
Introduction of a p‐nitro substituent in the D‐phenylalanine moiety of NBDP did not decrease lipophilicity but lowered affinity and potency by more than 30 fold.
Introduction of a p‐amino substituent in the D‐phenylalanine moiety of NBDP (N‐benzoyl‐p‐amino‐D‐phenylalanine, NBADP) reduced lipophilicity and lowered affinity and potency by about 10 fold. This loss of affinity and potency was compensated for by formation of the phenylpropionic acid derivative of NBADP. A similar difference in affinity was observed for the sulphonylurea carbutamide and its phenylpropionic acid derivative.
Replacing the benzene ring in the D‐phenylalanine moiety of NBDP by a cyclohexyl ring increased lipophilicity, and the KD and EC50 values were slightly lower than for NBDP. Exchange of both benzene rings in NBDP by cyclohexyl rings further increased lipophilicity without altering affinity and potency.
This study shows that N‐acylphenylalanines interact with the sulphonylurea receptor of pancreatic β‐cells in a stereospecific manner. Their potency depends on lipophilic but not aromatic properties of their benzene rings. As observed for sulphonylureas, interaction of N‐acylphenylalanines with the sulphonylurea receptor does not induce complete inhibition of KATP‐channel activity in the absence of inhibitory cytosolic nucleotides.

British Journal of Pharmacology (1998) 123, 1023–1030; doi:10.1038/sj.bjp.0701686
Induction of nitric oxide synthase by protein synthesis inhibition in aortic smooth muscle cells
Authors: Marczin Nándor , Go Carolyn Y. , Papapetropoulos Andreas , Catravas John D. 
Published: January 2005

The role of de novo protein synthesis in inducible NO synthase (iNOS) activation was investigated in vitro by evaluating the effects of protein synthesis inhibitors cycloheximide (CH) and anisomycin (ANI) on iNOS activity, protein and mRNA levels in rat aortic smooth muscle cells (RASMC).
As determined by cyclic GMP accumulation, substrate (L‐arginine)‐ and inhibitor (NG‐monomethyl‐L‐arginine, NMMA)‐sensitive iNOS activity was significantly elevated in CH‐ or ANI‐treated RASMC after 24 h.
Lipopolysaccharide (LPS) produced a time‐dependent increase in cyclic GMP levels with maximal stimulation at 6 h and a decline to near baseline at 24 h. CH attenuated LPS‐induced cyclic GMP accumulation at 3 and 6 h. However, cyclic GMP levels were superinduced at later times by CH. The concentration‐dependence of cyclic GMP stimulation by cycloheximide was biphasic both in the absence and presence of LPS, with maximal stimulation at 10 μM and inhibition at higher concentrations.
Increased iNOS activity by CH was associated with elevated levels of immunoreactive iNOS protein as judged by Western blotting in LPS‐ and CH‐treated cells.
CH‐induced iNOS activity and superinduction of iNOS by CH in cells treated with LPS were both significantly inhibited by actinomycin D, a transcription inhibitor.
RT‐PCR revealed elevated iNOS mRNA levels after 12 h of exposure to CH. The combination of LPS and CH caused a significant increase in iNOS gene expression relative to LPS‐ or CH stimulation alone.
These results show that partial protein synthesis inhibition by CH alone upregulates iNOS mRNA and superinduces iNOS mRNA in cytokine‐treated RASMC, which is translated to the functional enzyme generating biologically active NO. Thus iNOS activation in these cells not only requires new protein synthesis but it also appears to be negatively regulated by newly synthesized proteins.

British Journal of Pharmacology (1998) 123, 1000–1008; doi:10.1038/sj.bjp.0701687
Phosphodiesterase profile of human B lymphocytes from normal and atopic donors and the effects of PDE inhibition on B cell proliferation
Authors: Gantner Florian , Götz Christine , Gekeler Volker , Schudt Christian , Wendel Albrecht , Hatzelmann Armin 
Published: January 2005

CD19+ B lymphocytes were purified from the peripheral blood of normal and atopic subjects to analyse and compare the phosphodiesterase (PDE) activity profile, PDE mRNA expression and the importance of PDE activity for the regulation of B cell function.
The majority of cyclic AMP hydrolyzing activity of human B cells was cytosolic PDE4, followed by cytosolic PDE7‐like activity; marginal PDE3 activity was found only in the particulate B cell fraction. PDE1, PDE2 and PDE5 activities were not detected.
By cDNA‐PCR analysis mRNA of the PDE4 subtypes A, B (splice variant PDE4B2) and D were detected. In addition, a weak signal for PDE3A was found.
No differences in PDE activities or mRNA expression of PDE subtypes were found in B cells from either normal or atopic subjects.
Stimulation of B lymphocytes with the polyclonal stimulus lipopolysaccharide (LPS) induced a proliferative response in a time‐ and concentration‐dependent manner, which was increased in the presence of interleukin‐4 (IL‐4). PDE4 inhibitors (rolipram, piclamilast) led to an increase in the cellular cyclic AMP concentration and to an augmentation of proliferation, whereas a PDE3 inhibitor (motapizone) was ineffective, which is in accordance with the PDE profile found. The proliferation enhancing effect of the PDE4 inhibitors was partly mimicked by the cyclic AMP analogues dibutyryl (db) cyclic AMP and 5,6‐dichloro‐1‐β‐D‐ribofuranosylbenzimidazole‐3′,5′‐cyclic monophosphorothioate, Sp‐isomer (dcl‐cBIMPS), respectively. However, at concentrations exceeding 100 μM db‐cyclic AMP suppressed B lymphocyte proliferation, probably as a result of cytotoxicity. Prostaglandin E2 (PGE2, 1 μM) and forskolin (10 μM) did not affect B cell proliferation, even when given in combination with rolipram.
Inhibition of protein kinase A (PKA) by differentially acting selective inhibitors (KT 5720, Rp‐8‐Br‐cyclic AMPS) decreased the proliferative response of control cells and reversed the proliferation enhancing effects of rolipram.
Importantly, PDE4 activity in LPS/IL‐4‐activated B lymphocytes decreased by about 50% compared to unstimulated control values.
We conclude that an increase in cyclic AMP, mediated by down‐regulation of PDE4 activity, is involved in the stimulation of B cell proliferation in response to LPS/IL‐4. B cell proliferation in response to a mitogenic stimulus can be further enhanced by pharmacological elevation of cyclic AMP.

British Journal of Pharmacology (1998) 123, 1031–1038; doi:10.1038/sj.bjp.0701688
Possible mechanism of the negative inotropic effect of α1‐adrenoceptor agonists in rat isolated left atria after exposure to free radicals
Authors: Peters Stephan L M. , Batink Harry D. , Michel Martin C. , Pfaffendorf Martin , van Zwieten Pieter A. 
Published: January 2005

This study was designed to investigate the mechanism(s) of the negative inotropic effects of α1‐adrenoceptor agonists observed in rat isolated left atria after exposure to free radicals.
Ouabain and calphostin C were used in contraction experiments to block the sodium pump and protein kinase C. Methoxamine‐induced phospholipase C and Na+/K+ ATPase activities were measured.
Methoxamine (300 μM) increased contractile force by 1.6±0.2 mN in control atria but decreased contractile force in electrolysis‐treated atria by 2.0±0.1 mN (P<0.05), as determined 10 min after methoxamine addition. In contrast, the positive inotropic effects of endothelin‐1 (30 nM) and isoprenaline (10 μM) were reduced from 2.6±0.3 to 1.3±0.1 mN and from 2.6±0.3 to 1.7±0.2 mN, respectively, by electrolysis treatment (P<0.05), but not converted into a negative inotropic action.
In an inositol phosphate assay we observed that the stimulation of phospholipase C by methoxamine was attenuated by electrolysis when the (electrolyzed) medium from the organ bath was used, but the phospholipase C responses were restored by the use of fresh medium. However, fresh medium did not counteract the negative inotropic effect of methoxamine. Accordingly, the negative inotropic effect of methoxamine is not directly related to the impaired phospholipase C responses seen in atria subjected to electrolysis.
Ouabain (10 μM) and the protein kinase C inhibitor calphostin C (50 nM), completely prevented the negative inotropic effect of 300 μM methoxamine in electrolysis‐treated atria.
Measurement of the Na+/K+ ATPase activity, revealed that in control atria, α1‐adrenoceptor stimulation with 300 μM methoxamine, decreased the Na+/K+ ATPase activity by 14.4±7.7%. In contrast, methoxamine increased the Na+/K+ ATPase activity by 48.8±8.9% (P<0.05) in electrolysis‐treated atria. Interestingly, this increase in Na+/K+ ATPase activity was completely counteracted by calphostin C (1.4±0.1% over basal).
These results indicate that the negative inotropic effects of α1‐adrenoceptor agonists, observed in rat isolated left atria exposed to free radicals, are likely to be caused by protein kinase C‐mediated phosphorylation and subsequent activation of the Na+/K+ ATPase.

British Journal of Pharmacology (1998) 123, 952–958; doi:10.1038/sj.bjp.0701689
S‐nitrosothiols and the nitrergic neurotransmitter in the rat gastric fundus: effect of antioxidants and metal chelation
Authors: De Man Joris G. , De Winter Benedicte Y. , Moreels Tom G. , Herman Arnold G. , Pelckmans Paul A. 
Published: January 2005

The effects of the antioxidants ascorbic acid and α‐tocopherol and of the metal chelator ethylenediaminetetraacetic acid (EDTA) were studied on relaxations in response to S‐nitrosothiols, authentic nitric oxide (NO) and nitrergic non‐adrenergic non‐cholinergic stimulation of the rat gastric fundus.
The S‐nitrosothiols S‐nitrosocysteine (1–100 nM), S‐nitrosoglutathione (0.01–3 μM) and S‐nitroso‐N‐acetylpenicillamine (0.01–3 μM) induced concentration‐dependent relaxations of the rat gastric fundus muscle strips, which were precontracted with prostaglandin F2α. The relaxations to all S‐nitrosothiols were concentration‐dependently enhanced by the antioxidants ascorbic acid (0.1–3 μM) and α‐tocopherol (3–30 μM) and inhibited by the metal chelator EDTA (26 μM).
Ascorbic acid and α‐tocopherol alone did not induce a relaxation of the precontracted rat gastric fundus muscle strip. However, when ascorbic acid (1 μM) or α‐tocopherol (1 μM) were injected in the organ bath 1 minute after S‐nitrosoglutathione (0.1 μM) or after S‐nitroso‐N‐acetylpenicillamine (0.1 μM), they induced an immediate, sharp and transient relaxation. This relaxation was inhibited by the superoxide generator pyrogallol (2 μM). Such a relaxation to ascorbic acid or α‐tocopherol was not observed in the presence of S‐nitrosocysteine (10 nM).
Electrical field stimulation (0.5–4 Hz) of the precontracted rat gastric fundus strips induced frequency‐dependent nitrergic relaxations which were mimicked by authentic NO (3–300 nM) and by acidified sodium nitrite NaNO2 (0.3–10 μM). Ascorbic acid (0.3–3 μM), α‐tocopherol (3–30 μM) or EDTA (26 μM) did not affect the relaxations to nitrergic stimulation, NO or NaNO2.
In summary, relaxations to S‐nitrosothiols in the rat gastric fundus are enhanced by the antioxidants ascorbic acid and α‐tocopherol and inhibited by the metal chelator EDTA. However, relaxations to nitrergic stimulation of the rat gastric fundus or those to authentic NO were not affected by the antioxidants or by the metal chelator. These results indicate that antioxidants and metal chelators have a different effect on the biological activity of S‐nitrosothiols and on that of the nitrergic neurotransmitter. Therefore, our results suggest that S‐nitrosothiols do not act as intermediate compounds in nitrergic neurotransmission in the rat gastric fundus.

British Journal of Pharmacology (1998) 123, 1039–1046; doi:10.1038/sj.bjp.0701692