The importance of potassium channels in the mechanism of the relaxing effect of pentoxifylline on isolated rat uteri

Background. Pentoxifylline is a methylxanthine derivative that is used to treat peripheral vascular disease. One of the mechanisms of action of pentoxifylline is the vasodilatation of blood vessels. This study examined the effect of increasing pentoxifylline concentrations on the contractility of isolated rat uteri in the presence of a potassium channel antagonist. Methods. The uteri were isolated from virgin Wistar rats (180-220 g) and suspended in an isolated organ bath chamber containing De Jalon's solution and aerated with 95% O2 and 5% CO2. The temperature was maintained at 37єC. Isometric contractions were recorded using an isometric force transducer (Ugo Basile). The preload of the preparation was approximately 1 g. The uteri were allowed to contract spontaneously or in the presence of Ca2+ (0.018 and 0.36 mM) and acetylcholine (ACh) and were treated with pentoxifylline. Results. Pentoxifylline caused concentration-dependent inhibition of spontaneous rhythmic uterine activity and uterine activity caused by calcium Ca2+ (0.018 mM and 0,36 mM). We showed that the inhibitory effects of pentoxifylline depend on the type of muscle contractions activated and that the inhibitory effect is significantly stronger for spontaneous rhythmic activity and forin Ca2 -induced contractions of isolated rat uteri+. The relaxing effect of pentoxifylline depends on the calcium concentration in the medium. Pentoxifylline exerted the weakest relaxant effects on contractions induced by acetylcholine. In contrast to methylene blue, tetraethylammonium, or 4-aminopyridine, glibenclamide did not antagonise the relaxing effect of pentoxifylline on isolated rat uteri. Conclusion. The results obtained suggest that the mechanism of action of pentoxifylline does not lead to the opening of KATP channels. However, the opening of BKCa and voltage-dependent Ca2+ channels had some role, but to varying degrees, in the mechanism of the relaxing effect of pentoxifylline on the spontaneous rhythmic activity and calcium-induced contractions of isolated rat uteri. Our results provide additional confirmation of the dominance of the NO/cGMP signaling pathway in the mechanism of the relaxing effect of pentoxifylline (because the presence of methylene blue significantly antagonised this effect) in relation to the opening of potassium channels, especially KATP channels. These results indicate that pentoxifylline could be a potential tocolytic drug.Cilj. Pentoksifilin, koji se koristi za lečenje perifernih vaskularnih oboljenja, je derivat metilksantina. Jedan od načina delovanja pentoksifilina je prouzrokovanje vazodilatacije krvnih sudova. U ovom radu ispitivali smo efekat rastućih koncentracija pentoksifilina na kontraktilnost izolovanog uterusa pacova, u prisustvu blokatora kalijumskih kanala. Metode. Uterusi, koji su izolovani od neparenih ženki pacova Wistar soja (180-220 g), držani su u kupatilu za izolovane organe na temperaturi od 37oC, u De Jalon-ovom rastvoru kroz koji je propuštana smeša gasova od 95% kiseonika i 5% ugljendioksida. Izometrijske kontrakcije su registrovane korišćenjem izometrijskog transdjusera Ugo Basile, pri opterećenju preparata od 1 g. Ispitivan je efekat pentoksifilina na kontrakcije za vreme spontane ritmičke aktivnosti i u prisustvu kalcijuma, Ca2+ (0.018 and 0.36 mM)) i acetilholina (ACh). Rezultati. Pentoksifilin je prouzrokovao koncentracijski zavisnu inhibiciju spontane ritmičke aktivnosti, kao i fazne aktivnosti prouzrokovane kalcijumom. Inhibicijski efekt pentoksifilina zavisio je od tipa aktivacije glatkog mišića uterusa. On je ispoljio značajno jači relaksirajući efekt na spontanu ritmičku aktivnost i kontrakcije prouzrokovane sa 0.018 mM kalcijuma. Njegov relaksirajući efekt zavisi i od koncentracije Ca2+ u medijumu..Najslabiji relaksirajući efekat pentoksifilina je zabeležen na acetilholinskom tipu aktivacije. Nasuprot metilenskom plavilu, 4-aminopiridinu i tetraetilamonijumu, glibenklamid ne antagonizuje relaksirajući efekat pentoksifilina na izolovanom uterusu pacova. Zaključak. Dobijeni rezultati sugerišu da u mehanizmu relaksantnog delovanja pentoksifilina nije zastupljeno otvaranje ATP kalijumskih kanala. Međutim, otvaranje BKCa i voltažno zavisnih Ca2+ kalijumskih kanala ima izvestan značaj, ali u različitom stepenu, u mehanizmu relaksirajućeg delovanja pentoksifilina na spontanu ritmičku aktivnost i kontrakcije prouzrokovane kalcijumom. Naši rezultati su dodana potvrda o dominaciji NO/cGMP signalinih puteva kojima pentoksifilin prouzrokuje relaksaciju glatkih mišičnih ćelija uterusa (jer metilensko plavilo značajno antagonizuje njegov efekt), u odnosu na otvaranje kalijumskih kanala, posebno ATP zavisnih kalijumskih kanala. Rezultati ukazuju da bi pentoksifilin mogao da bude potencijalni tokolitički lek.nul

Efekat MnSOD (E. coli) Na relaksaciju izolovane renalne arterije pacova izazvanu natrijum-nitro-prusidom

In this study themolecular foundation of nitric oxide induced relaxation of arteries, with or without endothelium, of normotensive and spontanously hypertensive ratswas re-examined. With this purpose in mind, the effects of the nitric oxide donor sodium nitroprusside (NaNP), with and without manganese containing superoxide dismutase (MnSOD E.C. 1.15.1.1), on rat renal artery relaxation was strudied. The results show that the relaxation effect of NaNP is two times higher in normotensive, compared to spontaneously hypertensive rats. Similar differences exist in the relaxation effects of NaNP on isolated renal arteries without endothelium, indicating that besides the difference in the function of an endothelium, concerning basal NO production in normotensive and hypertensive rats, there is a differencewith respect to NO relaxation in the smoothmuscle that is induced by hypertension. MnSOD decreased the relaxation effect of NaNP in all the examined renal arteries, more in normotensive than in hypertensive ones regardless of the presence of an endothelium. These results show that MnSOD, by modifying the chemical versatility of NO into redox active forms - nitrosonium (NO+) and nitroxyl (NO-), produces different relaxation effects in normotensive and hypertensive arteries of rats, with or without an endothelium, potentiating the role of nitroxyl induced relaxation in sponteneously hypertensive rats. The results prove the need for the synthesis of complex NO donors, as the mechanisms of artery relaxation are different due to an endothel and smooth mouscle changes in hypertensive, as compared to normotensive rats.U ovom radu smo pokušali da detaljnije ispitamo molekulsku osnovu azotoksid indukovane relaksacije arterija, sa i bez endotela, normotenzivnih i spontano hipertenzivnih pacova. U tu svrhu ispitivan je efekat azot-oksid (NO) donora natrijum- nitroprusida (NaNP), bez i u prisustvu superoksid-dismutaze koja sadrži mangan (MnSOD, EC 1.15.1.1) na relaksaciju renalne arterije. Rezulati pokazuju dvostruko veći relaksantni efekat NaNP kod normotenzivnih, u odnosu na spontano hipertenzivne pacove. Slična razlika postoji i u relaksantnom efektu NaNP na izolovane renalne arterije normotenzivnog i hipertenzivnog pacova kada je odstranjen endotel što ukazuje da, pored razlike u funciji endotela u odnosu na bazalnu NO produkciju kod normotenzivnih i spontano hipertenzivnih pacova, postoje i promene u glatkim mišićima indukovane hipertenzijom, u odnosu na NO relaksaciju. MnSOD kod svih grupa smanjuje relaksantni efekat SNP i to više kod normotenzivnih nego kod hipertenzivnih nezavisno od prisustva endotela. Ovi rezultati pokazuju da MnSOD menjajući hemijsku prirodu iz NaNP oslobođenog NO u redoks aktivne forme – nitrozonijum (NO+) i nitroksil (NO–) – ostvaruje različit relaksantni efekat kod normotenzivnih i kod hiperetenzivnih pacova sa i bez endotela potencirajući značaj nitroksilom indukovane relaksacije kod spontano-hipertenzivnih pacova. Dobijeni rezultati potvrđuju potrebu sinteze NO donora koji daju različite redoks-aktivne forme NO budući da se primarni mehanismi relaksacije razlikuju u zavisnosti od karakteristika endotela i glatkih mišića, kod hipertenzije u odnosu na normotenzivno stanje

Effect of MnSOD (E. coli) on the relaxation caused by sodium nitroprusside on isolated rat renal artery

In this study themolecular foundation of nitric oxide induced relaxation of arteries, with or without endothelium, of normotensive and spontanously hypertensive ratswas re-examined. With this purpose in mind, the effects of the nitric oxide donor sodium nitroprusside (NaNP), with and without manganese containing superoxide dismutase (MnSOD E.C. 1.15.1.1), on rat renal artery relaxation was strudied. The results show that the relaxation effect of NaNP is two times higher in normotensive, compared to spontaneously hypertensive rats. Similar differences exist in the relaxation effects of NaNP on isolated renal arteries without endothelium, indicating that besides the difference in the function of an endothelium, concerning basal NO production in normotensive and hypertensive rats, there is a differencewith respect to NO relaxation in the smoothmuscle that is induced by hypertension. MnSOD decreased the relaxation effect of NaNP in all the examined renal arteries, more in normotensive than in hypertensive ones regardless of the presence of an endothelium. These results show that MnSOD, by modifying the chemical versatility of NO into redox active forms - nitrosonium (NO+) and nitroxyl (NO-), produces different relaxation effects in normotensive and hypertensive arteries of rats, with or without an endothelium, potentiating the role of nitroxyl induced relaxation in sponteneously hypertensive rats. The results prove the need for the synthesis of complex NO donors, as the mechanisms of artery relaxation are different due to an endothel and smooth mouscle changes in hypertensive, as compared to normotensive rats.U ovom radu smo pokušali da detaljnije ispitamo molekulsku osnovu azotoksid indukovane relaksacije arterija, sa i bez endotela, normotenzivnih i spontano hipertenzivnih pacova. U tu svrhu ispitivan je efekat azot-oksid (NO) donora natrijum- nitroprusida (NaNP), bez i u prisustvu superoksid-dismutaze koja sadrži mangan (MnSOD, EC 1.15.1.1) na relaksaciju renalne arterije. Rezulati pokazuju dvostruko veći relaksantni efekat NaNP kod normotenzivnih, u odnosu na spontano hipertenzivne pacove. Slična razlika postoji i u relaksantnom efektu NaNP na izolovane renalne arterije normotenzivnog i hipertenzivnog pacova kada je odstranjen endotel što ukazuje da, pored razlike u funciji endotela u odnosu na bazalnu NO produkciju kod normotenzivnih i spontano hipertenzivnih pacova, postoje i promene u glatkim mišićima indukovane hipertenzijom, u odnosu na NO relaksaciju. MnSOD kod svih grupa smanjuje relaksantni efekat SNP i to više kod normotenzivnih nego kod hipertenzivnih nezavisno od prisustva endotela. Ovi rezultati pokazuju da MnSOD menjajući hemijsku prirodu iz NaNP oslobođenog NO u redoks aktivne forme – nitrozonijum (NO+) i nitroksil (NO–) – ostvaruje različit relaksantni efekat kod normotenzivnih i kod hiperetenzivnih pacova sa i bez endotela potencirajući značaj nitroksilom indukovane relaksacije kod spontano-hipertenzivnih pacova. Dobijeni rezultati potvrđuju potrebu sinteze NO donora koji daju različite redoks-aktivne forme NO budući da se primarni mehanismi relaksacije razlikuju u zavisnosti od karakteristika endotela i glatkih mišića, kod hipertenzije u odnosu na normotenzivno stanje.Projekat ministarstva br. 166

POPULATION PHARMACOKINETICS OF 2-OXO-CLOPIDOGREL IN PATIENTS WITH ACUTE CORONARY SYNDROME

The aim of the study was to develop a population pharmacokinetic (PK) model for clearance of 2-oxo-clopidogrel in patients with acute coronary syndrome (ACS). Population pharmacokinetic analysis was performed by using 72 plasma concentrations from the same number of patients (mean age of 60.82±10.76 years; total body weight (TBW) of 73.63±9.67 kg) with ACS using non-linear mixed-effect modeling (NONMEM). Validation of the final PPK model was carried out through the bootstrap analysis with 200 runs and it was used to estimate the predictive performance of the pharmacokinetic model. The typical mean value for 2-oxo-clopidogrel clearance (CL), estimated by the base model (without covariates), in our population was 39.2 l h−1.The value of aspartate transaminase and co-medication with digoxin were determinants of a derived population model. The final regression model for the clearance of 2-oxo-clopidogrel was the following: CL (lh-1) = 1.7 + 1.31*AST + 115*DIGOXIN. The derived PK model describes the clearance of 2-oxo-clopidogrel in patients with ACS, showing that the value of aspartate transaminase and co-medication with digoxin are the most important covariate. This finding will provide the basis for future PK studies

The role of nitric oxide (NO) in gastrointestinal tract (GIT)

NO je veoma bitan inhibitorni neurotransmiter u GIT-u koji se oslobađa iz neadrenergičkih i neholinergičkih neurona. NO deluje kao unutar i ekstraćelijski signalni molekul u vaskularnim i glatkim mišićnim ćelijama GIT-a, i važan je medijator u brojnim fiziološkim i patofiziološkim stanjima. NO se sintetiše iz L-arginina pod dejstvom enzima azot monoksid sintetaze (NOS). NOS je lokalizovana u nitrergičkim neuronima u GIT-u, ali može se sintetisati i u drugim ćelijskim tipovima, kao što su glatke mišićne ćelije. Najšire prihvaćeno delovanje NO-a u crevu je relaksacija glatkih mišića putem aktivacije solubilne forme guanilil ciklaze i akumulacije cikličnog guanozin monofosfata (cGMP). U glatkim mišićima GIT-a, NO i donori NO izazivaju različite efekte: kontrakciju, relaksaciju praćenu kontrakcijom i kontrakciju praćenu relaksacijom, što zavisi od tipa NO donora, tkiva i vrste. Ekscitatorni efekt NO-a obuhvata holinergičke motorne neurone. Višestruko delovanje NO-a sugeriše da se manipulacijom NO sistema može uticati na enteričke motorne reflekse, kao i na peristaltiku tankog creva. Visoke koncentracije NO-a mogu oštetiti intestinalne epitelne ćelije, za šta je odgovoran cGMP. Odsustvo nitrergičke inervacije je pokazano u gastrointestinalnim tkivima pacijenata sa različitim bolestima, kao što su Ahalazija i Hirschsprungsova bolest.NO is a very important inhibitory neurotransmitter in the GIT, released from non-adrenergic non-cholinergic neurons. NO acts as an intra- and extracellular signalling molecule in vascular and smooth muscle GIT cells and it is an important mediator in numerous physiological and pathophysiological conditions. NO is synthesized from L-arginine by the action of NO synthase (NOS) that is present in nitrergic neurons of GIT. NO could be also synthesized in other cell types such as smooth muscle cells. The most widely reported action of NO in the gut is relaxation of smooth muscle through activation of the soluble guanylate cyclase and the accumulation of the cyclic guanosine 3’, 5’-monophosphate (cGMP). In GIT smooth muscle cells, NO and NO donors evoke different responses including contractile effects, relaxations followed by contractions or contractions followed by relaxations, which depends on the type of NO donor, tissue and species. The excitatory effect of NO involves cholinergic motor neurons. Multiple actions of NO suggest that manipulation of the NO system may influence enteric motor reflexes and peristalsis of small intestine. Large amounts of NO can damage the intestine epithelial cells, and cGMP is responsible for this damage. Deficiency of nitrergic innervations has been shown in gastrointestinal tissues of patients with various diseases such as are Achalasia and Hirschsprung’s disease.Available at: [http://www.tmg.org.rs/v320107.htm

Sodium nitroprusside regulates the relaxation of the longitudinal muscle in the gut

Nitric oxide (NO) has been shown to mediate nonadrenergic-noncholinergic relaxation in gastrointestinal (GI) smooth muscle cells. As GI smooth muscles relaxations are partly dependent on NO, we decided to investigate the effect of sodium nitroprusside (SNP) on the longitudinal muscle contraction of the isolated guinea pig ileum. Increasing concentrations of SNP (10(-10)M, 10(-9)M, 10(-8)M, 10(-7)M, 10(-6)M and 10(-5)M) reduced ileum contractions stimulated by electrical stimulation (ES) (8-76%; p < 0.05) and by acetylcholine (Ach) (23-62%; p < 0.05) significantly and in a concentration-dependent manner. Furthermore, treatment with an inhibitor of the soluble guanylate cyclase, methylene blue (10 mM), antagonized significantly the relaxing effect of SNP (0-39%; p < 0.05, p < 0.01, p < 0.001 for ES- and 4-27%; p < 0.05 for Ach-induced contractions). The results show that treatment with 1 mu M manganese-containing superoxide dismutase (MnSOD) and 10 mu M L-arginine (L-arg) caused a significant decrease in SNP induced relaxations (6-55%; p < 0.05, p < 0.001 and 2-46%; p < 0.05, p < 0.01 for ES- and 15-28%; p < 0.05, p < 0.01, p < 0.001 and 12-32%; p < 0.05, p < 0.01 for Ach-induced contractions, respectively). In conclusion, our data suggest that SNP, which releases NO, is able to depress longitudinal muscle contraction of the isolated guinea pig ileum, suggesting that exogenous application of NO inhibits intestinal contractions of smooth muscle cells and that cGMP mediates the response to NO. In addition, MnSOD and L-arg decreased the relaxing effect of SNP on the isolated ileum of the guinea pig.nul