V. Cupic

Immunomodulatory effect of xylazine, an α2 adrenergic agonist, on rat spleen cells in culture

Abstract Xylazine is an adrenergic α2 agonist, which is used in veterinary medicine as a sedative and anesthetic agent. In this work we found that xylazine administered in vivo at a dose of 2.5 mg/kg enhanced spleen cell proliferation and interleukin 2 (IL-2) production in cultures stimulated with concanavalin A (Con A), whereas doses of 10 and 25 mg/kg were inhibitory. A similar stimulatory (10 μM) and inhibitory (50–500 μM) effect on splenocyte proliferation and IL-2 production was observed in vitro. Clonidine, another α2 adrenergic agonist, only had a stimulatory proliferative effect on splenocytes. Yohimbine, an α2 adrenergic antagonist, abrogated the stimulatory action of both clonidine and xylazine, but not the suppressive proliferative activity of xylazine in vitro. The inhibited proliferation of splenocytes to Con A correlated with increased apoptosis of T cells. The apoptosis was not blocked by yohimbine or antibodies to Fas and Fas-L. N-Nitro- l -arginine methyl ester ( l -NAME), an inhibitor of nitric oxide (NO) synthase, enhanced proliferation of splenocytes to Con A, partly abrogated the inhibitory effect of xylazine in the proliferation assay, and, only at high concentration (1000 μM), partly suppressed apoptosis of lymphocytes. The enhancing effect of l -NAME on the Con A-induced proliferation of splenocytes correlated with decreased NO production. However, decreased NO production observed in cultures with xylazine was followed by both decreased lymphocyte proliferation and apoptosis. Cumulatively, these results suggest that the immunosuppressive properties of xylazine on splenocytes in vitro are due to increased apoptosis of lymphocytes, predominantly involve NO-independent pathways, and are probably independent of its action through α2 adrenoreceptors.

Downregulation of nicotinic and muscarinic receptor function in rats after subchronic exposure to diazinon

Highlights • We studied the development of tolerance to subchronic p.o. administration of DZN in rats, under both in vivo and in vitro conditions• As a consequence of AChE inhibition, ACh neurotransmitters are being accumulated in over stimulated nicotinic and muscarinic receptors.• With isolated diaphragm and ileum, we examined the down regulation of nicotinic and muscarinic receptor function through EFS technique.• The results of our research could be useful in forensic diagnostics of organophosphate poisoning.

The Novel Phenylpropiophenone Derivates Induced Relaxation of Isolated Rat Aorta

Our aim was to define how different chemical properties of newly developed phenylpropiophenone derivates (PhPds) influenced their potency and efficacy to relax rat aorta. A contribution of ion channels in the PhPds and propafenone mechanism of vasodilatation was tested. PhPds were syntethysed by substitution in the benzyl moiety with -F, -CH3 or -CF3 groups on the ortho or para position. The vasodilatation by PhPds was examined on the rings of rat aorta precontracted with phenylephrine. In order to test involvement of voltage-gated Na+ and K+ channels and L-type Ca2+ channels in a mechanism of action of PhPds, we used their blockers: lidocaine, nifedipine and 4-aminopiridine, respectively. Aorta was more sensitive to 5-ortho-trifluoromethyl derivate than to propafenone and other PhPds. The 5-para-methyl derivate had lower potency and efficacy than propafenone and other PhPds. Lidocaine did not influenced relaxation induced by PhPds, but slightly inhibited the effect of propafenone. The 4-aminopiridine only inhibited relaxation induced by 5-para-methyl derivate. Nifedipine inhibited relaxation of the rat aorta induced by 5-ortho-trifluoromethyl derivate and by propafenone. Introduction of 5-ortho-trifluoromethyl and 5-para-methyl group in the benzyl moiety of propafenone molecule changed its potency, efficacy and mechanism of action in the rat aorta. The 4-aminopiridine- and nifedipine sensitive ion channels are involved in mechanism of action of 5-para-methyl and 5-ortho-trifluoromethyl derivate. The introduction of other tested groups in the benzyl moiety does not affect pharmacological properties of the PhPds in relation to propafenone.

The effect of a new nitro-aspirin on apoptosis of neutrophil granulocytes.

Apoptosis of neutrophil granulocytes is a critical event in the resolution of inflammation. Neutrophils have a short lifespan which can be modulated by aspirin. In this work we studied the effect of a nitroaspirin (NCX4040) on apoptosis of inflammatory granulocytes. This nitro-aspirin has been synthesized in attempt to reduce the side effects of aspirin in the gastrointestinal tract. Inflammatory granulocytes have been isolated from polyvinyl sponges implanted under the skin of Albino Oxford (AO) rats. Inflammatory cells that were isolated 20 hours later were about 95% neutrophil granulocytes. The cells were cultivated 24h with different concentrations of NCX4040 ranging from 0.01 μM to 10μM. After that period, apoptosis of neutrophils was performed by using morphological criteria, as well as by flow cytometry (after staining the cells with propidium iodide). We found that NCX4040 at concentrations from 0.25 to 10 μM induced the apoptosis of rat inflammatory granulocytes in a dose-dependent manner. Also, in these concentrations NCX4040 decreased production of nitric oxide in the cells culture supernatants. In conclusion, our results suggest that antiinflammatory properties of NO-aspirins are additionally potentiated by their proapoptotic effect on granulocytes, which could be a novel mechanism of their action.

The influence of the induction of farrowing on live birth, body mass, appearance of dystocia, mortality and surviving of neonatal pigs in litter during the first ten days

* Rad primljen za štampu 11.03.2015. ** Dr sc. vet. med. Jović Slavoljub, van. profesor, dr sc. vet. med. Ćupić Vitomir, red. profesor, Fakultet veterinarske medicine, Univerzitet u Beogradu; Ristić Gordana, dr vet. med. spec., Delta Agrar, Beograd, dr sc. vet. med. Vakanjac Slobodanka, vanr. profesor, dr sc. vet. med. Dimitrijević Blagoje, docent, Fakultet veterinarske medicine, Univerzitet u Beogradu, Miladinović Dejana, dr vet. med., doktorant, Fakultet veterinarske medicine, Univerzitet u Beogradu, dr sc. biol. Živković Lada, docent, Farmaceutski fakultet, Univerzitet u Beogradu ORIGINALNI RAD / ORIGINAL PAPER

The effect of acetylsalicylic acid and meloxicamon hematological parameters in rats

In this work there was investigated the effect of two nonsteroidal antiinflammatory drugs (NSAIDs), acetylsalicylic acid or aspirin (nonselective cyclooxygenase inhibitor - COX1 i COX2) and meloxicam (selective cyclooxygenase inhibitor - COX2) on certain hematological parameters in rats. The objective of the work was to determine whether (and to which extent), these drugs, after multiple peroral application, influence erythrocyte number, concentration of hemoglobin, hematological indices (mean corpuscular value - MCV; mean concentration of hemoglobin in erythrocytes - MCH; mean corpuscular hemoglobin concentration - MCHC), hematocrit, number of platelets, leukocytes, neutrophilic leukocytes, lymphocytes and monocytes. The experiment was conducted in in vivo conditions on 70 clinically healthy Wistar strain male rats, 10 to 12 weeks of age and body weight 250 to 300 g. The rats were divided into seven groups and they were daily perorally (by probe) given aspirin (ASCOPIR) at doses of 30, 40 and 80 mg/kg b.m. (I, II and III groups), or meloxicam (METACAM) at doses of 100, 125 and 250 μg/kg b.m. (IV, V and VI groups), for seven days. The seventh group was a control one and they were given only saline. The obtained results showed that: acetylsalicylic acid in maximum dose tested (80 mg/kg b.m.) statistically significantly reduced the number of platelets (p<0,05), the number of leukocytes (p<0,05), the number of lymphocytes (p<0,05) and the number of monocytes (p<0,05), while on the other side, meloxicam in maximum dose tested (250 μg/kg), statistically significantly reduced the mean corpuscular value (MCV), and increased the number of platelets (p<0,05), relative to the control value.

The effect of combined use of prostaglandin and oxytocin in sows, on the duration of partus, number of pigs and period between two expulsions of piglets

* Rad primljen za štampu 11. 03. 2015. godine ** Dr sc. vet. med. Vitomir Ćupić, red. profesor, dr sc. vet. med. Slavoljub Jović, docent, Fakultet veterinarske medicine, Univerzitet u Beogradu; Gordana Ristić, dr vet. med. spec., Delta Agrar, Beograd; dr sc. vet. med. Slobodanka Vakanjac, vanr. profesor, dr sc. vet. med. Blagoje Dimi trijević, docent, Dejana Ćupić-Miladinović, dr vet. med., istraživač pripravnik, Fakultet veterinarske medicine, Univerzitet u Beogradu ORIGINALNI RAD / ORIGINAL PAPER

Potassium channels opener pinacidil have multiple effects on KCl-elicited contractions of isolated non-pregnant rat uterus.

The effects of K+ channel opener, pinacidil on contractions provoked by contraction-stimulating KCl were investigated on isolated uterus of non-pregnant rats in oestrus. Pinacidil produced a more potent inhibition of 20 mM KCl-elicited contractions (pD2 = 6.57 μM) than of 40 or 80mMKCl-elicited contractions (pD2=5.11 and 5.19 μM, respectively). Glibenclamide, a selective blocker of adenosine triphosphate (ATP)-sensitive K+ (KATP) channels, antagonized the pinacidil-induced inhibition of contractions elicited by 20 mM KCl in a competitive manner. However, the pinacidil-induced inhibition of contractions provoked by 40 and 80 mM KCl glibenclamide was unable to prevent them. Pinacidil ability to completely relax the non-pregnant uterus pre-contracted with K+-rich solution suggests that K+ channelindependent mechanism(s) also plays a part in its relaxant effect.

CHARACTERISTICS OF THE ANTIVASOCONSTRICTOR EFFECT OF PINACIDIL ON ISOLATED RADIAL ARTERY

Pinacidil, a prevously studied potassium channel opener (PCO), is a potent antihypertensive agent in animals and humans. Its mechanism of action is not completly defined. The aim of our study was to investigate the antivasoconstricting effect of pinacidil on the isolated RA and to study whether this effect is endothelium-dependent. Contractions of isolated RA rings with intact endothelium were provoked by electrical field stimulation (EFS, 20 Hz) or exogenously applied noradrenaline (NA, 10 μM). Pinacidil (10 nM-0.1 mM) produced a concentration-dependent inhibition of both EFS- and NA-evoked contractions (p>0.05). NO synthesis inhibitor, L-NAME (10 μM) and the guanylate cyclase inhibitor, methylene blue (10 μM) did partly antagonize NA-evoked contractions and were without effect on EFSinduced contractions. Thus, the antivasoconstrictor effect of pinacidil on RA is partly endothelium-dependent and probably mediated via cGMP-dependent NO-pathway.