Cicatriz miocárdica arritmogénica localizada mediante el uso combinado de la SPECT con 123I-MIBG y la RMN cardíaca / Arrhythmogenic myocardial scar localized through the combined use of 123I-MIBG SPECT and cardiac MRI

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Methylamine-dependent release of nitric oxide and dopamine in the CNS modulates food intake in fasting rats.

BACKGROUND AND PURPOSE: Methylamine is an endogenous aliphatic amine exhibiting anorexigenic properties in mice. The aim of this work was to show whether methylamine also modifies feeding behaviour in rats and, if so, to identify the mediator(s) responsible for such effects. EXPERIMENTAL APPROACH: Microdialysis experiments with the probe inserted in the periventricular hypothalamic nucleus were carried out in 12 h starved, freely moving rats. Collected perfusate samples following methylamine injection (i.c.v.) were analysed for nitric oxide by chemiluminescence and for dopamine and 5-hydroxytryptamine content by HPLC. Kv1.6 potassium channel expression was reduced by antisense strategy and this decrease quantified by semi-quantitative RT-PCR analysis. KEY RESULTS: Methylamine showed biphasic dose-related effects on rat feeding. At doses of 15-30 microg per rat, it was hyperphagic whereas higher doses (60-80 microg) were hypophagic. Methylamine stimulated central nitric oxide (+115% vs. basal) following hyperphagic and dopamine release (60% over basal values) at hypophagic doses, respectively. Treatment with L-N(G)-nitro-L-arginine-methyl ester (i.c.v. 2 microg 10 microl(-1)) or with alpha-methyl-p-tyrosine (i.p. 100 mg kg(-1)) before methylamine injection, reduced nitric oxide output and hyperphagia, or dopamine release and hypophagia respectively. Moreover, hypophagia and hyperphagia, as well as nitric oxide and dopamine release were significantly reduced by down-regulating brain Kv1.6 potassium channel expression. CONCLUSIONS AND IMPLICATIONS: The effects of methylamine on feeding depend on the hypothalamic release of nitric oxide and dopamine as a result of interaction at the Kv1.6 channels. The study of methylamine levels in the CNS may provide new perspectives on the physiopathology of alimentary behaviour.

The polysaccharide from Tamarindus indica (TS-polysaccharide) protects cultured corneal-derived cells (SIRC cells) from ultraviolet rays.

The aim of this work was to investigate the possible protective effect of a new viscosising agent, TS-polysaccharide, on corneal-derived cells (SIRC) exposed to ultraviolet-B rays. To verify this, SIRC cells were first exposed, in the absence or in the presence of TS-polysaccharide (1% w/v), for 9 s at the UV-B source and then post-incubated for 45 min at 37 degrees C. After this period the hydrogen peroxide (H(2)O(2)) accumulated in the medium and the concentration of 8-hydroxy-2'-deoxy-guanosine (8-OHdG) in cell DNA was measured. In addition, the amount of (3)H-methyl-thymidine incorporated in cellular DNA was evaluated after 18 h from irradiation. Our results show that cells exposed to UV-B rays accumulate H(2)O(2), and have higher levels of 8OHdG and a lower amount of (3)H-methyl-thymidine incorporated in DNA than control cells. In the presence of TS-polysaccharide, the H(2)O(2) and 8-OHdG accumulation, and the (3)H-methyl-thymidine incorporation were significantly reduced with respect to the values measured in cells exposed in the absence of the polysaccharide. We propose a protective role of the polysaccharide in reducing UV-B derived DNA damage to eye cells. This finding could be of some clinical importance when the polysaccharide is used as a delivery system for ophthalmic preparations.

The reduction of food intake induced in mice by benzylamine and its derivatives.

Characterisation of the pharmacological profile of non-physiological amine oxidase substrates could help to identify the endogenous role of this class of enzymes. Previous studies have suggested that benzylamine, a common non-physiological substrate for monoamine and tissue-bound or soluble benzylamine oxidases, could behave as a potassium channel blocking agent. Potassium channel blockers are known to modify several forms of animal behaviour including food consumption. To characterise further the pharmacological profile of benzylamine and the role of amine oxidases, we have studied the effect of benzylamine on mice food intake. Our results confirm that benzylamine produces a reduction in mice feeding in a similar manner to that obtained by amphetamine. The anorectic effect of benzylamine and amphetamine in mice was potentiated by pretreatment with amine oxidase inhibitors. In addition, the introduction of substituents in the aromatic ring of benzylamine did not produce compounds with a higher anorectic potency than the one measured with benzylamine.

Methylamine and benzylamine induced hypophagia in mice: modulation by semicarbazide-sensitive benzylamine oxidase inhibitors and aODN towards Kv1.1 channels.

1. In starved mice, the anorectic activity of methylamine (MET) and benzylamine (BZ), both substrates of semicarbazide-sensitive benzylamine oxidases (Bz-SSAO), was compared with that of the potassium channel blocking agents charybdotoxin (ChTX), tetraethylammonium (TEA), gliquidone (GLI), ammonium chloride (NH(4)(+)) and of the anoressants amphetamine (AMPH) and nicotine (NIC). After i.c.v. administration, an approximate ranking order of potency was: ChTX> or =AMPH>NIC=TEA> or =GLI> or =MET>BZ>NH(4)(+). 2. Clorgyline (2.5 mg kg(-1) i.p.) or deprenyl (10 mg kg(-1) i.p.) potentiated the anorectic effect of i.c.v.-administered BZ, NIC and AMPH. The effect of TEA was increased only by deprenyl, while MET, NH(4)(+), ChTX and GLI were not affected by either of the inhibitors. 3. The Bz-SSAO inhibitors alpha-aminoguanidine (50 mg kg(-1) i.p.), B24 (100 mg kg(-1) i.p.) and MDL 72274 (2.5 mg kg(-1) i.p.) potentiated the effect of i.p., but not of i.c.v.-administered MET. 4. Antisense oligodeoxyribonucleotides (aODN) to Kv1.1 potassium channels abolished the effect of BZ and TEA, but was ineffective in reducing the activity of MET and other compounds. 5. These results suggest that MET is endowed with peculiar hypophagic effects at dosage levels that are not able to affect gross behaviour in mice. The effect of MET, differently from BZ, seems unrelated to an increase in the central release of monoaminergic mediators, as well as to a Kv1.1 blocking activity. Through a reduction of the endogenous breakdown of MET, Bz-SSAO inhibitors enhance the central pharmacological activity of this amine.

Selective inhibition of amine oxidases differently potentiate the hypophagic effect of benzylamine in mice.

In mice deprived of food for 12 h, the i.c.v. or i.p. administration of benzylamine, a substrate common to both monoamine oxidase B and semicarbazide-sensitive benzylamine oxidases, dose-dependently inhibited feeding. This effect was significantly potentiated by selective monoamine oxidase A and B inhibition, suggesting that central monoamines, known to be substrates of these enzymes may be released. The i.p. administration of semicarbazide-sensitive benzylamine oxidase inhibitors, B24 (3,5-ethoxy-4-aminomethylpyridine) and MDL 72274 ((E)-2-phenyl-3-chloroallylamine) strongly potentiated the effect of i.p. but not i.c.v.-administered benzylamine. The hypophagic effect of benzylamine was evaluated following i.c.v. administration, in comparison with the effect of the sympathomimetic compound amphetamine or the K(+) channel blocker tetraethylammonium, as reference compounds. Our results make it possible to define benzylamine as a centrally acting hypophagic compound devoid of amphetamine-like motor stimulatory effects and point to a role of B24 and MDL 72274 as specific peripheral enhancers of the pharmacological effects of benzylamine.

Possible different fates for the hydrogen peroxide produced by rat white adipocyte amine oxidases.

Monoamine oxidase (MAO) and benzylamine oxidases (Bz-SSAO) of rat white adipocytes, deaminating benzylamine and tyramine produce hydrogen peroxide at different cellular levels. The peroxide produced by their activity has a very short half-life in adipocyte suspension. The addition of a catalase inhibitor allows for the recovery of the intact peroxide within the first 10-min of its production. Thus, benzylamine and tyramine-dependent peroxide recovery is different, suggesting that the fate of the peroxide produced by the two amine oxidases might be different depending on the cellular site of its production.

Sedum telephium L. polysaccharide content affects MRC5 cell adhesion to laminin and fibronectin.

In traditional medicine the fresh leaves and juice of Sedum telephium L. are used as wound-healing promoters. Cell adhesion represents a primary event in wound repair and in tissue homeostasis, and therefore we have investigated the effect of Sedum juice and its main fractions, polysaccharides and flavonols, on human fibroblast (MRC5) adhesion to fibronectin and laminin. Our findings revealed that total Sedum juice strongly inhibited cell adhesion to laminin and fibronectin (EC50 1.03+/-0.12 mg mL(-1)). This anti-adhesive feature was concentrated mainly in the two polysaccharide fractions (EC50 values comprised between 0.09 and 0.44 mg mL(-1)). The flavonol fractions did not seem to contribute to this effect. A first attempt to elucidate the polysaccharide-related anti-adhesive feature of Sedum juice was also performed. The results confirmed that natural polysaccharides, with chemical structures different from heparin, were able to interfere with integrin-mediated cell behaviour and they contributed to the outstanding effects of Sedum juice and to the role of polysaccharides in cell-matrix interaction.

Hydrogen peroxide generation by monoamine oxidases in rat white adipocytes: role on cAMP production.

In rat, white adipocytes monoamine oxidases (EC 1.4.3.4.) generate hydrogen peroxide (H(2)O(2)). Recent studies suggested that, in addition to its toxic features, H(2)O(2) may behave as a cell second messenger. In the present study, using fluorimetric and chemiluminescence (CL) assays, we showed that tyramine degradation by monoamine oxidases in intact adipocytes resulted in the concentration-dependent generation of H(2)O(2). In addition, we found that, in the presence of low tyramine concentrations, forskolin-dependent cAMP production was significantly increased as compared to that of the control and this increase was prevented by the monoamine oxidase inhibitor pargyline or by the H(2)O(2) trapping system homovanillic acid-peroxidase. Finally, we demonstrated that tyramine degradation by monoamine oxidases increased the ability of isoproterenol to induce cell lipolysis. Taken together, these data suggest that H(2)O(2) produced during substrate degradation by monoamine oxidases may participate in the regulation of adipocyte metabolism.

Effect of xyloglucan (tamarind seed polysaccharide) on conjunctival cell adhesion to laminin and on corneal epithelium wound healing.

PURPOSE: To explore the role of a natural polysaccharide extracted from tamarind seed (xyloglucan, or tamarind seed polysaccharide, TSP) on the integrin-substrate recognition system and on repair of corneal wounds. METHODS: a) Cultured human conjunctival cells were labeled by addition of a tritiated amino acid mixture. Their adhesion to laminin-coated culture wells in the absence or presence of TSP was checked by radioactivity count. b) The corneal epithelium of albino rabbits was damaged by applying a paper disc soaked with n-heptanol. The eyes were then treated with TSP, with a hyaluronate reference formulation and with normal saline solution (controls). The diameter of corneal wounds was measured daily, after fluorescein staining. RESULTS: Compared to hyaluronate, TSP slightly but significantly increased the wound healing rate. TSP 1.0% exerted a positive influence on cell adhesion to laminin, up to a certain laminin concentration. CONCLUSIONS: The ability of the polysaccharide to promote corneal wound healing might depend on its influence on the integrin recognition system.

Benzylamine-related compounds stimulate rat vas deferens neurotransmission and potentiate memory in the mouse acting as potassium channel blockers.

In stimulated rat vas deferens, the new compound 2, 6-dibutylbenzylamine (B25) and some related benzylamines, first potentiated then completely inhibited electrically-induced twitch response, showing the biphasic effect previously observed in unstimulated preparations. To verify if this effect could be referred to as a modulation of potassium channels the activity of some benzylamines, KCl, tetraetylammonium (TEA), BaCl(2), 4-aminopyridine (4-AP), glibenclamide (GLI), charibdotoxin (ChTX) and apamin (APA) has been compared. While KCl and benzylamine-related derivatives induced biphasic effects, TEA, 4-AP, BaCl(2), GLI stimulated but were unable to inhibit the twitches. The pretreatment with stimulating concentrations of TEA, 4-AP, GLI, APA or ChTX and B25, as reference compound in the benzylamine series, dose-dependently reduced the stimulatory effect of KCl but were unable to modify the inhibitory effect induced by this ion. Both KCl and B25 potentiated each others own inhibitory effect suggesting that, unlike other potassium channel blockers, they could modulate in an opposite way voltage-dependent potassium channels in order to facilitate and then depress neurotransmission. In other experiments, benzylamines, KCl, TEA, 4-AP and GLI reverted the inhibitory effect of cromakalim and omega-conotoxin GVIA (omega-CTX). This effect further supports a common mechanism of action (potassium channel blockade) probably inducing the opening of Ca(2+)channels different from N or L in the preparation. Finally, the prevention of minoxidil-induced amnesia in the mouse by B25 and related benzylamines, comparable to the same effect shown by TEA and 4-AP, indicates that these compounds are endowed with potential pharmacological activity in the CNS as well.

Amplification of alpha 1D-adrenoceptor mediated contractions in rat aortic rings partially depolarised with KCl.

Partial depolarisation of smooth muscle in endothelium-denuded rat aortic ring preparations, by increasing physiological buffer KC1 concentrations from 4.7 to 14.7 mM, produced a leftward shift of concentration response curves (CRCs) to the alpha 1-adrenoceptor agonist noradrenaline (NA), phenylephrine and methoxamine, without changing maximal responses, whereas maximal responses to clonidine (CLON), also an alpha 1-agonist in this tissue were considerably increased. Partial depolarisation did not alter responses to 10 nM NA or 100 nM CLON in Ca2+(-free) buffer, but significantly increased the contractions obtained on adding Ca2+ back in the presence of the agonists. The potentiation of NA (2.5 and 5 nm) contractions by partial depolarisation was prevented by the voltage-operated Ca2+ channel (VOCC) antagonist nifedipine (NIF, 1 microM). NIF did not significantly affect NA CRCs in 4.7 mM KCl, whereas responses in 14.7 mM KCl were significantly decreased, indicating VOCC recruitment by NA only in the latter condition. Initial depletion of intracellular Ca2+ stores with 1 microM thapsigargin (THAP) in Ca2+(-free) buffer did not alter NA CRCs subsequently obtained in normal Ca2+. However, after THAP-pretreatment, these NA responses (in both 4.7 and 14.7 mM KC1) were attenuated by NIF, indicating that VOCCs were activated by NA in THAP-treated tissues. SKF 96365 (SKF, 30 microM), which can block VOCC and non-VOCC routes of extracellular Ca2+ influx, inhibited NA responses in 4.7 mM and 14.7 mM KCl, possibly implying a role for both types of Ca2+ entry in contractions. However, the greater inhibitory effects of SKF in THAP-pretreated tissues, probably reflected the mobilisation of VOCCs by NA following THAP exposure, because SKF was shown separately to block VOCC-mediated contractions in tissues depolarised with 100 mM KCl alone. 10 microM niflumic acid, an inhibitor of Ca2+(-activated) Cl- channels, did not affect responses to NA in 4.7 mM or 14.7 mM KC1, suggesting that VOCC opening induced by NA in 14.7 mM KCl was not due to depolarisation produced by alpha 1-adrenoceptor induced Cl- efflux. CRCs for NA were unaffected by pretreatment of rings with 100 ng ml-1 pertussis toxin (PT), suggesting a lack of involvement of PT-sensitive G proteins in the contractions obtained either in 4.7 or 14.7 mM KCl. BMY 7378 (100 microM), a selective antagonist for alpha 1D-adrenoceptors, competitively inhibited NA contractions with apparent pKB values of 8.7 +/- 0.2 and 8.4 +/- 0.1 in 4.7 mM and 14.7 mM KCl, respectively. Pretreatment of rings with chloroethylclonidine (100 microM), an irreversible antagonist of alpha 1B-and alpha 1D-adrenoceptors, produced similar rightward shifts in CRCs to NA by 3.2 +/- 0.2 and 3.7 +/- 0.3 log concentration units in 4.7 mM and 14.7 mM KCl, respectively, without changing maximal responses. Inositol phosphate (IP) turnover produced by NA in aortic rings was not significantly different in 4.7 mM compared with 14.7 mM KCl. As a whole, these results suggest that partial depolarisation of the rat aorta with KCl enhances alpha 1-adrenoceptor mediated contractions predominantly via the alpha 1D-subtype, and by a mechanism to be identified which allows greater recruitment of VOCCs by NA. In addition, the ability of THAP-pretreatment also to enhance VOCC activation by NA suggests that Ca2+ release from, or prevention of its reuptake into, intracellular stores may contribute to those processes leading to VOCC opening.

Immunosuppressive activity of 13-cis-retinoic acid in rats: aspects of pharmacokinetics and pharmacodynamics.

Pharmacokinetics and pharmacodynamics of 13-cis-retinoic acid (13-cRA) administered at doses that suppress experimental autoimmune encephalomyelitis (EAE) have been investigated in rats. Serum concentration of the drug measured following oral administration of 37 mg/kg/12 h reached a peak of 1.8 x 10(-5) M in 2 h and linearly declined to 7.8 x 10(-7) M at hour 12. When spleen cells (SC) collected from 13-cRA-administered animals were cultured in vitro, their proliferative response to the T-cell mitogen concanavalin A (ConA) was suppressed and this effect was dependent on in vivo serum concentrations of the drug. In addition, in vitro exposure of antigen-specific T-cell lines to 13-cRA concentrations equivalent to those observed in vivo caused a dose-dependent suppression of the proliferation induced by the antigen as well as by T-cell mitogens. On a molar basis, 13-cRA showed a stronger in vitro immunosuppressive activity than two immunosuppressive agents used in human therapy, cyclosporin A and 6-mercaptopurin.

Some histamine-related compounds interacting with the benzylamine-oxidizing activity of rat white adipocytes.

In rat white adipocytes histamine is oxidized by a semicarbazide-sensitive amine oxidase which has benzylamine or preferential substrate (Bz-SSAO). To determine whether Bz-SSAO could control the extracellular levels of histamine and other histamine-related compounds active in lipid mobilization, a series of histaminergic compounds was screened as possible substrates or inhibitors of Bz-SSAO activity. Histaminergic compounds with imidazolo or thiazolo groups are oxidized by rat white-adipocyte Bz-SSAO whereas S-isothiourea derivatives, with two- or three-carbon-atom alkyl chains between the isothiourea and the N,N-dimethyl residue are, instead, inhibitors of the enzyme. Amtamine has been identified as a selective, high affinity substrate for rat white adipocyte Bz-SSAO. This enzymatic degradation might represent a catabolic pathway for the drug. These results show that the histaminase property of the rat white-adipocyte enzyme Bz-SSAO also extends to other histamine derivatives active at histamine receptors.

The histaminase activity of rat white adipocytes.

OBJECTIVE AND DESIGN: Investigate the reaction conditions which allow the measurement of high affinity histamine oxidation. MATERIAL: Isolated rat white adipocytes. METHODS: The histaminase activity of rat white adipocytes and its inhibition by B24 and MDL 72274 was measured fluorimetrically or radiochemically in different experimental conditions. RESULTS: Histamine oxidation by rat white adipocytes is enhanced by elevated pH and by the presence of bicarbonate ions. Under these conditions the oxidative deamination of histamine by white adipocytes, becomes comparable to that of other histaminases, suggesting that this amine oxidase activity may play a role in the catabolism of histamine in vivo. The specific semicarbazide-sensitive amine oxidase (SSAO) inhibitors MDL 72274 and B24 inhibit the oxidation of both histamine and benzylamine by the adipocyte preparation. CONCLUSIONS: Although there are kinetic differences between the behaviour of these two amine substrates, these results would be consistent with a SSAO being responsible for both activities.

Increased desensitization by picomolar phorbol ester of the endothelium-mediated effect of histamine in the perfused rat mesenteric bed.

The vasodilatatory, endothelium-mediated, effect of histamine (H), through H1 receptor, in the isolated and perfused mesenteric bed of the rat, undergoes strong desensitization during perfusion or repetitive injections of noradrenaline (NA) and H. The mesenteric bed completely desensitized to H is responsive to carbachol (C) and this latter compound does not affect the H desensitization. The homologous desensitization to C effect is very small, attaining less than 10% after 30 min of continuous perfusion. In this work the effect of inhibitors or activators of protein-kinase(s)-C (PKC) are studied during continuous perfusion of H or C in preparations preconstricted by NA. Staurosporine antagonizes the onset of the H desensitization, while the rate of desensitization in increased by phorbol-12-myristate-13-acetate (PMA). PMA, at a concentration from 10(-12) to 10(-10)M, selectively enhances the homologous desensitization of H, while at 10(-8)M it also produces a desensitization to C. At least two different PKC isoenzymes might be involved in the desensitization of the vasodilatatory effect of H and C in the isolated and perfused rat mesenteric bed.

Semicarbazide-sensitive amine oxidase activity in white adipose tissue of the insulin-deficient rat.

We have investigated whether the effects in white adipose tissue due to insulin deficiency might also be related to an alteration of histamine levels which are regulated by semicarbazide-sensitive amine oxidase. The lack of circulating insulin induced by streptozotocin produced, in rat white adipose tissue, a loss of affinity of the semicarbazide-sensitive amine oxidase for histamine oxidation. In parallel, a decrease of cell transformation, measured by glycerol-3-phosphate dehydrogenase activity and an augmented sensitivity to histamine lipolysis were observed. These findings could contribute to the understanding of histamine metabolism and function in diabetic rats and to the knowledge concerning amine oxidases in this animal pathology.

The role of semicarbazide-sensitive amine oxidase with a high affinity for benzylamine (Bz. SSAO) in the catabolism of histamine in the mesenteric arterial bed of the rat.

In the mesenteric arterial bed of the rat the semicarbazide-sensitive amine oxidase with a high affinity for benzylamine (Bz. SSAO) (E.C. 1.4.3.6) is also able to oxidize histamine. In the perfused mesenteric arterial bed of the rat the complete inhibition of the Bz. SSAO obtained with a specific inhibitor, B24, almost completely reduces the efflux of imidazole acetic acid and increases the relaxing effect of histamine. Bz. SSAO appears to be the only enzyme present in these blood vessels able to catabolize histamine.