Maribel Antonello Rubin

Polyamines reduces lipid peroxidation induced by different pro-oxidant agents.

Polyamines, among other functions, are considered to act as a free radical scavenger and antioxidant. The quinolinic acid (QA), sodium nitroprusside (SNP) and iron (Fe+2) stimulate production of free radicals and lipid peroxidation. In the present study, we investigated the free radical and/or aldehyde scavenger effects of polyamines spermine and spermidine on thiobarbituric acid reactive species (TBARS) production induced by QA, SNP, Fe+2/EDTA system and free Fe2+ in rat brain. Spermine and spermidine inhibited QA-induced TBARS production; however spermine was a better antioxidant than spermidine. Spermine also inhibited SNP-, Fe+2/EDTA- and free Fe2+-induced TBARS production, but had a modest effect. Spermidine, in turn, also discretely inhibited SNP-, Fe+2/EDTA- and free Fe2+-induced TBARS production. In the presence of MK-801, QA-induced TBARS production was considerably more inhibited by polyamines. In addition, arcaine does not affect the reducer effect of polyamines. The present findings suggest that the observed effects of polyamines are not related to the activation of NMDA receptor but with their antioxidant and free radical scavenger properties.

Antioxidant Properties of New Chalcogenides Against Lipid Peroxidation in Rat Brain

Ebselen (2-phenyl- 1,2-benzisoselenazole-3 (2H)-one) is a seleno-organic compound with antioxidant properties, and anti-inflammatory actions. Recently, ebselen improved the outcome of acute ischemic stroke in humans. In the present study, the potential antioxidant capacity of organochalcogenide compounds diphenyl diselenide (PhSe)2, diphenyl ditelluride (PhTe)2, diphenyl disulfide (PhS)2, p-Cl-diphenyl diselenide (pCl-PhSe)2, bis-[S-4-isopropyl 2-phenyl oxazoline] diselenide (AA-Se)2, bis-[S-4-isopropyl 2-phenyl oxazoline] ditelluride (AA-Te)2 and bis-[S-4-isopropyl 2-phenyl oxazoline] disulfide (AA-S)2 was compared with that of ebselen (a classical antioxidant). Spontaneous and quinolinic acid (QA)- (2 mM) and sodium nitroprusside (SNP)- (5 μM)-induced thiobarbituric reactive species (TBARS) production by rat brain homogenates was determined colorimetrically. TBARS formation was reduced by ebselen, (PhSe)2, (PhTe)2, (AA-Se)2, (AA-S)2 and (pCl- PhSe)2 to basal rates. The concentrations of these compounds needed to inhibit TBARS formation by 50% (lC50) are 1.71 μM, 3.73 μM, 1.63 μM, 9.85 μM, > 33.3 μM, 23.2 μM and 4.83 μM, respectively for QA. For TBARS production induced by SNP the lC50 was 2.02 μM, 12.5 μM, 2.80 μM, > 33.3 μM, 24.5 μM and 7.55 μM, respectively. The compounds (AA-Te)2 and (PhS)2 have no antioxidant activity and pro-oxidant activity, respectively. These results suggest that (AA-Se)2 and (AA-S)2 can be considered as potential pharmaceutical antioxidant agents.

Resveratrol prevents memory deficits and the increase in acetylcholinesterase activity in streptozotocin-induced diabetic rats

The objective of the present study was to investigate the effect of the administration of resveratrol (RV) on memory and on acetylcholinesterase (AChE) activity in the cerebral cortex, hippocampus, striatum, hypothalamus, cerebellum and blood in streptozotocin-induced diabetic rats. The animals were divided into six groups (n=6-13): Control/saline; Control/RV 10 mg/kg; Control/RV 20 mg/kg; Diabetic/saline; Diabetic/RV 10 mg/kg; Diabetic/RV 20 mg/kg. One day after 30 days of treatment with resveratrol the animals were submitted to behavioral tests and then submitted to euthanasia and the brain structures and blood were collected. The results showed a decrease in step-down latency in diabetic/saline group. Resveratrol (10 and 20 mg/kg) prevented the impairment of memory induced by diabetes. In the open field test, no significant differences were observed between the groups. In relation to AChE activity, a significant increase in diabetic/saline group (P<0.05) was observed in all brain structures compared to control/saline group. However, AChE activity decreased significantly in control/RV10 and control/RV20 (P<0.05) groups in cerebral cortex, hippocampus and striatum, while no significant differences were observed in diabetic/RV10 and diabetic/RV20 groups in all brain structures compared to control/saline group. Blood AChE activity increased significantly in diabetic/saline group (P<0.05) decreased in control/RV10, control/RV20 and diabetic/RV20 groups (P<0.05) compared to control/saline group. In conclusion, the present findings showed that treatment with resveratrol prevents the increase in AChE activity and consequently memory impairment in diabetic rats, demonstrating that this compound can modulate cholinergic neurotransmission and consequently improve cognition.

Ebselen blocks the quinolinic acid-induced production of thiobarbituric acid reactive species but does not prevent the behavioral alterations produced by intra-striatal quinolinic acid administration in the rat

Ebselen (EBS) is a seleno-organic compound with glutathione peroxidase-like activity which is neuroprotective in acute stroke ischemia. In this study, we investigated the effect of EBS on quinolinic acid (QA)-induced neurotoxicity. EBS inhibited QA-induced production of thiobarbituric acid reactive species (TBARS) by striatal homogenates in vitro with an IC(50) of 1.85 microM. Intra-striatal injection of QA (360 nmol) increased striatal content of TBARS and induced convulsions and contralateral rotational behavior. Intra-striatal pre-injection of EBS (10 nmol) 15 min before QA abolished QA-induced TBARS production but did not alter QA-induced behavioral effects. The present findings suggest that EBS acts on post-receptor events, neutralizing free radicals produced by overstimulation of N-methyl-D-aspartate receptors.

N-acetylcysteine prevents memory deficits, the decrease in acetylcholinesterase activity and oxidative stress in rats exposed to cadmium

The present study investigated the effect of the administration of N-acetylcysteine (NAC), on memory, on acetylcholinesterase (AChE) activity and on lipid peroxidation in different brain structures in cadmium (Cd)-exposed rats. The rats received Cd (2 mg/kg) and NAC (150 mg/kg) by gavage every other day for 30 days. The animals were divided into four groups (n=12-13): control/saline, NAC, Cd, and Cd/NAC. The results showed a decrease in step-down latency in the Cd-group, but NAC reversed the impairment of memory induced by Cd intoxication. Rats exposed to Cd and/or treated with NAC did not demonstrate altered shock sensitivity. Decreased AChE activity was found in hippocampus, cerebellum and hypothalamus in the Cd-group but NAC reversed this effect totally or partially while in cortex synaptosomes and striatum there was no alteration in AChE activity. An increase in TBARS levels was found in hippocampus, cerebellum and hypothalamus in the Cd-group and NAC abolished this effect while in striatum there was no alteration in TBARS levels. Urea and creatinine levels were increased in serum of Cd-intoxicated rats, but NAC was able to abolish these undesirable effects. The present findings show that treatment with NAC prevented the Cd-mediated decrease in AChE activity, as well as oxidative stress and consequent memory impairment in Cd-exposed rats, demonstrating that this compound may modulate cholinergic neurotransmission and consequently improve cognition. However, it is necessary to note that the mild renal failure may be a contributor to the behavioral impairment found in this investigation.

Antinociceptive effect of novel trihalomethyl-substituted pyrazoline methyl esters in formalin and hot-plate tests in mice.

The aim of the present study was to evaluate the antinociceptive potential of four novel pyrazoline methyl ester compounds on chemical and thermal models of pain in mice. The following 5-trihalomethylated-4,5-dihydro-1H-pyrazole methyl ester compounds were tested: 3-methyl-5-trifluoromethyl-(MPF3), 4-methyl-5-trifluoromethyl-(MPF4), 3-methyl-5-trichloromethyl-(MPCl3) and 4-methyl-5-trichloromethyl-(MPCl4). MPF3, MPF4, MPCl3 and MPCl4 (0.03-1.0 mmol/kg) given intraperitoneally decreased neurogenic and inflammatory phases of nociception in the formalin test. Moreover, MPF3, MPF4, MPCl3, MPCl4 (0.1-1.0 mmol/kg) and dipyrone (1.5 mmol/kg) also produced a dose-dependent antinociceptive effect in the hot-plate test. However, MPF3, MPF4, MPCl3 and MPCl4 did not impair motor coordination in the rotarod test or spontaneous locomotion in the open field test. The antinociceptive effect of MPF4 (1.0 mmol/kg, i.p.) was reversed by the opioid receptor antagonist naloxone (2 mg/kg, i.p.), but not by the alpha(2)-adrenergic receptor antagonist yohimbine (0.15 mg/kg, i.p.) or by p-chlorophenylalanine ethyl ester (PCPA, 300 mg/kg, i.p.) treatment. In contrast to morphine (5 mg/kg, i.p.), MPF4 given daily for up to 8 days did not generate a tolerance to its antinociceptive effect. However, similar to morphine (11 mg/kg, i.p.), MPF4 reduced gastrointestinal transit in mice. Taken together these results demonstrate that these novel pyrazoline methyl esters tested may be promising prototypes of additional mild analgesics.

Intra-Amygdala Administration of Polyamines Modulates Fear Conditioning in Rats

Amygdalar NMDA receptor activation has been implicated in the acquisition of fear memories in rats. However, little is known about the role of endogenous modulators of the NMDA receptor, such as polyamines, in pavlovian fear-conditioning learning. Therefore, in the present study we investigated whether the immediate pretraining or post-training bilateral infusion of arcaine, an antagonist of the NMDA receptor polyamine-binding site, or spermidine, an agonist of the NMDA receptor polyamine-binding site, into the amygdala affected classical fear conditioning in rats. Bilateral microinjections of arcaine (0.0002-0.2 nmol) decreased, whereas spermidine (0.002-20 nmol) increased, contextual and auditory fear conditioning. Arcaine coadministration, at a dose that had no effect per se, reversed the facilitatory effect of spermidine. These results provide evidence that endogenous and exogenous polyamines modulate the acquisition or early consolidation (or both) of the fear-conditioning task in the amygdala.

Intrastriatal methylmalonic acid administration induces rotational behavior and convulsions through glutamatergic mechanisms

The effect of intrastriatal administration of methylmalonic acid (MMA), a metabolite that accumulates in methylmalonic aciduria, on behavior of adult male Wistar rats was investigated. After cannula placing, rats received unilateral intrastriatal injections of MMA (buffered to pH 7.4 with NaOH) or NaCl. MMA induced rotational behavior toward the contralateral side of injection and clonic convulsions in a dose-dependent manner. Rotational behavior and convulsions were prevented by intrastriatal preadministration of MK-801 and attenuated by preadministration of succinate. This study provides evidence for a participation of NMDA receptors in the MMA-induced behavioral alterations, where succinate dehydrogenase inhibition seems to have a pivotal role.

Antinociceptive effect of novel pyrazolines in mice

The antinociceptive effect of six novel synthetic pyrazolines (3-ethoxymethyl-5-ethoxycarbonyl-1H-pyrazole (Pz 1) and its corresponding 1-substituted methyl (Pz 2) and phenyl (Pz 3) analogues, and 3-(1-ethoxyethyl)-5-ethoxycarbonyl-1H-pyrazole (Pz 4) and its corresponding 1-substituted methyl (Pz 5) and phenyl (Pz 6) analogues) was evaluated by the tail immersion test in adult male albino mice. The animals (N = 11-12 in each group) received vehicle (5% Tween 80, 10 ml/kg, sc) or 1.5 mmol/kg of each of the pyrazolines (Pz 1-Pz 6), sc. Fifteen, thirty and sixty minutes after drug administration, the mice were subjected to the tail immersion test. Thirty minutes after drug administration Pz 2 and Pz 3 increased tail withdrawal latency (vehicle = 3.4 +/- 0.2; Pz 2 = 5.2 +/- 0.4; Pz 3 = 5.9 +/- 0.4 s; mean +/- SEM), whereas the other pyrazolines did not present antinociceptive activity. Dose-effect curves (0.15 to 1.5 mmol/kg) were constructed for the bioactive pyrazolines. Pz 2 (1.5 mmol/kg, sc) impaired motor coordination in the rotarod and increased immobility in the open-field test. Pz 3 did not alter rotarod performance and spontaneous locomotion, but increased immobility in the open field at the dose of 1.5 mmol/kg. The involvement of opioid mechanisms in the pyrazoline-induced antinociception was investigated by pretreating the animals with naloxone (2.75 micro mol/kg, sc). Naloxone prevented Pz 3- but not Pz 2-induced antinociception. Moreover, naloxone pretreatment did not alter Pz 3-induced immobility. We conclude that Pz 3-induced antinociception involves opioid mechanisms but this is not the case for Pz 2.

Intrahippocampal spermidine administration improves inhibitory avoidance performance in rats.

Polyamines are polycations present at high concentrations in the mammalian brain. We investigated the effect of an intrahippocampal infusion of spermidine, a polyamine agonist, immediately post‐training on the inhibitory avoidance learning paradigm in rats. Bilateral intrahippocampal microinjection of spermidine (0.02–20 nmol) caused an increase in test step‐down latencies at low concentrations. Administration of arcaine (0.002–0.2 nmol), an antagonist of the N ‐methyl‐d‐aspartate (NMDA) receptor polyamine binding site, did not modify the test step‐down latencies. In contrast, co‐administration of arcaine and spermidine completely reversed the facilitatory effect of spermidine on the test step‐down latencies. These results provide evidence that polyamines may be involved in learning and memory modulation.