Biomedical subjects

Josefa León

Publications and source records attributed to Josefa León.

At least 19 recordsLinked to original sources

PubMed2006

Inhibition of neuronal nitric oxide synthase activity by N1-acetyl-5-methoxykynuramine, a brain metabolite of melatonin.

We assessed the effects of melatonin, N(1)-acetyl-N (2)-formyl-5-methoxykynuramine (AFMK) and N(1)-acetyl-5-methoxykynuramine (AMK) on neuronal nitric oxide synthase (nNOS) activity in vitro and in rat striatum in vivo. Melatonin and AMK (10(-11)-10(-3) m), but not AFMK, inhibited nNOS activity in vitro in a dose-response manner. The IC(50) value for AMK (70 microm) was significantly lower than for melatonin (>1 mm). A 20% nNOS inhibition was reached with either 10(-9) m melatonin or 10(-11) m AMK. AMK inhibits nNOS by a non-competitive mechanism through its binding to Ca(2+)-calmodulin (CaCaM). The inhibition of nNOS elicited by melatonin, but not by AMK, was blocked with 0.05 mm norharmane, an indoleamine-2,3-dioxygenase inhibitor. In vivo, the potency of AMK to inhibit nNOS activity was higher than that of melatonin, as a 25% reduction in rat striatal nNOS activity was found after the administration of either 10 mg/kg of AMK or 20 mg/kg of melatonin. Also, in vivo, the administration of norharmane blocked the inhibition of nNOS produced by melatonin administration, but not the inhibition produced by AMK. These data reveal that AMK rather than melatonin is the active metabolite against nNOS, which may be inhibited by physiological levels of AMK in the rat striatum.

Animals↗

PubMed2006

Melatonin counteracts inducible mitochondrial nitric oxide synthase-dependent mitochondrial dysfunction in skeletal muscle of septic mice.

Mitochondrial nitric oxide synthase (mtNOS) produces nitric oxide (NO) to modulate mitochondrial respiration. Besides a constitutive mtNOS isoform it was recently suggested that mitochondria express an inducible isoform of the enzyme during sepsis. Thus, the mitochondrial respiratory inhibition and energy failure underlying skeletal muscle contractility failure observed in sepsis may reflect the high levels of NO produced by inducible mtNOS. The fact that mtNOS is induced during sepsis suggests its relation to inducible nitric oxide synthase (iNOS). Thus, we examined the changes in mtNOS activity and mitochondrial function in skeletal muscle of wild-type (iNOS(+/+)) and iNOS knockout (iNOS(-/-)) mice after sepsis. We also studied the effects of melatonin administration on mitochondrial damage in this experimental paradigm. After sepsis, iNOS(+/+) but no iNOS(-/-) mice showed an increase in mtNOS activity and NO production and a reduction in electron transport chain activity. These changes were accompanied by a pronounced oxidative stress reflected in changes in lipid peroxidation levels, oxidized glutathione/reduced glutathione ratio, and glutathione peroxidase and reductase activities. Melatonin treatment counteracted both the changes in mtNOS activity and rises in oxidative stress; the indole also restored mitochondrial respiratory chain in septic iNOS(+/+) mice. Mitochondria from iNOS(-/-) mice were unaffected by either sepsis or melatonin treatment. The data suggest that inducible mtNOS, which is coded by the same gene as that for iNOS, is responsible for mitochondrial dysfunction during sepsis. The results also suggest the use of melatonin for the protection against mtNOS-mediated mitochondrial failure.

Animals↗

PubMed2006

Pharmacological utility of melatonin in the treatment of septic shock: experimental and clinical evidence.

Sepsis is a major cause of mortality in critically ill patients and develops as a result of the host response to infection. In recent years, important advances have been made in understanding the pathophysiology and treatment of sepsis. Mitochondria play a central role in the intracellular events associated with inflammation and septic shock. One of the current hypotheses for the molecular mechanisms of sepsis is that the enhanced nitric oxide (NO) production by mitochondrial nitric oxide synthase (mtNOS) leads to excessive peroxynitrite (ONOO-) production and protein nitration, impairing mitochondrial function. Despite the advances in understanding of its pathophysiology, therapy for septic shock remains largely symptomatic and supportive. Melatonin has well documented protective effects against the symptoms of severe sepsis/shock in both animals and in humans; its use for this condition significantly improves survival. Melatonin administration counteracts mtNOS induction and respiratory chain failure, restores cellular and mitochondrial redox status, and reduces proinflammatory cytokines. Melatonin clearly prevents multiple organ failure, circulatory failure, and mitochondrial damage in experimental sepsis, and reduces lipid peroxidation, indices of inflammation and mortality in septic human newborns. Considering these effects of melatonin and its virtual absence of toxicity, the use of melatonin (along with conventional therapy) to preserve mitochondrial bioenergetics as well as to limit inflammatory responses and oxidative damage should be seriously considered as a treatment option in both septic newborn and adult patients. This review summarizes the data that provides a rationale for using melatonin in septic shock patients.

Animals↗

PubMed2005

Kynurenamines as neural nitric oxide synthase inhibitors.

To find new compounds with potential neuroprotective activity, we have designed, synthesized, and characterized a series of neural nitric oxide synthase (nNOS) inhibitors with a kynurenamine structure. Among them, N-[3-(2-amino-5-methoxyphenyl)-3-oxopropyl]acetamide is the main melatonin metabolite in the brain and shows the highest activity in the series, with an inhibition percentage of 65% at a 1 mM concentration. The structure-activity relationship of the new series partially reflects that of the previously reported 2-acylamido-4-(2-amino-5-methoxyphenyl)-4-oxobutyric acids, endowed with a kynurenine-like structure. Structural comparisons between these new kinurenamine derivatives, kynurenines, and 1-acyl-3-(2-amino-5-methoxyphenyl)-4,5-dihydro-1H-pyrazole derivatives also reported confirm our previous model for the nNOS inhibition.

Animals↗

PubMed2005

Identification of an inducible nitric oxide synthase in diaphragm mitochondria from septic mice: its relation with mitochondrial dysfunction and prevention by melatonin.

Sepsis provokes an induction of inducible nitric oxide synthase (iNOS) and melatonin down-regulates its expression and activity. Looking for an inducible mtNOS isoform, we induced sepsis by cecal ligation and puncture in both normal and iNOS knockout mice and studied the changes in mtNOS activity. We also studied the effects of mtNOS induction in mitochondrial function, and the role of melatonin against induced mtNOS and mitochondrial dysfunction. The activity of mtNOS and nitrite levels significantly increased after sepsis in iNOS+/+ mice. These animals showed a significant inhibition of the respiratory chain activity and an increase in mitochondrial oxidative stress, reflected in the disulfide/glutathione ratio, glutathione redox cycling enzymes activity and lipid peroxidation levels. Interestingly, mtNOS activity remained unchanged in iNOS-/- septic mice, and mitochondria of these animals were unaffected by sepsis. Melatonin administration to iNOS+/+ mice counteracted mtNOS induction and respiratory chain failure, restoring the redox status. The results support the existence of an inducible mtNOS that is likely coded by the same gene as iNOS. The results also suggest that sepsis-induced mtNOS is responsible for the increase of mitochondrial impairment due to oxidative stress in sepsis, perhaps due to the high production of NO. Melatonin treatment prevents mitochondrial failure at the same extend as the lack of iNOS gene.

Animals↗

PubMed2005

Melatonin reduces prostate cancer cell growth leading to neuroendocrine differentiation via a receptor and PKA independent mechanism.

BACKGROUND: Melatonin, the main secretory product of the pineal gland, inhibits the growth of several types of cancer cells. Melatonin limits human prostate cancer cell growth by a mechanism which involves the regulation of androgen receptor function but it is not clear whether other mechanisms may also be involved. METHODS: Time-course and dose-dependent studies were performed using androgen-dependent (LNCaP) and independent (PC3) prostate cancer cells. Cell number, cell viability, and cell cycle progression were studied. Neuroendocrine differentiation of these cells was evaluated by studying morphological and biochemical markers. Finally, molecular mechanisms including the participation of melatonin membrane receptors, intracellular cAMP levels, and the PKA signal transduction pathway were also analyzed. RESULTS: Melatonin treatment dramatically reduced the number of prostate cancer cells and stopped cell cycle progression in both LNCaP and PC3 cells. In addition, it induced cellular differentiation as indicated by obvious morphological changes and neuroendocrine biochemical parameters. The role of melatonin in cellular proliferation and differentiation of prostate cancer cells is not mediated by its membrane receptors nor related to PKA activation. CONCLUSIONS: The treatment of prostate cancer cells with pharmacological concentrations of melatonin influences not only androgen-sensitive but also androgen-insensitive epithelial prostate cancer cells. Cell differentiation promoted by melatonin is not mediated by PKA activation although it increases, in a transitory manner, intracellular cAMP levels. Melatonin markedly influences the proliferative status of prostate cancer cells. These effects should be evaluated thoroughly since melatonin levels are diminished in aged individuals when prostate cancer typically occurs.

Adjuvants, Immunologic↗

PubMed2005

Melatonin mitigates mitochondrial malfunction.

Melatonin, or N-acetyl-5-methoxytryptamine, is a compound derived from tryptophan that is found in all organisms from unicells to vertebrates. This indoleamine may act as a protective agent in disease conditions such as Parkinson's, Alzheimer's, aging, sepsis and other disorders including ischemia/reperfusion. In addition, melatonin has been proposed as a drug for the treatment of cancer. These disorders have in common a dysfunction of the apoptotic program. Thus, while defects which reduce apoptotic processes can exaggerate cancer, neurodegenerative disorders and ischemic conditions are made worse by enhanced apoptosis. The mechanism by which melatonin controls cell death is not entirely known. Recently, mitochondria, which are implicated in the intrinsic pathway of apoptosis, have been identified as a target for melatonin actions. It is known that melatonin scavenges oxygen and nitrogen-based reactants generated in mitochondria. This limits the loss of the intramitochondrial glutathione and lowers mitochondrial protein damage, improving electron transport chain (ETC) activity and reducing mtDNA damage. Melatonin also increases the activity of the complex I and complex IV of the ETC, thereby improving mitochondrial respiration and increasing ATP synthesis under normal and stressful conditions. These effects reflect the ability of melatonin to reduce the harmful reduction in the mitochondrial membrane potential that may trigger mitochondrial transition pore (MTP) opening and the apoptotic cascade. In addition, a reported direct action of melatonin in the control of currents through the MTP opens a new perspective in the understanding of the regulation of apoptotic cell death by the indoleamine.

Animals↗

PubMed2005

Selective CCK-A but not CCK-B receptor antagonists inhibit HT-29 cell proliferation: synergism with pharmacological levels of melatonin.

Some data suggest that cholecystokinin (CCK) receptor agonists stimulate the growth of colon cancer. Melatonin, an endogenous indoleamine with strong antioxidant properties, displays antiproliferative and proapoptotic properties both in vivo or in vitro in several types of tumors. We used HT-29 human colon cancer cells, expressing CCK receptors, to test the antiproliferative effects of several antagonists of CCK-A and/or CCK-B and their possible synergism with melatonin. HT-29 cells were cultured in RPMI 1640 medium supplemented with fetal bovine serum at 37 degrees C. Cell proliferation was assessed by the incorporation of [3H]-thymidine into DNA. Annexin V-FITC plus propidium iodine were used for flow cytometry apoptosis/necrosis evaluation. The following drugs were tested: gastrin (CCK-B agonist); CCK-8s (CCK-A agonist); proglumide (CCK-A plus CCK-B antagonist); lorglumide (CCK-A antagonist); PD 135,158 (CCK-B antagonist and weak CCK-A agonist); devazepide or L 364,718 (CCK-A antagonist); L 365,260 (CCK-B antagonist), and melatonin. The results shown a lack of effects of gastrin on HT-29 cell proliferation, whereas CCK-8s induced proliferation at high doses. The order of the antiproliferative effect of the other drugs was devazepide > lorglumide > proglumide. These drugs produce cell death mainly inducing apoptosis. Melatonin showed strong antiproliferative effect at millimolar concentrations, and it induced apoptotic cell death. Melatonin generally enhanced the antiproliferative effects of devazepide, lorglumide and proglumide and increased the proglumide-induced apoptosis. These results suggest that melatonin and CCK-A antagonists are useful for controlling human colon cancer cell growth in culture and in combined therapy significantly increases their efficiency.

Antineoplastic Agents↗

PubMed2005

Physiological ischemia/reperfusion phenomena and their relation to endogenous melatonin production: a hypothesis.

Ischemia/reperfusion is a frequently encountered phenomenon in organisms. Prolonged ischemia followed then by reperfusion results in severe oxidative injury in tissues and organs; however, some species can tolerate such events better than others. In nature, arousal from hibernation and resurfacing from diving causes animals to experience classic ischemia/reperfusion and, somehow, these animals cope well with the potential oxidative stress. It has been documented that during these physiological ischemia/reperfusion events, the activities of several antioxidant enzymes and the levels of some small-molecular-weight antioxidants become elevated. For example, the potent small-molecular-weight antioxidant melatonin often attains especially high levels during these physiological ischemia/reperfusion events including during arousal from hibernation or in the newborns during delivery. Highly elevated melatonin production during these physiological ischemia/reperfusion episodes exhibits several features. First, this high melatonin production is transient and fits well with the time schedule of the physiological ischemia/reperfusion period; therefore, it is not related to the normal endogenous melatonin rhythm. Yet, this transient peak protects the animals from destructive oxidative processes that occur during these transition periods. Second, these high levels of melatonin seem to derive from several organs since pinealectomy does not totally reduce circulating levels of this agent. Third, high melatonin production present at arousal from hibernation or in the newborns at birth does not appear to be controlled by light, i.e., it occurs both during the day and at night, and the amplitudes of elevated melatonin levels are equivalent at these times. The significance of these findings is discussed herein. Based on currently available data, we hypothesize that melatonin plays an important role in the physiological ischemia/reperfusion, i.e., as a member of antioxidant defense system, to protect against the potential oxidative injury induced by the physiological ischemia/reperfusion.

Animals↗

PubMed2005

Melatonin and nitric oxide: two required antagonists for mitochondrial homeostasis.

The presence of nitric oxide (NO* ) in the mitochondria led to analysis of its source and functions in mitochondrial homeostasis. Studies have revealed the existence of a mtNOS isoform with similar features to nNOS, with some post-traslational modifications, although without the typical signal peptide responsible for addressing proteins to mitochondrion. This isoform may account for the physiological production of NO* related to the respiratory control. During inflammatory conditions there is an excess of NO* in the mitochondria responsible for an increase in reactive oxygen and nitrogen species in sufficient amounts to compromise mitochondrial function. These conditions led to the discovery of the presence of an inducible mtNOS isoform with kinetic properties similar to iNOS. Experiments with knockout mice lacking either nNOS or iNOS further confirmed the existence of these two mtNOS isoforms in mitochondria. Although the increase in NO* in sepsis by inducible mtNOS may have important regulatory functions including the redistribution of oxygen into other pathways under hypoxia, it causes the production of excess NO* that is deleterious for the cell. Melatonin, an endogenous antioxidant, regulates mitochondrial respiration and bioenergetics and protects mitochondria from excess NO* by controlling the activity of mtNOS.

Animals↗

PubMed2004

4,5-dihydro-1H-pyrazole derivatives with inhibitory nNOS activity in rat brain: synthesis and structure-activity relationships.

In an attempt to find new compounds with neuroprotective activity, we have designed, synthesized and characterized 19 new nNOS inhibitors with a 4,5-dihydro-1H-pyrazole structure. Compounds 11r [1-cyclopropanecarbonyl-3-(2-amino-5-chlorophenyl)-4,5-dihydro-1H-pyrazole] and 11e [1-cyclopropanecarbonyl-3-(2-amino-5-methoxyphenyl)- 4,5-dihydro-1H-pyrazole] show the highest activities with inhibition percentages of 70% and 62%, respectively. A structure-activity relationship for the nNOS inhibition can be established from the structural comparison of these new pyrazole derivatives and the described synthetic kynurenines 10.

Aniline Compounds↗

PubMed2004

Changes in iNOS activity, oxidative stress and melatonin levels in hypertensive patients treated with lacidipine.

OBJECTIVE(S): To study the changes in macrophage inducible nitric oxide synthase (iNOS) activity, plasma levels of nitrite, lipid peroxidation (LPO) and melatonin in human essential hypertension before and 6 months after 4 mg/day lacidipine treatment. DESIGN: The study was carried out in a total of 25 subjects--11 healthy subjects and 14 hypertensive patients. Blood pressure and peripheral blood samples were taken before and after 6 months of lacidipine treatment (4 mg/day). METHODS: Systolic (SBP) and diastolic blood pressure (DBP), renal function, lipid and carbohydrate metabolism, renin, aldosterone and catecholamine levels were measured by routine methods. The activity of macrophage iNOS and plasma nitrite, LPO and melatonin levels were also measured. CONCLUSIONS: Besides reducing blood pressure, lacidipine treatment significantly decreased plasma LPO and macrophage iNOS activity, without changes in NO. Melatonin significantly increases in hypertensive patients, returning to control after lacidipine. Thus, lacidipine reduced blood pressure and free radicals, avoiding the oxidative damage to endothelium. It is suggested that administration of lacidipine plus melatonin may enhance the beneficial effects of each drug in essential hypertension.

Adult↗

PubMed2004

Effects of some synthetic kynurenines on brain amino acids and nitric oxide after pentylenetetrazole administration to rats.

We have previously proven that some synthetic kynurenines behave as antagonists of the N-methyl-d-aspartate receptor inhibiting neuronal subtype of nitric oxide synthase activity. We now investigate the anticonvulsant activity of four of these kynurenines in pentylenetetrazole (PTZ)-treated rats. The rats were treated with each kynurenine (10-160 mg/kg, s.c.) 30 min before PTZ administration (100 mg/kg, s.c.). Then, latency, duration and intensity of the first seizure and the percent animal survival were noted. PTZ-induced death was counteracted by high doses of kynurenines. Latency of the first seizure was significantly increased and its intensity reduced at the same doses, whereas the duration of the first seizure significantly decreased with doses of 20 mg/kg in most of the kynurenines tested. Three hours after PTZ administration, the surviving animals were sacrificed and the levels of brain amino acids and nitrite were measured. PTZ administration increased glutamate, glutamine, serine and taurine levels in different brain areas. High doses of kynurenines generally counteracted the effects of PTZ on excitatory amino acids, but they also reduced inhibitory aminoacids. However, the most consistent effect of kynurenines was the dose-dependent reduction of brain nitrite levels induced by PTZ. These results reveal a new family of anticonvulsant drugs that affect mainly to nitric oxide production in the brain.

Amino Acids↗

PubMed2003

Melatonin counteracts lipopolysaccharide-induced expression and activity of mitochondrial nitric oxide synthase in rats.

Mitochondrial nitric oxide synthase (mtNOS) is expressed constitutively, although it might be induced. Nitric oxide (NO) is a physiological regulator of mitochondrial respiration. Melatonin prevents mitochondrial oxidative damage and inhibits iNOS expression induced by bacterial lipopolysaccharide (LPS). The loss of melatonin with age may be related to the age-dependent mitochondrial damage. Thus, we examined the protective role of melatonin against the effects of LPS on mtNOS and on respiratory complexes activity in liver and lung mitochondria from young and old rats. The activity of mtNOS in control lung was low and did not change with age. LPS administration (10 mg/kg, i.v.) significantly increased mtNOS expression and activity and NO production in lung mitochondria, and the effect was greater in old rats. LPS administration also reduced the age-dependent decrease of the respiratory complexes I and IV. Melatonin administration (60 mg/kg, i.p.) prevented the LPS toxicity, decreasing mitochondrial NOS activity and NO production. Melatonin also counteracted LPS-induced inhibition of complexes I and IV. In general, the actions of melatonin were stronger in older animals than in younger ones. The results suggest that an inducible component of mtNOS, together with mitochondrial damage, occurs during sepsis, and melatonin prevents the mitochondrial failure that occurs during endotoxemia.

Aging↗

PubMed2003

Mitochondrial regulation by melatonin and its metabolites.

Our results show that melatonin and N-acetyl-5-methoxykynurenamine (aMK) physiologically regulate both the electron transport chain (ETC) and OXPHOS, increasing the electron transport and ATP synthesis by normal mitochondria. Melatonin also counteracts mitochondrial oxidative damage induced by t-butyl hydroperoxide, recovering glutathione levels and ATP production. However, the effects of melatonin not only depend of its antioxidant properties, since the indoleamine specifically interacts with complex I and IV of the ETC increasing their activity. Experiments in vivo showed that melatonin administration prevents sepsis-induced ETC damage decreasing the activity and expression of INOS and mtNOS, thus reducing intramitochondrial nitric oxide (NO) and peroxynitrite (ONOO-) levels. Consequently, mitochondrial ETC ad ATP production recovered to normal conditions. The presence of specific binding of melatonin in mitochondrial matrix led us to explore the genomic role of the indoleamine in these organelles. In vivo and in vitro experiments showed that administration of melatonin increased mtONA transcriptional activity of the subunits 1-3 of the complex IV. These effects correlated well with the effects of melatonin on complex IV activity. The data suggest a new rate for melatonin to regulate mitochondrial homeostasis. Due to the relationships between mitochondrial damage, aging and neurodegenerative diseases, the effects of melatonin here described further support its antiaging and neuroprotective properties.

Adenosine Triphosphate↗

PubMed2003

Characterization of melatonin high-affinity binding sites in purified cell nuclei of the hamster (Mesocricetus auratus) harderian gland.

In the present paper, binding of melatonin to purified cell nuclei from harderian glands of male and female hamsters was assessed. Binding of 125I-melatonin to cell nuclei fulfills the criteria for binding to a receptor site. Binding kinetics exhibit properties such as dependence on time and temperature as well as reversibility, saturability, high affinity and specificity. The dissociation constants (K(D)) and the number of binding sites (B(max)) for the binding of 125I-melatonin to harderian gland nuclei were 260 +/- 56 pm and 12.2 +/- 0.8 fmol/mg protein in male glands, and 280 +/- 43 pm and 9.8 +/- 0.6 fmol/mg protein in female glands, respectively. Competition experiments showed IC50 values for melatonin of 250 +/- 45 pm and 290 +/- 68 pm in male and female glands, respectively. Other indoleamines such as N-acetylserotonine and 5-metoxytryptamine showed IC50 values in the micromolar range, suggesting that the binding sites are specific for melatonin. Hill analyses of the data show nH values of 0.96-0.98, suggesting the existence of a single class of binding sites. These data indicate that specific 125I-melatonin binding sites exist in the cell nuclei of Harderian glands in male as well as in female hamsters, without significant differences between them. The K(D) and B(max) values obtained from the binding in both sexes correlates well with the concentration of melatonin described in these respective Harderian glands. It is hypothesized that the nuclear binding sites of melatonin here described could be a physiological melatonin receptor, which may be involved in the genomic-dependent antioxidant effects of melatonin on hamster Harderian glands elsewhere reported.

Animals↗

PubMed2003

Specific binding of melatonin to purified cell nuclei from mammary gland of swiss mice: day-night variations and effect of continuous light.

Melatonin binding sites were characterized in the nuclei of mouse mammary glands. The specific binding of 2-125I-iodomelatonin by homogenates of purified mammary gland cell nuclei was found to be rapid, reversible and saturable. Binding of 125I-melatonin exhibited day-night variations with the highest binding affinity observed during the dark period and the lowest affinity at midday. However, when the animals were maintained under continuous light exposure, binding of 125I-melatonin to cell nuclei showed a higher affinity than the daytime values found in animals maintained in normal photoperiod. Scatchard analysis of the binding data revealed the existence of a significant night-day variation in the binding kinetics, compatible with the existence of two classes of binding sites: a high-affinity binding site expressed during the night, with KD = 185 +/- 36 pm and a binding capacity of 6.24 +/- 0.4 fmol/mg protein, and a low-affinity site with KD = 562 +/- 57 nm and Bmax = 7.56 +/- 0.49 pmol/mg protein during the day. Interestingly, after 2 wk of continuous exposure to light, the animals killed at an equivalent midday time showed a significant increase in binding affinity, with KD = 1.43 +/- 0.2 nm and Bmax = 24.75 +/- 3.5 fmol/mg protein. Displacement experiments show an IC50 value compatible with the affinity constants obtained in the saturation experiments. These results indicate that the low-affinity binding site may be saturated by the high levels of melatonin found in the mouse mammary gland, and sustain the hypothesis of a circadian regulation of these melatonin binding sites by the photoperiod.

Animals↗

PubMed2003

Changes in brain amino acids and nitric oxide after melatonin administration in rats with pentylenetetrazole-induced seizures.

We examined the effect of melatonin on brain levels of amino acids and nitric oxide (NO) after pentylenetetrazole (PTZ)-induced seizures in rats. Animals were treated with melatonin (10-160 mg/kg, i.p.) 30 min before PTZ administration (100 mg/kg, s.c.), and were killed 3 hr later. At the dose of 80 mg/kg, melatonin significantly increased the latency (5.7-12.7 min) and decreased the duration (31.2-18.4 s) of the first seizure, reducing PTZ induced mortality from 87.5 to 25%. After kill, brains were removed and neurotransmitters and nitrite levels measured in prefrontal cortex (PF), parieto-temporal cortex (PF), striatum (ST), hippocampus (HP) and brain stem (BS) by high performance liquid chromatography. PTZ treatment increased glutamine levels in all brain areas studied, without changes in glutamate, gamma-amino butyric acid (GABA) and glycine. Aspartate and taurine increased in PF and PT and in HS and PT, respectively. Melatonin administration displayed a dose-dependent effect. At doses of 10-40 mg/kg, melatonin counteracted the PTZ-induced glutamine increase and reduced both glutamate and aspartate levels in the studied areas, with minor changes in GABA and glycine content. At doses of 80 and 160 mg/kg, the levels of glutamine, and glutamate, and to a lesser extent aspartate increased, whereas serine levels did not change. These two doses of melatonin also increased taurine, GABA and glycine in most brain areas studied. Treatment with melatonin (40-160 mg/kg) significantly decreased nitrite content in PT cortex, ST and BS areas of epileptic rats, without changes in the other brain regions. The results suggest that the anticonvulsant property of melatonin involves a modulation of both brain amino acids and NO production.

Amino Acids↗