Antonio Muñoz-Hoyos

Pharmacological actions of melatonin in oxygen radical pathophysiology.

Melatonin, the chief secretory product of the pineal gland, was recently found to be a free radical scavenger and antioxidant. This review briefly summarizes the published reports supporting this conclusion. Melatonin is believed to work via electron donation to directly detoxify free radicals such as the highly toxic hydroxyl radical. Additionally, in both in vitro and in vivo experiments, melatonin has been found to protect cells, tissues and organs against oxidative damage induced by a variety of free radical generating agents and processes, e.g., the carcinogen safrole, lipopolysaccharide, kainic acid, Fenton reagents, potassium cyanide, L-cysteine, excessive exercise, glutathione depletion, carbon tetrachloride, ischemia-reperfusion, MPTP, amyloid beta (25-35 amino acid residue) protein, and ionizing radiation. Melatonin as an antioxidant is effective in protecting nuclear DNA, membrane lipids and possibly cytosolic proteins from oxidative damage. Also, melatonin has been reported to alter the activities of enzymes which improve the total antioxidative defense capacity of the organism, i.e., superoxide dimutase, glutathione peroxidase, glutathione reductase, glucose-6-phosphate dehydrogenase, and nitric oxide synthase. Most studies have used pharmacological concentrations or doses of melatonin to protect against free radical damage; in a few studies physiological levels of the indole have been shown to be beneficial against oxidative stress. Melatonins function as a free radical scavenger and antioxidant is likely assisted by the ease with which it crosses morphophysiological barriers, e.g., the blood-brain barrier, and enters cells and subcellular compartments. Whether the quantity of melatonin produced in vertebrate species is sufficient to significantly influence the total antioxidative defense capacity of the organism remains unknown, but its pharmacological benefits seem assured considering the low toxicity of the molecule.

Melatonin prevents changes in microsomal membrane fluidity during induced lipid peroxidation

We tested the effect of melatonin on membrane fluidity in microsomes of a rat liver model in which lipid peroxidation was induced by the addition of FeCl3, ADP and NADPH. Membrane fluidity was monitored using fluorescence spectroscopy and lipid peroxidation was estimated by quantifying malonaldehyde (MDA)+4‐hydroxyalkenals (4‐HDA) concentrations following the induction of lipid peroxidation with and without pre‐incubation with melatonin (1 μM–3 mM). Membrane rigidity increased during induced lipid peroxidation while melatonin reduced in a concentration‐dependent manner both membrane rigidity and MDA+4‐HDA generation. Melatonins protective effect may relate to its known ability to scavenge free radicals and function as an antioxidant.

Utility of high doses of melatonin as adjunctive anticonvulsant therapy in a child with severe myoclonic epilepsy: two years' experience.

Molina‐Carballo A, Muñoz‐Hoyos A, Reiter RJ, Sánchez‐Forte M, Moreno‐Madrid F, Rufo‐Campos M, Molina‐Font JA, Acuña‐Castroviejo D. Utility of high doses of melatonin as adjunctive anticonvulsant therapy in a child with severe myoclonic epilepsy: Two years experience. J. Pineal Res. 1997; 23:97–105.

Melatonin treatment normalizes plasma pro‐inflammatory cytokines and nitrosative/oxidative stress in patients suffering from Duchenne muscular dystrophy

Abstract:u2002 Duchenne muscular dystrophy (DMD), a lethal disorder characterized by dystrophin absence, courses with chronic inflammation, sarcolemmal damage, and skeletal muscle degeneration. Among the multiple pathogenic mechanisms proposed for DMD, oxidative stress and inflammation are directly involved in the dystrophic process. Unfortunately, there is no current treatment for DMD, and the inflammatory process is an important target for therapies. Based on the antioxidant and anti‐inflammatory properties of melatonin, we investigated whether melatonin treatment may reduce the dystrophic process. Ten DMD patients aged 12.8u2003±u20030.98u2003yr, were treated with melatonin (60u2003mg at 21:00u2003hr plus 10u2003mg at 09:00u2003hr), and plasma levels of lipid peroxidation (LPO), nitrites (NOx), interleukin (IL)‐1β, IL‐2, IL‐6, tumor necrosis factor‐α, interferon‐γ, and plasma markers of muscle injury, were determined at 3, 6 and 9u2003months of treatment. Healthy age‐ and sex‐matched subjects were used as controls. The results show a significant increase in LPO, NOx, and cytokine levels in plasma of DMD patients compared with controls. Melatonin administration reduced these values to control levels at 3u2003months of treatment, decreasing further 9u2003months later. In parallel, melatonin also reduced plasma levels of creatine kinase (CK; 50%), lactate dehydrogenase (28%), aspartate aminotransferase (28%), alanine aminotransferase (20%), and myoglobin (13%). These findings strongly support the conclusion that melatonin administration significantly reduced the hyperoxidative and inflammatory process in DMD patients, reducing the muscle degenerative process.

Melatonin and vitamin E limit nitric oxide-induced lipid peroxidation in rat brain homogenates.

We have investigated the level of lipid peroxidation (LPO) in rat brain homogenates in the presence of nitric oxide (NO) which was released by the addition of sodium nitroprusside (SNP) and compared it with that induced by H2O2. We also examined the effect of melatonin and vitamin E on the NO-induced LPO. The concentration of malonaldehyde (MDA) plus 4-hydroxyalkenals (4-HDA) was used as an index of LPO. While both H2O2 and SNP increased MDA + 4-HDA production in brain homogenates in a concentration-dependent manner, SNP was more potent than H2O2 at all concentrations tested. Both melatonin or vitamin E reduced NO-induced LPO in a dose-dependent manner in concentrations ranging from 10 microM to 10 mM. Under the in vitro conditions of this experiment, vitamin E was more efficient than melatonin in limiting NO-induced LPO in rat brain homogenates.

Melatonin's role as an anticonvulsant and neuronal protector: Experimental and clinical evidence

The pineal gland classically has been considered as a vestigial and mystic organ. In the last decades, and with the incorporation of new methodologic procedures, it could be proved that it also has physiologic actions that vary depending on the level of the phylogenetic scale. Its best-known secretion, melatonin, has been related to many different actions, such as sleep promotion, control of biologic rhythms, hormonal inhibition, and an inhibiting action on central nervous system regulation mechanisms. In animal experimentation, there are papers even accepting an anticonvulsant effect. In humans, evidence is reduced to few experiences. In addition to this clinical experience, there is other evidence that clearly relates melatonin to convulsive phenomena. This relationship must be mediated by the following mechanisms attributed to melatonin: altered brain GABAergic neurotransmission, its known interaction with benzodiazepinic brain receptors, through tryptophan metabolite activity (kynurenine, kynurenic acid), or even by its efficacy as a free-radical scavenger. (J Child Neurol 1998;13:501-509).

Day-night variations in melatonin secretion by the pineal gland during febrile and epileptic convulsions in children

The pineal gland is a complex neuroendocrine organ with pronounced effects on central nervous system activity. Because previous studies in animals and humans have suggested an anticonvulsant role for the pineal hormone melatonin, we studied the day-night variations in plasma melatonin in normal children and children with febrile or epileptic convulsions. We found significant changes in day-night melatonin levels during convulsions, consistent with the hypothesis that melatonin has an inhibitory function on central nervous system activity.

Normalization of the sleep–wake pattern and melatonin and 6-sulphatoxy-melatonin levels after a therapeutic trial with melatonin in children with severe epilepsy

Abstract:u2002 This study evaluated the sleep–wake pattern, plasma melatonin levels and the urinary excretion of its metabolite, 6‐sulphatoxy‐melatonin among children with severe epileptic disorders, before and after a therapeutic trial with melatonin. Ten paediatric patients, suffering from severe epileptic disorders, were selected and given a nightly dose of 3u2003mg of a placebo, for 1u2003wk; for the next 3u2003months, the placebo was replaced with a nightly dose of 3u2003mg of melatonin. At the end of each treatment period, the urinary excretion of 6‐sulphatoxy‐melatonin (for the intervals 09.00 – 21:00u2003hr or 21:00–09:00u2003hr) and plasma levels of melatonin (recorded at 01:00, 05:00, 09:00, 13:00, 17:00 and 21:00u2003hr) were recorded, over a period of 24u2003hr; an actigraph record was also kept. Sleep efficiency among patients who received melatonin was significantly higher than among those given the placebo, with fewer night‐time awakenings. Periodic plasma melatonin levels were regained and a better control gained of convulsive episodes, in that the number of seizures decreased. We conclude that melatonin is a good regulator of the sleep–wake cycle for paediatric patients suffering from severe epilepsy, moreover, it to a better control of convulsive episodes.

Melatonin protects against lipid peroxidation and membrane rigidity in erythrocytes from patients undergoing cardiopulmonary bypass surgery

Abstract: The first aim of this study was to test whether there are changes in erythrocyte lipid peroxidation products and membrane fluidity during cardiac surgery involving cardiopulmonary bypass. Secondly, in vitro tests were performed to examine whether melatonin alters induced lipid peroxidation and reduced membrane fluidity in erythrocytes from these patients. Fifteen patients undergoing cardiac surgery involving cardiopulmonary bypass (CPB) were selected. Five blood samples were taken at different times during surgery for analysis of thiobarbituric acid reactive substances (TBARS) content and membrane fluidity of the erythrocytes. TBARS are an index of the level of lipid peroxidation. The results revealed an increase in TBARS levels and a parallel decrease in erythrocyte membrane fluidity (increased membrane rigidity) after the onset of CPB with respect to the reference sample. Likewise, in vitro induction of lipid peroxidation in the erythrocyte samples from CPB patients followed a similar pattern. The changes in TBARS levels and membrane fluidity were suppressed by pre‐incubation of erythrocytes with melatonin prior to the induction (by 70u2003μm Fe2+u2003+u2003250u2003μm ascorbate) of lipid peroxidation in a concentration‐dependent manner. The results constitute a persuasive argument for the use of melatonin for preventive and therapeutic purposes during CPB.

Melatonin concentration in the umbilical artery and vein in human preterm and term neonates and neonates with acute fetal distress

Abstract: In order to assess the existence of a rhythmic secretion of melatonin (aMT) in newborns and whether this rhythm is affected by neonatal stress, we studied 112 newborns classified in three groups: normal babies delivered at term, preterm infants born before the 38th week, and babies with fetal distress. Melatonin was measured by RIA in the umbilical artery and vein at the time of birth. Melatonin levels in umbilical arterial and venous blood showed a diurnal rhythm in all groups. Melatonin levels in umbilical cord artery and vein were closely related. Nocturnal melatonin levels were increased in newborns with acute fetal distress in comparison with normal term and preterm neonates. These results suggest that (1) a rhythm of aMT secretion exists in newborns, although it cannot be determined whether this rhythm is of maternal or fetal origin and (2) neonatal stress (acute fetal distress) increases aMT production during the night in comparison with normal term and preterm neonates.