Aerobic cells use oxygen for the production of 90-95% of the total amount of ATP that they use. This amounts to about 40 kg ATP/day in an adult human. The synthesis of ATP via the mitochondrial respiratory chain is the result of electron transport across the electron transport chain coupled to oxidative phosphorylation. Although ideally all the oxygen should be reduced to water by a four-electron reduction reaction driven by the cytochrome oxidase, under normal conditions a small percentage of oxygen may be reduced by one, two, or three electrons only, yielding superoxide anion, hydrogen peroxide, and the hydroxyl radical, respectively. The main radical produced by mitochondria is superoxide anion and the intramitochondrial antioxidant systems should scavenge this radical to avoid oxidative damage, which leads to impaired ATP production.

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.

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We recently described that melatonin and some kynurenines modulate the N-methyl-D-aspartate-dependent excitatory response in rat striatal neurons, an effect that could be related to their inhibition of nNOS. In this report, we studied the effect of melatonin and these kynurenines on nNOS activity in both rat striatal homogenate and purified rat brain nNOS. In homogenates of rat striatum, melatonin inhibits nNOS activity, whereas synthetic kynurenines act in a structure-related manner. Kynurenines carrying an NH(2) group in their benzenic ring (NH(2)-kynurenines) inhibit nNOS activity more strongly than melatonin itself. However, kynurenines lacking the NH(2) group or with this group blocked do not affect enzyme activity. Kinetic analysis shows that melatonin and NH(2)-kynurenines behave as noncompetitive inhibitors of nNOS. Using purified rat brain nNOS, we show that the inhibitory effect of melatonin and NH(2)-kynurenines on the enzyme activity diminishes with increasing amounts of calmodulin in the incubation medium. However, changes in other nNOS cofactors such as FAD or H(4)-biopterin, do not modify the drugs' response. These data suggest that calmodulin may be involved in the nNOS inhibition by these compounds. Studies with urea-polyacrylamide gel electrophoresis further support an interaction between melatonin and NH(2)-kynurenines, but not kynurenines lacking the NH(2) group, with Ca(2+)-calmodulin yielding Ca(2+)-calmodulin-drug complexes that prevent nNOS activation. The results show that calmodulin is a target involved in the intracellular effects of melatonin and some melatonin-related kynurenines that may account, at least in part, for the neuroprotective properties of these compounds.

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Degeneration of nigrostriatal dopaminergic neurons is the major pathogenic substrate of Parkinson's disease (PD). Inhibitors of monoamine oxidase B (MAO-B) have been used in the treatment of PD and at least one of them, i.e., deprenyl, also displays antioxidant activity. Dopamine (DA) autoxidation produces reactive oxygen species implicated in the loss of dopaminergic neurons in the nigrostriatal pathway. In this study we compared the effects of melatonin with those of deprenyl and vitamins E and C in preventing the hydroxyl radical (8OH) generation during DA oxidation. The rate of production of 2,3-dihydroxybenzoate (2,3-DHBA) in the presence of salicylate, an *OH scavenger, was used to detect the in vitro generation of *OH during iron-catalyzed oxidation of DA. The results showed a dose-dependent effect of melatonin, deprenyl and vitamin E in counteracting DA autoxidation, whereas vitamin C had no effect. Comparative analyses between the effect of these antioxidants showed that the protective effect of melatonin against DA autoxidation was significantly higher than that of the other compounds tested. Also, when melatonin plus deprenyl were added to the incubation medium, a potentiation of the antioxidant effect was found. These findings suggest that antioxidants may be useful in brain protection against toxicity of reactive oxygen species produced during DA oxidation, and melatonin, alone or in combination with deprenyl, may be an important component of the brain's antioxidant defenses to protect it from dopaminergic neurodegeneration.

The recently described 'hydrogen hypothesis' invokes metabolic symbiosis as the driving force for a symbiotic association between an anaerobic, strictly hydrogen-dependent organism (the host) and an eubacterium (the symbiont) that is able to respire, but which generates molecular hydrogen as an end product of anaerobic metabolism. The resulting proto-eukaryotic cell would have acquired the essentials of eukaryotic energy metabolism, evolving not only aerobic respiration, but also the cost of oxygen consumption, i.e., generation of reactive oxygen species (ROS) and oxidative damage. Mitochondria contain their own genome with a modified genetic code that is highly conserved among mammals. Control of gene expression suggests that transcription of certain mitochondrial genes may be regulated in response to the redox potential of the mitochondrial membrane. Mitochondria are involved in energy production and conservation, and they have an uncoupling mechanism to produce heat instead of ATP. Also, mitochondria are involved in programmed cell death. Increasing evidence suggests the participation of mitochondria in neurodegenerative and neuromuscular diseases involving alterations in both nuclear (nDNA) and mitochondrial (mtDNA) DNA. Melatonin is now known as a powerful antioxidant and increasing experimental evidence shows its beneficial effects against oxidative stress-induced macromolecular damage and diseases, including those in which mitochondrial function is affected. This review summarizes the data and mechanisms of action of melatonin in relation to mitochondrial pathologies.

The existence of dopamine (DA)-melatonin (aMT) relationships is well documented in several brain areas of the mammalian central nervous system such as the retina and hypothalamus or the nigrostriatal system. For instance, aMT tempers 1 methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced nigrostriatal damage in C57BL/6 mice. In this mouse strain however, rhythmic production of aMT and its possible interaction with striatal DA is still unclear. In the present work we investigated circadian variations in pineal production of aMT and striatal DA levels in C57BL/6 mice. Effects of pinealectomy and aMT administration were also assessed. Intact, pinealectomized and pinealectomized + aMT-treated mice and their respective control groups were sacrificed at six different times during the 24-hour period. In control animals, aMT displayed a circadian rhythm with a narrow peak at midnight. The peak of aMT coincided with the nadir of the DA rhythm present in the striatum. Shortly after the decrease of DA levels, an increase in 3,4-dihydroxyphenylacetic acid (DOPAC), the main DA metabolite, was observed. The rhythmic changes of DA and DOPAC levels in the striatum were blunted by pinealectomy, whereas administration of aMT (0.1–10 mg/kg) during 6 days to pinealectomized mice restored the rhythms in a dose-dependent manner. Striatal levels of 3-methoxytyramine and homovanillic acid did not change during the 24-hour cycle. The serotonergic system, assessed by the determination of 5-hydroxytryptamine and 5-hydroxyindole-3-acetic acid concentration in striatum, did not show significant time-dependent changes in control animals and was not affected by pinealectomy or aMT treatment. These data substantiate the existence of a link between pineal function, melatonin secretion and DA circadian rhythm in the mouse striatum.