Melatonin Regulates Nitric Oxide Synthase Expression in Ischemic Brain Injury

Abstract

ABSTRACT. Nitric oxide (NO) is produced by three NO synthases (NOS), iNOS, eNOS, and nNOS. Production of NO by iNOS plays key roles in neurodegeneration, while eNOS is a protective enzyme. This study investigated the neuroprotective effect of melatonin and the levels of NOS isoforms induced by melatonin in ischemic brain injury. Adult male rats were treated with melatonin (5 mg/kg) or vehicle prior to middle cerebral artery occlusion (MCAO). Brains samples were collected at 24 hr after the onset of occlusion. Results confirmed that melatonin significantly reduces infarct area. Western blot analysis was used to evaluate the expression levels of iNOS, eNOS, and nNOS. The level of iNOS and nNOS increased in vehicle-treated animals, while melatonin prevented injury-induced increase of iNOS. In contrast to iNOS levels, eNOS levels decreased in vehicle-treated animals, while melatonin prevented the injury-induced decrease of eNOS. This study provides further evidence that melatonin exerts neuroprotective effects, and the regulation of NOS isoforms by melatonin may contribute to the neuroprotective effects. KEY WORDS: melatonin, neuroprotection, NOS.J. Vet. Med. Sci. 70 (7): [747][748][749][750] 2008 Melatonin has a number of physiological functions, including the regulation of circadian rhythms, the inhibitin of the oxidation of biomolecules, and the removal of free radicals [14,20]. Moreover, melatonin plays a neuroprotective role against transient or permanent ischemic brain injury [8,10,16]. In clinical studies, a decrease of melatonin is related to neurodegenerative diseases such as Parkinson' disease and Alzheimer's disease [3,18].Nitric oxide (NO) is a critical player in pathological and physiological processes in the central nervous system [19,22]. NO is generated by the activation of three distinct nitric oxide synthases (NOS), inducible NOS (iNOS), endothelial NOS (eNOS), and neuronal NOS (nNOS). NO exerts both neuroprotective and neurotoxic effects in focal cerebral ischemia [4,17]. iNOS produces NO with cytotoxic effects, whereas eNOS plays a protective role. Furthermore, melatonin has antioxidant activity and can inhibit free radical formation thereby decreasing NOS [2,12]. Although previous studies have elucidated the neuroprotective mechanism of melatonin, little data is available regarding the levels of NOS isoforms. Thus, this study investigated the neuroprotective effect of melatonin and its effect on the expression of NOS isoforms after ischemic brain injury.All procedures were carried out according to the Guide for the care and use of laboratory animals, published by the U.S. National Institutes of Health. Male Sprague-Dawley rats (230-250 g, n=30) were used in this study. Animals were maintained under controlled temperature (25°C) and lighting (12/12 light/dark cycle) and allowed to have free access to food and water. Animals were randomly divided into two groups, vehicle-treated group and melatonintreated group (n=15 per group). All experiments were performed during the day between 10:00 and 16...

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