Neuroprotective Effect of N-nitro-L-arginine Methylester Pretreatment on the Early Stage of Kainic Acid Induced Neuronal Degeneration in the Rat Brain

OBJECTIVE: Kainic acid(KA) enhances the expression of nitric oxide synthase, increases nitric oxide(NO), and thus evokes epileptic convulsion, which results in neuronal damage in the rat brain. NO may stimulate cyclooxygenase type-2 (COX-2) activity, thus producing seizure and neuronal injury, but it has also been reported that KA-induced seizure and neurodegeneration are aggravated on decreasing the COX-2 level. This study was undertaken to investigate whether the suppression of NO using the NOS inhibitor, N-nitro-L-arginine methyl ester(L-NAME), suppresses or enhances the activity of COX-2. METHODS: Silver impregnation and COX-2 immunohistochemical staining were used to localize related pathophysiological processes in the rat forebrain following KA-induced epileptic convulsion and L-NAME pretreatment. Post-injection survival of the rat was 1, 2, 3days and 2months, respectively. RESULTS: After the systemic administration of KA in rats, neurodegeneration increased with time in the cornu ammonis (CA) 3, CA 1 and amygdala, as confirmed by silver impregnation. On pretreating L-NAME, KA-induced neuronal degeneration decreased. COX-2 enzyme activities increased after KA injection in the dentate gyrus, CA 3, CA 1, amygdala and pyriform cortex, as determined by COX-2 staining. L-NAME pretreatment prior to KA-injection, caused COX-2 activities to increase compared with KA- injection only group by 1day and 2days survival time point. CONCLUSION: These results suggest that L-NAME has a neuroprotective effect on KA-induced neuronal damage, especially during the early stage of neurodegeneration.

Nitric Oxide-Mediated Neurotoxicity after Excimer Laser Photorefractive Keratectomy

The aim of this study was to evaluate neurotoxicity of Nitric oxide(NO) on cornea after excimer laser photorefractive keratectomy(PRK). PRK was performed on rabbit eyes. According to the time table, tear samples were collected with microcapillary tubes and corneal sensitivity was measured with a Cochet-Bonnet esthesiometer. No generation in the tear fluid was analyzed. To demonstrate NO Synthase(NOS), immunohistochemical localization was performed on frozen sections from rat eyeball tissue. Western blot analysis was used for detection of peroxynitrite, powerful oxidant of NO. NO generation was increased and reached to a maximum value(0.69+/-0.22micrometer/microgram) after 96 hours of PRK, as compared with in normal subjects(Mean: 0.30+/-0.08micrometer/microgram) and was not increased in the treated group with topical application of Ng-nitro-L-arginine methyl ester(L-NAME), a competitive inhibitor of constitutive NOS(cNOS) and inducible NOS(iNOS). Corneal sensitivity decreased below pretreatment levers after three postoperative days, but it was not observed in the L-NAME applied group. We have confirmed that a very strong iNOS and BNOS immunoreactivity was present in corneal keratocytes. Western blot analysis identifed the bands of nitrotyro-sine-proteins suggesting in vivo peroxynitrite toxicity. Our results suggested that NO generated from the enzyme after PRK decreased corneal sensitivity by damaging corneal sensory nerve through the NO and iths oxidant peroxitrite. Therefore topical application of a NOS inhibitor may be effective in maintaining corneal sensitivity.