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Y. Yomura, Y. Shoji, Y. Munemasa, Y. Hayashi, Y. Kitaoka, E. Murakami, H. Nakashima, S. Ueno; Direct Real–Time Monitoring of Intravitreal Nitric Oxide (NO) and Oxygen (pO2) Simultaneously in Endotoxin–Induced Uveitis (EIU) of Rabbits . Invest. Ophthalmol. Vis. Sci. 2006;47(13):4555.
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NO plays a critical role in the pathogenesis of EIU. Since NO is a labile free radical, it was difficult to access the time course of NO generation directly in vivo. Moreover, there were few reports that indicated interaction between NO and pO2. The purpose of study is to establish the direct monitoring system for NO and pO2 dynamics under EIU in vivo.
We developed a new microdialysis probe that combined a NO–selective microelectrode and an amperometric oxygen microelectrode for real–time monitoring of NO concentration and pO2 in vivo. The current (calculated concentration) of NO and pO2 in the vitreous of rabbits was monitored after the intravitreal injection of saline or 100ng lipopolysaccharide (LPS, from E.coli). Thereafter, these eyes were enucleated and immunostained with an anti–nitrotyrosine antibody.
NO and pO2 generation could be monitored simultaneously in the vitreous for several hours. Saline had no effect on either levels of NO or pO2 compared to basal levels. However, NO increased in waves 30∼45 minutes after LPS injection, and pO2 gradually increased soon after LPS injection. The current returned to respective baseline levels within 2 hours. Furthermore, pO2 moved in the opposite direction at the point of time whether increased or decreased in the NO levels, that we observed in this study. Immunohistochemical study showed that an increase of nitrotyrosine immunoreactivity was seen in the retinal ganglion cell layer, inner plexiform layer and photoreceptor layer after LPS injection.
This study demonstrated the feasibility of monitoring intravitreal NO and pO2 simultaneously in EIU of rabbits. Our direct real–time monitoring system showed not only the elevated NO levels in early phase following LPS injection but also the close relationship between NO production and pO2 in vivo. Furthermore, we confirmed the production of nitrotyrosine, as a marker of peroxynitrite. It seemed that NO and oxygen had reacted readily to produce the cytotoxic spiecies, peroxynitrite. This process may be related to the retinal damage in EIU. Acordingly, this direct monitoring system would provide useful information for EIU by dynamic variation of NO and pO2.
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