Anatomy and Neuroscience - Theses
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ItemThe role of the purinergic system in the normal and diseased retina( 2015)Extracellular adenosine 5’-triphosphate (ATP), its breakdown products and other related nucleotides are chemical transmitters mediating purinergic signaling within the central and peripheral nervous systems. In the retina, ATP has been shown to act as a neuro- and glio-transmitter that is important for visual processing and maintaining tissue homeostasis. However, the cellular expression and potential role of different components in the purinergic signaling pathway remains to be characterized in the retina. The mechanism by which purinergic signaling mediates the pathogenesis of retinal degeneration also remains ill-defined. The fundamental aim of this study was to investigate the role of purinergic signaling in the normal mammalian retina and to determine how this signaling pathway is involved in the mechanisms of retinal degeneration. The cellular localization of the vesicular nucleotide transporter (VNUT), the synaptic vesicles in which ATP is stored, was explored in the mammalian retina. With the use of a novel polyclonal antibody directed against VNUT, specific expression in dopaminergic interplexiform cells (IPCs) in the retina was demonstrated by fluorescence immunohistochemistry and confocal microscopy. Further investigations by three-dimensional reconstructions revealed that VNUT-expressing IPC distal varicosities were in close contact with horizontal cell processes and cone photoreceptor terminals in the outer retina, suggesting a role of vesicular ATP release in modulating outer retinal processing. The expression profile of the P2X4 receptor (P2X4-R) was examined in detail with reference to the specific neuronal and glial cell classes in the retina. Fluorescence immunohistochemistry and pre-embedding immuno-electron microscropy together demonstrated post-synaptic expression of the P2X4-R in both plexiform layers of the retina. In the outer retina, P2X4-R expression was identified on horizontal cell somata and processes. In the inner retina, P2X4-R immunoreactivity on amacrine and ganglion cells was observed. Furthermore, P2X4-R expression was detected on all glial cell types in the retina. These data indicate that P2X4-Rs are likely to play a role in the lateral inhibitory pathways, as well as a role in macro- and micro-glial signaling in the retina. The role of purinergic signaling in the cellular and vascular responses of the retino-choroidal complex to laser-induced injury was investigated. Application of the nanosecond laser at suprathreshold energy setting in P2X7-R knockout (P2X7-KO) mice resulted in an attenuated inflammatory response at the RPE/choroidal layer, indicating a role for purinergic signaling in mediating inflammation in response to retinal injury. The safety and efficacy benefits of the nanosecond laser system over standard photocoagulation lasers were also demonstrated by the lack of post-treatment neovascularization. Taken together, these findings indicate a critical involvement of the purinergic system in modulating diverse signal transmissions and maintaining tissue integrity in the normal mammalian retina. This study also provides evidence that the purinergic system has an important role in mediating the inflammatory response to retinal pathology.