Hyperglycaemia in experimental glaucoma.

Abstract

Glaucoma refers to a family of optic neuropathies with multi-factorial aetiology. The pathogenesis of glaucoma remains unclear, but there is good evidence that the optic nerve head is involved early in the pathogenesis of the disease. Inadequate blood supply to the optic nerve head may play a role, at least in some types of glaucoma. Given that vasculopathy is a hallmark of diabetes, one would expect that diabetes might exacerbate glaucoma; however, in large epidemiological studies no clear association was found. The Ocular Hypertension Treatment Study even suggested that diabetes protected against the conversion of ocular hypertension to glaucoma. In this thesis, I attempted to investigate the effect of short-term hyperglycaemia on retinal ganglion cell death and optic nerve damage in an experimental rat model of chronic ocular hypertension, which consisted of laser photocoagulation of the trabecular meshwork. The thesis is made up of four papers. The first paper characterises the rat model for our laboratory and validates the laser parameters used, which, in comparison to the original publication describing the model, have been slightly modified to minimise the ocular complications. A combination of histology, immunohistochemistry, Western blotting and realtime polymerase chain reaction was used to portray the spatial and temporal nature of retinal ganglion cell pathology. The data provides robust support for the hypothesis that the optic nerve head is the pivotal site of retinal ganglion cell injury, with resulting anterograde degeneration of axons and retrograde injury and death of perikarya. It was found that disruption of axonal transport occurs very soon after ocular hypertension, prior to structural damage, substantiating the hypothesis that axonal dysfunction may be an important cause of retinal ganglion cell degeneration. Moreover, as a novel finding, restricted axonal regeneration were observed at the optic nerve head. The second and third papers address the issue of damage quantification in the optic nerve. Axon counting on semi-thin optic nerve cross-sections represents the gold standard to evaluate the extent of axonal injury. However, this method is very laborious and time consuming. In search for alternatives, I investigated the accuracy of different sampling methods to estimate optic nerve axon numbers on cross sections and the usefulness of immunohistochemical markers on longitudinal optic nerve sections. Random sampling of pictures for automated axon counting was sufficiently accurate and the microglial response proved very valuable and effective for quantification of optic nerve damage. The thesis culminates in the fourth paper, which presents a limited reproduction of the Ocular Hypertension Treatment Study in a laboratory environment. Unilateral ocular hypertension was induced in two groups (n=26 per group) of Sprague-Dawley rats. One group remained normoglycaemic; the other was rendered hyperglycaemic by intraperitoneal injection of streptozotocin. After two weeks of elevated intraocular pressure, axonal and retinal damage were compared using the quantification methods introduced in the previous papers. There was convincing evidence for delayed axonal degeneration and retinal ganglion cell death in the hyperglycaemic rats. Axonal loss was reduced by about 50%. Survival of retinal ganglion cell somata was increased to a similar extent in hyperglycaemic rats. Hence, energy substrate availability may play a role in glaucomatous optic neuropathy. Targeted manipulation of neuronal energy metabolism may delay optic nerve degeneration and may represent a novel neuroprotective strategy for neurodegenerative diseases of the visual system such as glaucoma.