Intraocular inflammatory disease is first recognized by breakdown of the blood aqueous barrier with appearnce of protein and leukocytes in the anterior chamber. Secondary problems include cataract, corneal clouding, glaucoma and anterior synechia. Major signs of inflammation are attributed to the release of inflammatory mediators, including the eicosanoids derived from arachidonic acid. The ling-term goal of this prject is to fully understand the role of eicosanoids in the mechanisms involved in the initiation, sustenance and resolution of ocular inflammation and also to contribute to the development of impproved therapy. The action of eicosanoids is mediated ghrough specific receptors located in the cell membrane. Our previous studies have demonstrated that the eye possesses a great diversity of prostanoid receptor subtypes in keeping with the multiple actions of prostanoids in the eye. We now hypothesize that the ocular effects of eicosanoids are modified by up and down- regulation of the receptors. To test this hypothesis we will (1) demonstrate that diminished levels of receptor agonists (in the presence of inhibitors of eicosanoid synthesis) evoke up-regulation of specific receptors in cultured human non-pigmented epithelial cells (EP2, EP4 receptors) and cultured human ciliary muscle cells (Ep1, Ep2, F2alpha receptors). Receptor levels will be evaluated in terms of radioligando binding data, stimulation of second messenger production and mRNa levels. We will also (2) demonstrate that down-regulation of specific receptors can be evoked by prolonged exposure to natural or synthetic receptor agonists. As a bridge to furture in vivo studies we will (3) use the cultured cell system to explore reeptor regulation under conditions which mimie those encountered in vivo. Receptor regulation will be muesured in the presence of ascorbic acid or hydrogen peroxide which can alter the oxidative tone the environment or in the presence of bactrial endotoxin, which is known to incite an experimental inflammatory response. Both oxidation and endotoxin can impact upon arachidonic acid metabolism. These studies will generate significant information and contribute to our understanding of prostaglandin actions in the normal and inflamed eye.
