The major purpose of this project is to investigate by means of molecular and biochemical pharmacological techniques the drugreceptor systems present in ocular ciliary processes, and to relate these systems to the control of aqueous humor secretion. A variety of receptor which are the site of action of several experimental drugs already known to lower intraocular pressure will be studied. These include receptors for dopamine, serotonin, alpha-adrenergic ligands, adenosine, and prostaglandins (PGF2o) which will be studied with respect to their recognition sites (by ligand binding), and the signal transduction/second messenger system they are coupled to (positive and negative regulation of adenylate cyclase, Ca2+/calmodulin, phospholipase C/phosphatidyl inositol turnover). In addition, the intracellular biochemical effectors that mediate cellular responses initiated by these five receptor types will be investigated (c-AMP dependent protein kinase, Ca2+/calmodulin dependent protein kinase, protein kinase C, and tyrosine kinase). Stimulusdependent phosphorylation of specific cellular proteins will also be determined. The proteins that undergo specific agonistmediated phosphorylation are responsible for the control of cell function and their identification for each type of receptor system is an ultimate objective of this work. The proposed studies on five receptor types, four signal transduction systems, and four kinase enzymes will involve a wide variety of drug probes. Many of these agents will also be investigated for their effects on aqueous humor dynamics by administration to the rabbit eye. Measurements of intraocular pressure and aqueous humor flow rates will be correlated with the biochemical and pharmacological investigations. Novel receptor agonists and antagonists, agents affecting Ca2+ or calmodulin (e.g. W-7, BAY K8644) and agents affecting kinase enzymes (e.g. H-7, phorbol esters) will be tested. The most promising agents will be investigated to a limited extent in the cynomolgus monkey to establish primate relevance. The long range goal of these studies is to establish the molecular bases for the regulation of aqueous humor formation by ciliary processes and to uncover potential new drug therapies for openangle glaucoma.
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