This project seeks ways to enhance aqueous humor outflow for glaucoma therapy. Specifically it will (1) determine the outflow enhancing potential of agents by examining their effects on cytoskeletal, cell junctional and extracellular matrix interaction profiles in vitro and effects on outflow facility in vitro and vivo, (2) determine the in vivo morphological and physiological effects on outflow of long-term therapy with cytoskeleton modulating agents (H-7 or latrunculin B) used in combination with common glaucoma therapies or resulting from over-expression of viral vector delivered transgenes encoding for cytoskeleton modulating proteins (caldesmon and C3), (3) create a new primate model of glaucoma by excess exposure to cochlin or TGFb2 which will be more relevant for studies of glaucoma targeting aqueous outflow enhancement, (4) determine the mechanisms of intraocular pressure (IOP) reduction of other classes of potential glaucoma therapeutic agents (cannabinoids, nitric oxide donors, prostaglandin EP4 agonists, angiotensin and chymase), and their interactions with existing glaucoma therapies. In vitro cell studies will be conducted in human trabecular meshwork and ciliary muscle cells using immunohistochemistry. Aqueous outflow will be determined by measuring outflow facility and uveoscleral outflow in monkeys. Trabecular outflow facility will be measured in monkey anterior segments in organ culture. Intraocular pressure (Goldmann applanation tonometry) and aqueous humor formation (fluorophotometry) will be measured in monkeys. Lentiviral vectors will deliver genes to the outflow pathways following intracameral injection. Glaucoma model progression will be monitored by IOP, nerve fiber layer thickness and contour, and retinal and central function by electrophysiology. Structural parameters in the anterior segment of intact eyes and organ cultured anterior segments will be evaluated by light/electron microscopy, immunohistochemistry and quantitative morphometry. This project should increase our knowledge of the physiological, pharmacological, neural and morphological mechanisms and responses governing aqueous humor drainage, and provide insights into the therapeutic and toxicological mechanisms of current and putative antiglaucoma drugs. Such knowledge will contribute to the development of new antiglaucoma drugs and perhaps to understanding the underlying biochemical, physiological, and morphological basis of the human open angle glaucomas. ? ? ?
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