Elevated intraocular pressure is a major risk factor for glaucomatous optic neuropathies. The glaucomas are a family of irreversible blinding diseases, which affect as many as 15 million persons in the United States and over 150 million persons world-wide. This proposal is focused on the basic question, how is intraocular pressure regulated in the normal eye. Much of intraocular pressure regulation resides in the trabecular meshwork, which is the source of the resistance to aqueous humor outflow. Thus the question simplifies to, how does the trabecular meshwork adjust the outflow resistance? The basic working hypothesis of this proposal is that the trabecular extracellular matrix is the primary source of resistance to aqueous humor outflow and that trabecular cells regulate outflow by changing the turnover and subsequent replacement biosynthesis of this extracellular matrix. This hypothesis will be tested by evaluating changes in the outflow facility of perfused anterior segment explant organ cultures exposed to conditions and agents which modulate the trabecular extracellular matrix. A family of matrix metalloproteinases, regulated by their tissue inhibitors, TIMPs, are primary candidates for initiating extracellular matrix turnover. Conditions and agents, which modify the activity or manipulate the expression of the metalloproteinases and TIMPs, will be used to perturb trabecular homeostasis. Consequent changes in the trabecular outflow resistance will then be assessed using perfused human anterior segment cultures. Among the perturbations to be used are, specific synthetic metalloproteinase inhibitors, recombinant and purified native TIMPs, recombinant and purified native metalloproteinases, several factors which induce expression of the metalloproteinases or TIMPs by trabecular cells, antisense oligonucleotides to metalloproteinase and TIMP transcripts, and gene transfer of the coding sequences for the metalloproteinases and TIMPs into trabecular cells to selectively overexpress these proteins within the outflow pathway. Extracellular matrix biosynthesis and turnover and component expression levels will be assessed as will trabecular metalloproteinase and TIMP activity and expression. These studies will provide a clear test of this working hypothesis and a basis for addressing the next questions, how does the meshwork know when and how much to regulate outflow resistance and what goes wrong in primary open-angle glaucoma?