INHIBITION OF RETINAL GROWTH CONE ACTIVITY BY SPECIFIC METALLOPROTEINASE INHIBITORS IN-VITRO

The developing neural retina expresses a set of extracellular protease s including plasminogen activator and gelatinases. Since neurites of r etina cells cultured on fluorescent gelatin digest the substrate in th eir paths, we have suggested that the proteases are used by the tips o f growing fibers to allow them to migrate within the mass of the tissu e in vivo. In order to obtain further information about relationships between extracellular proteases and fiber growth, we have examined the effects of the specific inhibitors HS-LFA (HS-Leu-Phenylala-Ala, enan tiomeric forms 1 and 2), bathophenanthroline sulfonate (BPS), phenylme thyl sulfonyl fluoride (PMSF), and relevant controls on the activity o f retinal growth cones in vitro, monitored by time lapse video microsc opy. Of the inhibitors tested, only the two enantiomeric forms of HS-L FA caused a reproducible cessation of both spike extension and filopod ial processes at the growth cone ruffling, while control media had no effect. In some cases, the growth cone swelled and exhibited small pro trusions. The behavior of growth cones was in sharp distinction to tha t of the cytoplasm of neural cells, and membrane ruffling of flat cell s, which continued in activity throughout. Growth cone activity return ed after several hours in the presence of the agent. BPS was toxic at concentrations above 2.5 mM. Below that, it had no effect. L-cysteine, PMSF, and control media had no effect. The relevance of these results to the possible role of proteases in fiber outgrowth from retinal cel ls is discussed. (C) 1994 Wiley-Liss, Inc.