Growth Cone Behavior at Borders between Different Extracellular Matrix Molecules

The reaction of growth cones in in vitro assays to substrate-bound molecules might yield important clues to the roles that these molecules play in growth cone guidance in vivo. Janusin and tenascin are glia-derived, extracellular matrix molecules that are expressed in the nervous system at times and in locations that suggest that they might act as barriers to neurite outgrowth. To test this hypothesis we have used video time-lapse microscopy to observe the behavior of growth cones, growing on a substrate permissive for neurite outgrowth, when they are confronted with janusin or tenascin as sharp, substrate boundaries. Here we describe the method for offering growth cones a choice between two substrates, in which the border between the two molecules can be clearly visualized in the phase-contrast microscope during the period of observation. We have learned from these observations that growth cones avoid advancing onto janusin or tenascin substrates, but do not undergo gross morphological changes, such as complete collapse, when they contact these molecules.