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Recent advances in our ability to microfabricate tissue culture environments (1), and to store and retrieve microscopy data in a digital format, have made it increasingly possible to study fundamental aspects of neuronal migration and response to environmental cues. Neuronal migration refers to the migration of the growth cone at the tip of an extending neurite, a process that guides the neurite to its destination during neural development and regeneration (2). This chapter describes techniques for the observation of neuronal migration on both plain and patterned surfaces through the use of high-resolution, phase-contrast videomicroscopy. These techniques are particularly appropriate for analyzing the dynamics of single growth cone behavior in the presence of two-dimensional environmental microfeatures similar in scale to the growth cone dimensions (1-50 µm). Growth cone response to topographical features (3) or other three-dimensional environments are beyond the scope of this chapter. Substrate preparation procedures are derived from microlithography techniques first used in the microelectronics industry and adapted to cell culture systems (4-7). Glass cover slips are used as the substrate support to provide the required optical clarity for high-resolution microscopy, and patterns are created using laminin or collagen. |
