The most active work on this project this year was aimed at characterizing differences in the adhesive patterns and associated growth characteristics of cells grown on defined substrates. Neurite formation by sympathetic and retinal neurons from embryonic chicks was observed using a technique (time-lapse laser scanning interference reflection microscopy) to show local distances of the cell membrane from substrates composed of either polyamino acids or laminin. The polyamino acids promoted close adhesion of both cell types over much of their surface and restricted their ability to produce neurites. In contrast, laminin promoted close attachment only in discrete patches and promoted rapid neurite extension. These results suggest that there is a critical level and, perhaps, pattern of adhesion of a growth cone to its substrate that are necessary for neurites to form and move forward. Substrates that are even more adhesive restrict rather than promote neurite growth. A comprehensive effort to develop methods for making and maintaining organotypic brain cultures continues (see also Project #ZOl NS 02610-08 LN). For the present project, the advantage of this preparation is that regions of the brain, such as the hippocampal and deep cerebellar cortices, would be exposed to structural analysis by rapid freezing techniques and laser scanning confocal microscopy. It has, however, proven difficult to consistently produce large expanses of mature brain. A new attempt to make orqanotypic cultures of cerebral cortex are currently being analyzed. A new initiative just started, is to apply the thin film structural techniques (see Projects #ZOl NS 02700-06 LN and NS 02836-01 LN) to analyze the structure of isolated postsynaptic densities from different parts of the brain.
