The effect of type I astrocytes on neuronal complexity: A fractal analysis

Embryonic, ventral spinal cord neurons were grown on ploy(D-lysine) (PDL) o r on a monolayer of type I astrocytes. At various times from 6 h to 2 weeks postplating, cells were fluorescently labeled and fixed with 4% paraformal dehyde. The cell surface immunoreaction allowed visualization of neurons in their entirety, namely, cell bodies and various membranous extensions that included lamellipodia, growth cones, axons, and dendrites. Outlines were d rawn for individual neurons and their fractal dimension (D) was calculated. Neurons on poly(D-lysine) reached a peak D at 3 days in vitro, 1 day later than neurons on astrocytes (2 days in vitro). The maximum D was greater fo r cells on poly(D-lysine) when compared with neurons on astrocytes. In a se cond experiment the maximum D was similar for neurons on both surfaces but neurons on PDL maintained a higher D for a much longer period than neurons on astrocytes. An examination of fluorescent images revealed that neurons o n poly(D-lysine) exhibited lamellipodia and large growth cones for several days and these structures were likely responsible for the high D seen in th ese cells. These structures were rarely observed in neurons plated on astro cytes. Interestingly, D on both surfaces decreased to a similar value at be tween :1 and 2 weeks in vitro. The trend for D in these cultures, an initia l increase to a peak value followed by a decrease to a stable value, is dis cussed in light of the chemical nature of the two surfaces and synapse form ation and stabilization.