A scanning electron microscope study of the in vitro development of dissociated hippocampal cells

Abstract

Cultures of dissociated hippocampal neurons from 18-day-old rat fetuses were examined by scanning electron microscopy after periods between one hour and 20 days following plating on a poly-L-lysine coated substrate. Cell attachment was virtually complete within one hour after plating, and at that stage many cells could be seen which had started to extend processes with broad growth cones. By four hours in culture, process formation was well advanced and some cells had already assumed a pyramidal configuration. After 16 hours in culture, numerous contacts were seen between neighboring growth cones, and this frequently led to fasciculation of the interacting fibers. During the next three weeks the cell bodies enlarged considerably and rounded-up, and two distinct types of processes became evident: large, rapidly tapering dendrite-like processes and finer, essentially uniform-diametered processes that resemble axons. In most of the older cultures a dense plexus of processes was formed, and many of the finer processes appeared to have "bouton-like" swellings as they traversed the upper surfaces of the neuronal perikarya. Non-neuronal elements, which comprised only about 5% of the cells initially plated, rapidly proliferated in our cultures and within three to six days formed a confluent monolayer beneath the neurons.