Single neuron cultivation of embryonic and perinatal rabbit or rat brains based on plasma clot technique

Isolated neuronal cells dissociated from the brain of embryonic rabbits on the sixteenth day of gestation and of perinatal rats (eighteenth embryonic day, to E18, thirteenth day postnatum, p.n. 13) were selectively cultured using a plasma clot technique. The cells grown were shown to be neurons by means of the neuron-specific synaptosomal plasma membrane antibody (SPM). They differentiated at a very high frequency from rounded cells lacking processes into different shapes characteristic for several neuronal cell types. Morphological differences could be distinguished even after 24 h in culture. The neurons differentiated in vitro for up to 11 days, apparently without need of any direct intercellular contact. Cells caught inside the plasma clot were prevented from decreasing in number. This provides the opportunity to culture few neurons even from an extremely small area of a single brain. As an example, different cell types are shown originating from rat cerebella aged E18 to p.n. 13. Their appearance apparently corresponds to the genesis of cerebellar cell types, as is known from the in vivo situation. The high degree of characteristic neuronal differentiation and the prevention of direct intercellular contacts indicate that this culture method may serve as an in vitro assay for genetically fixed properties acquired in vivo.