Abstract
Aim of this experimental study was to develop an in vitro model that simplifies the study of various factors regulating neuronal regeneration.
An in vitro-system that allows co-culture of slices from rat motorcortex and spinal cord (p4) was established. Two groups of cultures were investigated: In the first group, intact spinal cord slices were cultured adjacent to motorcortex slices, while in the second group the spinal cord slices were sagitally cut into halves, with the sectioned interface placed directly adjacent to the motorcortex, in order to prevent the spinal white matter from interference. Each group was further divided into two subgroups: The NT-3 group, where the culture medium contained 50 ng/ml NT-3 and the control group treated with normal culture medium. Motorcortex pyramidal neurons were anterogradely labelled with MiniRuby, a 10 kD biotinylated dextran amine.
After 4 days the co-cultures were propagated, and axonal sprouting occurred. The group of co-cultures treated with NT-3 showed an improved cortical cytoarchitecture, and sprouting axons were more frequently observed. In NT-3-treated co-cultures where spinal cord gray matter was directly opposed to cortical slices sprouting axons entered the adjacent spinal cord tissue. This phenomenon was not observed if spinal white matter was opposed to the cortical slices, or if NT-3 was absent.
Our data suggest that the absence of repellent factors such as white matter and the presence of neuro-trophic factors promote axonal sprouting. Co-cultures of motorcortex and spinal cord slices combined with anterograde axonal labelling could provide a valuable in vitro model for the simplified screening of factors influencing corticospinal tract regeneration
Correspondence should be addressed to Anastasia C. Tilentzoglou MD, General Secretary of the Board of Directors of HAOST, 20 A. Fleming Str. (N.Filothei), Gr. 15123 Maroussi, Athens Greece. E-mail: info@eexot.gr
