gms | German Medical Science

Objective: After spinal cord injury (SCI), primary and secondary damage inhibits axonal reconnection from supralesional structures and leads to misrouting of nerve fibers. For the in vitro study of SCI today several spinal in vitro lesion models are used to investigate neuronal differentiation, axonal growth and path finding. We report here about a new slice co-culture model for the study of neuronal regeneration.

Methods: Spinal cord (sc) from postnatal (P1-P2) C57black6 mice and motor cortex (mc) dissected from postnatal (P1-P2) mice expressing green fluorescent protein (GFP) under the control of the β-actin promoter were chopped either in a sagittal longitudinal plane for the sc or in a coronal plane for the mc. Afterwards the medial cortex zone was orientated to the rostral end of the spinal cord, placed either directly next to it or at a distance of 10 mm, mounted with collagen and incubated up to two weeks. Through the use of GFP expressing mice as mc donors we could easily distinguish ingrowing mc neurons and astrozytes from the nonfluorescent wildtype sc neurons. Furthermore axonal regrowth and newly established connections were analysed via confocal microscopy after immunhistochemical staining.

Results: We found a strong ingrowth with growth cones, reestablishment of cortical fibers and new synaptic connections from the motor cortex slices into the spinal cord slices, especially when placed directly next to each other.

Conclusions: This new slice co-culture model provides an important in vitro tool for a variety of questions in the field of neuronal regeneration and the establishment of new connections of the disconnected spinal cord after a SCI. We are now able to study the various factors increasing and decreasing axonal regrowth, the amount of occurring growth cones and newly established synaptic connections.