Distribution of non-phosphorylated and phosphorylated neurofilament proteins in the spinal cord of an anuran amphibian during development and regeneration.

Abstract

The changes in the neurofilament medium and high molecular weight subunits (NF150 and NF200) in the developing and transected spinal cords of bullfrog tadpoles were studied. A monoclonal antibody recognizing the nonphosphorylated epitope of NF150, NF150D, stained the neuronal cell bodies and axons, whereas other antibodies against the phosphorylated NFs, NF150P or NF200P, labeled chiefly the axons. During development, the intensity of axonal staining by the anti-NF150D in the ventral fasciculi in younger tadpoles appeared stronger than older animals, but the reverse was seen for NF150P and NF200P. Complete signal transection of stage IV tadpoles resulted in degeneration and then regeneration of the cord tissue of both cut ends. Each stump lengthened by about 350 microns in the 4 weeks after the lesion. In the proximal stumps, the levels of NF150P or NF200P in the ventral axons at 550-350 microns proximal to the transection site increased notably by about 24-73% of the control value 7-28 days post-transection; however, the content of NF150D was decreased. The densities of NF150D and NF150P protein spots on the Coomassie blue-stained two-dimensional gels of the normal and injured cords also displayed alterations similar to the immunocytochemical data. Intense labeling by the anti-NF150P or NF200P was present in the cell bodies of axotomized motor neurons in the ventral horn. The results suggest that central axonal regeneration may be accompanied by upregulated phosphorylated neurofilament proteins.