THE RETURN OF PHOSPHORYLATED AND NONPHOSPHORYLATED EPITOPES OF NEUROFILAMENT PROTEINS TO THE REGENERATING OPTIC-NERVE OF XENOPUS-LAEVIS

Neurofilament proteins of mammalian axotomized peripheral axons, which regenerate effectively, resemble those of embryonic axons. However, i njured centrally projecting mammalian axons, which fail to regenerate, have very different neurofilament compositions than during developmen t. If changes in neurofilament composition after injury reflect the ab ility of axotomized neurons to regenerate effectively, then the neurof ilaments of centrally projecting axons that can regenerate should more closely resemble those of developing axons. In this study, the neurof ilament compositions of injured optic axons of the frog, Xenopus laevi s, were examined, since these axons can regenerate a fully functional projection. Antibodies to phosphorylated and nonphosphorylated forms o f neurofilament proteins that had been used previously to study the ne urofilament composition of newly developing X. laevis optic axons were used in immunocytochemical studies to examine the return of neurofila ments to the optic nerve after an intraorbital nerve crush. Intraocula rly injected wheat germ agglutinin conjugated to horseradish peroxidas e was used to label the regenerating axons independently of their neur ofilaments. Neurofilament immunoreactivities disappeared rapidly from crushed axons during the first week after surgery. By nine days after surgery, antibodies to nonphosphorylated forms of middle (NF-M) and lo w molecular weight (NF-L) neurofilament proteins and the Xenopus neuro nal intermediate filament protein (XNIF) began to stain the nerve just beyond the lesion. By this time, however, growing axonal terminals ha d reached the optic chiasm. Antibodies to phosphorylated epitopes of N F-M began to stain axons at 15 days, just as growing axons began to ar rive at the optic tectum. Nonphosphorylated high molecular weight neur ofilament protein (NF-H) began to appear in axons between 18 and 21 da ys after surgery. Thus, the reappearance of neurofilaments during opti c axon regeneration resembled the general pattern seen during developm ent. The chief difference between development and regeneration was tha t neurofilament epitopes took longer to emerge during regeneration. On e possibility is that cues encountered along the optic pathway influen ce the neurofilament composition of retinal ganglion cell axons. Then, the greater distances travelled by regenerating axons could account f or the longer time taken for their neurofilament compositions to matur e. (C) 1994 Wiley-Liss, Inc.