Phosphorylated neurofilament antigen redistribution in intercostal nerve subsequent to retrograde axonal transport of diphtheria toxin.
A novel enzyme-linked immunosorbent assay technique using specific monoclonal antibodies has been used to examine the proximal-distal distribution of phosphorylated neurofilament proteins (pNF) in normal feline intercostal nerve, and to compare it with that following retrograde axonal transport of the ADP-ribosylating protein diphtheria toxin (DTX) to thoracic motoneurones. The molecular target of DTX is elongation factor 2 which resides solely in the cell body. Normal intercostal nerves exhibited significantly higher amounts of the 200-kDa pNF-H, 160-kDA pNF-M, and 68-kDA pNF-L in proximal nerve than in the distal nerve. The overall content of all three triplet pNF proteins decreased 3 days after injection of DTX, but the normal proximal-distal gradient was retained. By 8 days post DTX injection, the proximal-distal gradient had reversed, with proximal nerve starved of pNF-H and pNF-M and distal nerve showing abnormally high pNF-L content. Correlative immunocytochemistry of spinal cords from normal animals verified that pNF-H and pNF-M are confined to efferent axons in the spinal grey matter, and that motoneurones are only reactive for pNF-L. At 8 days following toxin treatment, motoneurones in the ipsilateral ventral horn were strongly immunoreactive for all pNF. Contralateral motoneurones were non-reactive. Onset of abnormal perikaryal pNF immunoreactivity at 3 days precedes onset of ultrastructural cytopathology. Together these results indicate an early deficit in transference to the axon of NF proteins synthesised prior to full toxicity, probably because of a toxin-induced failure in regulation of phosphorylation-dependent NF assembly and turnover immediately prior to entry into the proximal axon. Results are discussed in relation to diphtheritic motoneuronopathy.
Sunner
K
K
Sobell Department of Neurophysiology, Institute of Neurology, London, UK.
Pullen
A H
AH
D000818:Animals; D000906:Antibodies / Q000276:immunology; D000941:Antigens / Q000276:immunology*; D001369:Axons / Q000502:physiology*; D002415:Cats; D004167:Diphtheria Toxin / Q000494:pharmacology*; D004797:Enzyme-Linked Immunosorbent Assay; D007150:Immunohistochemistry; D007367:Intercostal Nerves / Q000502:physiology*; D007700:Kinetics; D016900:Neurofilament Proteins; D010766:Phosphorylation; D013997:Time Factors
D000906:Antibodies; D000941:Antigens; D004167:Diphtheria Toxin; D016900:Neurofilament Proteins