title: Persistence of botulinum neurotoxin action in cultured spinal cord cells.
abstract: Primary dissociated fetal mouse spinal cord cultures were used to study the mechanisms underlying the differences in persistence of botulinum neurotoxin A (BoNT/A) and botulinum neurotoxin/E (BoNT/E) activities. Spinal cord cultures were exposed to BoNT/A (0.4 pM) for 2-3 days, which converted approximately half of the SNAP-25 to an altered form lacking the final nine C-terminal residues. The distribution of toxin-damaged to control SNAP-25 remained relatively unchanged for up to 80 days thereafter. Application of a high concentration of BoNT/E (250 pM) either 25 or 60 days following initial intoxication with BoNT/A converted both normal and BoNT/A-truncated SNAP-25 into a single population lacking the final 26 C-terminal residues. Excess BoNT/E was removed by washout, and recovery of intact SNAP-25 was monitored by Western blot analysis. The BoNT/E-truncated species gradually diminished during the ensuing 18 days, accompanied by the reappearance of both normal and BoNT/A-truncated SNAP-25. Return of BoNT/A-truncated SNAP-25 was observed in spite of the absence of BoNT/A in the culture medium during all but the first 3 days of exposure. These results indicate that proteolytic activity associated with the BoNT/A light chain persists inside cells for > 11 weeks, while recovery from BoNT/E is complete in < 3 weeks. This longer duration of enzymatic activity appears to account for the persistence of serotype A action.
authors.affiliation: Pharmacology Division, U.S. Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, MD 21010-5400, USA. James.Keller@amedd.army.mil
authors.forename: J E
authors.initials: JE
authors.lastname: Keller
authors.forename: E A
authors.initials: EA
authors.lastname: Neale
authors.forename: G
authors.initials: G
authors.lastname: Oyler
authors.forename: M
authors.initials: M
authors.lastname: Adler
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001905:Botulinum Toxins / Q000378:metabolism* / Q000494:pharmacology; D019274:Botulinum Toxins, Type A / Q000378:metabolism* / Q000494:pharmacology; D002478:Cells, Cultured / Q000378:metabolism; D005260:Female; D008565:Membrane Proteins; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism; D013116:Spinal Cord / Q000166:cytology / Q000187:drug effects / Q000196:embryology / Q000378:metabolism*; D050825:Synaptosomal-Associated Protein 25
chemical_list: D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; C494209:Snap25 protein, mouse; D050825:Synaptosomal-Associated Protein 25; D010447:Peptide Hydrolases; D001905:Botulinum Toxins; D019274:Botulinum Toxins, Type A; C096326:botulinum toxin type E