Axolemmal repair requires proteins that mediate synaptic vesicle fusion

A damaged cell membrane Is repaired by a seal that restricts entry or exit of molecules and ions to that of the level passing through an undamaged mem brane. Seal formation requires elevation of intracellular Ca2+ and, very li kely, protein-mediated fusion of membranes. Ca2+ also regulates the interac tion between synaptotagmin (Syt) and syntaxin (Syx), which is thought to me diate fusion of synaptic vesicles with the axolemma, allowing transmitter r elease at synapses. To determine whether synaptic proteins have a role in s ealing axolemmal damage, we injected squid and crayfish giant axons with an antibody that inhibits squid Syt from binding Ca2+, or with another antibo dy that inhibits the Ca2+-dependent interaction of squid Syx with the Ca2+- binding domain of Syt, Axons injected with antibody to Syt did not seal, as assessed at axonal cut ends by the exclusion of extracellular hydrophilic fluorescent dye using confocal microscopy, and by the decay of extracellula r injury current compared to levels measured in uninjured axons using a vib rating probe technique. In contrast, axons injected with either denatured a ntibody to Syt or preimmune IgG did seal. Similarly, axons injected with an tibody to Syx did not seal, but did seal when injected with either denature d antibody to Syx or preimmune IgG, These results indicate an essential inv olvement of Syt and Syx in the repair (sealing) of severed axons. We sugges t that vesicles, which accumulate and interact at the injury site, re-estab lish axolemmal continuity by Ca2+-induced fusions mediated by proteins such as those involved in neurotransmitter release. (C) 2000 John Wiley & Sons, Inc.