Plasmalemmal repair of severed neurites of PC12 cells requires Ca2+ and synaptotagmin

E. R. Detrait, S. Yoo, C. S. Eddleman, M. Fukuda, G. D. Bittner, H. M. Fishman

Research output: Contribution to journalArticlepeer-review

51 Citations (Scopus)

Abstract

Ca2+ and synaptotagmin (a Ca2+-binding protein that regulates axolemmal fusion of synaptic vesicles) play essential roles in the repair of axolemmal damage in invertebrate giant axons. We now report that neurites of a rat pheochromocytoma (PC12) cell line transected and maintained in a serum medium form a dye barrier (exclude an external hydrophilic fluorescent dye) and survive for 24-hr posttransection (based on morphology and retention of another hydrophilic dye internally loaded at 6-hr posttransection). Some (25%) transected neurites that form a dye barrier regrow. Most (83%) neurites transected in a saline solution containing divalent cations (PBS++) also exclude entry of an externally placed hydrophilic fluorescent dye at 15-min posttransection. In contrast, only 14 or 17% of neurites maintained in a divalent cation-free solution (PBS(=)) or in PBS(=) + Mg2+, respectively, form a dye barrier. Neurites that do not form a dye barrier do not survive for 24 hr. When PC12 neurites are loaded with an antibody to squid synaptotagmin, most (81%) antibody-loaded neurites do not form a dye barrier, whereas most (≥81%) neurites loaded with heat-inactivated antibody or preimmune IgG do form a barrier. These data show that: 1) transected neurites of PC12 cells have mechanism(s) for plasmalemmal repair (dye barrier formation and survival); 2) Ca2+ is necessary for dye barrier formation, which occurs minutes after transection and is necessary for survival and regrowth; and 3) synaptotagmin is an essential mediator of barrier formation. The similarity in the requirements for plasmalemmal repair in this mammalian cell preparation with those reported previously for invertebrate axons suggests that mechanisms necessary for plasmalemmal repair have been conserved phylogenetically. (C) 2000 Wiley-Liss, Inc.

Original languageEnglish
Pages (from-to)566-573
Number of pages8
JournalJournal of Neuroscience Research
Volume62
Issue number4
DOIs
Publication statusPublished - 2000 Nov 15
Externally publishedYes

Keywords

  • Axotomy
  • Ca-binding synaptic protein
  • Exocytosis
  • Mammalian cell culture
  • Neurite regrowth
  • Plasma membrane repair

ASJC Scopus subject areas

  • Cellular and Molecular Neuroscience

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