Localization of voltage-sensitive Ca2+ fluxes and neuropeptide Y immunoreactivity to varicosities in SH-SY5Y human neuroblastoma cells differentiated by treatment with the protein kinase inhibitor staurosporine

Eur J Neurosci. 1997 Jan;9(1):140-50. doi: 10.1111/j.1460-9568.1997.tb01362.x.

Abstract

The distribution of voltage-sensitive elevations of the level of Ca2+ in untreated SH-SY5Y cells and cells that had been induced to differentiate with staurosporine was investigated by monitoring fura-2 fluorescence in cell suspensions, and by using microfluorometry and quantitative fluorescence imaging on cell bodies and on cellular processes. Cell bodies of both types of cells displayed small Ca2+ elevations, which were composed of transient and sustained components. Elevations were partially sensitive to the L- and N-channel blockers nifedipine (1 microM) and omega-conotoxin GVIA (100 nM) respectively. Up to ten times Ca2+ elevations were observed in varicosities of treated cells than in cell bodies of treated and cells. These elevations were insensitive to compounds known to release Ca2+ from intracellular stores. Elevations of Ca2+ were sustained, and they were insensitive to 5 microM nifedipine, 100 nM omega-agatoxin IVA and 100 nM omega-conotoxin GVIA, and partially sensitive to 2 microM omega-conotoxin GVIA, indicating predominance of non-L-type, non-N-type, non-P-type channel activity. The intracellular localization of neuropeptide Y, a marker of differentiation in these cells, was also investigated by fluorescence immunocytochemistry. Varicosities of treated cells displayed marked fluorescence when viewed in a confocal microscope. These findings show that the varicosities of staurosporine-treated cells exhibit some of the functional properties of nerve terminals. The varicosities resemble boutons en passant nerve endings and they seem to express Ca2+ channels different from those in the cell body.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Brain Neoplasms / metabolism*
  • Brain Neoplasms / ultrastructure*
  • Calcium Channel Blockers / pharmacology
  • Calcium Channels / drug effects
  • Calcium Channels / metabolism*
  • Cell Differentiation / drug effects
  • Culture Media
  • Electrophysiology
  • Enzyme Inhibitors / pharmacology*
  • Fluorescent Dyes
  • Fura-2
  • Humans
  • Immunohistochemistry
  • Ion Channel Gating / drug effects
  • Neuroblastoma / metabolism*
  • Neuroblastoma / ultrastructure*
  • Neuropeptide Y / metabolism*
  • Protein Kinase Inhibitors*
  • Staurosporine / pharmacology*

Substances

  • Calcium Channel Blockers
  • Calcium Channels
  • Culture Media
  • Enzyme Inhibitors
  • Fluorescent Dyes
  • Neuropeptide Y
  • Protein Kinase Inhibitors
  • Staurosporine
  • Fura-2