| Abstrakt: |
Bradykinin and caffeine were used as two different agonists to study inositol 1,4,5-trisphosphate (IP)-sensitive and caffeine/ryanodine-sensitive intracellular Ca release in the outgrowing neurites of nerve-growth-factor (NGF)-treated rat phaeochromocytoma cells (PC12). Changes in neuritic intracellular free Ca ([Ca]) in single cells were measured after loading with a 1:1 mixture of the acetoxymethyl (AM) ester of the Ca-sensitive dyes Fura-red and Fluo-3, in combination with confocal microscopy. Bradykinin-induced Ca release was blocked by U73211, a specific phospholipase C inhibitor. Caffeine-induced Ca release was very low in neurites at rest. It increased after the cells were pre-loaded with Ca. The Ca signal evoked at high concentrations of bradykinin (> 500 nM) arose from a trigger zone in the proximal part of the neurite, as a bi-direction-al wave towards the growth cone and cell body. The speed of neuritic Ca waves was reduced in cells loaded with the Ca chelator l,2-bis(2-aminophenoxy)ethane-tetraacetic acid/AM. Preloading of Ca stores led to increased bradykinin-induced Ca release, as seen for caffeine, and faster Ca wave speeds. Caffeine evoked a simultaneous [Ca] rise along the neurites of Ca pre-loaded cells. Higher Ca signal amplitudes and faster Ca wave speeds, but no longer-lasting IP-induced [Ca] signals, correlated with increased caffeine-induced Ca release in the neurites. At low concentrations of bradykinin (< 1.0 nM), the Ca signals ceased to propagate as complete Ca waves. Instead, repetitive stochastic Ca release events (neuritic Ca puffs) were observed. Neuritic Ca puffs spread across only a few microns, at a slower speed than neuritic Ca waves. These Ca puffs represent elementary Ca release units, whereby the released Ca ions form these elementary events into the shape of a Ca wave. [ABSTRACT FROM AUTHOR] |
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