Confocal imaging of redox mechanisms
The key intracellular variables mitochondrial membrane
potential (Dym), ROS and reduced glutathione (GSH) were
monitored with fluorescent probes (TMRE, CM-DCF and
MCB); the NAD(P)H redox status was imaged using
autofluorescence (Fig. 5). A significant increase in Dym
was evident after treatment with 1 mM AA as compared to
controls or exposure or 5 mg GPE ml21 (Fig. 5i). A pattern
of effects characteristic of oxidative stress was induced (i.e.
high ROS, and a low level of GSH) was induced by addition
of 1 mM AA (the IC50 value for this compound) to earlystationary-phase
cells (Fig. 5). Addition of 5 mg GPE ml21
caused a similar increase in ROS and depletion of GSH, but a
decrease in NADH autofluorescence was also observed. A
similar response was exhibited by exponential-phase cells
(not shown). Low levels of ROS were localized in the
mitochondria in control cells (Fig. 5aii); increased CMDCF
fluorescence was observed in both mitochondrial
and cytoplasmic compartments after treatment with either
GPE or AA (Fig. 5bii and 5cii).