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(6) DYNAMICS OF MITOCHONDRIAL MEMBRANE POTENTIAL IN NEURONS AND ASTROCYTES J.F. Buckman and I.J. Reynolds
Mitochondrial involvement in the fate of neurons and astrocytes through both apoptotic and necrotic injury processes has become increasingly evident. In intact cells, the measurement of mitochondrial membrane potential (Dym) can be utilized as a marker of mitochondrial status. We monitored Dym in primary cultures of forebrain neurons and astrocytes loaded with the Dym-sensitive fluorescent dyes, JC-1 or TMRM. In neurons, JC-1- and TMRM-loaded mitochondria exhibited spontaneous oscillations in fluorescence intensity in small regions of the processes, indicative of partial, transient mitochondrial depolarizations. Dym oscillations appeared more rapid and intense in TMRM-loaded mitochondria, corresponding to the known differences between TMRM and JC-1. In astrocytes, JC-1-loaded mitochondria transiently and nearly synchronously depolarize sending waves of fluorescence across the cell. TMRM-loaded mitochondrial depolarizations appear asynchronous and much more rapid. This difference in fluorescence pattern may also be associated with the different kinetics of these dyes. Moreover, the transient depolarizations in TMRM-loaded mitochondria appear to be associated with phototoxicity and may represent a model for studying mitochondrial reactive oxygen species generation. When light exposure was minimized, no Dym oscillations were observed. However, following a short, intense period of illumination, Dym became unstable and oscillations were immediately observed. We are currently investigating the nature and significance of mitochondrial Dym oscillations to determine the relationship of this phenomenon to reactive oxygen species generation, cellular and synaptic signaling, ATP availability and usage, and low-amplitude permeability transition Supported by USAMRMC, Scaife Family Foundation and T32NS07391.
For further information contact...Carmen Mannella: carmen@wadsworth.org |
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