Spontaneous Changes in Mitochondrial Membrane Potential in Cultured Neurons

Using the mitochondrial membrane potential (ΔΨm)-sensitive fluorescent dyes 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolocarbocyanine iodide (JC-1) and tetramethylrhodamine methyl ester (TMRM), we have observed spontaneous changes in the ΔΨm of cultured forebrain neurons. These fluctuations in ΔΨm appear to represent partial, transient depolarizations of individual mitochondria. The frequency of these ΔΨmfluctuations can be significantly lowered by exposure to a photo-induced oxidant burden, an ATP synthase inhibitor, or a glutamate-induced sodium load, without changing overall JC-1 fluorescence intensity. These spontaneous fluctuations in JC-1 signal were not inhibited by altering plasma membrane activity with tetrodotoxin or MK-801 or by blocking the mitochondrial permeability transition pore (PTP) with cyclosporin A. Neurons loaded with TMRM showed similar, low-amplitude, spontaneous fluctuations in ΔΨm. We hypothesize that these ΔΨmfluctuations are dependent on the proper functioning of the mitochondria and reflect mitochondria alternating between the active and inactive states of oxidative phosphorylation.

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