Second, the conclusion that cells expressing the triple mutant, which we have now shown to be effectively nonfunctional at physiological membrane potentials, cannot grow photoheterotrophically is consistent with previous data showing that cells lacking a functional BR can likewise not sustain photoheterotrophic growth.
For evaluation of the membrane potential, fibroblast cells were grown in 3 cm plastic dishes with glass coverslips (Nunc) and the procedure was performed as described previously (Alirol et al, 2006).
One can note that the current is an almost exponentially growing function of membrane potential.
Measurements of the membrane potential of individual mitochondria in the growing axons of chick sensory neurons using the dye TMRM (tetramethylrhodamine methyl ester) revealed no major differences among mitochondria along the length of the axon, and no differences in membrane potential in stationary versus moving mitochondria (Verburg and Hollenbeck, 2008).
In spite of the reduced mitochondrial membrane potential of Δ mca promastigotes, they were viable, grew at rates comparable to those of WT and had an intact mitochondrion as assessed by tetramethylrhodamine methyl ester perchlorate (TMRM) labelling (data not shown).
Despite the lower rates of respiration, the membrane potential was maintained at the same magnitude as fast growing cells.
The lipophilic cationic probe 5,5',6,6'-tetrachloro-1,1',3,3'- tetraethylbenzinidazolylcarbocyanine iodide JC-1 dye (Molecular ProbesTM, Invitrogen) was used to measure mitochondrial inner membrane potential (ΔΨm) in drug treated or untreated cells (5×105) grown in 35 mm tissue plates.
Human mesangial (HM) cells grown in high glucose show largely increased mitochondrial membrane potential and ROS production [13].
(B ) Plasma membrane potential as indicated by DiOC6 staining of WT and sir2 ∆ cells grown in glucose or ethanol medium (*p < 0.005).
This effect was observed with yeast grown in either glycerol or dextrose-containing media, but the degree of membrane potential loss was greater in the glycerol-containing media (data not shown).
