Biomaterial Database

Assessment of mitochondrial polarization status in living cells based on analysis of the spatial heterogeneity of rhodamine 123 fluorescence staining. 2000

E C Toescu, and A Verkhratsky

Dept. Physiology, The University of Birmingham, Edgbaston, UK. e.c.toescu@bham.ac.uk

The mitochondrial membrane potential (psimito) is an important parameter not only of mitochondrial but also of cellular status. Prolonged mitochondrial depolarization is associated with various forms of neuronal death. Assessment of mitochondrial depolarization can take advantage of the specific properties of the lipophilic dyes that distribute in a potentiometrically determined ratio across membranes. Using conventional imaging, we showed that rhodamine 123 accumulated in the mitochondria, generating a highly heterogeneous pattern of spatial distribution of fluorescence across the cell body. Collapse of the psimito following exposure to a protonophore, carbonylcyanide p-chloromethoxyphenylhydrazone (CCCP), released rhodamine 123 from mitochondria into the cytosol. Under acutely changed conditions, this increased the overall intensity of the fluorescence signal and significantly decreased the degree of spatial heterogeneity of the signal. If mitochondrial depolarization was sustained chronically, the intensity of the signal decreased, but the increase in the spatial homogeneity of the fluorescent signal was maintained. Image analysis showed that the level of spatial heterogeneity of the signal can be assessed by calculating, for each individual neurone, the spread of pixel intensities values around the mean. This spread is defined by the coefficient of variation (CV), which is a measure of the standard deviation normalized to the average, and was inversely related to mitochondrial depolarization measured under different conditions. Thus, the degree of spatial heterogeneity of the rhodamine 123 signal measured from a neurone is a reliable indicator for the assessment of mitochondrial depolarization and can be used in experiments to monitor psimito over shorter or longer periods.

UI	MeSH Term	Description	Entries
D007425	Intracellular Membranes	Thin structures that encapsulate subcellular structures or ORGANELLES in EUKARYOTIC CELLS. They include a variety of membranes associated with the CELL NUCLEUS; the MITOCHONDRIA; the GOLGI APPARATUS; the ENDOPLASMIC RETICULUM; LYSOSOMES; PLASTIDS; and VACUOLES.	Membranes, Intracellular,Intracellular Membrane,Membrane, Intracellular
D007476	Ionophores	Chemical agents that increase the permeability of biological or artificial lipid membranes to specific ions. Most ionophores are relatively small organic molecules that act as mobile carriers within membranes or coalesce to form ion permeable channels across membranes. Many are antibiotics, and many act as uncoupling agents by short-circuiting the proton gradient across mitochondrial membranes.	Ionophore
D007700	Kinetics	The rate dynamics in chemical or physical systems.
D008564	Membrane Potentials	The voltage differences across a membrane. For cellular membranes they are computed by subtracting the voltage measured outside the membrane from the voltage measured inside the membrane. They result from differences of inside versus outside concentration of potassium, sodium, chloride, and other ions across cells' or ORGANELLES membranes. For excitable cells, the resting membrane potentials range between -30 and -100 millivolts. Physical, chemical, or electrical stimuli can make a membrane potential more negative (hyperpolarization), or less negative (depolarization).	Resting Potentials,Transmembrane Potentials,Delta Psi,Resting Membrane Potential,Transmembrane Electrical Potential Difference,Transmembrane Potential Difference,Difference, Transmembrane Potential,Differences, Transmembrane Potential,Membrane Potential,Membrane Potential, Resting,Membrane Potentials, Resting,Potential Difference, Transmembrane,Potential Differences, Transmembrane,Potential, Membrane,Potential, Resting,Potential, Transmembrane,Potentials, Membrane,Potentials, Resting,Potentials, Transmembrane,Resting Membrane Potentials,Resting Potential,Transmembrane Potential,Transmembrane Potential Differences
D008856	Microscopy, Fluorescence	Microscopy of specimens stained with fluorescent dye (usually fluorescein isothiocyanate) or of naturally fluorescent materials, which emit light when exposed to ultraviolet or blue light. Immunofluorescence microscopy utilizes antibodies that are labeled with fluorescent dye.	Fluorescence Microscopy,Immunofluorescence Microscopy,Microscopy, Immunofluorescence,Fluorescence Microscopies,Immunofluorescence Microscopies,Microscopies, Fluorescence,Microscopies, Immunofluorescence
D008928	Mitochondria	Semiautonomous, self-reproducing organelles that occur in the cytoplasm of all cells of most, but not all, eukaryotes. Each mitochondrion is surrounded by a double limiting membrane. The inner membrane is highly invaginated, and its projections are called cristae. Mitochondria are the sites of the reactions of oxidative phosphorylation, which result in the formation of ATP. They contain distinctive RIBOSOMES, transfer RNAs (RNA, TRANSFER); AMINO ACYL T RNA SYNTHETASES; and elongation and termination factors. Mitochondria depend upon genes within the nucleus of the cells in which they reside for many essential messenger RNAs (RNA, MESSENGER). Mitochondria are believed to have arisen from aerobic bacteria that established a symbiotic relationship with primitive protoeukaryotes. (King & Stansfield, A Dictionary of Genetics, 4th ed)	Mitochondrial Contraction,Mitochondrion,Contraction, Mitochondrial,Contractions, Mitochondrial,Mitochondrial Contractions
D009474	Neurons	The basic cellular units of nervous tissue. Each neuron consists of a body, an axon, and dendrites. Their purpose is to receive, conduct, and transmit impulses in the NERVOUS SYSTEM.	Nerve Cells,Cell, Nerve,Cells, Nerve,Nerve Cell,Neuron
D011189	Potassium Chloride	A white crystal or crystalline powder used in BUFFERS; FERTILIZERS; and EXPLOSIVES. It can be used to replenish ELECTROLYTES and restore WATER-ELECTROLYTE BALANCE in treating HYPOKALEMIA.	Slow-K,Chloride, Potassium
D002258	Carbonyl Cyanide m-Chlorophenyl Hydrazone	A proton ionophore. It is commonly used as an uncoupling agent and inhibitor of photosynthesis because of its effects on mitochondrial and chloroplast membranes.	CCCP,Carbonyl Cyanide meta-Chlorophenyl Hydrazone,Carbonylcyanide 4-Chlorophenylhydrazone,Propanedinitrile, ((3-chlorophenyl)hydrazono)-,Carbonyl Cyanide m Chlorophenyl Hydrazone,4-Chlorophenylhydrazone, Carbonylcyanide,Carbonyl Cyanide meta Chlorophenyl Hydrazone,Carbonylcyanide 4 Chlorophenylhydrazone
D002531	Cerebellum	The part of brain that lies behind the BRAIN STEM in the posterior base of skull (CRANIAL FOSSA, POSTERIOR). It is also known as the "little brain" with convolutions similar to those of CEREBRAL CORTEX, inner white matter, and deep cerebellar nuclei. Its function is to coordinate voluntary movements, maintain balance, and learn motor skills.	Cerebella,Corpus Cerebelli,Parencephalon,Cerebellums,Parencephalons

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