Membrane potential-sensitive probes

In primary cultures of cerebellar granule cells palytoxin produced a concentration-dependent depolarization of the cell membrane, as assessed with the membrane potential sensitive probe bis-oxonol (Fig. 6.1). Significant differences between control and paly toxin-treated cells were found at palytoxin concentrations equal or higher than 5 nM. 

Botana and collaborators also evaluated the effect of ostreocin-D and palytoxin in an excitable cellular model (human neuroblastoma cells). Both toxins caused an important loss of filamentous actin associated to morphological alterations (Fig. 6.7). In neuroblastoma cells, the variation of ion fluxes caused by ostreocin-D and palytoxin was studied by using the membrane potential sensitive probe bis-oxonol and the Ca sensitive dyes fluo-3 and fura red. Ostreocin-D modifies the membrane potential, causing depolarization, although more slight than that produced by palytoxin (Fig. 6.8a). 

Coclet-Ninin, J. Rochat, T. Poitry, S. Chanson, M. Discrimination between cystic fibrosis and CFTR-corrected epithelial cells by a membrane potential-sensitive probe. Exp. Lung Res. 2002, 28, 181-199. 

A number of dyes belonging to the polyene class - oxonols, merocyanine, rho-damines, and cyanines - have been used in 31P-NMR as potential-sensitive probes to investigate their effects on the membrane and their location in PC vesicles [115], A substantial broadening of the 31P resonances and a reduction in spin-lattice and... 

Baxter DF, Kirk M, Garcia AF et al. (2002) A novel membrane potential-sensitive fluorescent dye improves cell-based assays for ion channels. J Biomol Screen 7(1) 79—85 Epps DE, Wolfe ML, Groppi V (1994) Characterization of the steady-state and dynamic fluorescence properties of the potential-sensitive dye bis-(l,3-dibutylbarbituric acid)trimethine oxonol (BiBAC4(3)) in model systems and cells. Chem Phys Lipids 69(2) 137—150 Gonzalez JE, Maher MP (2002) Cellular fluorescent indicators and voltage/ion probe reader (VIPR(TM)) tools for ion channel and receptor drug discovery. Recept Channels 8(5—6) 283—295... 

The membrane or interfacial potential, particularly in biological applications, is often determined from the change in fluorescence of added carbocyanine dyes (12, 13). The fluorescence intensity of the dyes depends on the solvent in which the dyes are present. When the dyes are used as potential-sensitive probes, their fluorescent intensity is a function of the interfacial potential across the membrane. We studied the behavior of dye transport on a phase boundary between water and nitrobenzene to better understand the principles of the potential dye partitioning as a function of interfacial potential (141... 

A variety of probes for Na , K and Cr ion channels and carriers have been described. Ion flux that affects the cell membrane potential, can be indicated with potential-sensitive probes. 

Dragsten, P. R., and Webb, W. W., 1978, Mechanism of the membrane potential sensitivity of the fluorescent membrane probe mero-cyanine 540, Biochemistry 17 5228-5240. 

Wolff, C. Fuks, B. Chatelain, P. Comparative study of membrane potential-sensitive fluorescent probes and their use in ion channel screening assays. J. Biomol. Screening 2003, 8, 533-543. 

Electric field sensitive dyes respond to changes in electrical membrane potential by a variety of different mechanisms with widely varying response times depending on their chemical structure and their interaction with the membrane. An understanding of the mechanisms of dye response and their response mechanisms is important for an appropriate choice of a probe for a particular application. The purpose of this chapter is, therefore, to provide an overview of the dyes presently available, how they respond to voltage changes, and give some examples of how they have been applied. Finally, because there is still scope for the development of new dyes with improved properties, some directions for future research will be discussed. 

The most important reason for the laige number of technical applications of polymethine dyes is their relatively low electron-transition energies and their highly intense and narrow spectral bands. Indeed, polymethines display strong light absorption and emission, between 300 and 1600 nm. In the 1990s, these dyes are mainly used as photographic sensitizers and desensitizers (11,90), as laser dyes (12,13,91), as probes of membrane potentials (14), and in other applications where the theoretical aspects of polymethines are useful. 

Fluorescent, voltage-sensitive dyes can be employed to investigate the activity of ion channels by reflecting the cellular membrane potential (Epps et al. 1994 Plasek and Sigler 1996). This approach is suitable for use in conjunction with different measurement systems including Fluorometric imaging plate readers (FLIPR), Voltage/Ion probe readers (VIPR) or conventional fluorescence readers. 

Fluorescence is very sensitive to the chemical environment and may be utilized to provide information about the microenvironment surrounding the probe (5, 6). The fluorescence intensity (quantum yield) (1-3), the maximum emission wavelength (1-3), the fluorescence lifetime (6-9), or the polarization (10-12) may all be monitored for specific changes that are induced as a result of changes in polarity, pH, ion concentration, membrane potential, or ligand binding. 

The use of potential-sensitive fluorescent probes to monitor the electrical potential across a cell membrane permits an accurate, noninvasive measurement of membrane potential changes in a wide variety of cells, vesicles, and organelles without the external electrical or mechanical manipulation required by micro-... 

To determine if a dye will be a useful potentiometric probe in biological applications, information on its chemical, physical, spectral, and toxicological properties must be gathered in addition to data on the sensitivity to membrane potential. A number of model membrane systems are employed to characterize dye properties as well as some simple biological preparations. 

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