Synthesis, spectra, delivery and potentiometric responses of new styryl dyes with extended spectral ranges

Wuskell, Joseph P.; Boudreau, David; Wei, Mei; Liu, Jin; Engl, Reimund; Ravikrishna, Chebolu; Bullen, Andrew; Hoffacker, Kurt D.; Kerimo, Josef; Cohen, Lawrence B.; Żochowski, Michał; Loew, Leslie M.

doi:10.1016/j.jneumeth.2005.07.013

Cited by 77 publications

(80 citation statements)

References 42 publications

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“…5 Although increasing the polyenic chain-length is a good strategy for enhancing the molecular first hyperpolarizability 6 (a key factor for efficient SHG imaging), extended polyenes suffer from poor chemical and photochemical stability. 7 Thus there is a growing interest in new amphiphilic push-pull membrane probes based on other π-conjugated units. We recently reported that porphyrin 8 dyes can be engineered for SHG imaging of cell membranes, while Andraud and co-workers have used 5 fluorethynyl π-conjugated bridge to construct probes for TPEF and SHG imaging.…”

Section: Introductionmentioning

confidence: 99%

Thiophene-based dyes for probing membranes

López‐Duarte

Chairatana

et al. 2015

Org. Biomol. Chem.

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We report the synthesis of four new cationic dipolar push-pull dyes, together with an evaluation of their photophysical and photobiological characteristics pertinent for imaging membranes by fluorescence and second harmonic generation. All four dyes consist of an N,N-diethylaniline electron-donor conjugated to a pyridinium electron-acceptor via a thiophene bridge, with either vinylene (-CH=CH-) or ethynylene 10 (-CC-) linking groups, and with either singly-charged or doubly-charged pyridinium terminals. The absorption and fluorescence behavior of these dyes was compared to a commercially available fluorescent membrane stain, the styryl dye FM4-64. The hyperpolarizabilities of all dyes were compared using hyper-Rayleigh scattering at 800 nm. Cellular uptake, localization, toxicity and phototoxicity were evaluated using tissue cell cultures (HeLa, SK-OV-3 and MDA-231). Replacing the central alkene bridge 15 of FM4-64 with a thiophene does not substantially change the absorption, fluorescence or hyperpolarizability, whereas changing the vinylene-links to ethynylenes shifts the absorption and fluorescence to shorter wavelengths, and reduces the hyperpolarizability by about a factor of two. SHG and fluorescence imaging experiments in live cells showed that the doubly-charged thiophene dyes localize in plasma membranes, and exhibit lower internalization rates compared to FM4-64, resulting in 20 less signal from the cell cytosol. At a typical imaging concentration of 1 µM, the doubly-charged dyes showed no significant light or dark toxicity, whereas the singly-charged dyes are phototoxic even at 0.5 µM, and toxic in the dark at concentrations above 5 µM. The doubly-charged dyes showed phototoxicity at concentrations greater than 10 µM, although they do not generate singlet oxygen, indicating that the phototoxicity is type I rather than type II. Our data demonstrate that the doubly-charged thiophene dyes 25 are more effective than FM4-64 as nonlinear optical imaging agents for live cells.

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Section: Introductionmentioning

confidence: 99%

Thiophene-based dyes for probing membranes

López‐Duarte

Chairatana

et al. 2015

Org. Biomol. Chem.

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“…Loew & Wuskell developed a family of styryl dyes which excite at 600-650 nm and emit >750 nm, but have restricted application for cardiac tissue study. 9,26 While these dyes have improved spectral ranges, they exhibit limited photostability and low S/N ratio signals. The hydrophilic anchor in these dyes is the quaternary ammonium group, which can be drawn through the plasma membranes of cells with large negative membrane potential and thus is not effective for anchoring the dyes at the cell surface.…”

Section: Discussionmentioning

confidence: 99%

“…recently reported the synthesis of VSDs with similar chromophores utilizing quaternary ammonium anchors. 9,21 The sulfonic acid anchors used here in dyes PGH-1 to PGH-8 produced zwitterionic dyes with the positive charge of the chromophore delocalized between the nitrogen atoms. The benzoxazole dye, PGH-5, was produced; however, the dye rapidly decomposed after chromatographic purification upon drying under mild conditions.…”

Section: Sulfonated Dyesmentioning

confidence: 99%

“…7 Effective imaging deep within tissues requires VSDs with high molar extinction coefficients at red wavelengths and near-IR emission with high quantum yields to avoid intrinsic tissue absorption and fluorescence. 6,8,9 Styryl dyes are a class of fluorescent VSDs widely used to measure membrane potentials because of their ability to follow voltage changes on a millisecond timescale. 3,6,[10][11][12] Their structures consist of an electron-rich aminophenyl group linked to a quaternized nitrogen heterocycle by a conjugated polymethine chain.…”

Section: Introductionmentioning

confidence: 99%

“…Most styryl VSDs dyes require an additional reagent such as DMSO, Pluronics or cyclodextrins for transport through the aqueous medium to the tissues. 9,15 Labeling conditions are determined for each dye, which can be a time-consuming, trial and error process. 4 Attachment of an uncharged hydrophilic anchoring group to the dye structure may eliminate the need for such agents.…”

Section: Introductionmentioning

confidence: 99%

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Enhanced aqueous solubility of long wavelength voltage-sensitive dyes by covalent attachment of polyethylene glycol

et al. 2007

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Long wavelength voltage-sensitive dyes (VSDs) called Pittsburgh (PGH) dyes were recently synthesized by coupling various heterocyclic groups to a styryl-thiophene intermediate forming extended, partially rigidized chromophores. Unlike most styryl VSDs, dyes with a sulfonic acid anchor directly attached to the chromophore showed no solvatochromic absorption shifts. The limited water solubility of many long wavelength VSDs requires the use of surfactants to transport the dye through aqueous media and effectively label biological membranes. Here, we tested the chemical substitution of the sulfonic acid moiety with polyethyleneglycol (PEG) chains ranging from MW 750 to 5000, to overcome the poor solubility of VSDs while retaining their properties as VSDs. The chemical synthesis of PGH dyes and their PEG derivatives are described. The PEGderivatives were soluble in aqueous solutions (> 1 mM) and still reported membrane potential changes. In frog and mouse hearts, the voltage sensitivity (ΔF/F per action potential) and spectral properties of PEG dyes were the same as the sulfonated analogs. Thus, the solubility of VSDs can be considerably improved with small polyethyleneglycol chains and can provide an effective approach to improve staining of excitable tissues and optical recordings of membrane potential.

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Functional Imaging of Nervous Systems Using Voltage‐Sensitive Dyes

Salzberg

2009

Encyclopedia of Life Sciences

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Voltage‐sensitive dyes are small molecules that bind to cell membranes and change their optical properties, absorption or fluorescence, in response to changes in membrane potential, the voltage across the membrane. These optical changes are extremely rapid and tend to be linear reporters of local electrical events. Thus, these ‘potentiometric probes’ behave as molecular voltmeters and can be used, with a high‐speed camera or other image dissector, for functional imaging of electrical activity including action potentials, synaptic potentials, and passive (electrotonic) signals. No physical contact with the object is required and spatial and temporal resolution is limited primarily by signal‐to‐noise considerations. Such millisecond time‐resolved functional imaging can be of single cells, of regions of cells or of nervous systems, of cardiac muscle or of any other electrically active tissue. In the nervous system, functional imaging of electrical activity can lead to an understanding of the spatio‐temporal relationships between neurons and of their functional connectivity. Key Concepts: In electrically excitable cells, the membrane potential can change extremely rapidly. Some coloured molecules (dyes) change their optical properties in response to changes in the local electric field. Changes in the optical properties of the molecules that stain‐cell membranes can be used to follow rapid changes in membrane potential. A high‐speed camera, usually with a microscope, can be used together with these dyes to monitor the electrical activity at hundreds or even thousands of sites, virtually simultaneously. Functional imaging of electrical activity can lead to an understanding of the spatio‐temporal relationships in nervous systems, and to the functional connectivity among neurons.

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Synthesis, spectra, delivery and potentiometric responses of new styryl dyes with extended spectral ranges

Cited by 77 publications

References 42 publications

Thiophene-based dyes for probing membranes

Thiophene-based dyes for probing membranes

Enhanced aqueous solubility of long wavelength voltage-sensitive dyes by covalent attachment of polyethylene glycol

Functional Imaging of Nervous Systems Using Voltage‐Sensitive Dyes

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