New Voltage Sensitive Dyes

Abstract

To improve the photostability of voltage sensitive dyes (VSDs) we have explored structural modifications such as fluorination, trifluoromethylation, cyanation, and rigidification, on the classic ANEP (aminonaphthylethenylpyridinium) chromophore. Some new dyes show significantly improved photostability in confocal imaging of brain slices and in a photobleaching assay with a suspension of lipid vesicles. Most show high sensitivities to membrane potential changes when tested on a voltage-clamped hemispherical lipid bilayer (HLB) apparatus. The modifications also induce large spectral shifts, up to 50 nm in either blue or red direction, depending on the nature and position of modification. The high sensitivity and photostability of the new VSDs have already produced new applications, such as imaging of electrical signals in single dendritic spines. Because of the range of excitation and emission wavelengths covered by this array of new VSDs, they may be especially useful in multiplexed imaging of membrane potentials together with other fluorescent indicators in live cells and tissues. (Supported by NIH grant EB001963).