How DASPMI reveals mitochondrial membrane potential : fluorescence decay kinetics and steady-state anisotropy in living cells

  • Spectroscopic responses of the potentiometric probe 2-(4-(dimethylamino)styryl)-1-methylpyridinium iodide (DASPMI) were investigated in living cells by means of a time- and space-correlated single photon counting technique. Spatially resolved fluorescence decays from single mitochondria or only a very few organelles of XTH2 cells exhibited three-exponential decay kinetics. Based on DASPMI photophysics in a variety of solvents, these lifetimes were attributed to the fluorescence from the locally excited state, intramolecular charge transfer state, and twisted intramolecular charge transfer state. A considerable variation in lifetimes among mitochondria of different morphologies and within single cells was evident, corresponding to high physiological variations within single cells. Considerable shortening of the short lifetime component ({tau}1) under a high-membrane-potential condition, such as in the presence of ATP and/or substrate, was similar to quenching and a dramatic decrease of lifetime in polar solvents. Under these conditions {tau}2 and {tau}3 increased with decreasing contribution. Inhibiting respiration by cyanide resulted in a notable increase in the mean lifetime and a decrease in mitochondrial fluorescence. Increased DASPMI fluorescence under conditions that elevate the mitochondrial membrane potential has been attributed to uptake according to Nernst distributions, delocalization of {pi}-electrons, quenching processes of the methyl pyridinium moiety, and restricted torsional dynamics at the mitochondrial inner membrane. Accordingly, determination of anisotropy in DASPMI-stained mitochondria in living cells revealed a dependence of anisotropy on the membrane potential. The direct influence of the local electric field on the transition dipole moment of the probe and its torsional dynamics monitor changes in mitochondrial energy status within living cells.

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Author:Radhan Ramadass, Jürgen Bereiter-HahnORCiDGND
Pubmed Id:
Parent Title (English):Biophysical journal
Publisher:Cell Press ; Biophysical Society
Place of publication:Cambridge, Mass. ; Bethesda, Md.
Document Type:Article
Date of Publication (online):2008/11/07
Date of first Publication:2008/10/15
Publishing Institution:Universitätsbibliothek Johann Christian Senckenberg
Release Date:2008/11/07
Page Number:9
First Page:4068
Last Page:4076
© 2008 The Biophysical Society. This is an Open Access article distributed under the terms of the Creative Commons-Attribution Noncommercial License (, which permits unrestricted noncommercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Source:Biophysical Journal 95:4068-4076 (2008) doi:10.1529/biophysj.108.135079
Institutes:Biowissenschaften / Biowissenschaften
Exzellenzcluster / Exzellenzcluster Makromolekulare Komplexe
Dewey Decimal Classification:5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie
Sammlungen:Sammlung Biologie / Sondersammelgebiets-Volltexte
Licence (German):License LogoCreative Commons - Namensnennung-Nicht kommerziell 2.0