Development of an image-based system for measurement of membrane potential, intracellular Ca(2+) and contraction in arteriolar smooth muscle cells.

ABSTRACT:

Objective

Changes in smooth muscle cell (SMC) membrane potential (Em) are critical to vasomotor responses. As a fluorescent indicator approach would lessen limitations of glass electrodes in contracting preparations, we aimed to develop a Forster (or fluorescence) resonance energy transfer (FRET)-based measurement for Em.

Methods

The FRET pair used in this study (donor CC2-DMPE [excitation 405 nm] and acceptor DisBAC(4) (3)) provide rapid measurements at a sensitivity not achievable with many ratiometric indicators. The method also combined measurement of changes in Ca(2+) (i) using fluo-4 and excitation at 490 nm.

Results

After establishing loading conditions, a linear relationship was demonstrated between Em and fluorescence signal in FRET dye-loaded HEK cells held

SUBMITTER: Ella SR 

PROVIDER: S-EPMC4826593 | biostudies-literature | 2010 Nov

REPOSITORIES: biostudies-literature