Abstract
A system is described that can simultaneously record cytosolic Ca2+ concentration ([Ca2+](i)), cell length, and either membrane potential or current in single cardiac myocytes loaded with the fluorescent Ca2+ indicator indo-1. Fluorescence is excited by epi-illumination with 3.8-μs flashes of 350 ± 5 nm light from a xenon arc. Indo-1 fluorescence is measured simultaneously in spectral windows of 391-434 nm and 457-507 nm, and the ratio of indo-1 emission in the two bands is computed as a measure of [Ca2+](i) for each flash. With cells loaded with the permeant acetoxymethyl ester of indo-1, quantitation of [Ca2+](i) is not precise, owing to subcellular compartmentation of indo-1; however, the instrument would allow full quantitation if indo-1 free acid was introduced by microinjection. Simultaneously, cell length is measured on-line from the bright-field image of the cells. Because fluorescence collection is time gated during the brief flash, and red light (650-750 nm) is used for the bright-field image, cell length and [Ca2+](i) measurements are obtained simultaneously without cross talk. Membrane potential or current can be recorded simultaneously with indo-1 fluorescence and cell length via standard patch-clamping techniques.
Original language | English (US) |
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Pages (from-to) | H574-H586 |
Journal | American Journal of Physiology - Heart and Circulatory Physiology |
Volume | 258 |
Issue number | 2 27-2 |
State | Published - Jan 1 1990 |
Externally published | Yes |
Keywords
- cytosolic calcium ions
- excitation-contraction coupling
- fluorescent calcium ion indicators
- indo-1
ASJC Scopus subject areas
- Physiology
- Cardiology and Cardiovascular Medicine
- Physiology (medical)