Aequorin measurements of free calcium in single heart cells

Abstract

The performance of the heart depends on the concentrations of free calcium ions in the cytoplasm of the myocytes. However, direct evidence for changes in free Ca concentration in physiological events during response to drugs and in pathogenesis has been difficult to obtain because of technical problems in measuring free Ca at 10(-7) M in cells with a volume of only a few picolitres. Here we describe measurements made with the Ca-sensitive photoprotein aequorin in single ventricular myocytes isolated from rat heart. We have detected signals from resting and contracting cells, and from cells exposed to media of altered ionic composition (raised K, lowered Na), ouabain and metabolic inhibitors. We report that free Ca in metabolically-poisoned myocytes is remarkably stable and that severe injury to the cell occurs before the free Ca concentration rises above 1-3 X 10(-7) M, hence cell damage seems to be a cause, not a consequence, of a rise in free Ca. The technique used here should help to resolve many uncertainties regarding free Ca in heart function, and should be particularly valuable for investigating the role of free Ca in ischaemic pathogenesis.