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Aequorin measurements of free calcium in single heart cells
Insights
Measuring free calcium ions (Ca) in heart cells is crucial for understanding heart function and disease. This study used a novel technique to show that cell damage, not high Ca levels, may cause heart problems.
Area of Science:
- Biochemistry
- Cell Biology
- Cardiology
Background:
- Heart function relies on intracellular free calcium ion (Ca) concentrations in myocytes.
- Measuring Ca in picoliter-volume cells at 10(-7) M is technically challenging, hindering research into drug responses and pathogenesis.
Purpose of the Study:
- To develop and apply a method for measuring free Ca in single rat ventricular myocytes.
- To investigate the role of free Ca in cellular injury and heart function under various conditions.
Main Methods:
- Utilized the Ca-sensitive photoprotein aequorin for measurements in isolated single rat ventricular myocytes.
- Exposed cells to altered ionic compositions, ouabain, and metabolic inhibitors to simulate physiological and pathological conditions.
Main Results:
- Detected measurable free Ca signals in resting, contracting, and stressed myocytes.
- Found that free Ca in metabolically poisoned myocytes remained stable, with cell injury preceding significant Ca concentration increases (above 1-3 X 10(-7) M).
Conclusions:
- Cellular damage appears to be a cause, rather than a consequence, of elevated free Ca in heart cells.
- The aequorin technique offers a valuable tool for resolving uncertainties in heart function and investigating the role of free Ca in ischemic pathogenesis.
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.