Use of chlorotetracycline fluorescence to demonstrate Ca2+-induced release of Ca2+ from the sarcoplasmic reticulum of skinned cardiac cells

Abstract

It has been proposed that the trans-sarcolemmal influx of Ca2+ occurring during the plateau of the mammalian cardiac action potentials is insufficient in itself to activate the myofilaments, but can trigger a release of Ca2+ from the sarcoplasmic reticulum (SR) which is sufficient for activation. The demonstration of this Ca2+-induced release of Ca2+ relied entirely on experiments in which the tension developed by the myofilaments was used as a sensor of the changes of myoplasmic free Ca2+ concentration ([free Ca2+]) in segments of single cardiac cells from which the sarcolemma had been removed by microdissection (skinned cardiac cells). The small size of these preparations has previously prevented the use of more direct methods for the detection of myoplasmic Ca2+ movements. The present study is a direct demonstration of Ca2+-induced release of Ca2+ from the SR of skinned cardiac cells treated with chlorotetracycline (CTC), a fluorescent chelate probe which enables changes in the amount of Ca2+ bound to a variety of biological membranes or micelles to be monitored. The fluorescence increases when more Ca2+ is bound.