Strips of rat ventricle were treated with EGTA (5 mM) for 24 h at 4°C and used to perform isotopical measurements of transport and release of Ca2+ in the SR in situ.45Ca accumulated by these preparations showed dependence on time of incubation until it reached saturation after 30 min. Rate and capacity of Ca2+ accumulation were calculated in 0.075 nmol mg ww−1 min−1 and 0.402 nmol mg ww−1, respectively. These values were increased by a factor of 2.6 and 8.6 when K oxalate was present in the incubation media as could be expected for Ca2+ transported by SR. Ca2+ release was assayed on45Ca desaturation curves at three free Ca2+ concentrations: 0.3, 1 and 10 μM. Significant increases in the velocity of Ca2+ efflux and net release of Ca2+ were induced only by 1 μM free Ca2+, and the Ca2+ release could be inhibited by 75% when 50 μM of ruthenium red was included in the washout solution. These results are in agreement with those obtained in assessing the SR function by mechanical measurements in skinned cardiac cells or by biochemical determinations in isolated cardiac SR vesicles. In spite of the fact that the resolution time is not as high as that required for the physiological handling of Ca2+ by SR, this methodology looks promising for approaching the SR function in cardiac pathologies as well as the effects of drugs on transport and release of Ca2+ by cardiac SR.Centro de Investigaciones Cardiovasculares