Suspensions of cardiac cells have been prepared from fragments of hearts from adult rats by prolonged incubation with trypsin and collagenase. In these preparations, about 50 percent of the viable cells routinely exhibit spontaneous contractions ranging from 20 to 120 beats per minute at room temperature. The contractile activity is essentially lost at 2 degrees C but returns to normal as the temperature is increased. The rate of cellular contractions and the percentage of total cells exhibiting contractility are influenced by experimental conditions of cell preparation and the composition of the suspending medium.