1. The influence of load and activation on relaxation of heart muscle has been studied. 2. Cardiac cells devoid of functioning sarcolemma were isolated from rat and cat ventricular myocardium. Pretreatment with the detergent Brij-58 destroyed residual sarcoplasmic reticulum function. In order to analyse the mechanical properties of relaxation in these cells, a new miniature transducer was designed which could measure force by feedback sensing (resolution of 1 microgram). Contraction was induced by ionophoretically released calcium ions. Activation, sequestration of calcium and loading conditions could be controlled independently. 3. The time course of relaxation was shown to be governed by the amount of calcium released, and unlike intact preparations from rat or cat heart (but like those from frog), to be independent of load and of alterations in load. 4. We conclude that relaxation of the cardiac contractile system is determined basically by an activation-dependent mechanism, which is masked by load dependence in intact muscle preparations with a well developed calcium sequestering membraneous system.