Free Ca2+ in the cytosol ([Ca2+]i) of individual rat ventricle cells injected with aequorin was measured under anoxia. In glucose-free medium myocytes spontaneously shortened after about 60 min, although [Ca2+]i was still at or near resting levels. However, within minutes a net inward movement of Ca2+ across the sarcolemma developed and [Ca2+]i began to rise. Provided oxygen was readmitted before [Ca2+]i exceeded 2-3 microM, cells were able to restore [Ca2+]i to resting levels through caffeine-sensitive sequestration of Ca2+ in the sarcoplasmic reticulum. We suggest that Ca2+-independent shortening of anoxic cardiomyocytes reflects onset of rigor which triggers loss of [Ca2+]i homoeostasis.