To determine whether development and regression of cardiac hypertrophy are accompanied by changes in heart volume and to learn whether a change in heart volume is associated with changes in the myocardial connective tissue, cardiac hypertrophy was induced in rats by administration of thyroxine. Rats were given L-thyroxine for 4 weeks. Heart volume was estimated radiologically in vivo at the start of the experiment and at 1- or 2-week intervals for 7 weeks. At each of these stages a number of rats were killed, their hearts were weighed and determinations were made of the myocardial contents of DNA, of collagen measured as hydroxyproline, and of glycosaminoglycans, measured as uronic acid. After thyroxine treatment the ratio of left heart ventricle weight to body weight and of heart volume to body weight rose significantly. The increase in heart weight was greater than the increase in heart volume. At the same time, there was a significant decrease in the concentration of hydroxyproline. After discontinuation of thyroxine treatment heart volume, heart weight and the concentration of myocardial collagen returned to normal within 2 weeks. However, the total amount of myocardial collagen was still less than normal at 2 weeks. The results suggest that the decrease in the amount of myocardial collagen associated with thyroxine-induced cardiac hypertrophy--because it results in a weakening of the supporting properties of the myocardial connective tissue framework--might contribute to a slight increase in in vivo heart volume.