1. The lipids of the rat heart have been studied with regard to amount, classes present and fatty acid composition of free fatty acids, triglycerides and phospholipids. Myocardial lipid contained 300mumoles of total fatty acid/g. dry wt. of which only 2-4mumoles were free; the remainder was esterified, chiefly as phospholipid. Neutral esters, of which triglyceride was the principal form, made up 15% of the total fatty acids. 2. When normal hearts were perfused with a nutrient-free medium until exhaustion, the triglyceride concentration declined from 43 to 13mumoles/g. dry wt. The content of phospholipids, partial glycerides and cholesteryl esters did not change. When the lipids of the rat heart were labelled with [1-(14)C]palmitate before perfusion with non-nutrient medium, radioactivity disappeared from the triglyceride, diglyceride and free fatty acid fractions, but not from the phospholipid or other ester classes. 3. These experiments support the view that only a small fraction of the total cardiac lipid, principally triglycerides and to a smaller extent diglycerides, is available as a source of fuel in the absence of exogenous substrate.