Precise estimation of cellular water content is a necessary basis for quantitative studies of metabolic control in the heart; however, marked discrepancies in water spaces of heart tissue are found in the literature. Reasons for this wide diversity are analyzed, and the conclusion is that the most probable value of total intracellular water content is 615 ml H(2)O/kg of wet mass (wm) and intracellular content of dry substance is 189 g/kg wm in intact in vivo rat heart. An extracellular water of 174 ml per kg wm and 22 g of dry mass per kg wm in vascular and interstitium spaces account for the rest of the tissue mass. These values can be directly related to normoosmotic saline perfused hydrated hearts, characterized by water accumulation in the extracellular spaces. Due to essentially intact heart cells, the experimentally determined dry mass, water and metabolite contents of these hydrated hearts can be extrapolated to the original morphological configuration of an intact heart muscle before the onset of edema. Such an 'extrapolated' heart is defined as a standardized perfused heart (SPH). SPH is the heart in its original morphological configuration, characterized by cell density and cellular water contents of the intact heart, but with perfusate in the extracellular spaces. The total cellular water is distributed in the cell compartments of SPH and intact hearts according to volumes of particular compartments and density of their dry mass. The volumes of bulk water phases in different organelles, accessible to diffusion of low molecular metabolites, were obtained after corrections for the fraction of 'bound' water of 0.3 g per g of compartmental dry mass content. The diffusible water spaces are proposed to be 321, 55, 153, 21 and 8 ml/kg wm for myofibrils, sarcoplasm, mitochondria, sarcoplasmic reticulum and nuclei, respectively. The SPH model allows direct comparison of metabolic data for intact and perfused hearts. We used this model to analyze the penetration of extracellular marker into cells of intact and hydrated perfused rat hearts.