Variations in blood volume in the myocardium through the cardiac cycle have previously been considered constant. More recent studies have indicated a considerably variation from end diastole to end systole. These studies were nearly all performed under non-physiological conditions using muscle preparations or ex situ cardiac preparations. This study was designed to assess the dynamic changes of the intramyocardial blood volume in the intact animal under normal flow conditions using single photon emission computed tomography (SPECT). Radiolabeled, 15 μm diameter, microspheres were emoblized in the myocardial microcirculation of dogs with subsequent scans in a TRIAD single-photon-emission-computed- tomography scanner. Gated images were obtained at 63 msec intervals encompassing the entire heart. Transmural voxel (= volume element) brightness was measured in all tomographic images reflecting global and regional count density in the myocardium. There was a significant decrease in the blood volume from end diastole to end systole (10.8 cc/100 mL muscle volume; p < 0.00001). The decrease from diastole (ED) to systole (ES) in image brightness at the apex, mid ventricle, and base were: -5.7% (p < 0.01, apex vs. base), -4.7% (p < 0.01, mid ventricle vs. base) and +2.2%, respectively. Conclusions: (1) respiratory and ECG gated SPECT images allow measurement of intramyocardial blood volume changes throughout the cardiac cycle in the intact animal; (2) myocardial blood content is maximum at ED; (3) these findings progressively diminished in magnitude from apex to base.