Mice are now commonly used as models of human cardiovascular diseases and conditions, but it is challenging to measure blood flow velocity in small vessels such as coronary arteries. Accordingly, we have developed a method using a 2 mm diameter 20 MHz pulsed Doppler probe applied to the chest of anesthetized mice to measure left main coronary blood flow velocity noninvasively. We also found that coronary flow velocity could be increased from baseline (B) to hyperemic (H) levels by changing the concentration of isoflurane gas anesthesia from 1% to 2.5% in oxygen. We used the ratio B/H to estimate coronary flow reserve (CFR) in young, adult, and old mice and in mice with obesity, atherosclerosis, pressure overload hypertrophy, and coronary artery occlusion. We found that B/H increases with age from 2.4 (young) to 3.6 (old) and is decreased to as low as 1.1 by all forms of heart and vascular disease studied. We conclude that CFR can be measured noninvasively and serially in mice as their cardiovascular systems adapt and remodel to various imposed or natural conditions, and that coronary flow reserve may be a good index of overall cardiac function in mice and potentially in man.