Arterial spin labeling has been developed and used for the quantitative and completely noninvasive assessment of myocardial perfusion in vivo. Here we propose a novel arterial spin labeling method called cine-ASL, which is based on an electrocardiogram-gated steady-pulsed labeling approach combined with simultaneous readout over the cardiac cycle using cine-fast low-angle shot. This method led to shorter acquisition times than the previously used Look-Locker flow-sensitive alternating inversion recovery gradient-echo technique while preserving spatial resolution and robustness with respect to cardiac motion. High resolution perfusion mapping (in-plane resolution = 195 μm × 391 μm) was carried out with both techniques at 4.7 T in a group of 14 healthy mice. Mean perfusion values were 5.0 ± 0.8 mL g−1 min−1 with cine-ASL and 5.9 ± 1.4 mL g−1 min−1 with Look-Locker flow-sensitive alternating inversion recovery. In one animal, physiological stress was induced with higher anesthetic concentration to evaluate the response of both methods under vasodilation. Global myocardial perfusion increased from 5.6 to 16.0 mL g−1 min−1 with cine-ASL and from 6.3 to 18.7 mL g−1 min−1 with Look-Locker flow-sensitive alternating inversion recovery. Although this original scheme requires a separate T1 measurement to be fully quantitative, it improves arterial spin labeling sensitivity while maintaining compatibility with motion constraints in cardiac MRI in small rodents. Magn Reson Med 70:1389–1398, 2013. © 2012 Wiley Periodicals, Inc.