Objective: By using colored microsphere (CMS) technique in newborn rats, we explored the relationship between injection sites and blood flow distribution patterns in open-chest and in closed-chest models, with hopes that percutaneous left ventricle injection can be applicable to measure relative blood flow distribution to the brain. Methods: Seven-day-old Wistar rats were used. In open-chest models (n = 30), we exposed the heart and injected CMS (15 μm diameter, 125 000 spheres in 0.05 mL) to left and right ventricles, respectively. In closed-chest models (n=12), we percutaneously punctured the left ventricle to administer microspheres to see the difference in blood flow distribution between the right and left side of organs. Microsphere counts were compared between the left and right side of each organ by unpaired t-test (mean ± SD). Results: Open-chest showed that right ventricle injection resulted in exclusive entrapment in the lungs, while left ventricle injection resulted in systemic distribution. According to this, one of 12 injections in the closed-chest was judged as right ventricle injection. The other 11 showed insignificant differences between the right and left side in the cortex, lung, and kidney. Conclusion: Right ventricle injection is differentiated from left ventricle injection and microsphere counts are the same between right and left side of the organs, suggesting that percutaneous microsphere injection is applicable for relative blood flow distribution in 7-day-old rat models.