To characterize the differences in photosynthate distribution and transport between nitrogen(N)-tolerant and N-susceptible sweetpotato cultivars under different N conditions, three N levels, including 0 (N0), 120 (N120), and 240 kg ha−1 (N240), were used in field experiments with the Jishu26 (J26) and Xushu32 (X32) cultivars in 2015 and 2016. The results from both years revealed that high N application reduced the tuberous root yield, the tuber/vine rate of carbon-13 (13C), and top-to-base (three equal segments of stem divided from the fifth opened leaf of the shoot tip to the main stem, defined as the top, middle, and base parts, respectively) gradients such as sucrose, ammonia N and potassium along the stem. ‘J26’ showed a higher yield than ‘X32’ under N0 but lower yield than ‘X32’ under N120 and N240. It also exhibited a higher 13C distribution to tuberous roots compared with that of ‘X32’ under N0, and the opposite trend was observed under N120 and N240. Under N0, ‘J26’ showed a steep top-to-base amino acid gradient and a significantly lower top-to-base sucrose increase along the stem in the late growth stage. Under N120 and N240, ‘X32’ exhibited a greater top-to-base decrease in the ammonia N along the stem during the main growth stages, a steep top-to-base sucrose gradient along the stem in the early growth stage, and a lower top-to-base sucrose increase along the stem in the middle and late growth stages. The formation of a reasonable photosynthate distribution structure attributed to high yield was related to a desirable sucrose, ammonia N or K+ gradient downward along the stem. These results might help provide farmers with sweetpotato cultivars using less or no N fertilizer in soils of different fertility and enhance the knowledge of yield-related physiology.