In high-intensity interval training the interval duration can be adjusted to optimize training results in oxygen uptake, cardiac output, and local oxygen supply. This study aimed to compare these variables in two interval trainings (long intervals HIIT3m: 3 min work, 3 min active rest vs. short intervals HIIT30s: 30 s work, 30 s active rest) at the same overall work rate and training duration. 24 participants accomplished both protocols, (work: 80% power output at VO2peak, relief: 85% power output at gas exchange threshold) in randomized order. Spirometry, impedance cardiography, and near-infrared spectroscopy were used to analyze the physiological stress of the cardiopulmonary system and muscle tissue. Although times above gas exchange threshold were shorter in HIIT3m (HIIT3m 1669.9 ± 310.9 s vs. HIIT30s 1769.5 ± 189.0 s, p = 0.034), both protocols evoked similar average fractional utilization of VO2peak (HIIT3m 65.23 ± 4.68% VO2peak vs. HIIT30s 64.39 ± 6.78% VO2peak, p = 0.261). However, HIIT3m resulted in higher cardiovascular responses during the loaded phases (VO2p textless 0.001, cardiac output p textless 0.001). Local hemodynamics were not different between both protocols. Average physiological responses were not different in both protocols owning to incomplete rests in HIIT30s and large response amplitudes in HIIT3m. Despite lower acute cardiovascular stress in HIIT30s, short submaximal intervals may also trigger microvascular and metabolic adaptions similar to HIIT3m. Therefore, the adaption of interval duration is an important tool to adjust the goals of interval training to the needs of the athlete or patient.