Purpose We evaluated whether repeated high-intensity interval exercise (HIIE) influences plasma oxytocin (OT) concentration in healthy men, and, given that OT is mainly synthesized in the hypothalamus, we assessed the concentration difference between the arterial (OT_ART) versus the internal jugular venous OT concentration (OT_IJV). Additionally, we hypothesized that an increase in cerebral OT release and the circulating concentration would be augmented by repeated HIIE. Methods Fourteen healthy men (age = 24 ± 2 yr; mean ± SD) performed two identical bouts of HIIE. These HIIE bouts included a warm-up at 50%-60% maximal workload (W_max) for 5 min followed by four bouts of exercise at 80%-90% W_max for 4 min interspersed by exercise at 50%-60% W_max for 3 min. The HIIE bouts were separated by 60 min of rest. OT was evaluated in blood through radial artery and internal jugular vein catheterization. Results Both HIIE bouts increased both OT_ART (median [IQR], from 3.9 [3.4-5.4] to 5.3 [4.4-6.3] ng·mL^-1 in the first HIIE, P < 0.01) and OT_IJV (from 4.6 [3.4-4.8] to 5.9 [4.3-8.2] ng·mL^-1, P < 0.01), but OT_ART-IJV was unaffected (from -0.24 [-1.16 to 1.08] to 0.04 [-0.88 to 0.78] ng·mL^-1, P = 1.00). The increased OT levels were similar in the first and second HIIE bouts (OT_ART P = 0.25, OT_IJV P = 0.36). Conclusions Despite no change in the cerebral OT release via the internal jugular vein, circulating OT increases during HIIE regardless of the accumulated exercise volume, indicating that OT may play role as one of the exerkines.