One or two days after single injection with HCG (500 IU/kg) and chum salmon pituitary homogenate (7 mg/kg) combinedly, spermiation was successfully induced in three-year-old cultured tiger puffer Takifugu rubripes with fully grown testis. In experiment 1, milt was sampled from five males every day after hormone injection, and spermatocrit and motility of spermatozoa (time until all spermatozoa cease their forward movement) were monitored for 30 days. Spermatocrit maintained low values (30-59%) for early 10 days, thereafter gradually increased and exceeded 80% after 23 days. In contrast, motility of spermatozoa showed stable values around 60 second during experimental period. In experiment 2, milt was sampled successively from spermiated 10 males at short time intervals; 2, 4, 8, 12, 16, 20 and 24 hours after first sampling (0 hour). Spermatocrit showd a wide range (29-83%), but individual spermatocrit maintained stable value during the experimental period. Motility of spermatozoa also maintained stable values between 60 and 80 second. The results obtained indicate that a single injection with HCG and chum salmon pituitary combinedly is efficient for inducing and synchronizing spermiation of cultured tiger puffer. Milt stripped out the fish after such hormonal treatment showed stable motility instead of spermatocrit, and it may be useful for artificial fertilization over a long period of time.