Repetitive activation of perch (Perca fluviatilis L.) sperm motility was investigated in this study. The first phase of sperm motility activation was initiated by dilution in a 260 mM glucose solution (75% motility). The second phase of motility was achieved by adding water to previously activated sperm, so that the glucose concentration dropped to 220 mM (24% motility). Finally, the third phase was obtained by further addition of water (down to 90 mM glucose) to the activated sperm suspension (15% motility). Parallel measurements of sperm ATP content were also made. The median value for nonactivated sperm was 43.9 nmol ATP/10(9) spermatozoa. The ATP concentration decreased significantly from 35 to 7 nmol ATP/10(9) spermatozoa after successive activations of motility in the above glucose solutions. Sperm velocity ranged in value from 25 to 330 microm/sec at 10 sec postactivation, from 10 to 290 microm/sec at 30 sec, and from 0 to 200 microm/sec at 45 sec. A model postulating several classes in the population of spermatozoa is developed, tentatively accounting for such successive activation. Possible further application of multiple sperm activation is discussed.