Osmolality and composition of the activating solution affects motility of fresh and frozen Prochilodus lineatus sperm differently

•Activating solution osmolality affects motility of fresh and frozen prochilod sperm.•Activating solution composition affects motility of fresh but not frozen sperm.•Prochilod sperm show short period of motility which decreases fast after activation. Osmolality and composition of the activating solution on motility of fresh and frozen Prochilodus lineatus sperm were evaluated. Sperm was triggered in 11 solutions prepared with reverse osmosis (RO) water (∼0mOsmkg−1), and glucose or NaCl adjusted to 50, 100, 150, 200 and 250mOsmkg−1. Sperm motility rate and velocities (curvilinear=VCL, among others) were evaluated in fresh sperm at 10, 30 and 50s post-activation (spa), and in frozen sperm at 10 spa only. Sperm was frozen under a standardized methodology for this species. Fresh sperm motility was higher in samples triggered in RO (91%), in glucose at all osmolalities (90–92%) and in 50–150mOsmkg−1 NaCl (88–91%) than that in 200–250mOsmkg−1 NaCl (74–80%). Motility decreased (P