Movie 1

Simultaneous live monitoring of motility parameters (composite orientation (bottom left), head and flagellum beating time-series -frequency and amplitude-) and [Ca²⁺]i changes with intercalated fluorescence for a free swimming thapsigargin treated spermatozoon (before and after), extracted with the segmentation-free method (see also Fig. 1). After a significant delay of ~75 s from the thapsigargin addition (at ~45 s), the cell briefly displays a hyperactivated type of swimming before stopping, as the rate of [Ca²⁺]i increase slows before approaching the plateau. Thereafter, when [Ca²⁺]i is no longer increasing during the plateau, the sperm re-initiates motility displaying a hyperactivated type mode behavior which becomes more exacerbated as [Ca²⁺]i continues increasing.