Characterization of extracellular Ca2+‐dependent full‐type hyperactivation in ejaculated boar spermatozoa preincubated with a cAMP analog

Ejaculated boar spermatozoa exhibit two types of hyperactivation: full and non‐full. Full‐type hyperactivation is characterized by the asymmetrical bending of the entire middle piece‐principal piece and a twisting/figure‐eight‐like trajectory, and can be induced by simple incubation with CaCl 2 after preincubation with a cAMP analog (Sp‐5,6‐dichloro‐1‐β‐D‐ribofuranosyl‐benzimidazole‐3′,5′‐cyclic monophosphorothioate [cBiMPS]). Here, we compared the sperm flagellar motility after treatments with elevators of [Ca 2+ ] i (cBiMPS/CaCl 2 , thimerosal, procaine, and 4‐aminopyridine) to characterize the regulatory mechanism of extracellular Ca 2+ ‐dependent, full‐type hyperactivation in ejaculated boar spermatozoa, and examined the possible involvement of Transient receptor potential cation channel subfamily C member 3 (TRPC3) in this event using the specific inhibitor Pyr3. Full‐type hyperactivation was induced by a 60‐min incubation with CaCl 2 following a 180‐min preincubation with cBiMPS but without Ca 2+ . Thimerosal‐treated spermatozoa exhibited full‐type hyperactivation in a manner independent of extracellular Ca 2+ ; conversely, this was not observed in procaine‐ or 4‐aminopyridine‐treated spermatozoa. A 20‐min treatment with Pyr3 between preincubation with cBiMPS and incubation with CaCl 2 , significantly suppressed the normal phenotype. These observations indicated that mechanisms underlying full‐type hyperactivation in spermatozoa incubated with CaCl 2 after preincubation with cBiMPS are different from those in the thimerosal‐treated spermatozoa. Furthermore, indirect immunofluorescence localized TRPC3 in the upper segment of the middle piece, which bends asymmetrically during full‐type hyperactivation but not in non‐full‐type hyperactivation, suggesting that TRPC3 may be involved in the extracellular Ca 2+ ‐dependent full‐type hyperactivation in ejaculated boar spermatozoa.