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BOR - Papers in Press, published online ahead of print May 4, 2005.
Biol Reprod 2005, 10.1095/biolreprod.105.040733
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BIOLOGY OF REPRODUCTION 73, 519–526 (2005)
DOI: 10.1095/biolreprod.105.040733
© 2005 by the Society for the Study of Reproduction, Inc.

Calcium/Calmodulin and Calmodulin Kinase II Stimulate Hyperactivation in Demembranated Bovine Sperm1

George G. Ignotz , and Susan S. Suarez 2 

Department of Biomedical Sciences, Cornell University, Ithaca, New York 14853

Hyperactivated motility is observed among sperm in the mammalian oviduct near the time of ovulation. It is characterized by high-amplitude, asymmetrical flagellar beating and assists sperm in penetrating the cumulus oophorus and zona pellucida. Elevated intracellular Ca2+ is required for the initiation of hyperactivated motility, suggesting that calmodulin (CALM) and Ca2+/CALM-stimulated pathways are involved. A demembranated sperm model was used to investigate the role of CALM in promoting hyperactivation. Ejaculated bovine sperm were demembranated and immobilized by brief exposure to Triton X-100. Motility was restored by addition of reactivation medium containing MgATP and Ca2+, and hyperactivation was observed as free Ca2+ was increased from 50 nM to 1 µM. However, when 2.5 mM Ca2+ was added to the demembranation medium to extract flagellar CALM, motility was not reactivated unless exogenous CALM was readded. The inclusion of anti-CALM IgG in the reactivation medium reduced the proportion hyperactivated in 1 µM Ca2+ to 5%. Neither control IgG, the CALM antagonist W-7, nor a peptide directed against the CALM-binding domain of myosin light chain kinase (MYLK2) inhibited hyperactivation. However, when sperm were reactivated in the presence of CALM kinase II (CAMK2) inhibiting peptides, hyperactivation was reduced by 75%. Furthermore, an inhibitor of CAMK2, KN-93, inhibited hyperactivation without impairing normal motility of intact sperm. CALM and CAMK2 were immunolocalized to the acrosomal region and flagellum. These results indicate that hyperactivation is stimulated by a Ca2+/CALM pathway involving CAMK2.

calcium, gamete biology, male reproductive tract, sperm motility and transport

1 Supported by grant MCB-0421855 from the National Science Foundation.

2 Correspondence: Susan S. Suarez, Department of Biomedical Sciences, Cornell University, T5-002B Veterinary Research Tower, College of Veterinary Medicine, Cornell University, Ithaca, NY 14853. FAX: 607 253 3541; sss7{at}cornell.edu






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Copyright © 2005 by the Society for the Study of Reproduction.