Hyperactivated Motility of Bull Sperm Is Triggered at the Axoneme by Ca2+ and Not cAMP

Abstract

Hyperactivated motility, a swimming pattern of mammalian sperm in the oviduct, is essential for fertilization in vivo. It is characterized by high-amplitude flagellar waves and, usually, highly asymmetrical flagellar beating. It had been suggested, but not tested, that Ca2+ and cAMP switch on hyperactivation by directly affecting the flagellar axoneme. In this study, the direct affects of these agents on the axoneme were tested by using detergent-demembranated bull sperm. As confirmed by TEM, treatment of sperm with 0.2% Triton X-100 disrupted the plasma, acrosomal, and inner mitochondrial membranes, leaving axonemes intact. In the presence of 2 mM ATP, the percentage of reactivated sperm that were hyperactivated increased to 80% when free Ca2+ was increased from 50 to 400 nM. The effect of the Ca2+ in this range was to increase beat asymmetry by increasing the curvature of the principal bend. No additional increases were observed above 400 nM free Ca2+, but motility was suppressed at 1 mM. The ability of Ca2+ to produce hyperactivation depended on ATP availability, such that more ATP was required to produce the high amplitude flagellar bends characteristic of hyperactivated motility than to produce activated motility. Cyclic AMP was not required for reactivation, nor for hyperactivation. Production of hyperactivated motility also required an alkaline environment (pH 7.9-8.5). These results suggest that, provided sufficient ATP is present and pH is sufficiently alkaline, Ca2+ switches on hyperactivation by enabling curvature of the principal bends to increase.

FGFR-1 signaling is involved in spermiogenesis and sperm capacitation

“…In vivo fertilization in the absence of global tyrosine phosphorylation has not been reported. Hyperactivation, which is required for sperm to penetrate the outer vestments of the oocyte in vivo, is generally considered a functional extension of capacitation however, data particularly from the bull and Catsper knockout mice, suggest it is that possible to uncouple capacitation and hyperactivation (Carlson et al, 2003;Ho et al, 2002;Marquez and Suarez, 2004;Okunade et al, 2004).…”

Flagellar ion channels of sperm: similarities and differences between species

“…This asymmetrical, whip-like bending of the flagellum, commonly referred to as hyperactivation, is essential for the ability of mammalian sperm to overcome the protective vestments of the oocyte. The propagation of a Ca 2+ -induced wave produced from the opening of Ca 2+ channels along the flagella is a necessary milestone in sperm maturation and makes hyperactivated motility possible [4,54,55]. Additionally, [Ca 2+ ] i plays a pivotal role in acrosome exocytosis [56][57][58][59][60].…”

Characterization of 3-hydroxyisobutyrate dehydrogenase, HIBADH, as a sperm-motility marker

“…Reverse transcription-polymerase chain reactions (RT-PCR) for the synthesis of complementary DNA (cDNA), 12 μL aliquots of master mixture containing 100 ng of human testes RNA, 1 μL of 500 ng/μL oligo(dT) [12][13][14][15][16][17][18] primer (Invetrogen, Grand Island, NY, USA), and 9 μL of diethylpyrocarbonate-treated water were heated to 70°C for 10 min and put on ice. RT reactions were performed in 20 μL containing master mixture, 4 μL of 5X first-strand synthesis buffer, 0.1 M dithiothreitol, 10 mM of each dNTP, and 200 units of Superscript TM II RNase H -reverse transcriptase (Invetrogen, Grand Island, NY, USA).…”

“…Calcium signaling is also involved in sperm motility. Sperm express various voltage-gated Ca 2+ channels [13,14]. Certain voltagegated Ca 2+ channels have been shown to be critical for sperm motility by knockout experiments in mice [15,16].…”