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doi: 10.1242/10.1242/dev.00353

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Bicarbonate actions on flagellar and Ca²⁺-channel responses: initial events in sperm activation

¹ Department of Physiology and Biophysics, Box 357290, University of Washington, Seattle, WA 98195-7290, USA
² Department of Anatomy and Cell Biology, Philipps University Marburg, 35037 Marburg, Germany
³ Department of Obstetrics and Gynecology, Box 356460, University of Washington, Seattle, WA 98195-6460, USA

^* Author for correspondence (e-mail: donner{at}u.washington.edu)

Accepted 16 December 2002

At mating, mammalian sperm are diluted in the male and female reproductive fluids, which brings contact with HCO₃^- and initiates several cellular responses. We have identified and studied two of the most rapid of these responses. Stop-motion imaging and flagellar waveform analysis show that for mouse epididymal sperm in vitro, the resting flagellar beat frequency is 2-3 Hz at 22-25°C. Local perfusion with HCO₃^- produces a robust, reversible acceleration to 7 Hz or more. At 15 mM the action of HCO₃^- begins within 5 seconds and is near-maximal by 30 seconds. The half-times of response are 8.8±0.2 seconds at 15 mM HCO₃^- and 17.5±0.4 seconds at 1 mM HCO₃^-. Removal of external HCO₃^- allows a slow return to basal beat frequency over 10 minutes. Increases in beat symmetry accompany the accelerating action of HCO₃^-. As in our past work, HCO₃^- also facilitates opening of voltagegated Ca²⁺ channels, increasing the depolarization-evoked rate of rise of intracellular Ca²⁺ concentration by more than fivefold. This action also is detectable at 1 mM HCO₃^- and occurs with an apparent halftime of 60 seconds at 15 mM HCO₃^-. The dual actions of HCO₃^- respond similarly to pharmacological intervention. Thus, the phosphodiesterase inhibitor IBMX promotes the actions of HCO₃^- on flagellar and channel function, and the protein kinase A inhibitor H89 blocks these actions. In addition, a 30 minute incubation with 60 µM cAMP acetoxylmethyl ester increases flagellar beat frequency to nearly 7 Hz and increases the evoked rates of rise of intracellular Ca²⁺ concentration from 17±4 to 41±6 nM second^-1. However, treatment with several other analogs of cAMP produces only scant evidence of the expected mimicry or blockade of the actions of HCO₃^-, perhaps as a consequence of limited permeation. Our findings indicate a requirement for cAMP-mediated protein phosphorylation in the enhancement of flagellar and channel functions that HCO₃^- produces during sperm activation.

Key words: Capacitation, sAC, cAMP-AM, PKA, Motility, Shear angle, Tan angle, Asymmetry

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