Revisiting the Role of H⁺ in Chemotactic Signaling of Sperm

Chemotaxis of sperm is an important step toward fertilization. During chemotaxis, sperm change their swimming behavior in a gradient of the chemoattractant that is released by the eggs, and finally sperm accumulate near the eggs. A well established model to study chemotaxis is the sea urchin Arbacia punctulata. Resact, the chemoattractant of Arbacia, is a peptide that binds to a receptor guanylyl cyclase. The signaling pathway underlying chemotaxis is still poorly understood. Stimulation of sperm with resact induces a variety of cellular events, including a rise in intracellular pH (pH_i) and an influx of Ca²⁺; the Ca²⁺ entry is essential for the chemotactic behavior. Previous studies proposed that the influx of Ca²⁺ is initiated by the rise in pH_i. According to this proposal, a cGMP-induced hyperpolarization activates a voltage-dependent Na⁺/H⁺ exchanger that expels H⁺ from the cell. Because some aspects of the proposed signaling pathway are inconsistent with recent results (Kaupp, U.B., J. Solzin, J.E. Brown, A. Helbig, V. Hagen, M. Beyermann, E. Hildebrand, and I. Weyand. 2003. Nat. Cell Biol. 5:109–117), we reexamined the role of protons in chemotaxis of sperm using kinetic measurements of the changes in pH_i and intracellular Ca²⁺ concentration. We show that for physiological concentrations of resact (<25 pM), the influx of Ca²⁺ precedes the rise in pH_i. Moreover, buffering of pH_i completely abolishes the resact-induced pH_i signal, but leaves the Ca²⁺ signal and the chemotactic motor response unaffected. We conclude that an elevation of pH_i is required neither to open Ca²⁺-permeable channels nor to control the chemotactic behavior. Intracellular release of cGMP from a caged compound does not cause an increase in pH_i, indicating that the rise in pH_i is induced by cellular events unrelated to cGMP itself, but probably triggered by the consumption and subsequent replenishment of GTP. These results show that the resact-induced rise in pH_i is not an obligatory step in sperm chemotactic signaling. A rise in pH_i is also not required for peptide-induced Ca²⁺ entry into sperm of the sea urchin Strongylocentrotus purpuratus. Speract, a peptide of S. purpuratus may act as a chemoattractant as well or may serve functions other than chemotaxis.