Calcium Signaling Required for the Fast Polyspermy Block in Xenopus Laevis

Abstract

Preventing polyspermy is essential for the normal embryonic development of most animals. Polyspermy, or the fertilization of an egg by more than one sperm, causes severe chromosomal defects and embryonic mortality. The eggs of nearly all externally fertilizing species employ a fast block to polyspermy in the form of a fertilization signaled depolarization. In the African clawed frog, Xenopus laevis, the fast block requires an increase in intracellular calcium that activates a Ca2+-activated Cl- channel; an efflux of chloride from the egg then depolarizes the membrane. Although increased Ca2+ is crucial for the fast block, the source of this Ca2+ remains unknown. Here we tested the hypothesis that Ca2+ enters the egg to signal the fast block in X. laevis using whole cell recordings on eggs inseminated in the nominal absence of Ca2+or in the presence of 10 μM Gd3+. We recorded normal depolarizations, as well as normal embryonic development, from eggs fertilized in both treatments thereby suggesting that Ca2+ is not entering from the extracellular environment. To test the importance Ca2+ released from the endoplasmic reticulum (ER) in the fast block, the membrane potential was recorded from eggs that had inhibited inositol trisphophate (IP3) synthesis or IP3 receptor binding. Our data show that inseminated eggs under both of these conditions failed to depolarize. Taken together, these data suggest that the primary source of calcium needed for the fast block to polyspermy is the ER.