Store-Operated Ca2+ Entry Sustains the Fertilization Ca2+ Signal in Pig Eggs

The role of store-operated Ca2+ entry (SOCE) in the maintenance of sperm-induced Ca2+ oscillations was investigated in porcine eggs. We found that 10 mu M gadolinium (Gd3+), which is known to inhibit SOCE, blocked Ca2+ entry that was triggered by thapsigargin-induced store depletion and also caused an abrupt cessation of the fertilization Ca2+ signal. In a similar manner 3,5-bis(trifluoromethyl)pyrazole 2 (20 mu M), and tetrapandin-2 (10 mu M), potent SOCE inhibitors, also blocked thapsigargin-stimulated Ca2+ entry and disrupted the Ca2+ oscillations after sperm-egg fusion. The downregulation of Stim1 or Orai1 in the eggs did not alter the Ca2+ content of the intracellular stores, whereas co-overexpression of these proteins led to the generation of irregular Ca2+ transients after fertilization that stopped prematurely. We also found that thapsigargin completely emptied the endoplasmic reticulum, and that the series of Ca2+ transients stopped abruptly after the addition of thapsigargin to the fertilized eggs, indicating that the proper reloading of the intracellular stores is a prerequisite for the maintenance of the Ca2+ oscillations. These data strengthen our previous findings that in porcine eggs SOCE is a major signaling cascade that is responsible for sustaining the repetitive Ca2+ signal at fertilization.