Gamete Biology

Store-Operated Calcium Entry in Human Oocytes and Sensitivity to Oxidative Stress

Abstract
Calcium signaling is a cellular event that plays key roles at many steps of fertilization and early development. However, little is known regarding the contribution of extracellular Ca²⁺ influx into the cell to this signaling in gametes and early embryos. To better know the significance of calcium entry on oocyte physiology, we have evaluated the mechanism of store-operated calcium entry (SOCE) in human MII oocytes and its sensitivity to oxidative stress, one of the major factors implicated in the outcome of in vitro fertilization (IVF) techniques. We show that depletion of intracellular Ca²⁺ stores through inhibition of sarco(endo)plasmic Ca²⁺-ATPase with thapsigargin triggers Ca²⁺ entry in resting human oocytes. Ba²⁺ and Mn²⁺ influx was also stimulated following inhibition, and Ca²⁺ entry was sensitive to pharmacological inhibition, since the SOCE blocker 2-aminoethoxydiphenylborate (2-APB) reduced calcium and barium entry. These results support the conclusion that there is a plasma membrane mechanism responsible for the capacitative divalent cation entry in human oocytes. Moreover, the Ca²⁺ entry mechanism described in MII oocytes was found to be highly sensitive to oxidative stress. Hydrogen peroxide, at micromolar concentrations that could mimic culture conditions in IVF, elicited an increase of [Ca²⁺]_i that was dependent on the presence of extracellular Ca²⁺. This rise was preventable by 2-APB, indicating that it was mainly due to the enhanced influx through store-operated calcium channels. In sum, our results demonstrate the occurrence of SOCE in human MII oocytes, and that oxidative stress can modify this pathway with possible consequences in IVF.

Key words: Calcium • hydrogen peroxide • oocyte • oxidative stress • store-operated calcium entry