Impact of anisosmotic conditions on structural and functional integrity of cumulus–oocyte complexes at the germinal vesicle stage in the domestic cat

Abstract

During cryopreservation, the immature oocyte is subjected to anisosmotic conditions potentially impairing subsequent nuclear and cytoplasmic maturation in vitro. In preparation for cryopreservation protocols and to characterize osmotic tolerance, cat cumulus-oocyte complexes (COC) at the germinal vesicle (GV) stage were exposed for 15 min to sucrose solutions ranging from 100 to 2,000 mOsm and then examined for structural integrity and developmental competence in vitro. Osmolarities > or =200 and < or =750 mOsm had no effect on incidence of oocyte nuclear maturation, fertilization success, and blastocyst formation compared to control COC (exposed to 290 mOsm). This relatively high osmotic tolerance of the immature cat oocyte appeared to arise from a remarkable stability of the GV chromatin structure as well as plasticity in mitochondrial distribution, membrane integrity, and ability to maintain cumulus-oocyte communications. Osmolarities <200 mOsm only damaged cumulus cell membrane integrity, which contributed to poor nuclear maturation but ultimately had no adverse effect on blastocyst formation in vitro. Osmolarities >750 mOsm compromised nuclear maturation and blastocyst formation in vitro via disruption of cumulus-oocyte communications, an effect that could be mitigated through 1,500 mOsm by adding cytochalasin B to the hyperosmotic solutions. These results (1) demonstrate, for the first time, the expansive osmotic tolerance of the immature cat oocyte, (2) characterize the fundamental role of cumulus-oocyte communications when tolerance limits are exceeded, and (3) reveal an interesting hyperosmotic tolerance of the immature oocyte that can be increased two-fold by supplementation with cytochalasin B.