REGULATION OF BLASTOCOELE FORMATION BY INTRACELLULAR CALCIUM-RELEASE IS MEDIATED THROUGH A PHOSPHOLIPASE C-DEPENDENT PATHWAY IN MICE

Calcium signaling plays a critical role in the regulation of mouse pre implantation development, in part through the activation of calmodulin , Calcium transients in mouse morulae appear to be generated predomina ntly through the production of inositol 1,4,5-trisphosphate (IP3) by p hospholipase C (PLC), IP3 receptors predominate in mouse embryos as re gulators of calcium release, Exposure to the PLC inhibitors ET-18-OCH3 or U73122 resulted in a dose-dependent, reversible inhibition of cavi tation, while the inactive analogue U73343 did not alter the rate of c avitation, as compared to that in controls, U73122 inhibited the relea se of calcium from intracellular stores after exposure to ethanol or l ysophosphatidic acid, suggesting that PLC is required for blastocoele formation in the mouse and that calcium signaling may be PLC-dependent , In addition to IF,, PLC activation generates diacylglycerol, which s timulates protein kinase C (PKC) and could also alter the rate of embr yonic development, However, activation of PKC with phorbol ester or sy nthetic diacylglycerol was not sufficient to accelerate development, a lthough embryo culture in medium containing PKC inhibitors did delay c avitation, Exposure to a PKC inhibitor during ethanol-induced calcium signaling did not attenuate the ensuing acceleration of cavitation. Th ese results demonstrate that a PLC-mediated signaling pathway is requi red for blastocoele formation and that the generation of IF,, but not diacylglycerol, is critical for accelerating preimplantation developme nt.