Molecular reproduction and development
The present study was conducted to investigate the mechanisms underlying fluid movement across the trophectoderm during blastocyst formation by determining whether aquaporins (AQPs) are expressed during early mammalian development. AQPs belong to a family of major intrinsic membrane proteins and function as molecular water channels that allow water to flow rapidly across plasma membranes in the direction of osmotic gradients. Ten different AQPs have been identified to date. Murine preimplantation stage embryos were flushed from the oviducts and uteri of superovulated CD1 mice. Reverse transcription-polymerase chain reaction (RT-PCR) methods employing primer sets designed to amplify conserved sequences of AQPs (1-9) were applied to murine embryo cDNA samples. PCR reactions were conducted for up to 40 cycles involving denaturation of DNA hybrids at 95 degrees C, primer annealing at 52-60 degrees C and extension at 72 degrees C. PCR products were separated on 2% agarose gels and were stained with ethidium bromide. AQP PCR product identity was confirmed by sequence analysis. mRNAs encoding AQPs 1, 3, 5, 6, 7, and 9 were detected in murine embryos from the one-cell stage up to the blastocyst stage. AQP 8 mRNAs were not detected in early cleavage stages but were present in morula and blastocyst stage embryos. The results were confirmed in experimental replicates applied to separate embryo pools of each embryo stage. These results demonstrate that transcripts encoding seven AQP gene products are detectable during murine preimplantation development. These findings predict that AQPs may function as conduits for trophectoderm fluid transport during blastocyst formation.