Suppression of oocyte glycine transporter activity in mouse cumulus-oocyte complexes before resumption of meiosis

Abstract Glycine is a key regulator of cell volume in early preimplantation mouse embryos and supports embryo viability. Its accumulation is initiated when the GLYT1 glycine transporter (SLC6A9) is activated in oocytes at about the same time the oocyte is released from meiotic arrest at the germinal vesicle (GV) stage. The mechanism by which GLYT1 is maintained in an inactive state before ovulation is triggered is unknown. Here, we have shown that GLYT1 activity can remain suppressed in isolated cumulus oocyte complexes (COCs) under defined culture conditions that include keeping COCs physically separated and using the physiological mediator of GV arrest, Natriuretic Peptide Precursor C (NPPC). When GV arrest is instead maintained in oocytes within COCs by inhibiting phosphodiesterase 3A (PDE3A) or cyclin-dependent kinase 1 (CDK1), GLYT1 similarly remains inactive. However, GLYT1 becomes activated in isolated GV oocytes similarly maintained in GV arrest, indicating that cumulus cells are required for suppressing GLYT1 activity. This implied that meiotic arrest was necessary but not sufficient for preventing GLYT1 activation and that an inhibitory factor likely arising from the cumulus was also required. Finally, we found that pyrrophenone, a selective inhibitor of arachidonic acid production by cytoplasmic phospholipase A alpha (cPLAα), caused GLYT1 to become activated in oocytes within COCs despite maintenance of meiotic arrest of the oocyte. Since arachidonic acid levels decrease in oocytes after release from GV arrest, we propose that arachidonic acid may be a candidate for the inhibitory factor in COCs that regulates GLYT1 activity.