OUABAIN SENSITIVITY AND EXPRESSION OF NA K-ATPASE ALPHA-SUBUNIT AND BETA-SUBUNIT ISOFORM GENES DURING BOVINE EARLY DEVELOPMENT/

The fluid movements that arise during blastocyst formation (cavitation ) are, at least in part, driven by the Na/K-ATPase. in this study, the reverse transcriptase-polymerase chain reaction (RT-PCR) was used to survey bovine pre-attachment embryos far transcripts encoding known is oforms of the Na/K-ATPase alpha- and beta-subunits, including isoforms not previously detected during the first week of mammalian developmen t. Transcripts encoding the Na-K-ATPase alpha 1, alpha 2, alpha 3 and beta 2 isoforms were detected throughout bovine preattachment developm ent. This is the first indication that alpha 2, alpha 3 and beta 2 mRN As are expressed during this early developmental interval. As in the m ouse, beta 1-subunit transcripts were not detected until the morula st age and were also present in blastocysts. Thus, in two mammalian speci es an increase in abundance of pi isoform transcripts in the morula st age is coincident with the onset of cavitation. Transcripts encoding t he recently characterized alpha 4 isoform were not detected. The sensi tivity of bovine blastocysts to ouabain (a potent inhibitor of Na/K-AT Pase) was determined by assessing the ability of bovine blastocysts to recover in ouabain supplemental culture medium following cytochalasin -induced blastocyst collapse. Re-expansion of bovine blastocysts was i nhibited in all ouabain concentrations down to 10(-9) M. Mouse blastoc ysts, in contrast, were sensitive to ouabain at or above 10(-3) M. The se results have established that transcripts encoding multiple isoform s of both the alpha and beta subunits of the Na/K-ATPase are expressed throughout early bovine development and that bovine blastocysts displ ay a greater sensitivity to ouabain than murine blastocysts. Future an alysis will determine the possible individual and collective roles of these isoforms during blastocyst formation. (C) 1997 Wiley-Liss, Inc.