ANALYSIS OF GLUTAMINASE ACTIVITY AND RNA EXPRESSION IN PREIMPLANTATION MOUSE EMBRYOS

Glutamine is utilized as an energy substrate in preimplantation mouse embryos. Glutaminase is the enzyme responsible for the conversion of g lutamine to glutamic acid, which then enters the trichloro acetic acid (TCA) cycle as alpha-ketoglutarate. Glutaminase enzyme activity was a ssessed in preimplantation embryos that developed in vivo, and glutami nase RNA expression was examined in embryos that developed in vivo or were cultured in CZB medium to various preimplantation stages between 1-cell and blastocyst. Glutaminase activity in 1-8-cell-stage mouse em bryos that developed in vivo ranged from 0.009-0.01 U/mg protein (2.39 -2.95 x 10(-7) U per embryo), and increased 3-4 Fold to 0.034 U/mg pro tein (8.13 x 10(-7) U per embryo) at the blastocyst stage. Relative st age-specific expression of glutaminase RNA was assessed by reverse tra nscription polymerase chain reaction (RT-PCR) in embryos that develope d both in vivo and in CZB culture. In vivo, glutaminase RNA was expres sed at the 1-cell stage, declined to 23% of 1-cell levels at the early 2-cell stage, and reaccumulated from late 2-cell through blastocyst s tage, where it reached a high of 204% of 1-cell levels. CZB-cultured e mbryos exhibited a similar pattern of developmental RNA expression, de clining to 30% of 1-cell levels at the early 2-cell stage, and increas ing RNA expression at the blastocyst stage to 191% of the 1-cell level . (C) 1997 Wiley-Liss, Inc.