Components of histotroph in the uterine lumen of pregnant ewes are required for survival and development ovine conceptuses (embryo and extraembryonic membranes). The hypothesis that arginine (Arg), leucine (Leu), glutamine (Gln) and glucose have differential effects on hypertrophy, hyperplasia and differentiated functions critical to ovine conceptus development is supported by results from our laboratory indicating significant increases in Arg, Leu, Gln and glucose in the uterine lumen between Days 10 and 15 of pregnant, but not cyclic ewes. These nutrients activate the phosphatidylinositol 3-kinase (PI3K)/proto-oncogenic protein kinase 1 (AKT1) and/or FK506 binding protein 12-rapamycin associated protein 1 (FRAP1; also known as mammalian target of rapamycin or mTOR)/ribosomal protein S6 kinase (RPS6K) signaling pathways in other cell types. Therefore, this study examined effects of these select nutrients on signal transduction pathways in ovine trophectoderm cells and conceptuses. In Study One, effects of Arg, Leu, Gln and glucose on PI3K-AKT1 and FRAP1-RPS6K signaling pathways were studied using mononuclear ovine trophectoderm cells (oTr) isolated from Day 15 conceptuses. The oTr cells were serum-starved for 24h in a customized medium containing one-tenth the normal amount of the select nutrients and then treated with physiological concentrations of either Arg (0.2 mM), Leu (0.2 mM), Gln (0.4 mM), or glucose (2 mM). Western blot analyses of whole cell extracts with antibodies to phosporylated (activated) target proteins found that Arg, Leu and glucose increased levels of phosphorylated AKT1 (p-AKT1) by 2.5- (P<0.01), 1.8- (P<0.05), and 1.4-fold, respectively, within 15 min. Arg, Leu and glucose also stimulated 2.2- (P<0.01), 1.7- (P<0.05) and 1.9-fold (P<0.01) increases, respectively, in phosphorylated-glycogen synthase kinase 3 beta (p-GSK3B) within 15 min as well as increases in p-FRAP1 (P<0.05) and p-RPS6K (P<0.001) protein between 0 and 15 min post-treatment that were sustained to 60 min. In Study Two, conceptuses (n=15) recovered from Day 16 of pregnancy were washed twice in PBS, and then divided equally (about 100 mg) for culture with Arg (0.2 mM), Leu (0.2 mM), Gln (0.4 mM) and glucose (2 mM) in 2 ml of customized medium. Western blot analyses of whole conceptus extracts with antibody to p-RPS6K found that Arg, Leu, Gln and glucose increased levels of p-RPS6K by 2.0- (P<0.01), 2.2- (P<0.01), 1.8- (P<0.05) and 1.4-fold. Collectively, these results indicate that Arg, Leu, Gln and glucose coordinately activate PI3K-AKT1 and FRAP1-RPS6K signaling pathways in ovine trophectoderm cells and conceptuses, and support the hypothesis that components of histotroph activate these cell signaling pathways to stimulate hypertrophy, hyperplasia, and/or migration of conceptus trophectoderm. (Supported by USDA Grant 2006-35203-17283).