RUNNING TITLE: Nitric oxide deficiency and conceptus development

Nitric oxide (NO) is a gaseous molecule that regulates angiogenesis and vasodilation via activation of the cGMP pathway. However, functional roles of NO during embryonic development from spherical blastocysts to elongated filamentous conceptuses (embryo and extra-embryonic membrane) during the peri-implantation period of pregnancy have not been elucidated in vivo. In order to assess roles of NO production in survival and development of the ovine conceptus, we conducted an in vivo morpholino antisense oligonucleotide (MAO)-mediated knockdown of nitric oxide synthase-3 (NOS3) mRNA, the major isoform of NO synthase in ovine conceptus trophectoderm (Tr). Translational knockdown of NOS3 mRNA results in small, thin and under-developed conceptuses, but normal production of interferon tau, the pregnancy recognition signal in sheep. MAO-NOS3 knockdown in conceptuses decreased the abundance of NOS3 (72%, P < 0.05) and arginine transporter SLC7A1 proteins in conceptus Tr. Further, the amounts of ornithine and polyamines were less (P < 0.01) in uterine fluid, while the amounts of arginine (58%, P < 0.01), citrulline (68%, P < 0.05), ornithine (68%, P < 0.001), glutamine (78%, P < 0.001), glutamate (68%, P < 0.05), and polyamines (P < 0.01) were less i