Marine elasmobranchs retain relatively high levels of urea to counterbalance the osmotic strength of seawater. Oviparous species, such as the little skate Raja erinacea, release encapsulated embryos that hatch after about 9 months on the seafloor. To study the ureosmotic capability of skate embryos, we measured a variety of possible osmolytes and ornithine-urea cycle (OUC) enzyme activities in little skate embryos, and determined their physiological response to dilute seawater (75% SW) exposure relative to controls (100% SW). The urea:trimethylamine oxide (TMAO) + other osmolytes ratio was 2.3-2.7:1. At the earliest stage of development investigated (4 months), there were significant levels of the key OUC enzyme, carbamoyl phosphate synthetase III, as well as ornithine transcarbamoylase, arginase and glutamine synthetase, providing evidence for a functional OUC. Embryos (4 and 8 months) survived and recovered from exposure to 5 days of 75% SW. There was a significant increase in the rate of urea excretion (five- to tenfold), no change in OUC enzyme activities, and significant decreases in the tissue content of urea, TMAO and other osmolytes in embryos exposed to 75% SW compared to 100% SW. Taken together, the data indicate that little skate embryos synthesize and retain urea, as well as a suite of other osmolytes, in order to regulate osmotic balance with the external environment. Interestingly, these ureosmotic mechanisms are in place as early as 4 months, around the time at which the egg capsule opens and the embryo is in more direct contact with the external environment.