The study was designed to test the effects of dietary supplementation with fish meal or specific n-3 fatty acids on ovarian activity and uterine responses in early lactating cows. From 5 to 50 d in milk (DIM), cows were fed diets that were isonitrogenous, isoenergetic, and isolipidic containing none (control), 1.25, 2.5, or 5% menhaden fish meal (FM) or 2.3% Ca salts of fish oil fatty acids (CaFOFA). Ovarian follicular dynamics were monitored along with plasma concentrations of estradiol and progesterone. Beginning at 23 DIM, cows were induced into a synchronized ovulatory cycle. On d 15 after ovulation (49 DIM), cows were injected with oxytocin and blood samples were collected to monitor uterine release of PGF(2alpha) (measured as 13, 14-dihydro-15-keto PGF(2alpha); PGFM). Uterine endometrial biopsies were collected for fatty acid analysis and cyclooxygenase-2 (COX-2) protein measurement. Ovarian follicular activities as well as plasma estradiol and progesterone concentrations were similar across diets. Endometrial fatty acid composition of eicosapentaenoic acid (C20:5, n-3) and docosahexaenoic acid (C22:6, n-3) were increased as much as 3-fold by supplementation with fish meal and CaFOFA. Conjugated linoleic acid (C18:2 cis-9, trans-11) in the endometrium was also increased; conversely, arachidonic acid (C20:4, n-6) percentage was decreased by 5% FM. Plasma PGFM response to oxytocin injection was not different among diets and endometrial COX-2 protein abundance did not differ. Results from this experiment demonstrate that dietary supplementation with fish meal or n-3 fatty acids in early lactating dairy cows significantly increased uterine n-3 fatty acid concentrations, but had no apparent effect on endometrial COX-2 or PGF(2alpha) production in response to oxytocin challenge.