Effects of duodenally infused casein on milk secretion were investigated by coupling measurements of mammary blood flow and arteriovenous differences. Four lactating cows were given a continuous duodenal infusion of 0, 177, 352, and 762 g of calcium caseinate/d according to a Latin square design. Diets, formulated to meet 90% of energy and protein recommendations, consisted of a ratio of corn silage:concentrate of 76:24 supplemented with dehydrated alfalfa. Yields of milk, protein, and fat increased linearly with casein infusions (by up to 2.6 kg/d, 126 g/d, and 133 g/d, respectively) as did milk protein content (by up to 2.2 g/kg). Mammary blood flow and insulin, prolactin, and growth hormone in plasma were not affected by added casein. Concentrations of arterial acetate, glucose, BHBA, and NEFA remained unchanged, but their uptake by the udder increased by 3, 18, 22, and 91%, respectively, between the control and high casein treatments. Plasma arterial concentrations and the mammary uptake of essential and nonessential AA increased linearly with casein infusions. Although arterial concentrations increased to a much greater degree for essential AA than for nonessential AA, the increased udder extraction was higher for nonessential AA. The main part of the added AA (90%) was extracted by the udder. However, a direct transfer of supplementary-extracted AA into milk did not occur, thus increasing AA availability for other metabolic pathways.