During lactation there is an increased demand for nutrients required for milk protein and lipid synthesis as the mammary gland undergoes functional differentiation from late pregnancy to early lactation. Insulin resistance in insulin-sensitive peripheral tissues channels nutrients to the mammary gland for milk synthesis in early lactation. Glucose uptake by the mammary gland is insulin-independent, with insulin showing little effect on mRNA expression of glucose transporters. In contrast, it has long been known that the lactogenic hormones hydrocortisone, insulin and prolactin are required for maximum expression of milk protein genes in the mammary gland. Recently it has been recognized that the regulation of protein translation may play a central role in determining milk protein production. The mammalian target of rapamycin (mTOR) signal transduction pathway has been identified as a master regulator of protein translation. Data indicating that hormones (i.e., insulin and IGF-I), nutrients (i.e., amino acids) and intracellular energy status interact to regulate the mTOR signaling pathway and thus protein synthesis in the mammary gland will be presented.