Regulation and Action of Leptin in Pregnant and Lactating Dairy Cows.

The original project had four specific objectives: (1) To complete the development of a radioimmunoassay for bovine leptin; (2) To characterize the leptin system in lactating dairy cows during the transition from pregnancy to lactation; (3) To identify endocrine factors regulating the production of leptin by bovine adipose tissue; (4) To study the actions of leptin on bovine adipose and mammary tissues in vitro. However, BARD funded only the development of the bovine leptin RIA (Objective 1) for a single year. This report describes our work in completing this objective. Leptin, a protein hormone secreted predominantly by white adipose tissue, plays a critical role in the regulation of energy metabolism. In rodents and humans, leptin informs the central nervous system of the size of the energy reserves, coordinates adaptations to periods of nutrient insufficiency, and regulates the metabolism of key tissues involved in the storage and dissipation of energy. However, almost nothing is known on the biology of leptin in cattle, in part because of the absence of a valid assay to measure bovine leptin. To remediate this situation, we have developed a radioimmunoassay capable of measuring bovine leptin with a high degree of sensitivity, accuracy and precision. First, we produced recombinant bovine leptin and used it to immunize rabbits, and to prepare bovine leptin trace and standards. A single antiserum with sufficient affinity and titer was identified. Using this antiserum, binding of 125I bovine leptin was displaced in a dose dependent manner by the addition of bovine or ovine leptin. Serial dilution of bovine and ovine plasma gave displacement curves that were parallel to that of bovine or ovine leptin. Recoveries of external addition of bovine leptin in ewe and cow plasma ranged between 94 and 104%. Plasma leptin concentration measured by this assay was increased by the plane of nutrition in growing calves and lambs. Finally, plasma leptin concentration was linearly related to the fat content of the empty carcass in growing cattle. We conclude that circulating leptin in sheep and cattle is increased by fatness and plane of nutrition, consistent with results in humans and rodents. This assay provides an important tool to investigate mechanisms that regulate plasma leptin in cattle and sheep.