The effects of energy utilization during lactation on adipose tissue metabolism were determined in 51 first lactation Holstein heifers producing between 5950 and 10,246 kg milk in 305 d. Net energy intake ranged from 18.3 to 40.6 Mcal/d during 28 to 140 d of lactation. Milk yield ranged from 13.5 to 47.4 kg/d and fat percent from 1.49 to 4.60 during 28 to 140 d, providing a range of 8.2 to 32.6 Mcal/d milk energy secretion. Calculated energy balance ranged from -16.4 to 11.5 Mcal/d. Weight change ranged from -70 to 143 kg during that 112-d period. Subcutaneous adipose tissue was biopsied nine times from 30 d prepartum to 15 d after lactation ceased. Adipose lipid synthesis measured prepartum was negatively related to subsequent milk energy secretion. Net energy intake, body weight, and body weight change were related positively to adipose lipid synthesis rates from 28 to 56 d, but those rates were related negatively to milk energy secretion. Lipolysis was positively related to milk energy secretion and body weight and negatively related to NE intake. At d 60 of lactation, adipose tissue lipid synthesis rates were a function of body weight, weight gain, and net energy intake. However, catecholamine-stimulated lipolysis rates were a function of body weight and milk energy secretion. After 140 d, lipid synthesis and lipolysis were elevated and more closely related to the previous peak rather than to concomitant milk energy secretion. These relationships demonstrate the effects of dietary energy content and genetic selection for milk production on adipose tissue metabolism.