Activated immune cells are obligate glucose utilizers; thus, study objectives were to estimate the quantity of whole body glucose utilization during an IV endotoxin challenge. Fasting lactating Holstein cows (718 ± 16 kg; 169 ± 7 DIM) were jugular catheterized and assigned 1 of 3 bolus treatments: control (CON; 5 mL saline; n = 6), lipopolysaccharide infused (LPS; 1.5 μg/kg BW; E. coli 055:B5; n = 6), and LPS + euglycemic clamp (LPS-Eu; 1.5 μg/kg BW; 50% dextrose infusion; n = 6). After infusion, blood glucose was determined every 10 min and dextrose infusion was adjusted in LPS-Eu cows to maintain euglycemia. Blood samples were obtained 3, 6, 9, and 12 h post-bolus for further analysis. Cows were milked 6 and 12 h post-bolus. Milk yield decreased in LPS and LPS-Eu cows relative to CON (80%, P < 0.01). Milk SCC was increased in LPS relative to CON (48%, P = 0.02) while LPS-Eu did not differ from either treatment. LPS and LPS-Eu cows were hyperglycemic for 3 h post-bolus, but thereafter glucose content decreased in LPS relative to LPS-Eu and CON cows (30%, P < 0.01). Circulating insulin was and tended to be increased in LPS-Eu (80%, P = 0.01) and LPS (72%, P = 0.06) relative to CON. Plasma NEFA, BHBA, and Ca were decreased (P < 0.01) in LPS and LPS-Eu relative to CON (46, 53, and 46%, respectively). Plasma haptoglobin and L-lactate were increased (P < 0.01) in LPS and LPS-Eu cows relative to CON (72 and 62%, respectively). Serum amyloid A was increased in LPS (47%, P = 0.01) and tended be increased in LPS-Eu (34%, P = 0.09) compared with CON cows. White blood cells were decreased in LPS and LPS-Eu relative to CON cows (P < 0.01). Total glucose deficit during the 12 h post-bolus was calculated as the decrease in the amount of glucose required to synthesize milk (relative to pre-infusion levels) plus the amount of glucose infused to maintain euglycemia (in LPS-Eu cows only). Glucose deficit for CON, LPS, and LPS-Eu cows was 483, 1259, and 1553 g, respectively. If this model is a proxy of glucose demands during an immune response, our data indicates an intensely activated immune system uses at least 90 g glucose/h and maintaining euglycemia does not rescue the decrease in milk synthesis.