The lactational response to essential amino acid (EAA), histidine and glucose supply was determined in 6 early lactation fistulated dairy cows (92 ± 17 DIM) fed a diet to provide an NE_L of 6.9 MJ/kg DM and 11.7% crude protein. Treatments consisted of abomasal infusions of saline (Sal), 1 kg/d glucose (Glc), 563 g/d of an EAA mix (equivalent to EAA in 1kg casein) with (EAA + Glc) and without (EAA) glucose, or EAA less histidine with (–His + Glc) and without glucose (–His) in a 6 × 6 Latin square design. Data were analyzed using the MIXED procedure in SAS where period and treatment were fixed effects and cow a random effect. Milk yield was significantly increased in response to EAA + Glc compared with Sal (P < 0.001), while the removal of histidine from EAA + Glc significantly decreased milk yield (P < 0.001). Compared with Sal, milk protein yield increased 146 g/d (P < 0.001) in response to EAA. The –His treatment decreased milk protein yield 266 g/d (P < 0.001) compared with EAA, and EAA + Glc generated an increase of 346 g/d over –His + Glc (P < 0.001). Milk protein concentration increased 0.21 and 0.23 percentage points during EAA and EAA + Glc infusion compared with Sal (P ≤ 0.003), while the imbalance created with the omission of histidine, with and without glucose, significantly reduced milk protein concentrations to below those observed for Sal (P ≤ 0.002). With the exception of the complete EAA treatment, milk fat yield was significantly greater for –His compared with all other treatments (P ≤ 0.03). Milk fat concentration was 0.99 and 1.08 percentage points greater for –His compared with Glc (P < 0.001) and EAA + Glc (P < 0.001). Supplemental glucose did not stimulate a milk protein response and amplified the negative effects of a histidine deficiency. Additionally, histidine deficiency generated a positive milk fat response while glucose supplementation inhibited milk fat production. These results demonstrate the effect of histidine and glucose supply on lactation performance.