The objective of this study was to determine the effect of dietary supplemental Zn source and evaporative cooling on intake, milk yield and composition, and the rate of leukocyte migration into the mammary gland following intramammary lipopolysaccharide (LPS) infusion. Multiparous Holstein cows (n = 72) were assigned to one of four treatments with a 2×2 factorial arrangement including two sources of supplemental Zn: 75 mg/kg Zn hydroxychloride or 35 mg/kg Zn hydroxychloride + 40 mg/kg Zn-Met complex (ZMC) each with or without evaporative cooling. The cooling system was implemented by the use of fans and misters over the freestall and feeding areas. On day 34 of the experiment, cows (n = 16; days in milk = 263 ± 63 d) received an infusion of 10 μg of LPS, or a saline control, in the left or right rear quarters. Individual milk samples from both quarters were collected at -12, -4, 0, 6, 12, 24, 48, 72, 96, 120, 144, and 168 h relative to infusion and analyzed for composition and bovine serum albumin. Rectal temperature and respiration rate were assessed and blood samples were collected at the same time points (with an additional sample at 3 h) for analyses of lactose and cortisol. Complete blood counts were performed on samples collected within the first 24 h post infusion. Intramammary LPS infusion reduced (P < 0.01) milk yield, DMI and feed efficiency regardless of dietary or cooling treatments. Non-cooled cows tended (P = 0.09) to have greater feed efficiency (=milk yield/DMI) at 1 d after infusion than those subjected to cooling. Intramammary LPS infusion dramatically increased (P < 0.01) milk somatic cell count (SCC) but treatments had no apparent impact on milk SCC. Compared with cooled cows, non-cooled cows had greater (P < 0.05) plasma lactose concentrations, but lower (P < 0.03) blood concentrations of neutrophils and lymphocytes at 3 h post infusion. This suggests a greater leukocyte migration into the mammary gland of heat-stressed cows. In conclusion, noncooled cows tended to maintain greater feed efficiency and appeared to have greater leukocyte migration into the mammary gland immediately after intramammary LPS infusion compared with cooled cows. Dietary supplemental Zn source had no impact on measures assessed after intramammary LPS infusion.