The advance of genetics is continually leading cattle to more efficient utilization of nutrients and improved performance, but also to more metabolic disorders. Environmental changes, i.e., higher ambient temperature and humidity contribute to changes in metabolic requirements. A completely randomized design was used to evaluate the effect of heat stress and cooling while feeding an −11 mEq/kg of dietary cation anion difference (DCAD) diet to characterize thermoregulatory and hematological changes of cows in the dry period (DP). Cows were dried-off ~46 d before parturition and randomly assigned to cooling (shade, fans and soakers; CL) or heat stress (shade; HT). In the DP, vaginal temperature (VT, °C; every 10 min), respiration rate (RR; breaths per min) and temperature humidity index (THI) were recorded to evaluate heat strain. On one day (THI ≥78.8 in CL and HT) blood samples were collected at 0600, 1200, 1800 and 2400 from cows (HT = 27.5 ± 12.8 d dry; CL = 24.3 ± 14.8 d dry; n = 12/group) to assess electrolytes, hematology, and blood gases. The model included the fixed effect of prepartum environment (HT vs. CL) in a completely randomized design with cow as random effect analyzed using repeated measures. Urine pH was recorded weekly and was similar for HT and CL cows (P = 0.36). Cows under HT had higher VT and RR (P < 0.01), lower blood pH (HT = 7.41 ± 0.003, CL = 7.43 ± 0.003, P < 0.05) and had higher iCa at 0600 (HT = 1.30 ± 0.02, CL = 1.25 ± 0.01, P < 0.05) relative to CL. There was a time effect, whereby iCa was lower at 1200 but higher at 2400, regardless of treatment (P < 0.01). CL cows had higher blood glucose at 1200 and 2400 relative to HT cows (66.6 ± 1.08 vs. 63.4 ± 1.06, P < 0.05; 68.3 vs. 64.1 ± 1.08 P < 0.01, respectively). There was also a time effect on K concentration, whereby K was lower at 1200 but higher at 0600, regardless of treatment (P < 0.05). HT cows had higher PCO2 relative to CL cows (P < 0.05). Comparing HT vs CL, no differences were found for Na, Hct, Hgb, HCO₃, pO_2, SO₂ and tCO₂. Thus, we observed different physiological responses of cows receiving the same level of negative DCAD, but under different environmental conditions.