Epidemiological research has demonstrated that low daily total water intake is associated with increased diagnosis of hyperglycemia. Possible mechanisms for this increase include hormones related to the hypothalamic pituitary axis as well as the renin-angiotensin-aldosterone system (RAAS). Therefore, the hypothesis of the present study was that acute low water intake would result in differential hormonal profiles and thus impaired blood glucose regulation during an oral glucose tolerance test (OGTT) in people with type 2 diabetes mellitus (T2DM). Nine men (53 ± 9 years, 30.0 ± 4.3 m∙kg−2, 32% ± 6% body fat) diagnosed with T2DM completed OGTTs in euhydrated (EUH) and hypohydrated (HYP) states in counterbalanced order. Water restriction led to hypohydration of −1.6% of body weight, with elevated plasma (EUH: 288 ± 4, HYP: 298 ± 6 mOsm·kg−1; P < .05) and urine (EUH: 512 ± 185, HYP: 994 ± 415 mOsm·kg−1; P < .05) osmolality. There was a significant main effect of condition for serum glucose (at time 0 minute 9.5 ± 4.2 vs 10.4 ± 4.4 mmol∙L−1 and at time 120 minutes 19.1 ± 4.8 vs 21.0 ± 4.1 mmol∙L−1 for EUH and HYP, respectively; P < .001) but not insulin (mean difference between EUH and HYP −12.1 ± 44.9 pmol∙L−1, P = .390). An interaction between time and condition was observed for cortisol: decrease from minute 0 to 120 in EUH (−85.3 ± 82.1 nmol∙L−1) vs HYP (−25.0 ± 43.0 nmol∙L−1; P = .017). No differences between conditions were found within RAAS-related hormones. Therefore, we can conclude that 3 days of low total water intake in people with T2DM acutely impairs blood glucose response during an OGTT via cortisol but not RAAS-mediated glucose regulation.