Background: Laminitis is a complex and debilitating disease of horses. Numerous predisposing factors contribute to laminitis development, however the exact pathogenesis remains undetermined. Serum T4, cortisol, and histamine are components of the innate stress response and could play a causative or contributory role. Stress hormone concentrations in laminitis are largely unknown. Objective: To evaluate parameters associated with stress response in horses with laminitis, and compare these to healthy horses and horses with gastrointestinal (GI) disease. Methods: Thirty-eight adult horses presenting for non-medical conditions, GI abnormalities, or clinical laminitis were prospectively enrolled. Horses were assigned to the appropriate disease group (healthy, GI disease, and laminitis) and had blood drawn on presentation to the hospital. Samples were analyzed for plasma endogenous adrenocorticotrophic hormone (eACTH), serum cortisol, serum thyroid hormone, and plasma histamine. Results: Stress hormone concentrations were significantly different between horses in the laminitis and GI disease groups. Plasma histamine levels were highest in horses with laminitis, compared with GI disease and controls. Both horses with laminitis and GI disease had increased plasma eACTH when compared to healthy horses. Horses with GI disease had higher serum cortisol concentrations than horses with laminitis or controls. Serum T4 was lower in horses with GI disease than in horses with laminitis and controls. Conclusions Horses with laminitis had relative increases in both plasma histamine and eACTH concentrations. Serum T4 and cortisol concentrations of horses with laminitis did not differ significantly when compared to healthy horses. The role of stress hormones in equine disease warrants further investigation.