Background: Hypercortisolism affects calcium and phosphate metabolism in dogs; however, the exact mechanisms are not completely understood. Objectives: To evaluate circulating concentrations of whole parathormone (wPTH), 25-hydroxyvitamin D (25-(OH)D), calcitriol, and fibroblast growth factor-23 (FGF-23) in dogs with naturally occurring hypercortisolism (NOHC) and healthy dogs, and their association with calcium and phosphate homeostasis. Animals: Twenty-three client-owned dogs with NOHC, and 12 client or staff-owned healthy dogs. Methods: Prospective cross-sectional study. The circulating concentrations of total calcium, ionized calcium (iCa), phosphate, wPTH, 25-(OH)D, calcitriol and FGF-23, and the urinary fractional excretion of phosphate (FEP) and calcium (FECa) were compared between dogs with NOHC before treatment and healthy dogs.Results: Dogs with NOHC had higher mean serum phosphate concentrations (4.81 mg/dL, SD ± 0.71 vs 3.86 mg/dL, SD ± 0.60; P < .001), median FECa (0.43%, range, 0.03-2.44 vs 0.15%, range, 0.06-0.35; P = .005), and median serum wPTH concentrations (54.6 pg/mL, range, 23.7-490 vs 24.6 pg/mL, range, 5.5-56.4; P = .003) as compared to the controls. Circulating concentrations of total calcium, iCa, and calcitriol and the FEP did not differ between groups, whereas the serum 25-(OH)D concentrations were lower in dogs with NOHC as compared to the controls (70.2 pg/mL, SD ± 42.3 vs 106.3 pg/mL, SD ± 35.3; P = .02). The dogs with NOHC had lower plasma FGF-23 concentrations than controls (316.6 pg/mL, range, 120.8-575.6 vs 448.7 pg/mL, range, 244.8-753; P = .03).Conclusions and Clinical Importance: Urine loss of calcium and hyperphosphatemia could contribute to the adrenal secondary hyperparathyroidism.