Although osteopenia has been associated with human diabetes mellitus, the pathogenesis of diabetic osteopenia is unclear. In the present study, we evaluated the effect of diabetes on histomorphometry, bone mineral density (BMD)-measured by dual-energy X-ray absorptiometry (DXA)-and biomarkers of bone metabolism in rats up to 120 days after the onset of experimental diabetes. Female Wistar rats with a regular estrous cycle were randomly divided into two groups: control rats (n = 15) and diabetic rats without insulin treatment (n = 25). Diabetes was induced by injection of alloxan and was confirmed by the determination of blood glucose concentration (>250 mg/dl). The results revealed an approximate threefold increase of femoral trabecular distance in diabetic rats compared to controls. Conversely, trabecular thickness and bone trabecular volume were reduced twofold and 77%, respectively. BMD in both the metadiaphyseal region and total area of the femur was found to be clearly reduced in diabetic animals, with no significant differences between the groups. Serum alkaline phosphatase (ALP) and tartarate-resistant acid phosphatase (TRAP) activities showed significant six- and twofold increases, respectively, in diabetic rats. There were significant decreases in serum calcium and albumin concentrations in diabetic rats, but no difference was observed in serum magnesium, phosphorus, or creatinine concentrations between the groups. Overall, our findings support the conclusion that the diabetic state is associated with alterations in bone turnover, resulting in the development of osteopenia, which is related to the time of evolution of the disorder.