24-Hydroxylase (CYP24) is an enzyme distributed in the target tissues of 1alpha,25-dihydroxyvitamin D3 [1alpha,25(OH)2D3]. Two functions for this enzyme have been reported: One is production of 24,25-dihydroxyvitamin D3 [24,25(OH)2D3] and the other is inactivation of 1alpha,25(OH)2D3. To elucidate other physiologic roles of CYP24 in vivo, we previously generated rats that constitutively express the CYP24 gene. These transgenic (Tg) rats developed unexpected phenotypes, such as low plasma levels of 24,25(OH)2D3, lipidemia, and albuminuria. In this study, we elucidated the mechanisms for inducing low plasma 24,25(OH)2D3 levels and bone loss. Tg rats excreted massive amounts of vitamin D binding protein (DBP), which coincided with the loss of albumin. In Tg rats, the renal expression pattern of megalin, which serves as an endocytotic receptor responsible for the reuptake of urinary proteins such as DBP and albumin, was identical to that of the wild-type rats. Excreted albumin appeared to compete for the binding and reabsorption of the DBP-25-hydroxyvitamin D3 [25(OH)D3] complex with megalin, resulting in a loss of 25(OH)D3 into the urine and subsequent reduction of plasma 24,25(OH)2D3. In this prominent rat model of nephritis, supplementation of 25(OH)D3 was effective in preventing bone loss in an early stage of renal insufficiency.