To clarify the differences in the action of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] and 24,25-(OH)2D3 in hypophosphatemic (Hyp) mice, a model for familial X-linked hypophosphatemic rickets in humans, we carried out histomorphometric examinations of the effects of these agents in the lumbar vertebra of these mice. The Hyp mice received 1-1000 micrograms/kg.day 24,25-(OH)2D3, 0.01-0.1 micrograms/kg.day 1,25.(OH)2D3, or vehicle alone given daily for 28 days by ip injection. Histomorphometrically, 1,25-(OH)2D3 and 24,25-(OH)2D3 showed similar effects on bone formation. The parameters of bone formation, mineralized bone volume/bone volume, mineral apposition rate, and bone formation rate/bone surface, were improved to a similar extent in a dose-dependent manner by 1,25-(OH)2D3 and 24,25-(OH)2D3, but there were remarkable differences in the indexes of the bone resorption between these two metabolites. In 24,25-(OH)2D3-treated Hyp mice, osteoclast number/bone perimeter and osteoclast surface/bone surface, the parameters of bone resorption, increased to control levels and did not change according to the dose of 24,25-(OH)2D3. However, in 1,25-(OH)2D3-treated Hyp mice, these values increased remarkably, exceeding the control level. That is, 24,25-(OH)2D3 normalized bone resorption in the rachitic mice, whereas 1,25-(OH)2D3 caused excessive stimulation of bone resorption. This qualitative difference between the two compounds contributes to the superior effects exerted by 24,25-(OH)2D3 in improving the bone lesion in Hyp mice. At doses from 1-1000 micrograms/kg.day, 24,25-(OH)2D3 had dose-dependent effects in increasing bone formation without promoting excessive bone resorption, as shown by histomorphometric analysis.