The study investigates the mechanism of action of alendronate, a bisphosphonate, in rats. Alendronate binds to bone particles with a dissociation constant of approximately 1 mM at pH 7 and a binding capacity of 100 nmol/mg bone. At lower pH, binding decreases, indicating that alendronate is released during acidification, which occurs in the resorption space beneath osteoclasts. This release increases local concentration, which inhibits bone resorption and prevents ruffled border formation without destroying the osteoclasts. In vitro, alendronate inhibits bone resorption by isolated osteoclasts when the amount on the bone surface is around 1.3 × 10⁻³ fmol/μm², leading to a concentration of 0.1–1 mM in the resorption space. This concentration causes membrane leakiness to calcium. In vivo, alendronate is localized to resorption surfaces in rat bone, with 72% of the osteoclastic surface, 2% of the bone forming surface, and 13% of all other surfaces densely labeled 1 day after injection. Six days later, the label is 600–1,000 μm away from the epiphyseal plate, indicating normal growth and matrix deposition. Osteoclasts from adult animals infused with parathyroid hormone-related peptide (PTHrP) and treated with alendronate lack ruffled borders but not clear zones. Alendronate binds to hydroxyapatite in bone, which accounts for its rapid uptake and selective action on bone. The binding is pH-dependent, with significant release at pH 3.5. In vitro, alendronate inhibits bone resorption by both rat and chicken osteoclasts with an EC50 of approximately 0.1 μM. Alendronate also increases permeability to calcium and other ions, which may contribute to its inhibitory effect on bone resorption. The findings suggest that alendronate binds to resorption surfaces, is locally released during acidification, and interferes with bone resorption and ruffled border formation without destroying the osteoclasts. These results support the hypothesis that alendronate's mechanism of action involves binding to hydroxyapatite, local release during acidification, and interference with osteoclast function. The study also highlights the importance of understanding the interaction between bisphosphonates and bone cells to develop effective treatments for bone diseases.The study investigates the mechanism of action of alendronate, a bisphosphonate, in rats. Alendronate binds to bone particles with a dissociation constant of approximately 1 mM at pH 7 and a binding capacity of 100 nmol/mg bone. At lower pH, binding decreases, indicating that alendronate is released during acidification, which occurs in the resorption space beneath osteoclasts. This release increases local concentration, which inhibits bone resorption and prevents ruffled border formation without destroying the osteoclasts. In vitro, alendronate inhibits bone resorption by isolated osteoclasts when the amount on the bone surface is around 1.3 × 10⁻³ fmol/μm², leading to a concentration of 0.1–1 mM in the resorption space. This concentration causes membrane leakiness to calcium. In vivo, alendronate is localized to resorption surfaces in rat bone, with 72% of the osteoclastic surface, 2% of the bone forming surface, and 13% of all other surfaces densely labeled 1 day after injection. Six days later, the label is 600–1,000 μm away from the epiphyseal plate, indicating normal growth and matrix deposition. Osteoclasts from adult animals infused with parathyroid hormone-related peptide (PTHrP) and treated with alendronate lack ruffled borders but not clear zones. Alendronate binds to hydroxyapatite in bone, which accounts for its rapid uptake and selective action on bone. The binding is pH-dependent, with significant release at pH 3.5. In vitro, alendronate inhibits bone resorption by both rat and chicken osteoclasts with an EC50 of approximately 0.1 μM. Alendronate also increases permeability to calcium and other ions, which may contribute to its inhibitory effect on bone resorption. The findings suggest that alendronate binds to resorption surfaces, is locally released during acidification, and interferes with bone resorption and ruffled border formation without destroying the osteoclasts. These results support the hypothesis that alendronate's mechanism of action involves binding to hydroxyapatite, local release during acidification, and interference with osteoclast function. The study also highlights the importance of understanding the interaction between bisphosphonates and bone cells to develop effective treatments for bone diseases.