The purpose of this study was to investigate the effect of zoledronate (ZA) on osteoclast functions and viability in the tibiae of 8-week-old male mice. After weekly intravenous administration of ZA (125 μg/kg body weight) for 8 weeks, the mice were fixed by transcardial perfusion of 4% paraformaldehyde under anesthesia, and their tibiae were extracted for histochemical analysis. Compared with the control group, many tartrate-resistant acidic phosphatase-positive osteoclasts were found on the surface of the trabecular bone, but cartilage cores were obviously increased in the metaphysis of the ZA group. Osteoclasts of both groups showed similar expression of cathepsin K and matrix metalloproteinase-9. However, hardly any expression of c-src, a gene necessary for ruffled border formation and bone resorption, was found in osteoclasts of the ZA group. Moreover, no expression of CD44 or osteopontin (OPN) was observed in osteoclasts of the ZA group. Taken together, our findings suggest that ZA administration decreases the bone resorption ability of osteoclasts by inhibiting c-src expression and suppressing osteoclast adhesion by interfering with CD44/OPN binding.