Radium 223 (Ra-223) is an α-emitting bone-homing radiopharmaceutical that targets tumor-induced osteoblasts and is used to reduce bone pain and prolong overall survival in men with bone-metastatic, castrate-resistant prostate cancer. However, increased fracture risk in skeletal sites with no bone metastasis has been observed in patients treated with Ra-223. Both luciferase- or green fluorescence protein (GFP)-labeled osteoblast reporter mice were used to monitor the effect of Ra-223 on resident osteoblasts and normal bone structure. Upon Ra-223 treatment, 70% of resident osteoblasts were reduced within 2 days, and the osteoblast reduction lasted for at least 18 weeks without detectable recovery, as measured by in vivo bioluminescent imaging. In GFP-labeled osteoblast reporter mice, Ra-223 mainly reduced osteoblasts localized in the trabecular bone areas; the osteoblasts in the growth plates were less affected. Micro-computed tomography analyses showed that Ra-223 significantly reduced bone mineral density and bone microstructure in the trabecular area of femurs but not in the cortical bone. Tumor-induced bone was generated by inoculating osteogenic TRAMP-BMP4 prostate cancer cells into the mouse femurs; Ra-223 treatment significantly reduced tumor-induced osteoblasts. Our study shows that Ra-223 affects bone structures that are not involved in bone metastasis. Strategies that improve bone health may reduce fracture risk in patients receiving Ra-223.