Peripheral quantitative CT (pQCT) is the main method of bone mineral density (BMD) measurement in small animals. However, pQCT is usually only available in specialized centers, while clinical multidetector computed tomography (MDCT) is much more widely available. This study investigated the feasibility of using clinical 64-slice MDCT for measuring the BMD of rat lumbar vertebrae. The lumbar vertebrae of 18 7-month-old female Sprague-Dawley rats were studied. Two MDCT protocols (General Electric LightSpeed), comprising single 2.5-mm and continuous 0.625-mm acquisitions, and a single pQCT protocol (Scanco Densiscan 2000), comprising 1-mm acquisitions, were performed. The following comparisons were carried out: 2.5-mm MDCT densitometry versus 0.625-mm MDCT densitometry; 0.625-mm MDCT densitometry compared to pQCT densitometry; same day repeatability of 0.625-mm MDCT densitometry; longitudinal repeatability of 0.625-mm MDCT densitometry on day 0 and day 28 and longitudinal 0.625-mm MDCT densitometry in ovariectomized rats on day 0 and day 28. Comparisons were made using intra-class correlation coefficient (ICC). Examination time per animal was 5 min for MDCT and 30 min for pQCT. Acquisitions of 2.5-mm MDCT had a larger coefficient of variation (CoV) than 0.625-mm acquisitions. MDCT densitometry had good agreement with pQCT densitometry (ICC = 0.85). Same-day MDCT densitometry with 0.625-mm acquisitions had a small CoV (1.61%). MDCT densitometry of non-ovariectomized animals at 28 days showed no BMD change, while MDCT densitometry of ovariectomized animals showed a 13.7 ± 6.7% BMD reduction at 28 days. Clinical MDCT can reliably and accurately measure rat lumbar vertebral BMD and is much faster than pQCT.