Purpose: We hypothesize that Approach: Female C57BL/6 mice were exposed to cigarette smoke or ambient air for 1, 3, or 6 months. Each mouse received a respiratory-gated micro-CT scan at baseline and another scan following the exposure period, while anaesthetized and free-breathing. Images were obtained representing end-expiration and peak inspiration, and measurements were performed to characterize the lung structure and compute functional metrics. Following the final micro-CT session, the mice were euthanized and the lungs prepared for histology. Results: Following 6 months of smoke-exposure, the mice exhibited larger increases in end-expiration lung volume and functional residual capacity, and a reduction in weight gain when compared with air-exposed mice. The histogram of CT numbers in the lung obtained during end-expiration also showed a shift to lower CT numbers following 6 months of smoke-exposure, indicating increased air content within the lungs. The metrics suggested air-trapping in the lung, which is consistent with emphysema. In the 3-month exposure group, only the reduction in weight gain was significant compared with the air-exposed group. Histological analysis confirmed that the 6-month smoke-exposed mice likely developed centrilobular emphysema as measured by the mean linear intercept. Conclusions: Respiratory-gated micro-CT imaging of free-breathing mice at multiple respiratory phases is noninvasive and provides additional information about lung structure and function that complements postmortem techniques and could be used to monitor changes over time.