Despite nearly a half century of studies, it has not been fully understood how pulmonary alveoli, the elementary gas exchange units in mammalian lungs, inflate and deflate during respiration. Understanding alveolar dynamics is crucial for treating patients with pulmonary diseases. In-vivo, real-time visualization of the alveoli during respiration has been hampered by active lung movement. Previous studies have been therefore limited to alveoli at lung apices or subpleural alveoli under open thorax conditions. Here we report direct and real-time visualization of alveoli of live intact mice during respiration using tracking X-ray microscopy. Our studies, for the first time, determine the alveolar size of normal mice in respiration without positive end expiratory pressure as 58 ± 14 (mean ± s.d.) μm on average, accurately measured in the lung bases as well as the apices. Individual alveoli of normal lungs clearly show heterogeneous inflation from zero to ~25% (6.7 ± 4.7% (mean ± s.d.)) in size. The degree of inflation is higher in the lung bases (8.7 ± 4.3% (mean ± s.d.)) than in the apices (5.7 ± 3.2% (mean ± s.d.)). The fraction of the total tidal volume allocated for alveolar inflation is 34 ± 3.8% (mean ± s.e.m). This study contributes to the better understanding of alveolar dynamics and helps to develop potential treatment options for pulmonary diseases.