It is not clear whether the turnover of ovarian follicles during the estrous cycle in cattle is continuous and independent of the phase of the cycle, or whether waves of follicular growth occur at specific times of the cycle. To clarify this controversy, the pattern of growth and regression of ovarian follicles was characterized during a complete estrous cycle in ten heifers by daily ultrasonographic examinations. Follicles greater than or equal to 5 mm were measured and their relative locations within the ovary were determined in order to follow the sequential development of each individual follicle. Results indicated the presence of either two (n = 2 heifers), three (n = 7), or four (n = 1) waves of follicular growth per cycle. Each wave was characterized by the development of one large (dominant) follicle and a variable number of smaller (non-dominant) follicles. In the most common pattern observed (three waves/cycle), the first, second, and third waves started on Days 1.9 +/- 0.3, 9.4 +/- 0.5, and 16.1 +/- 0.7 (X +/- SEM), respectively. The dominant follicle in the third wave was the ovulatory follicle. The maximal size and the growth rate of the dominant follicle in the second wave were significantly lower than in the other waves, but no significant difference was observed between the first and third waves. For the two heifers that had two follicular waves/cycle, the waves started on Days 2 and 11, whereas in the remaining heifer (four waves/cycle), the waves began on Days 2, 8, 14, and 17, respectively. At 0, 1, 2, 3, and 4 days before estrus, the ovulatory follicle was the largest follicle in the ovaries in 100%, 95%, 74%, 35%, and 25% of follicular phases monitored, respectively. The relative size of the preovulatory follicle at the completion of luteolysis (progesterone less than 1 ng/ml) was negatively correlated (r = -0.90; p less than 0.0001) with the interval of time between the end of luteolysis and the luteinizing hormone surge, suggesting that the length of proestrus is determined by the size of the pre-ovulatory follicle at the beginning of proestrus. In conclusion, this study shows that the development of ovarian follicles greater than or equal to 5 mm in heifers occurs in waves and that the most common pattern is three waves per estrous cycle.