The effects of hypophysectomy (HX) on folliculogenesis, oocyte growth, and ovarian FSH and LH receptors were examined by histology, topical autoradiography, and in vitro incubation of isolated follicles with [³H]thymidine. At random stages of the estrous cycle, mice were HX (Day 0), and they were killed from 0 to 20 days thereafter. Four days after HX, the number of preantral (stages 1–3) and antral (stages 4–5) follicles per ovary was reduced by 40% to 60% compared to Day 0 values, and stage 6 (large preovulatory) follicles had vanished. By Day 20 after HX, the numbers of follicles at stages 1, 2, and 4 were comparable to Day 4 values, whereas stage 3 and 5 follicles were still further diminished to 30% and 15% of Day 0 values, respectively. The number of atretic follicles from Days 0 to 20 after HX did not differ for follicles of stages 3–5, and stage 1 and 2 follicles were all healthy. However, oocyte growth was not coordinated with follicular growth after HX; the oocytes of stage 2–4 follicles were significantly larger than Day 0 oocytes. The enlarged oocytes appeared normal as judged by light and electron microscopy. DNA synthesis for stage 1–3 follicles declined significantly from Days 4 to 20 compared with Day 0 values, but did not change for stage 4 and 5 follicles, since atretic follicles at these stages were still labeled by [³H]thymidine and this was rare for Day 0 follicles. Topical autoradiography for [¹²⁵I]-labeled FSH and hCG showed that follicular receptors for both hormones declined by Day 4 of HX and remained undetectable thereafter, except for trace amounts of [¹²⁵I]-hCG binding in the interstitium. The decrease in follicular DNA synthesis and the greater diameter of oocytes in preantral follicles after HX suggest that longer transit times are required for one stage to progress to the next because of the anhormonal environment of the growing follicle. These results provide concrete evidence that pituitary gonadotropins are essential in the mouse for initiation, proliferation, and differentiation of all stages of growing follicles.