By the use of high-performance liquid chromatography, the content of norepinephrine (NE) and epinephrine (E) in the theca of the largest and the second largest preovulatory follicle of the ovary of the hen was found to increase from 15 h (NE) or 18 h (E) to reach a peak 9 h (NE) or 9-6 h (E) before ovulation of the largest follicle. The content of progesterone in the granulosa of these follicles and the concentration of LH in the peripheral plasma of the same hen, both of which were measured by radioimmunoassays, showed a peak 6 h before ovulation. The results suggest that the catecholamines are involved in some events occurring in both follicles some several hours before ovulation of the largest follicle during the ovulatory cycle.