Anti-Muellerian hormone (AMH) is synthesized only by granulosa cells in female mammals as an inhibiting factor for early follicular recruitment. Especially in preantral and small antral follicles, expression of AMH is very intensive. However, we hypothesize that AMH affects also the development of large follicles by an estrogenesis modification. This current investigation provides for the first time information about concentrations of AMH in equine follicular fluid and demonstrates their relationship to functional characteristics of follicle development. Therefore, fluids of follicles in different developmental stages were obtained by repeated, transvaginal ultrasound guided-follicle aspirations from 8 mares during dioestrus and from preovulatory follicles (after hCG-inducted ovulation) during oestrus. Concentrations of AMH, 1 713-estradiol-, progesterone- and GE-i were measured in the follicular fluids. Data showed significant negative correlations between intra follicular concentrations of AMH and 1 713-estradiol (r = -0.58; p < 0.0001) as well as between AMH and follicle diameter (r = -0.76; p < 0.001), but not between AMH and progesterone or IGF-1. Based on the observed dynamic changes of AMH secretion in large antral follicles during follicle development, a regulatory involvement of AMH in equine follicle development is likely. Due to species-specific features, the results are discussed considering their potential relevance to controlled superovulation in mares.