Aromatase activity of ovine follicular walls: technical validation and physiological control

Abstract

Since aromatase activity quickly disappears in cultured sheep granulosa cells, its control is poorly understood. As a result, an aromatase assay was developed using cultures of follicular walls and measuring the amount of 3H2O generated from 3H-testosterone. Chromatography and mass spectrometry analysis demonstrated that 3H2O production was indeed associated with the production of oestradiol-17 beta. Optimization of the assay demonstrated: (1) a steady increase in the amount of 3H2O produced over at least 12 h; and (2) highly significant correlations between the amounts of 3H2O measured and the weight of the follicular wall or the amount of 3H-testosterone provided. Furthermore, a highly significant correlation (r = 0.82) was observed between the amount of 3H2O produced and the concentration of oestradiol in the same samples. The effects of follicle-stimulating hormone (FSH), luteinizing hormone (LH) and oestradiol on aromatization by follicles at two specific stages of maturation (recruitment, 12 h after luteolysis; dominance, 36 h after luteolysis) were then assessed. At recruitment and dominance, FSH was able to modulate aromatase activity similarly, increasing and decreasing the activity at low concentrations and high concentrations respectively. At recruitment and dominance, oestradiol had no stimulatory effect on basal aromatase activity and even blocked the stimulatory effects of FSH on aromatase at recruitment. LH significantly inhibited the FSH-stimulated aromatase activity of dominant follicles. It is concluded that: (1) FSH may induce the recruitment of follicles by increasing aromatase activity; and (2) neither oestradiol nor LH stimulate the aromatase activity of follicles which could explain maintenance of the dominant follicle.