LEVELS OF MESSENGER-RIBONUCLEIC-ACID FOR CYTOCHROME-P450 17-ALPHA-HYDROXYLASE AND P450 AROMATASE IN PREOVULATORY BOVINE FOLLICLES DECREASE AFTER THE LUTEINIZING-HORMONE SURGE

During the transition from the follicular to the luteal phase, follicu lar steroid secretion shifts from predominantly 17beta-estradiol and a ndrogen production before the LH surge to decreased androgen and estro gen and increased progesterone production after the LH surge. Our obje ctive was to determine if changes in 17beta-estradiol and androgen pro duction by the preovulatory follicle are effected via changes in mRNA levels for the steroidogenic enzymes cytochrome P450 17alpha-hydroxyla se/C17,C20-lyase (P450-17alpha) and cytochrome P450 aromatase (P450aro m). Heifers were injected with prostaglandin F2alpha (PGF2alpha) durin g the luteal phase to initiate luteolysis and a follicular phase. Preo vulatory follicles were obtained at two times before the LH surge, in the early follicular phase (24 h after PGF2alpha injection) and in the midfollicular phase (48 h after PGF2alpha), and at one time in the la te follicular phase, after the LH surge but before ovulation (20 h aft er the onset of estrus, about 20 h after the LH surge). Granulosa cell s and theca interna were isolated from preovulatory follicles obtained during the early follicular phase and cultured with or without ovine LH (100 ng/ml; n = 4 follicles isolated 24 h after PGF2alpha). Total R NA extracted from granulosa cells and theca interna, either immediatel y after cell isolation during the early (n = 5), mid (n = 4)-, or late follicular phase (n = 5) or after 24 or 72 h of culture, was subjecte d to Northern analysis for P450-17alpha and P450arom mRNA. Autoradiogr aphs were scanned densitometrically, and values were adjusted for load ing and transfer efficiency.P450-17alpha mRNA was highly abundant in t heca interna before the LH surge, but decreased by 96% after the LH su rge (P < 0.001). In contrast, P450-17alpha mRNA in the granulosa cell fraction was very low at all three times of follicle isolation and mos t likely was due to a slight contamination of the granulosa cell prepa ration with theca interna. P450arom mRNA was abundant in granulosa cel ls isolated in the early and midfollicular phase, but decreased by 94% after the LH surge (P < 0.001). In contrast, P450arom mRNA was undete ctable in theca interna. Levels of androstenedione and 17beta-estradio l in follicular fluid were high during the early and midfollicular pha se and decreased dramatically after the LH surge (P < 0.05 and P < 0.0 01, respectively). P450-17alpha and P450arom mRNA were undetectable af ter 24 h of culture in the presence or absence of LH and thereafter. T hese data suggest that the decreased secretion of androgen and estroge n previously observed in cultures of theca interna and granulosa cells obtained after the LH surge can at least in part be accounted for by a decrease in mRNA for P450-17alpha and P450arom.