Studies on Tissue Culture of Equine Ovarian Cell Types: Effect of Gonadotrophins and Stage of Cycle on Steroidogenesis

Abstract

Granulosa cells were harvested from follicles of mares at various stages of the oestrous cycle and maintained in a tissue culture medium containing 15% horse serum, 30% medium '199' and 55% Hanks's solution. Between days 4 and 10 of culture the granulosa cells harvested from small follicles (1-2 cm. diam.) of mares in the midluteal phase of the cycle secreted an average of 0\m=.\36 pg. progesterone/cell/day. Cells harvested from large follicles of mares in the late and/or early oestrous stage of the cycle secreted an average of 29\m=.\5 pg. progesterone cell/day; the cells harvested from the large vascular follicles found at oestrus secreted an average of 173 pg./cell/ day. The small, poorly vascularized follicles found adjacent to the large vascular follicles of mares in oestrus yielded cells which secreted less progesterone than those from the larger follicles.Addition of 5 to 10 i.u. human chorionic gonadotrophin (HCG)/ml. at each medium change (every 2\p=n-\3 days) or for the first 4 days of culture brought about a marked stimulation of progesterone secretion in cultures of ' mi d\x=req-\ luteal phase' cells which was maximal after 4 to 7 days.Pregnenolone was converted primarily to progesterone, 20\g=a\-hydroxypregn-4-en-3-one and 17-hydroxyprogesterone; the metabolism was not significantly altered by the addition of a mixture of 10 i.u. HCG plus 10 i.u. pregnant mare serum gonadotrophin (PMSG). Cells harvested from mares in oestrus converted pregnenolone to progesterone in a higher yield compared with cells harvested from mares in the midluteal phase of the cycle. Addition of 10 i.u. HCG/ml. or PMSG plus HCG (10 i.u. each/ml.) stimulated aromatization of testosterone by 'midluteal phase' cultures but not by 'oestrous phase' cell cultures.These results demonstrate that the in vivo environment as well as the in vitro conditions influence the steroidogenic activity of equine granulosa cell cultures.

Effect of methotrexate and leucovorin on female reproductive tract of albino rats

“…The transition to reproductive senescence often is accompanied by elevated FSH concentrations, delayed LH surges with altered pulse frequencies, and decreased serum estradiol 104. FSH and LH levels reflect the state of ovarian steroidogenesis, ovarian functions including the final phase of follicular maturation, ovulation, and in maintaining the corpus luteum 105–108. A marked decrease in ovarian weights, an increase in atresia of large follicles, decrease in currently formed corpus luteum as MTX treatment related changes in the current study was observed.…”

Effect of testosterone and dibutyryl cAMP on the spontaneous maturation of pig oocytes

“…The mechanism(s) by which testosterone and dbcAMP act to inhibit oocyte maturation remains undetermined. Since, however, oestradiol has been shown to inhibit porcine oocyte maturation reversibly (McGaughey, 1977a; and since granulosa cells of many species (pig: Bjersing & Carstensen, 1967; Anderson et al, 1979; mare: Channing, 1969;cow: Lacroix et al, 1974;rabbit: Erickson & Ryan, 1975; sheep: Moor, 1977; rat: Fortune & aromatize androgens to oestrogen, the possibility exists that oestradiol of granulosa cell origin mediated the inhibitory action of testosterone and dbcAMP on the intact oocyte. Although the maturation of denuded oocytes was also inhibited by testosterone and dbcAMP, our results show that this inhibition was due to a direct action of the cAMP derivative on the oocyte, with testosterone playing a purely passive role.…”