Higher gonadotrophin surge-attenuating factor bioactivity is found in small follicles from superovulated women

Abstract

Ovine and rat pituitary bioassays for gonadotrophin surge-attenuating factor (GnSAF) were utilized to determine whether the level of GnSAF bioactivity in pooled human follicular fluid (hFF) from superovulated women varied according to follicle diameter (< or = 11 mm, 12-15 mm and 16-21 mm follicles examined using the ovine bioassay, or < or = 10 mm, 11-13 mm, 14-17 mm, 18-20 mm, 21-24 mm and > or = 25 mm follicles examined using the rat bioassay). When tested using dispersed ovine pituitary cells, GnSAF bioactivity, expressed in terms of the reduction in gonadotrophin-releasing hormone (GnRH)-induced LH secretion, was inversely related to follicle diameter (P < 0.01). In response to 5 microliters hFF/well from follicles of < or = 11, 12-15 and 16-21 mm diameter, GnRH-induced LH secretion was reduced to 40.5 +/- 6.9%, 65.2 +/- 6.6% and 83.7 +/- 7.9% of control respectively. A similar inverse relationship was observed when a second batch of hFF samples from different sized follicles was tested using rat pituitary cell monolayers. Expressing GnSAF bioactivity in terms of the dose required to suppress GnRH-induced LH secretion by rat pituitary cells to 50% of the maximal suppression observed (ED50), the three smallest follicle size pools contained the most GnSAF (ED50 values of 0.13, 2.79 and 5.36 microliters hFF/well from follicles of < or = 10, 11-13 and 14-17 mm respectively). The ED50 values for follicles of 18-20, 21-24 and > or = 25 mm were 8.81, 27.1 and 60.0 microliters hFF/well respectively. Thus hFF from follicles < or = 11 mm was over 450 times more potent than hFF from follicles > or = 25 mm in suppressing GnRH-induced LH release. The ED50 values for inhibin bioactivity (measured as the suppression of basal FSH secretion from rat pituitary monolayers) were much less variable than those for GnSAF bioactivity (between 0.85 and 0.13 microliters hFF/well). Inhibin immunoreactivity, measured by a two-site immunoradiometric assay, followed the same pattern as inhibin bioactivity with lowest concentrations in the smallest follicles (41.96 ng/ml) and highest concentrations in the three largest follicle size groups (56.48-64.48 ng/ml). The specific effects of inhibin on GnRH-induced LH and basal FSH release in these pituitary bioassays were determined by incubating culture dishes with pure recombinant human inhibin at doses of 0.025-25 ng/well. In both the sheep and rat pituitary monolayers, basal FSH was suppressed (ED50 = 0.02 and 0.16 ng/well respectively).(ABSTRACT TRUNCATED AT 400 WORDS)

Activity of gonadotrophin surge-attenuating factor during the luteal phase in superovulated women

“…However, another possibility is that some of the smaller follicles that do not have the capacity to respond to hCG and luteinize continue producing GnSAF during the early luteal phase. A previous study in vitro has shown higher activity of GnSAF in the follicu¬ lar fluid of smaller compared with larger follicles (Fowler et al, 1990b …”

“…The effect of GnSAF is exerted at least at the pituitary level where it reduces luteinizing-hormonereleasing hormone (LHRH)-stimulated gonadotrophin secretion (Messinis and Templeton, 1990a). Activity of GnSAF has been detected during the follicular phase of superovulated cycles (Messinis and Templeton, 1990b). In that respect, the role of GnSAF is not limited to the time of the midcycle LH surge, but is also extended at least to the first half of the cycle when it controls LHRH-induced gonadotrophin release by a negative effect (Messinis and Templeton, 1991a).…”

The cultured rodent follicle as a model for investigations of gonadotrophin surge-attenuating factor (GnSAF) production

“…Dispersion and culture of the pituitary cells was carried out as described in Fowler et al (1994Fowler et al ( , 2002 and only preparations with . 75% viability of dispersed cells were used for bioassay.…”

“…Primary pituitary cells were cultured at 30 000 viable cells/200 ml culture medium per well in 96-well tissue culture plates. The cells were cultured under sterile conditions for 24 h at 37 8C in a watersaturated atmosphere of 5% CO 2 /95% air mixture with serum-free defined culture medium (SFDM) as defined in Fowler et al (1994). Media and non-attached cells were removed by aspiration and the wells were washed with 500 ml SFDM.…”

Non-steroidal factors in bovine follicular fluid inhibit or facilitate the action of pulsatile administration of GnRH on LH release in the female rat

“…The antagonistic interaction between GnRH and the ovarian protein, gonadotrophin surge-inhibiting factor or attenuating factor (GnSIF/AF), may be responsible for this tightly controlled LH release (de Koning et al 1979, 1987, 1989, Busbridge et al 1990, Knight et al 1990, Koppenaal et al 1993, Fowler et al 1994, de Koning 1995, van Dieten & de Koning 1995. It was postulated that after each pulse of GnRH, GnSIF/AF neutralizes the potential self-priming action of GnRH before it can become effective, thereby maintaining a state of low LH responsiveness of the gonadotrophs to the action of the next GnRH pulse (de Koning 1995, van Dieten & de Koning 1995.…”