Is GnRH self-priming an obligatory feature of the reproductive cycle?

J. de Koning1,,5, C.B. Lambalk2, F.M. Helmerhorst3 and M.N. Helder4

1 Division of Medical Pharmacology, Leiden/Amsterdam Center for Drug Research, Sylvius Laboratories, University of Leiden, PO Box 9503, 2300 RA Leiden, 2 Research Institute for Endocrinology, Reproduction and Metabolism, Division of Reproductive Endocrinology and Fertility, Division of Obstetrics and Gynaecology, University Hospital Vrije Universiteit, Amsterdam, 3 Department of Obstetrics, Gynaecology and Reproductive Medicine, Leiden University Medical Center, Leiden, and 4 Department of Gynaecology, University Hospital Groningen, The Netherlands

Abstract

Insufficient suppression of LH (premature elevation) and FSH (prolonged release) give rise to blood concentrations which may cause damaging effects on oocyte viability and too many follicles respectively. During the surge, LH rises from low to high threshold values to initiate processes from initiation of the resumption of oocyte meiosis to the induction of ovulation. In general, it is thought that a dramatic increase in LH concentration is required to attain the high threshold for ovulation. A self-priming mechanism, by which gonadotrophin-releasing hormone (GnRH) enhances the LH (and FSH) responses to its own action, was thought to be responsible. However, normal LH surges in rats consist of <2–7% of the maximal pituitary releasing capacity. The physiological roles of LH and FSH favour a control mechanism that restrains their blood concentrations during most of the cycle. Ovarian proteins, e.g. inhibin and putative gonadotrophin-surge-inhibiting factor/attenuating factor (GnSIF/AF), are involved in this process. We argue that the increased pituitary LH responsiveness during the mid-cycle surge is not the result of a self-priming process that `dramatically' increases the LH releasing capacity of the pituitary gland. This is probably due to elimination by GnRH of the inhibitory action of the putative ovarian proteins GnSIF/AF.

Key words: functional antagonism/GnSAF/GnSIF/gonadotrophins/LH hypersecretion

Introduction

A striking feature in the endocrinology of reproduction is the phenomenon of self-potentiation or self-priming that was introduced as a process whereby gonadotrophin-releasing hormone (GnRH) induces potentiation of its own action on the LH response of pituitary gonadotrophs (Aiyer et al., 1974Go; Bremner and Paulsen, 1974Go). This mechanism of action of GnRH was deduced from the so-defined biphasic pituitary LH response to GnRH. It was, and still is, the general opinion that self-priming plays an important role in the generation of the pre-ovulatory LH surge. The putative ovarian proteins, gonadotrophin-surge inhibiting and attenuating factors (GnSIF/AF), were defined as inhibitors of GnRH-induced LH and FSH release (Littman and Hodgen, 1984Go; de Koning et al., 1979Go, 1994Go; Messinis and Templeton, 1991Go; de Koning, 1995Go; Fowler and Templeton, 1996Go). It is believed that the hypothalamus (via GnRH) and the ovaries (via GnSIF/AF) control concentrations of gonadotrophins in circulation via a functional antagonistic interaction. In addition, secretion of the gonadotrophins is controlled by the ovarian steroids, oestradiol and progesterone (see below), and particularly FSH by other ovarian protein hormones from the inhibin family. The latter family of proteins will not be discussed here, although an intrinsic GnSIF/AF-like bioactivity of inhibin may contribute to the effects described here for GnSIF/AF (Culler, 1992Go; Tio et al., 1994Go, 1998Go; de Koning, 1995Go).

Here, we speculate that, in the pituitary, LH (and FSH) response during the mid-cycle surges is not the result of a self-priming process, but is due to elimination by GnRH of the inhibitory action of GnSIF/AF. As well as the interaction between GnRH and GnSIF/AF, the arguments are based on: (i) the specific biphasic patterns of LH release in response to GnRH; (ii) the relatively low release of LH during the surge; and (iii) the observation that during most of the reproductive cycle, LH and FSH concentrations have to be suppressed to prevent damaging effects of their premature elevation on oocyte viability (see deKoning, 1995Go) and to limit the follicle number (Lambalk et al., 1998Go). In view of this, we defend the view that instead of GnRH self-priming causing protracted gonadotrophin secretion, the action of GnSIF/AF to prevent premature rises in blood hormone concentrations, is the dominant regulatory mechanism in the reproductive cycle.

Actions of GnRH: gonadotrophin release and the classic self-priming process

The GnRH-induced release of LH and FSH involves at least two pathways (de Koning et al., 1976Go, 1979Go, 1980Go; Pickering and Fink, 1976Go; Fink, 1996Go): (i) the releasing pathway whereby GnRH `directly' causes the secretion of these hormones; and (ii) the self-priming pathway whereby GnRH increases the potential pituitary LH and FSH responsiveness to its own action. The latter pathway involves GnRH-induced de-novo synthesis of RNA and of proteins. Several candidate proteins have been proposed (Fink, 1996Go). However, these are not LH or FSH (de Koning et al., 1976Go). These pituitary proteins act as permissive or rate-limiting factors in the releasing pathway. In response to an increased GnRH stimulus, the initial release of LH is thus rate-limited by the relatively low concentration of these proteins (Figures 1 and 2GoGo). The new synthesis of these proteins induced by GnRH is concentration-dependent and requires time: a lag-phase of 0.5–2 h during which no increase in the rate of LH release takes place (an unprimed phase) (de Koning et al., 1977Go, 1980Go). A similar lag-phase between the occurrence of the endogenous GnRH surge and of the mid-cycle LH surge is found in the ewe (Moenter et al., 1991Go). After the lag-phase, the increased expression of proteins permits an enhanced rate of GnRH-induced LH and FSH release via the releasing pathway (a primed phase). This constitutes the well-known biphasic gonadotrophic hormone secretion patterns in response to GnRH and displays the classic self-priming phenomenon (Aiyer et al., 1974Go; Bremner and Paulsen, 1974Go; de Koning et al., 1976Go, 1979Go, 1994Go; Pickering and Fink, 1976Go; Schuiling et al., 1976Go; Hoff et al., 1977Go; Baldwin et al., 1983Go; Waring and Turgeon, 1983Go; Fink, 1996Go).



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Figure 1. In intact female rats, the self-priming phenomenon is demonstrated in response to two equal pulses of gonadotrophin-releasing hormone (GnRH) (vertical arrows in the most left panel) and to a GnRH surge or infusion (horizontal arrow in the second left panel). The initial responses are limited and after protein synthesis (during the lag-phase of ~1 h) the responses are increased. In long-term ovariectomized animals no such self-priming is demonstrated. This might be due to the absence of gonadotrophin-surge-inhibiting factor/attenuating factor (GnSIF/AF). The responses to GnRH equal pulses (second right panel) and to a GnRH surge or infusion (most right panel) are high (not limited by the protein synthesis dependent step) from the beginning of stimulation onwards; the gonadotrophs were `primed' already. The latter responses are similar to those of other hypothalamic releasing hormones on their respective target cells.

 


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Figure 2. Pre-ovulatory surge and the maximal LH releasing capacity of the pituitary gland. The (almost) maximal LH releasing capacity of the pituitary gland, in response to a high concentration gonadotrophin-releasing hormone (GnRH; 400 pmol/h) infusion, compared with the spontaneous pre-ovulatory LH surge in the rat. The mean area under the curves of the spontaneous LH surges is ~8% of that of the GnRH-induced LH surges. (Taken from de Koning et al., 1994.)

 
Rationale of the classic GnRH self-priming process

Generally, the self-priming mechanism was thought to be necessary to achieve a high ovulatory LH surge. However, the extent of a normal LH surge in rats is <8% of the maximal pituitary LH responsiveness to pharmacological concentrations of GnRH (Figure 2Go) (de Koning et al., 1994Go; de Koning, 1995Go). Taken together, our results and those of other authors (Peluso, 1990Go) have shown that ~25–85% of the LH surge is needed for ovulation (see also Shoham et al., 1995Go). This is only a relatively small amount of <2–7% of the maximal pituitary capacity.

Furthermore, implicating self-priming in the LH release process suggests that the underlying mechanism of action of GnRH is different from that of other releasing hormones. Addition of the latter hormones almost instantly stimulates the release of pituitary hormones at high rates, whereas the effect of GnRH is retarded due to time-consuming protein synthesis (see above). However, a rapid, high LH response of the gonadotrophs to GnRH can be achieved in ovariectomized rats (de Koning et al., 1976Go). In women with imminent ovarian failure, increased LH and FSH blood amplitudes and responses to GnRH are observed. Blood concentrations of oestradiol in these patients are normal (de Koning et al., 2000Go). In these cases, the increased pituitary LH responsiveness might be the result of the absence or decrease of GnSIF/AF bioactivity.

GnRH-self-priming: fact or fiction?

In previous experiments we titrated the biological effect of endogenous GnSIF/AF by varying the GnRH pulse amplitude and frequency in intact phenobarbital-blocked female rats (Van Dieten and de Koning, 1995Go). The applied frequencies were based on the patterns of endogenous LH pulses in the rat (Gallo, 1981aGo,bGo). The results showed that hourly GnRH pulses of 25 pmol/kg body weight maintains normal basal blood concentrations of LH. This means that a balance exists between the action of GnRH and GnSIF/AF. Increasing the frequency of GnRH to three pulses per hour was sufficient to overrule the action of GnSIF/AF and to generate a pre-ovulatory-like LH surge. Thus, during a normal ovarian cycle only moderate amounts of GnRH and changes in its pulse frequency are sufficient to maintain basal LH concentrations and to increase LH to surge concentrations.

The results presented here and in the previous section raise two questions. Firstly, whether the endogenous release of GnRH (outside the time of the LH surge) is too low to increase the LH responsiveness to its own action, or whether the potential stimulatory effect of GnRH is prevented by an inhibitory substance like GnSIF/AF. Secondly, whether a modest LH surge (but still in excess of the amount needed for induction of ovulation) requires a self-priming mechanism that, as generally accepted, should induce the release of a `super' amount of LH.

The first question was answered by experiments in which the bioactivity of GnSIF/AF in rats was increased by injection of steroid-free bovine follicular fluid (bFF) or decreased by removal of the ovaries. Under the physiological-like conditions as pointed out above (hourly GnRH pulses of 25 pmol/kg body weight), removal of GnSIF/AF bioactivity (within hours after ovariectomy) allows an immediate self-priming-like increase in LH release. The addition of steroid-free bFF inhibits this effect. Also, injection of steroid-free bFF temporally prevents the effect of a regimen of three such GnRH pulses per hour in intact rats (Van Dieten and de Koning, 1995Go; Van Dieten et al., 1999Go). Thus, it seems that what is called a self-priming effect is the result of disinhibition by GnRH of the LH response inhibited by GnSIF/AF.

The second question was answered already by the data in which the spontaneous LH surge is compared with the total pituitary LH release capacity. From this, we conclude that there is no need for a self-priming mechanism that `dramatically' increases the LH releasing capacity of the pituitary gland to induce ovulation.

GnRH and GnSIF/AF: antagonistic control of LH blood concentrations

The above results demonstrate the functionally antagonistic control of LH blood concentrations by GnRH and GnSIF/AF. LH concentrations are kept low by a GnSIF/AF-controlled mechanism that limits the immediate pulsatile GnRH-induced LH release. Each endogenous pulse of GnRH may neutralize this inhibitory action of GnSIF/AF. However, the interval of 1 h between these pulses allows GnSIF/AF to overcome this attempt of GnRH and to keep the pituitary LH responsiveness low at the next GnRH pulse. To start the pre-ovulatory surges, these inhibitory actions by GnSIF/AF on LH and FSH release must be overruled. This is achieved by an increased GnRH pulse-frequency (Gallo, 1981aGo,bGo; Goodman, 1994Go; Hotchkiss and Knobil, 1994Go; de Koning, 1995Go; Van Dieten and de Koning, 1995Go; Van Dieten et al., 1999Go; see above). Since this process involves a protein synthesis-dependent step, it results in a biphasic LH release pattern that is indistinguishable from one that would have been the result of a self-priming phenomenon. However, this release pattern is the result of a GnRH-induced escape from a restrained state of the LH release machinery. Although an attenuated action of GnSIF/AF may participate in this process, our results do not show such a possibility (de Koning, 1995Go; Fowler and Templeton, 1996Go).

The physiological roles of LH and FSH are also in favour of a control mechanism that restrains their blood concentrations during most of the cycle. When LH exceeds certain threshold values (blood concentrations and/or integrated amounts over time) during the mid-cycle LH surge, it initiates the resumption of oocyte meiosis, then luteinization of the granulosa cells and finally the induction of ovulation (Peluso, 1990Go; Mattheij et al., 1994Go). The proper sequential attainment of these threshold concentrations of LH may be very important for optimal oocyte quality (Guerro and Rojas, 1975Go; Hunter et al., 1976Go; Regan et al., 1990Go; Mattheij et al., 1994Go). Premature elevation of LH concentrations in spontaneous or stimulated cycles for ovulation induction precipitates resumption of meiosis and, perhaps, luteinization. Ovulation and thus conception will take place only in response to the normal-timed LH surge. This increases the chance of early miscarriage of the aged fertilized oocyte(s) (Hunter et al., 1976Go; Mattheij et al., 1994Go; de Koning, 1995Go; Shoham et al., 1995Go). It is therefore, of paramount importance that LH concentrations are kept below the threshold required for the resumption of meiosis and luteinization until the start of the pre-ovulatory LH surge. The inhibitory effect of GnSIF/AF, with the relatively slow protein synthesis-dependent restoration of the LH response by GnRH, provides a very powerful control mechanism to stabilize low blood hormone concentrations until the start of the mid-cycle surges (de Koning, 1995Go; Van Dieten and de Koning, 1995Go).

Role of sex steroids

During the ovarian cycle, oestradiol and progesterone exert negative and positive feedbacks at both the central (release of GnRH) and the pituitary (LH and FSH responses to GnRH) level (Goodman and Knobil, 1981Go). These steroids inhibit the release of GnRH and LH during most of the ovarian cycle; once per cycle the inhibitory action on the hypothalamus declines (a disinhibitory or stimulatory effect of the action of oestradiol), leading to an increased GnRH release. This, in concert with the steroid-induced positive feedback at the pituitary level, results in the pre-ovulatory LH and FSH surges (see also Shoham et al., 1995Go). In humans, a similar process takes place, but a positive steroid feedback at the hypothalamic level may be less distinct (Adams et al., 1994Go; Hall et al., 1994Go). A lowered input of GnSIF/AF may participate in this process (de Koning, 1995Go; Fowler and Templeton, 1996Go). The biological action of GnSIF/AF in steroid-free bFF can be demonstrated in ovariectomized rats during pulsatile administration of GnRH, but only in animals injected with oestrogen. The final inhibitory effect of this bFF preparation on LH release was much stronger than the small stimulatory effect of oestradiol (Tijssen et al., 1997Go; Van Dieten et al., 1999Go). These results proved that GnSIF/AF acts only under an oestrogen background. Schuiling et al. (1999) concluded from their results that GnSIF/AF-like activity is anti-oestrogenic in nature.

Pituitary LH and FSH responsiveness to GnRH is already increased long before the start of the LH and FSH surges. Since oestradiol inhibits GnRH release, the blood concentrations of LH and FSH remain low. This situation anticipates the LH and FSH surges to come. Under such endocrine conditions, an increase in pulsatile GnRH release may be translated immediately into an enhanced LH release (expression of the positive feedback at the pituitary level) (Van Rees et al., 1983Go). Therefore, any small increase in GnRH release before the LH surge would cause a disturbance of this delicate balance and could lead to premature increases in LH concentrations. The inhibitory effect of GnSIF/AF allows GnRH to increase the rate of LH and FSH release only after the time-consuming process of protein synthesis. Thus, factors like GnSIF/AF underlie the effectiveness of the negative feedback of oestradiol and the correct timing of the LH surge.

On the purification and the molecular base of action of GnSIF/AF

From the above, it is clear that GnSIF/AF is well known as a biological entity that is active in many experimental designs, but only little is known about its molecular mechanism of action. So far, this is due to the failure to fully identify and characterize its molecular structure. Different groups have failed in their attempts to purify GnSIF/AF in such a way that the results can be used for molecular biological approaches to elucidate the final structure. The reasons for these failures are unknown, but GnSIF/AF may be a labile protein present in a very low concentration. In addition, the N-terminal part of the protein may be insensitive to the amino acid sequence procedure. Moreover, in the latter case, during reduced sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS–PAGE) to separate protein subunits on the basis of their molecular weight, a blocked GnSIF/AF subunit might be accompanied by a non-blocked protein, thus giving false-positive results. Finally, the failures may also be due to the applied biological assays. Namely, these assays are based on the inhibition of GnRH-induced release of LH by pituitary cells by GnSIF/AF present in biological fluids. The action of GnRH on the release of LH may be attenuated by inhibitors of cellular processes that are present in the (partly purified) biological fluids, again giving rise to false-positive results.

However, different protein forms of candidate GnSIF/AF have been reported to be purified and a few small fragments with different amino acid sequences were measured (Tio, et al., 1994Go; Danforth and Cheng, 1995Go; Pappa et al., 1999Go; M.N.Helder and J.de Koning, unpublished results). Judging from molecular weights and partial amino acid sequences, none of these results indicated that proteins from the inhibin family are present in the final purified preparations.

Actions of GnRH are mediated via second messengers like cyclic AMP [protein kinase A (PKA) pathway] and diacylglycerol [protein kinase C (PKC) pathway]. Also, the enzyme MAPKinase is involved in this release process (Mitchell et al., 1994Go). We have shown that increments in the release of LH induced by subsequent pulses of GnRH or cyclic AMP or diacylglycerol derivatives in vitro, can be counteracted by the continuous presence of steroid-free bFF (Tijssen et al., 1997Go). In addition, we were able to demonstrate that two different highly purified fractions of bovine GnSIF/AF and a common synthetic 26 amino acid peptide derived from these structures were able to antagonize the stimulation of MAPKinase activity by a single pulse of GnRH (Helder et al., 1997Go; M.N.Helder and J.de Koning, unpublished observations). These results indicate that the antagonistic interaction between GnRH and GnSIF/AF takes place in the gonadotrophs downstream from PKA and PKC activation, but upstream or at the level of the enzyme MAPKinase.

Conclusions

In contrast to other hormonal regulated systems, the female reproductive system consists of well-timed changes in hormones and tissue activity repeated during successive cycles. Specific changes in LH and FSH concentrations are only required during defined periods of the cycle. To control cyclic changes in the reproductive tract, this system therefore must not respond to acute changes or requirements, but must respond at appropriate times. This makes the reproductive system different from other hormonally-regulated systems that must usually respond in mostly acute situations of changes in the internal and/or external environment. The functional antagonism between GnRH and GnSIF/AF underlies the unique responses of the gonadotrophs to prevent sudden changes in LH and FSH blood concentrations, but also to allow mid-cycle surges. This antagonistic interaction between GnRH and GnSIF/AF and its implication in the control of LH release under pathological conditions (LH hyper- and hyposecretion) has been discussed previously (de Koning, 1995Go; Fowler and Templeton, 1996Go).

At an appropriate time of the cycle, oestradiol increases the GnRH pulse frequency and enhances the amplitude of the LH and FSH pulses in response to GnRH. These processes initiate the mid-cycle surges. In view of the above, the common view of GnRH self-potentiation in the generation of a large LH surge to induce ovulation should be seen in another light. The typical biphasic release of the gonadotrophins following GnRH administration may reflect an alternative mechanism. Rather than GnRH potentiating the LH response to its own action, primary GnSIF/AF suppresses the release of LH and FSH during the main part of the cycle. Once every cycle, the oestradiol-induced increased GnRH pulse frequency neutralizes this inhibitory action and restores the intrinsic releasing capacity of the gonadotrophs, allowing the LH and FSH surge and ovulation. This alternative mechanism does not undermine the importance of studies on the mechanisms underlying the `self-priming action', which actually show how GnRH overcomes the inhibitory effect of GnSIF/AF.



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Figure 3. Initially, the increase in the gonadotrophin-releasing hormone (GnRH) pulse frequency results in a minor increase in blood LH concentrations via the releasing pathway. After some time an LH surge will occur. The lag-phase (hatched area) is the time elapsed from the moment of the first increase in the GnRH pulse frequency and the more pronounced start of the LH surge. During this time, GnRH overcomes the inhibitory action of gonadotrophin-surge-inhibiting factor/attenuating factor (GnSIF/AF) via de-novo protein synthesis.

 
Acknowledgments

The authors wish to thank Dr Grant Montgomery (The Queensland Institute of Medical Research, Department of Genetic Epidemiology, The Bancroft Centre, Herston, Brisbane, Australia) for his helpful comments.

Notes

5 To whom correspondence should be addressed. E-mail: j.koning{at}lacdr.leidenuniv.nl Back

References

Adams, J.M., Taylor, A.E., Schoenfeld, D.A. et al. (1994) The midcycle gonadotropin surge in normal women occurs in the face of an unchanging gonadotropin-releasing hormone pulse frequency. J. Clin. Endocrinol. Metab., 79, 858–864.[Abstract]

Aiyer, M.S., Chiappa, S.A. and Fink, G. (1974) A priming effect of luteinizing hormone releasing factor on the anterior pituitary gland in the female rat. J. Endocrinol., 62, 573–588.[ISI][Medline]

Baldwin, D.M., Ramey, J.W. and Wilfinger, W.W. (1983) Characterization of the luteinizing hormone response to continuous infusion of gonadotropin releasing hormone using perifused pituitaries from intact, ovariectomized and steroid-treated rats. Biol. Reprod., 29, 99–111.[Abstract]

Bremner, W.J. and Paulsen, C.A. (1974) Two pools of luteinizing hormone in the human pituitary: evidence from constant administration of luteinizing hormone releasing hormone. J. Clin. Endocrinol. Metab., 39, 8110–8115.

Culler, M.D. (1992) Inhibin suppresses luteinizing hormone (LH)-releasing hormone self-priming: direct action on follicle-stimulating hormone secretion and opposition of estradiol-enhanced LH secretion. Endocrinology, 130, 1605–1614.[Abstract]

Danforth, D.R. and Cheng, C.Y. (1995) Purification of a candidate gonadotropin surge inhibiting factor from porcine follicular fluid. Endocrinology, 136, 1658–1665.[Abstract]

de Koning, J. (1995) Gonadotrophin surge-inhibiting/attenuating factor governs luteinizing hormone secretion during the ovarian cycle: physiology and pathology. Hum. Reprod., 10, 2854–2861.[Abstract]

de Koning, J., Van Dieten, J.A. and Van Rees, G.P. (1976) LH-RH-dependent synthesis of protein necessary for LH release from rat pituitary glands in vitro. Mol.Cell. Endocrinol., 5, 151–160.[ISI][Medline]

de Koning, J., Van Dieten, J.A. and Van Rees, G.P. (1977) Effect of pre-incubation with different concentrations of LH-RH on subsequent LH release caused by supramaximally active amounts of LH-RH; role of LH-RH-induced protein synthesis. Life Sci., 21, 1621–1627.[ISI][Medline]

de Koning, J., Van Dieten, J.A., Tijssen, A.M. et al. (1979) Studies on a protein synthesis dependent step in LH release by LH-RH. Acta Endocrinol. Copenh., 92, 648–657.[Medline]

de Koning, J., Van Dieten, J.A.M.J. and Van Rees, G.P. (1980) The pattern of LH release of rat pituitary glands during long-term exposure to LH-RH in vitro. In Jutisz, M. and McKerns, K.W. (eds), Synthesis and Release of Adenohypophyseal Hormones. Plenum Publishing Corporation, New York, NY, pp. 639–657.

de Koning, J., Westhoff, W.E., Koppenaal, D.W. et al. (1994) On the dynamics between gonadotrophin surge-inhibiting factor and gonadotrophin releasing hormone (GnRH): role of self-priming and desensitization in the luteinizing hormone response to GnRH after follicle stimulating hormone treatment. Hum. Reprod., 9, 1600–1606.[Abstract]

de Koning, C.H., Popp-Snijders, C., Schoemaker, J. et al. (2000) Elevated FSH concentrations in imminent ovarian failure are associated with higher FSH and LH pulse amplitude and response to GnRH. Hum. Reprod., 15, 1452–1456.[Abstract/Free Full Text]

Fink, G. (1996) Commentary: The self-priming effect of LHRH: An unique servomechanism and possible cellular model for memory. Front. Neuroendocrinol., 16, 183–190.[ISI]

Fowler, P.A. and Templeton, A. (1996) The nature and function of putative gonadotropin surge-attenuating/inhibiting factor (GnSAF/IF). Endocr. Rev., 17, 103–120.[ISI][Medline]

Gallo, R.V. (1981a) Pulsatile LH release during the ovulatory LH surge on proestrus in the rat. Biol. Reprod., 24, 100–104.[ISI][Medline]

Gallo, R.V. (1981b) Pulsatile LH release during periods of low level LH secretion in the rat estrous cycle. Biol. Reprod., 24, 771–777.[ISI][Medline]

Goodman, R.L. (1994) Neuroendocrine control of the ovine estrous cycle. In Knobil, E., Neill, J.D., Greenwald, G.S. et al. (eds), The Physiology of Reproduction. 2nd edn. Raven Press, New York, NY, pp. 613–658.

Goodman, R.L. and Knobil, E. (1981) The sites of action of ovarian steroids in the regulation of LH secretion. Neuroendocrinology, 32, 57–63.[ISI][Medline]

Guerro, R.J. and Rojas, O.I. (1975) Spontaneous abortion and ageing of human ova and spermatozoa. N. Eng. J. Med., 293, 573–575.[Abstract]

Hall, J.E., Taylor, A.E., Martin, K.A. et al. (1994) Decreased release of gonadotropin-releasing hormone during the preovulatory midcycle luteinizing hormone surge in normal women. Proc. Natl Acad. Sci. USA, 91, 6894–6898.[Abstract/Free Full Text]

Helder, M.N., van Eersel, S.E., van Heurn, J.-W. and de Koning J. (1997) Gonadotrophin surge-inhibiting factor inhibits GnRH-stimulated mitogen-activated protein kinase activation. Hum. Reprod., 12, 67–68.

Hoff, J.D., Lasley, B.L., Wang et al. (1977) The two pools of pituitary gonadotropin: regulation during the menstrual cycle. J. Clin. Endocrinol. Metab., 44, 302–312.[Abstract]

Hotchkiss, J. and Knobil, E. (1994) The menstrual cycle and its neuroendocrine control. In Knobil, E., Neill, J.D., Greenwald, G.S. et al. (eds), The Physiology of Reproduction. 2nd edn. Raven Press, New York, NY, pp. 711–749.

Hunter, R.H.F., Cook, B. and Baker, T.G. (1976) Dissociation of response to injected gonadotrophin between the Graafian follicle and oocyte in pigs. Nature, 260, 156–158.[ISI][Medline]

Lambalk, C.B., Boomsma, D.I., De Boer, L. et al. (1998) Increased levels and pulsatility of follicle-stimulating hormone in mothers of hereditary dizygotic twins. J. Clin. Endocrinol. Metab., 83, 481–486.[Abstract/Free Full Text]

Littman, B.A. and Hodgen, G.D. (1984) Human menopausal gonadotropin stimulation in monkeys: blockade of the luteinizing hormone surge by a highly transient ovarian factor. Fertil. Steril., 41, 440–447.[ISI][Medline]

Mattheij, J.A., Swarts, J.J., Hurks, H.M. et al. (1994) Advancement of meiotic resumption in graafian follicles by LH in relation to preovulatory ageing of rat oocytes. J. Reprod. Fertil., 100, 65–70.[Abstract]

Messinis, I.E. and Templeton, A. (1991) Attenuation of gonadotrophin release and reserve in superovulated women by gonadotrophin surge attenuating factor (GnSAF). Clin. Endocrinol. Oxf., 34, 259–263.[ISI][Medline]

Mitchell, R., Sim, P.J., Leslie, T. et al. (1994) Activation of MAP kinase associated with the priming effect of LHRH. J. Endocrinol., 140, R15–R18.[Abstract]

Moenter, S.M., Caraty, A, Locatelli, A. et al. (1991) Pattern of gonadotropin-releasing hormone (GnRH) secretion leading up to ovulation in the ewe: existence of a preovulatory GnRH surge. Endocrinology, 129, 1175–1182.[Abstract]

Pappa, A., Seferiadis, K., Fotsis, T. et al. (1999) Purification of a candidate gonadotrophin surge attenuating factor from human follicular fluid. Hum. Reprod., 14, 1449–1456.[Abstract/Free Full Text]

Peluso, J.J. (1990) Role of the amplitude of the gonadotropin surge in the rat. Fertil. Steril., 53, 150–154.[ISI][Medline]

Pickering, A. and Fink, G. (1976) Priming effect of luteinizing hormone releasing factor: in-vitro studies with raised potassium ion concentrations. J. Endocrinol., 69, 453–454.[ISI][Medline]

Regan, L., Owen, E.J. and Jacobs, H.S. (1990) Hypersecretion of luteinising hormone, infertility, and miscarriage [see comments]. Lancet, 336, 1141–1144.[ISI][Medline]

Schuiling, G.A., de Koning, J., Zurcher, A.F. et al. (1976) Induction of LH surges by continuous infusion of LH-RH. Neuroendocrinology, 20, 151–156.[ISI][Medline]

Schuiling, G.A., Valkhof, N. and Koiter, T.R. (1999) FSH inhibits the augmentation by oestradiol of the pituitary responsiveness to GnRH in the female rat. Hum. Reprod., 14, 212–216.

Shoham, Z., Schachter, M., Loumaye, E. et al. (1995) The luteinizing hormone surge – the final stage in ovulation induction: modern aspects of ovulation triggering. Fertil. Steril., 64, 237–251.[ISI][Medline]

Tijssen, A.M.I, Helder, M.N., Chu, Z.W. et al. (1997) Intracellular antagonistic interaction between GnRH and gonadotrophin surge-inhibiting/attenuating factor bioactivity downstream of second messengers involved in the self-priming process. J. Reprod. Fertil., 111, 235–242.[Abstract]

Tio, S., Koppenaal, D., Bardin, C.W. et al. (1994) Purification of gonadotropin surge-inhibiting factor from Sertoli cell-enriched culture medium. Biochem. Biophys. Res. Commun., 199, 1229–1236.[ISI][Medline]

Tio, S., Van Dieten, J.A.M.J. and de Koning, J. (1998) Immunoneutralization of follicle-stimulating hormone does not affect gonadotrophin surge-inhibiting factor/attenuating factor bioactivity during the rat ovarian cycle. Hum. Reprod., 13, 2731–2737.[Abstract/Free Full Text]

Van Dieten, J.A.J.M. and de Koning, J. (1995) From basal luteinizing hormone (LH) concentrations to the pre-ovulatory LH surge: titration of the physiological effect of gonadotrophin surge-inhibiting/attenuating factor. Hum.Reprod., 10, 3110–3116.[Abstract]

Van Dieten, J.A.M.J., Helder, M.N., Van den Oever, C. et al. (1999) Non-steroidal factors in bovine follicular fluid inhibit or facilitate the action of pulsatile administration of GnRH on LH release in the female rat. J. Endocrinol., 161, 237–243.[Abstract/Free Full Text]

Van Rees, G.P., Van Dieten, J.A., de Koning, J. et al. (1983) Effect of pre-treatment of long-term ovariectomized rats with anti-LRH on the response of the pituitary gland in vitro to the analogue of LRH D-ser-(tBu)6-des-gly10-ethylamide-LRH (Buserelin). Acta Endocrinol. Copenh., 104, 272–278.[Medline]

Waring, D.W. and Turgeon, J.L. (1983) LHRH self priming of gonadotrophin secretion: time course of development. Am. J. Physiol., 244, C410–C418.[Abstract]