Negative Feedback Action Of Estradiol Research Articles

Melatonin implants do not alter estrogen feedback or advance puberty in gilts

Puberty in pigs is often delayed during late summer and autumn, with long daylength the most likely cause. We hypothesised (1) that gilts born around the shortest day would have a later release from the negative feedback actions of estradiol than gilts born around the spring equinox and (2) melatonin treatment would result in an earlier release from estradiol negative feedback and advance the onset of puberty in gilts born around the spring equinox. We first determined the optimal number of estradiol implants required to monitor the release from estradiol negative feedback in ovariectomised gilts. Secondly we determined whether melatonin implants altered negative feedback in 4 cohorts of ovariectomised gilts born between the winter solstice and spring equinox, and in the following year whether melatonin altered the time of the first ovulation in 5 cohorts of intact gilts born between the winter solstice and spring equinox. Plasma LH and FSH increased between 126 and 210d of age (P<0.001) in each cohort (season), but there was no effect of cohort, melatonin treatment or interactions (P>0.05). Age at first detection of elevated plasma progesterone in untreated, intact gilts decreased across the 4 cohorts (P<0.05). Melatonin treatment of intact gilts failed to advance the age of puberty irrespective of their season of birth (P>0.05). In conclusion, while we confirmed that estradiol sensitivity is decreased as gilts age, we failed to demonstrate any effects of season or melatonin on estradiol feedback or melatonin on puberty.

Positive, but not negative feedback actions of estradiol in adult female mice require estrogen receptor α in kisspeptin neurons.

Hypothalamic kisspeptin (Kiss1) neurons express estrogen receptor α (ERα) and exert control over GnRH/LH secretion in female rodents. It has been proposed that estradiol (E2) activation of ERα in kisspeptin neurons in the arcuate nucleus (ARC) suppresses GnRH/LH secretion (negative feedback), whereas E2 activation of ERα in kisspeptin neurons in the anteroventral periventricular nucleus (AVPV) mediates the release of preovulatory GnRH/LH surges (positive feedback). To test these hypotheses, we generated mice bearing kisspeptin cell-specific deletion of ERα (KERαKO) and treated them with E2 regimens that evoke either negative or positive feedback actions on GnRH/LH secretion. Using negative feedback regimens, as expected, E2 effectively suppressed LH levels in ovariectomized (OVX) wild-type (WT) mice to the levels seen in ovary-intact mice. Surprisingly, however, despite the fact that E2 regulation of Kiss1 mRNA expression was abrogated in both the ARC and AVPV of KERαKO mice, E2 also effectively decreased LH levels in OVX KERαKO mice to the levels seen in ovary-intact mice. Conversely, using a positive feedback regimen, E2 stimulated LH surges in WT mice, but had no effect in KERαKO mice. These experiments clearly demonstrate that ERα in kisspeptin neurons is required for the positive, but not negative feedback actions of E2 on GnRH/LH secretion in adult female mice. It remains to be determined whether the failure of KERαKO mice to exhibit GnRH/LH surges reflects the role of ERα in the development of kisspeptin neurons, in the active signaling processes leading to the release of GnRH/LH surges, or both.

KNDy (Kisspeptin/Neurokinin B/Dynorphin) Neurons Are Activated during Both Pulsatile and Surge Secretion of LH in the Ewe

KNDy (kisspeptin/neurokinin B/dynorphin) neurons of the arcuate nucleus (ARC) appear to mediate the negative feedback actions of estradiol and are thought to be key regulators of pulsatile LH secretion. In the ewe, KNDy neurons may also be involved with the positive feedback actions of estradiol (E(2)) to induce the LH surge, but the role of kisspeptin neurons in the preoptic area (POA) remains unclear. The goal of this study was to identify which population(s) of kisspeptin neurons is (are) activated during the LH surge and in response to the removal of E(2)-negative feedback, using Fos as an index of neuronal activation. Dual-label immunocytochemistry for kisspeptin and Fos was performed on sections containing the ARC and POA from ewes during the luteal phase of the estrous cycle, or before or after the onset of the LH surge (experiment 1), and from ovary-intact, short-term (24 h) and long-term (>30 d) ovariectomized (OVX) ewes in anestrus (experiment 2). The percentage of kisspeptin neurons expressing Fos in both the ARC and POA was significantly higher during the LH surge. In contrast, the percentage of kisspeptin/Fos colocalization was significantly increased in the ARC, but not POA, after both short- and long-term E(2) withdrawal. Thus, POA kisspeptin neurons in the sheep are activated during, and appear to contribute to, E(2)-positive feedback, whereas ARC kisspeptin (KNDy) neurons are activated during both surge and pulsatile modes of secretion and likely play a role in mediating both positive and negative feedback actions of E(2) on GnRH secretion in the ewe.

Differential Roles of GnRH-I and GnRH-II Neurons in the Control of the Primate Reproductive Axis

In vertebrates, gonadotropin-releasing hormone (GnRH) represents the primary neuroendocrine link between the brain and the reproductive axis, and in some species up to three different forms of GnRH have been detected. Until recently, it had been assumed that humans and non-human primates only express one form (GnRH-I), but it is now clear they also express a second form (GnRH-II). GnRH-II, like GnRH-I, is highly effective at stimulating gonadotropin release, both in vitro and in vivo, but the neurons that produce GnRH-II are completely distinct from those producing GnRH-I. Moreover, GnRH-II and GnRH-I producing neurons respond very differently to estradiol; specifically, estradiol stimulates GnRH-II gene expression in the former and inhibit GnRH-I gene expression in the latter. Consequently, the negative feedback action of estradiol may be mediated exclusively by the subpopulation of GnRH neurons that express GnRH-I, while the positive feedback action may be mediated exclusively by the subpopulation that expresses GnRH-II. Taken together, these findings raise the possibility that two completely different GnRH neuronal systems participate in the control of primate reproductive physiology. The primary role of GnRH-I neurons is likely to be focused on the maintenance and modulation of tonic pulsatile LH release, whereas the primary role of GnRH-II neurons is likely to be focused on the generation of the preovulatory LH surge. This functional segregation of the primate neuroendocrine reproductive axis lends itself for novel targeted approaches to fertility control and for treatment of human reproductive disorders.

Rapid direct action of estradiol in GnRH neurons: findings and implications.

Estradiol plays a pivotal role in the control of gonadotropin-releasing hormone (GnRH) neuronal function and female reproduction. While positive and negative feedback actions of estradiol that enhance and suppress release of GnRH and LH are primarily mediated through estrogen receptor alpha located in interneurons, a series of recent studies in our laboratory indicate that rapid excitatory actions of estradiol also directly modify GnRH neuronal activity. We observed this phenomenon in cultured primate GnRH neurons, but similar rapid direct actions of estradiol are also described in cultured GnRH neurons and green fluorescent protein-labeled GnRH neurons of mice. Importantly, rapid direct action of estradiol in GnRH neurons is mediated through membrane or membrane associated receptors, such as GPR30, STX-sensitive receptors, and ERβ. In this review, possible implications of this rapid estradiol action in GnRH neurons are discussed.

Luteinizing hormone secretion as influenced by age and estradiol in the prepubertal gilt

The aim of this study was to determine if there is an age related reduction in the sensitivity of the negative feedback action of 17β-estradiol (estradiol) on luteinizing hormone (LH) secretion in the prepubertal gilt. Ovariectomized gilts at 90 ( n = 12), 150 ( n = 11) or 210 ( n = 12) days of age received estradiol benzoate (EB) osmotic pump implants 6/group and the remaining animals received vehicle control (C) implants except for 150-day C ( n = 5) on Day 0. On Day 10 blood samples were collected every 15 min for 8 h and serum LH and estradiol concentrations were measured. Serum estradiol concentrations averaged 5 ± 1, 5 ± 1 and 7 ± 2 pg/ml for the 90-, 150- and 210-day-old gilts implanted with estradiol, respectively, whereas, serum estradiol concentrations was undetectable in C gilts. Mean serum LH concentrations, basal LH concentrations and serum LH pulse amplitude were less in EB-treated gilts at all ages compared to control animals. In contrast, LH pulse frequency initially was less in EB-treated gilts but subsequently increased ( P < 0.04) with age (from 0.8 ± 0.2 at 90 days to 5.2 ± 0.2/8 h at 210 days), and at 210 days of age the pulse frequency was similar to C gilts. These results demonstrate an age related reduction in the sensitivity to the negative feedback action of estradiol on LH secretion and support the idea that the gilt conforms to the gonadostat hypothesis.

Estradiol attenuates multiple tetrodotoxin-sensitive sodium currents in isolated gonadotropin-releasing hormone neurons

Secretion from gonadotropin-releasing hormone (GnRH) neurons is necessary for the production of gametes and hormones from the gonads. Subsequently, GnRH release is regulated by steroid feedback. However, the mechanisms by which steroids, specifically estradiol, modulate GnRH secretion are poorly understood. We have previously shown that estradiol administered to the female mouse decreases inward currents in fluorescently labeled GnRH neurons. The purpose of this study was to examine the contribution of sodium currents in the negative feedback action of estradiol. Electrophysiology was performed on GnRH neurons dissociated from young, middle-aged, or old female mice. All mice were ovariectomized; half were estradiol replaced. The amplitude of the sodium current underlying the action potential was significantly decreased in GnRH neurons from young estradiol-treated animals. In addition, in vivo estradiol significantly decreased the transient sodium current amplitude, but prolonged the sodium current inactivation time constant. Estradiol decreased the persistent sodium current amplitude, and induced a significant negative shift in peak current potential. In contrast to results obtained from cells from young reproductive animals, estradiol did not significantly attenuate the sodium current underlying the action potential in cells isolated from middle-aged or old mice. Sodium channels can modulate cell threshold, latency of firing, and action potential characteristics. The reduction of sodium current amplitude by estradiol suggests a negative feedback on GnRH neurons, which could lead to a downregulation of cell excitability and hormone release. The attenuation of estradiol regulation in peripostreproductive and postreproductive animals could lead to dysregulated hormone release with advancing age.

Arcuate Kisspeptin Neurons Play a Key Role in the Feedback Control of GnRH Secretion in Sheep.Robert L. Goodman, Ph.D.

In rodents, two populations of kisspeptin neurons, one in the arcuate nucleus (ARC) and the other in the preoptic area (POA) appear to contribute to the feedback control of GnRH. In large mammals including sheep and primates, the majority of kisspeptin neurons are found in the ARC. In sheep, these kisspeptin neurons have several important characteristics. Anatomically, the ARC kisspeptin neurons also contain dynorphin and neurokinin-B, whereas POA kisspeptin neurons do not. Functionally, ARC kisspeptin neurons appear to play important roles in both the anestrous and breeding season. In anestrus, they become more responsive to inhibition by estradiol and thus likely contribute to the annual changes in estradiol negative feedback that drive seasonal reproduction in ewes. During the breeding season, when estradiol suppresses LH pulse amplitude and progesterone inhibits pulse frequency, kisspeptin mRNA expression in the ARC correlates with estradiol inhibition of pulse amplitude. Progesterone has no effect on kisspeptin mRNA expression suggesting that kisspeptin is not critical for progesterone negative feedback. This conclusion is consistent with previous data indicating that progesterone most likely inhibits LH pulse frequency via dynorphin produced in these same ARC neurons. Based on these data, we propose that one subpopulation of ARC neurons mediate the negative feedback actions of estradiol and progesterone, but do so via different neurotransmitters. Thus ARC kisspeptin neurons appear to be key mediators of steroid negative feedback in ewes; we are currently exploring the possibility that they are also involved in the positive feedback action of estradiol.

Sexual Differentiation of the External Genitalia and the Timing of Puberty in the Presence of an Antiandrogen in Sheep

Testicular steroids during midgestation sexually differentiate the steroid feedback mechanisms controlling GnRH secretion in sheep. To date, the actions of the estrogenic metabolites in programming neuroendocrine function have been difficult to study because exogenous estrogens disrupt maternal uterine function. We developed an approach to study the prenatal actions of estrogens by coadministering testosterone (T) and the androgen receptor antagonist flutamide, and tested the hypothesis that prenatal androgens program estradiol inhibitory feedback control of GnRH secretion to defeminize (advance) the timing of the pubertal increase in LH. Pregnant sheep were either untreated or treated with T, dihydrotestosterone (DHT) (a nonaromatizable androgen), or T plus flutamide from d 30-90 of gestation. To study the postnatal response to steroid negative feedback, lambs were gonadectomized and estradiol-replaced, and concentrations of LH were monitored in twice-weekly blood samples. Although T and DHT produced penile and scrotal development in females, the external genitalia of T plus flutamide offspring remained phenotypically female, regardless of genetic sex. Untreated females and females and males treated with T plus flutamide exhibited a pubertal increase in circulating LH at 26.4+/-0.5, 26.0+/-0.7, and 22.4+/-1.6 wk of age, respectively. In females exposed to prenatal androgens, the LH increase was advanced (T: 12.0+/-2.6 wk; DHT: 15.0+/-2.6 wk). These results demonstrate the usefulness of combining T and antiandrogen treatments as an approach to increasing prenatal exposure to estradiol. Importantly, the findings support our hypothesis that prenatal androgens program sensitivity to the negative feedback actions of estradiol and the timing of neuroendocrine puberty.

Role of leptin in modulating the hypothalamic–pituitary axis and luteinizing hormone secretion in the prepuberal gilt

Three experiments (EXP) were conducted to test the hypothesis that leptin modulates LH, GnRH, and neuropeptide Y (NPY) secretion. In EXP I, prepuberal gilts received intracerebroventricular (i.c.v.) leptin injections and blood samples were collected. In EXP II, anterior pituitary cells from prepuberal gilts in primary culture were challenged with 10 −14, 10 −13, 10 −12, 10 −11, 10 −10, 10 −9, 10 −8, 10 −7, or 10 −6 M leptin individually or in combinations with 10 −10, 10 −9, and 10 −8 M GnRH. In EXP III, hypothalamic–preoptic area (HYP-POA) explants were placed in perfusion system and exposed to 0 ( n=5), 10 −12 M ( n=4), 10 −10 M ( n=4), 10 −8 M ( n=4), or 10 −6 M ( n=5) human recombinant leptin (LEP) for 30 min. In EXP I, serum LH concentrations were unaffected by leptin treatment. In EXP II, all doses of leptin increased LH secretion except for 10 −12 and 10 −7 M. Only 10 −7, or 10 −13 M leptin in combination with 10 −8 or 10 −9 M GnRH, respectively, suppressed LH secretion. In EXP III, prior to leptin, media GnRH concentrations were similar across treatments. Media GnRH concentrations increased after 10 −12, 10 −10, and 10 −8 M leptin compared to control. Leptin treatment failed to influence NPY secretion across treatments. These results indicate that components of the neuroendocrine axis that regulate GnRH and LH secretion are functional and leptin sensitive before the onset of puberty. Other neural peptides in addition to NPY may mediate the acute effects of leptin on the GnRH-LH system and lastly, the inability of i.c.v. leptin treatment to increase LH secretion may in part be related to stage of sexual maturation and associated change in negative feedback action of estradiol on LH secretion.

Estradiol increases multiunit electrical activity in the A15 area of ewes exposed to inhibitory photoperiods.

Seasonal anestrus in ewes results from an increase in response to the negative feedback action of estradiol (E(2)). This increase in the inhibitory effects of E(2) is controlled by photoperiod and appears to be mediated, in part, by dopaminergic neurons in the retrochiasmatic area of the hypothalamus (A15 group). This study was designed to test the hypothesis that E(2) increases multiunit electrical activity (MUA) in the A15 during inhibitory long days. MUA was monitored in the retrochiasmatic area of 14 ovariectomized ewes from 4 h before to 24 h after insertion of an E(2)-containing implant subcutaneously. In six of these ewes, MUA activity was also monitored before and after insertion of blank implants. Three of the 14 ewes were excluded from analysis because E(2) failed to inhibit LH. When MUA was recorded within the A15, E(2) produced a gradual increase in MUA that was sustained for 24 h. Blank implants failed to increase MUA in the A15 area, and E(2) did not alter MUA if recording electrodes were outside the A15. These data demonstrate that E(2) increases MUA in the A15 region of ewes and are consistent with the hypothesis that these neurons mediate E(2) negative feedback during long photoperiods.

Decreased lh pulsatility during initiation of gonadotropin superovulation treatment in the cow: Evidence for negative feedback other than estradiol and progesterone

LH pulse secretion is suppressed during superovulation of cattle. The objective of this study was to determine how soon after initiation of superovulation treatments this suppressive effect occurs, and to test the hypothesis that decreased LH pulsatility is not related to changes in circulating estradiol or progesterone. Heifers (n = 7/group) were injected with eCG (FOLLIGON®: a single injection of 2500 IU) or twice daily injections of decreasing doses of FOLLTROPIN®-V (total equivalent of 280 mg of NIH-FSH-P1) or F.S.H.-P® (total equivalent of 28 mg of Armour standard) or saline (time controls), starting on Day 10 (Day 0 = estrus). Blood samples were taken every 10 min for 12 h intervals on the day prior to first injection, at 8 to 20 h and 32 to 44 h after initiation of gonadotropin treatment, and also during prostaglandin (PG)-induced luteolysis. A simple method based on robust statistics and on graphical representations of time series was developed to characterize LH pulses. There was a significant interaction between time and treatment for mean LH, estradiol and progesterone when control and treated groups were analyzed together, and no interaction when only the gonadotropin groups were analyzed together. When compared to pretreatment values, pulse frequency of LH was significantly reduced (P<0.05) in each treatment group, 8 to 20 h and 32 to 44 h following initiation of gonadotropin treatment. Mean LH concentrations were also reduced 32 to 44 h following initiation of treatments (P<0.05). Mean estradiol concentrations increased two to threefold at 8 to 20 h following initiation of superovulation treatments (P<0.05). Progesterone concentrations also increased by 20 or 44 h. There was no significant correlation between estradiol or progesterone and LH pulse frequency, amplitude and mean concentrations at any time in control or superovulated animals. This study demonstrates that superovulation treatment in the cow causes a rapid decrease in pulsatile release of LH and suggests that this effect is not mediated through the negative feedback actions of estradiol and progesterone.

Suppression of Tonic Luteinizing Hormone Secretion and Norepinephrine Release near the GnRH Neurons by Estradiol in Ovariectomized Rats

One of the major neurotransmitters that controls pulsatile luteinizing hormone (LH) secretion is norepinephrine (NE). NE pulses detected in the median eminence of ovariectomized rhesus monkeys are highly correlated with both GnRH and LH pulses. In contrast, previous reports suggest that this is not the case in rats, thus it remains to be determined whether NE stimulates LH release on a pulse-by-pulse basis in that species. Further, a variety of indirect evidence supports the hypothesis that in rats, estradiol exerts its negative feedback action on LH secretion in part by inhibiting noradrenergic neurotransmission that is stimulatory to LH release, but there is no direct evidence to support this hypothesis. Therefore the following study was designed to test the hypothesis that estradiol suppresses NE release in the vicinity of the GnRH neurons after ovariectomy. In addition, we examined whether episodes of NE release are correlated with LH pulses in ovariectomized rats. Blood samples and microdialysates of the diagonal band of Broca/medial preoptic area (DBB/MPOA) were collected every 5 min from 09:00 to 14:00 h from untreated or estradiol-treated (4–5 days), long-term ovariectomized (1–4 months) rats for determination of plasma LH by RIA and NE release by HPLC. The results indicate that in both untreated and estradiol-treated ovariectomized rats, LH pulses are not correlated with episodes of NE. Thus, NE may play a permissive role in the control of pulsatile LH secretion in rats. Further, estradiol treatment leads to a suppression of both plasma LH levels and NE release in the DBB/MPOA, supporting the hypothesis that a decrease in NE neurotransmission that is stimulatory to LH release mediates the negative feedback action of estradiol on tonic LH secretion.

Neural systems mediating the negative feedback actions of estradiol and progesterone in the ewe.

The ewe shows a marked seasonal variation in the effects of ovarian steroids on pulsatile GnRH secretion. In the breeding season, progesterone inhibits GnRH pulse frequency, while estradiol suppresses pulse amplitude. In anestrus, both steroids inhibit pulse frequency. The effects of progesterone in both seasons are mediated by endogenous opioid peptides (EOP) that act in the preoptic area (POA) and medial basal hypothalamus (MBH). However, knife cut studies indicate that actions in the MBH are most important. Moreover, blockade of EOP receptors activates (e.g., induces Fos) GnRH perikarya in the MBH, but not those in the POA. Thus interactions between EOP and GnRH neurons within the MBH may be critical for progesterone negative feedback. The neural systems mediating estradiol suppression of GnRH pulse amplitude in the breeding season are largely unknown, although alpha-adrenergic neurons may be involved. The seasonal variation in inhibition of GnRH pulse frequency by estradiol is postulated to be mediated by a group of dopaminergic (DA) neurons that have three important properties: (1) they inhibit GnRH pulse frequency; (2) their activity is stimulated by estradiol; and (3) they are functional in anestrus, but not the breeding season. Recent work examining the effects of lesions of DA neurons and the ability of estradiol to induce Fos in DA cells strongly suggests that DA neurons in the retrochiasmatic area (A15) and POA (A14) have all three characteristics. We thus propose that these DA neurons are responsible for the seasonal variation in the ability of estradiol to inhibit GnRH pulse frequency.

Alpha adrenergic neurons inhibit luteinizing hormone pulse amplitude in breeding season ewes.

It is now clear that estradiol acts in the brain to inhibit LH pulse amplitude in breeding season ewes, but the neural mechanisms underlying this action of estradiol have yet to be determined. In this study, we performed four experiments to examine the role of alpha-adrenergic neurons in the control of LH pulse amplitude in the ewe. In the first experiment, implantation of the alpha-adrenergic antagonist phenoxybenzamine into the preoptic area (POA) during the follicular phase of the ovine estrous cycle significantly increased LH pulse amplitude. A similar stimulatory effect of POA implants of this antagonist was observed in estradiol-treated ovariectomized (OVX+E) ewes, but phenoxybenzamine implants had no effect in the absence of estradiol. In contrast, systemic administration of phenoxybenzamine decreased LH pulse amplitude in both OVX and OVX+E ewes. Finally, POA implants of the alpha 1-adrenergic antagonist, prazosin, increased LH pulse amplitude in OVX+E ewes, whereas the alpha 2-adrenergic antagonist, yohimbine, had no effect in these animals. These results suggest that a noradrenergic (or adrenergic) system acts in the POA via alpha 1-adrenergic receptors to inhibit LH pulse amplitude in the ewe. They also raise the possibility that this system may be involved in the negative feedback action of estradiol during the breeding season.

Estrogen receptors are expressed in a subset of tyrosine hydroxylase-positive neurons of the anterior preoptic region in the rainbow trout.

A double immunocytochemical procedure, with two different chromogens, was used to compare the respective distribution of estrogen receptor-immunoreactive cells and tyrosine hydroxylase-immunoreactive neurons on the same sections of the preoptic region of adult female rainbow trout (Oncorhynchus mykiss). Estrogen receptor-immunoreactive cells were observed in the anterior preoptic region surrounding the preoptic recess and its large lateral extensions. Tyrosine hydroxylase-immunoreactive cells were consistently detected in the ventral and ventrolateral walls of the preoptic recess, in an area that was named nucleus preopticus pars anteroventralis. Dopamine immunohistochemistry and Dil retrograde transport studies indicated that part of these catecholaminergic neurons are dopaminergic and could project to the pituitary. Double staining studies showed consistently that most estrogen receptor-positive cells located ventral to the large extensions of the preoptic recess are also tyrosine hydroxylase-positive, indicating that this region is a major target for estradiol feedback. The results are discussed in relation to the role of the nucleus preopticus pars anteroventralis in mediating the negative feedback actions of estradiol on the secretion of gonadotrophin (GTH2) secretion. A hypothesis is drawn in order to explain the synchronizing role of estradiol at the time of ovulation in rainbow trout.

Endogenous opioid peptides control the amplitude and shape of gonadotropin-releasing hormone pulses in the ewe.

This study was designed to test the hypothesis that endogenous opioid peptides (EOP) mediate the negative feedback action of estradiol on GnRH pulse size in breeding season ewes. If this hypothesis is correct, one would predict that an EOP antagonist should increase GnRH pulse size in estradiol-treated ovariectomized (OVX+E), but not in OVX, ewes. We, therefore, examined the effects of naloxone on GnRH pulse profiles in the hypophyseal portal blood of OVX and OVX+E ewes (n = 6/group). Samples were collected every 10 min for 6 h before, 6 h during, and 4 h after naloxone infusion. Estradiol treatment decreased GnRH pulse size and increased GnRH pulse frequency. Naloxone treatment had no effect on GnRH pulse frequency, but significantly increased GnRH pulse size. However, this stimulatory action of naloxone on GnRH pulse size was evident in both OVX and OVX+E ewes. These results are thus not consistent with the hypothesis that EOP mediate the negative feedback action of estradiol. Interestingly, naloxone not only increased GnRH pulse amplitude, but also prolonged the duration of GnRH release during a pulse. To obtain a more precise characterization of the effects of naloxone on the dynamics of GnRH release, pulse profiles in six OVX ewes were examined in hypophyseal portal blood sampled every minute for 4 h before and 4 h during naloxone infusion. Naloxone again increased both the amplitude and duration of GnRH pulses. The increase in GnRH pulse duration was caused by a prolongation of both the plateau and declining phases of the GnRH pulse. In addition to these effects on GnRH release during a pulse, naloxone increased the amount of GnRH collected between pulses in both experiments. The stimulatory effects of naloxone on GnRH release in OVX ewes indicate that the role of EOP in the control of GnRH is not limited to mediating the feedback actions of steroids. In particular, the dramatic effects of naloxone on GnRH pulse shape and interpulse GnRH levels raise the possibility that EOP play an important role in synchronizing the activity of the GnRH neurons involved in episodic GnRH secretion.

Reproductive Senescence in Female Japanese Monkeys (Macaca Fuscata): Age- and Season-Related Changes in Hypothalamic-Pituitary-Ovarian Functions And Fecundity Rates

Age- and season-related changes in gonadotropin-gonadal functions were studied in captive female Japanese monkeys (Macaca fuscata fuscata), with special reference to reproductive senescence. In experiment 1, a total of 57 nonlactating females at ages 3-32 yr were used. Blood samples were collected for at least one breeding season (September through March), and changes in plasma concentrations of LH, estradiol, and progesterone were examined. In experiment 2, two groups of females (older females, n = 5, over 21 yr old; mature females, n = 5, aged 8-12 yr) received s.c. injections of 50 micrograms/kg of estradiol benzoate during the mid-breeding season. In experiment 1, ovarian functions as well as fecundity rates were maintained in the animals aged 5-20 yr. A decline in ovarian activity became apparent at 21-25 yr of age, when animals exhibited a slight increase in plasma LH during the breeding season. Onset of menopause occurred at around 27 yr of age, near the end of the life span. Menopausal females exhibited a marked seasonal difference in plasma LH concentrations, with high levels during the breeding season. In experiment 2, estradiol treatment elicited an LH surge in both old and mature females. The marked seasonal difference in plasma LH, as well as the maintenance of LH response to estradiol in menopausal females, suggests a possibility that biannual changes in sensitivity of the hypothalamo-hypophysial axis to the negative feedback action of estradiol are maintained even after menopause.

Is the inhibitory action of estradiol on luteinizing hormone pulse frequency in anestrous ewes mediated by noradrenergic neurons in the preoptic area?

This study tested the hypothesis that estradiol inhibits luteinizing hormone (LH) pulse frequency in anestrous ewes by increasing the activity of an inhibitory noradrenergic (NE) system that acts in the ovine preoptic area (POA). The effects of estradiol on the release of NE and other neurotransmitters in the POA were determined using intracranial microdialysis. Microdialysis probes (5 mm membrane length) were inserted via chronic guide tubes into the POA. Ringer's solution was pumped through the probes at a rate of 2 microliters/min for 8 h, the alpha-adrenergic antagonist phenoxybenzamine (PBZ; 60 micrograms/ml of Ringer's solution) was then administered via the probe from hours 8 to 12, and Ringer's solution alone was given the last 4 h. The outflow from the dialysis probes was collected every 20 min from 2 to 16 h and concentrations of aminergic transmitters and gamma aminobutyric acid determined by high-performance liquid chromatography. Blood samples were collected every 10 min throughout the experiment and the LH pulse patterns determined. Dialysis was done in the same neural area twice in each ewe, without (ovariectomy only) and with estradiol treatment (ovariectomy and a 1.5-cm-long Silastic capsule filled with crystalline estradiol placed subcutaneously for 2 days); the order of dialysis was randomized. As expected, estradiol decreased the LH pulse frequency. This negative feedback action of estradiol was associated with a decrease in mean NE concentrations in dialysate samples and an increase in the intra-animal variability of NE. Estradiol had no effect on any other neurotransmitter measured.(ABSTRACT TRUNCATED AT 250 WORDS)

Mechanisms Controlling the Seasonal Breeding of Japanese Monkeys

This study outlines environmental and internal factors affecting the seasonal breeding of Japanese monkeys (Macaca fuscata). 1) Females housed indoors exhibited the prolonged breeding season as well as the reduced inhibition of ovarian functions during the nonbreeding season in comparison with those housed out-doors, as judged from reproductive endocrine profiles. 2) Artificial manipulation of photoperiod alone had no effect on the manifestation of annual reproductive cyclicity, whereas simultaneous manipulation of photoperiod and ambient temperature was at least to some extent effective in modifying the ovarian functions. 3) Estradiol-treated ovariectomized monkeys exhibited marked seasonal changes in response to the negative feedback action of estradiol on LH secretion. 4) Thyroidectomy resulted in significantly earlier termination of the breeding sea-son. 5) Analysis of hormonal profiles in socially housed male and female monkeys revealed that onset of the breeding season was preceded by the increase in plasma testosterone and inhibin in male monkeys, which sug-gested that social factor is also involved in the manifestation of the breeding seasonality. From these studies, it is concluded that seasonal breeding of Japanese monkeys is governed by biannual changes in the response of the hypothalamo-hypophysial axis to the negative feedback action of estradiol, and that multiple annually cyclic environmental factors appears to influence the seasonal breeding of this animal.