Hypothalamic GnRH Content Research Articles

Combined exposure of beta-cypermethrin and emamectin benzoate interferes with the HPO axis through oxidative stress, causing an imbalance of hormone homeostasis in female rats

The impact of pesticides on reproductive health has been increasingly recognized. β-cypermethrin (β-CYP) and emamectin benzoate (EMB) are commonly used with agricultural workers. There are few published studies on the effects of combined poisoning of these two pesticides on the reproductive system. This study investigated the toxic effects and mechanism of β-CYP and EMB on the reproductive system of female rats based on the hypothalamic-pituitary-ovarian (HPO) axis. The hypothalamic GnRH content tended to decrease, and Kiss-1 and GPR-54 mRNA and protein expression tended to increase in exposed rats. FSH content was elevated for the pituitary gland, and Kiss-1 and GPR-54 mRNA and protein expression were enhanced in all experimental groups compared with the control group. E2 content in rat ovaries and ERα mRNA and protein expression were reduced by β-CYP and EMB. Furthermore, there were interactive effects of β-CYP and EMB on FSH and E2 release, pituitary GPR-54 mRNA and protein, and ovarian ERα mRNA expression. To investigate causes of damage, oxidative damage indicators were tested and showed that exposure to β-CYP and EMB decreased GSH-Px and SOD activities in the HPO axis, increased MDA levels in the hypothalamus and ovary together with LDH activities in the HPO axis, with an interaction effect on GSH-Px and SOD activities in the hypothalamus and pituitary gland as well as on MDA in the ovary. The above results support the screening of sensitive molecular biomarkers and evaluation of the adverse effects of pesticide exposure in greenhouse operations on reproductive health.

Chronic Administration of the Metastin/Kisspeptin Analog KISS1-305 or the Investigational Agent TAK-448 Suppresses Hypothalamic Pituitary Gonadal Function and Depletes Plasma Testosterone in Adult Male Rats

Metastin/kisspeptin, a hypothalamic peptide, plays a pivotal role in controlling GnRH neurons. Here we studied the effect of chronic sc administration of two kisspeptin analogs, KISS1-305 and TAK-448, on hypothalamic-pituitary-gonadal function in male rats in comparison with a GnRH analogue leuprolide or bilateral orchiectomy (ORX). The prototype polypeptide, KISS1-305 (1-4 nmol/h), caused substantial elevations of plasma LH and testosterone, followed by abrupt reductions of both hormone levels. Notably, testosterone levels were reduced to castrate levels within 3 d and remained depleted throughout the 4-wk dosing period, an effect that was faster and more pronounced than leuprolide (1 nmol/h) dosing. KISS1-305 also reduced genital organ weight more profoundly than leuprolide. In mechanistic studies, chronic KISS1-305 administration only transiently induced c-Fos expression in GnRH neurons, suggesting that GnRH-neural response was attenuated over time. Hypothalamic GnRH content was reduced to 10-20% of control at 3 wk without any changes in Gnrh mRNA expression. Dosing with the investigational peptide TAK-448 was also studied to extend our understanding of hypothalamic-pituitary functions. Similar to ORX, TAK-448 (0.1 nmol/h) depleted testosterone and decreased GnRH content by 4 wk. However, in contrast to ORX, TAK-448 decreased gonadotropin levels in pituitary and plasma samples, implying the suppression of GnRH pulses. These results suggest that chronic administration of kisspeptin analogs disrupts endogenous kisspeptin signals to suppress intrinsic GnRH pulses, perhaps by attenuating GnRH-neural response and inducing continuous GnRH leakage from the hypothalamus. The potential utility of kisspeptin analogs as novel agents to treat hormone-related diseases, including prostate cancer, is discussed.

Passive transfer of maternal GnRH antibodies does not affect reproductive development in elk (Cervus elaphus nelsoni) calves

Gonadotropin-releasing hormone is intermittently released from the hypothalamus in consistent patterns from before birth to final maturation of the hypothalamic-pituitary-gonadal axis at puberty. Disruption of this signaling via GnRH vaccination during the neonatal period can alter reproduction at maturity. The objective of this study was to investigate the long-term effects of GnRH-antibody exposure on reproductive maturation and function in elk calves passively exposed to high concentrations of GnRH antibodies immediately after birth. Fifteen elk calves (eight males and seven females) born to females treated with GnRH vaccine or sham vaccine during midgestation were divided into two groups based on the concentration of serum GnRH antibodies measured during the neonatal period. Those with robust (>15 pmol 125I-GnRH bound per mL of serum) titers (N = 10; four females and six males) were designated as the exposed group, whereas those with undetectable titers (N = 5; three females and two males) were the unexposed group. Onset of puberty, reproductive development, and endocrine function in antibody-exposed and unexposed male and female elk calves were compared. Neonatal exposure to high concentrations of GnRH antibodies had no effect on body weight (P = 0.968), endocrine profiles (P > 0.05), or gametogenesis in either sex. Likewise, there were no differences between groups in gross or histologic structure of the hypothalamus, pituitary, testes, or ovaries. Pituitary stimulation with a GnRH analog before the second potential reproductive season induced substantial LH secretion in all experimental elk. All females became pregnant during their second reproductive season and all males exhibited similar mature secondary sexual characteristics. There were no differences between exposure groups in hypothalamic GnRH content (P = 0.979), pituitary gonadotropin content (P > 0.05) or gonadal structure. We concluded that suppressing GnRH signaling through immunoneutralization during the neonatal period likely does not alter long-term reproductive function in this species.

Leptin’s effect on puberty in mice is relayed by the ventral premammillary nucleus and does not require signaling in Kiss1 neurons

Studies in humans and rodents indicate that a minimum amount of stored energy is required for normal pubertal development. The adipocyte-derived hormone leptin is a key metabolic signal to the neuroendocrine reproductive axis. Humans and mice lacking leptin or the leptin receptor (LepR) (ob/ob and db/db mice, respectively) are infertile and fail to enter puberty. Leptin administration to leptin-deficient subjects and ob/ob mice induces puberty and restores fertility, but the exact site or sites of leptin action are unclear. Here, we found that genetic deletion of LepR selectively from hypothalamic Kiss1 neurons in mice had no effect on puberty or fertility, indicating that direct leptin signaling in Kiss1 neurons is not required for these processes. However, bilateral lesions of the ventral premammillary nucleus (PMV) of ob/ob mice blunted the ability of exogenous leptin to induce sexual maturation. Moreover, unilateral reexpression of endogenous LepR in PMV neurons was sufficient to induce puberty and improve fertility in female LepR-null mice. This LepR reexpression also normalized the increased hypothalamic GnRH content characteristic of leptin-signaling deficiency. These data suggest that the PMV is a key site for leptin's permissive action at the onset of puberty and support the hypothesis that the multiple actions of leptin to control metabolism and reproduction are anatomically dissociated.

Hypothalamic GnRH and pituitary gonadotroph relationships during rat fetal life.

The ontogeny of hypothalamic GnRH, of pituitary and gonadal receptors and of pituitary LH and FSH was studied in the fetal and neonatal rat. Hypothalamic, hypophyseal and gonadal primordia were dissected from animals ranging in age from postconceptual day 12 to birth. Immunoreactive GnRH was detectable in the hypothalamus from fetal day 12 onwards at a low level until day 17, whereafter hypothalamic GnRH content and concentration increased until birth. GnRH receptors were reliably detectable in the pituitary anlage from fetal day 16 onwards and increased progressively with advancing age whether expressed as content or concentration. Signs of pituitary LH synthesis were evident as early as fetal day 12 but intrapituitary LH levels remained low until fetal day 17 when levels increased progressively until the end of gestation. Pituitary FSH was undetectable until fetal day 19, thereafter rising dramatically until the end of gestation. GnRH binding to testicular and ovarian tissues was undetectable throughout the period of fetal development. The possible relations among the developmental changes in hypothalamic GnRH, pituitary GnRH receptors and gonadotrophins are discussed.

Effect of Bacterial Lipopolysaccharide on the Reproductive Axis of Prepubertal and Peripubertal Female Rats

The immune, endocrine and nervous systems are closely interrelated, which allows the organism to respond to different types of stress such as infection. Chronic infectious and inflammatory conditions are often accompanied by an impaired reproductive function. Leptin, a hormone produced by adipose tissue, exerts a regulatory function on the reproductive axis. It has homology with other proinflammatory cytokines and could be modified by lipopolysaccharide (LPS). Therefore, these studies were designed to investigate the effect of LPS administration on the neuroendocrine mechanisms involved in the regulation of the reproductive axis during sexual maturation. Fifteen- and 30-day-old female rats were injected with a single dose of LPS 250 µg/kg (i.p.) and then nitric oxide synthase (NOS) activity, hypothalamic excitatory/inhibitory amino acids and Gn-RH content, serum LH and leptin concentration were studied. In 15-day-old female rats LPS treatment did not modify hypothalamic inducible (iNOS) and constitutive (cNOS) NOS activity, Gn-RH, glutamate (GLU) and GABA content. Also serum LH and leptin levels were not modified. In 30-day-old rats LPS increased iNOS and cNOS activity (p < 0.001) and hypothalamic Gn-RH content (p < 0.001). At this age hypothalamic GABA content was significantly decreased (p < 0.001) without changes in GLU content, and serum LH (p < 0.001) and leptin (p < 0.0001) decreased significantly. In summary, current studies have demonstrated that LPS administration to 15- and 30-day-old female rats results in a different response of the hypothalamus-pituitary-gonadal axis and of the adipose tissue, demonstrating an ontogenic response of the immune-neuroendocrine system to LPS administration.

The bi-modal effects of estradiol on gonadotropin synthesis and secretion in female mice are dependent on estrogen receptor-α

Depending on the estrous/menstrual cycle stage in females, ovarian-derived estradiol (E(2)) exerts either a negative or a positive effect on the hypothalamic-pituitary axis to regulate the synthesis and secretion of pituitary gonadotropins, LH, and FSH. To study the role of estrogen receptor-alpha (ERalpha) mediating these effects, we assessed the relevant parameters in adult wild-type (WT) and ERalpha-null (alphaERKO) female mice in vivo and in primary pituitary cell cultures. The alphaERKO mice exhibited significantly higher plasma and pituitary LH levels relative to WT females despite possessing markedly high levels of circulating E(2). In contrast, hypothalamic GnRH content and circulating FSH levels were comparable between genotypes. Ovariectomy led to increased plasma LH in WT females but no further increase in alphaERKO females, while plasma FSH levels increased in both genotypes. E(2) treatment suppressed the high plasma LH and pituitary Lhb mRNA expression in ovariectomized WT females but had no effect in alphaERKO. In contrast, E(2) treatments only partially suppressed plasma FSH in ovariectomized WT females, but this too was lacking in alphaERKO females. Therefore, negative feedback on FSH is partially E(2)/ERalpha mediated but more dependent on ovarian-derived inhibin, which was increased threefold above normal in alphaERKO females. Together, these data indicate that E(2)-mediated negative feedback is dependent on functional ERalpha and acts to primarily regulate LH synthesis and secretion. Studies in primary cultures of pituitary cells from WT females revealed that E(2) did not suppress basal or GnRH-induced LH secretion but instead enhanced the latter response, indicating that the positive influence of E(2) on gonadotropin secretion may occur at the level of the pituitary. Once again this effect was lacking in alphaERKO gonadotropes in culture. These data indicate that the aspects of negative and positive effects of E(2) on gonadotropin secretion are ERalpha dependent and occur at the level of the hypothalamus and pituitary respectively.

Overexpression of glutamic acid decarboxylase-67 (GAD-67) in gonadotropin-releasing hormone neurons disrupts migratory fate and female reproductive function in mice.

gamma-Aminobutyric acid (GABA) inhibits the embryonic migration of GnRH neurons and regulates hypothalamic GnRH release. A subset of GnRH neurons expresses GABA along their migratory route in the nasal compartment before entering the brain, suggesting that GABA produced by GnRH neurons may help regulate the migratory process. To examine this hypothesis and the possibility that persistence of GABA production by GnRH neurons may affect subsequent reproductive function, we generated transgenic mice in which the expression of glutamic acid decarboxylase-67 (GAD-67), a key enzyme in GABA synthesis, is targeted to GnRH neurons under the control of the GnRH gene promoter. On embryonic d 15, when GnRH neurons are still migrating, the transgenic animals had more GnRH neurons in aberrant locations in the cerebral cortex and fewer neurons reaching the hypothalamic-preoptic region, whereas migration into the brain was not affected. Hypothalamic GnRH content in mutant mice was low during the first week of postnatal life, increasing to normal values during infantile development (second week after birth) in the presence of increased pulsatile GnRH release. Consistent with these changes, serum LH and FSH levels were also elevated. Gonadotropin release returned to normal values by the time steroid negative feedback became established (fourth week of life). Ovariectomy at this time demonstrated an enhanced gonadotropin response in transgenic animals. Although the onset of puberty, as assessed by the age at vaginal opening and first ovulation, was not affected in the mutant mice, estrous cyclicity and adult reproductive capacity were disrupted. Mutant mice had reduced litter sizes, increased time intervals between deliveries of litters, and a shorter reproductive life span. Thus, GABA produced within GnRH neurons does not delay GnRH neuronal migration, but instead serves as a developmental cue that increases the positional diversity of these neurons within the basal forebrain. In addition, the results suggest that the timely termination of GABA production within the GnRH neuronal network is a prerequisite for normal reproductive function. The possibility arises that similar abnormalities in GABA homeostasis may contribute to syndromes of hypothalamic amenorrhea/oligomenorrhea in humans.

Deletion of Exon 6 of the Neuronal Nitric Oxide Synthase Gene in Mice Results in Hypogonadism and Infertility

Nitric oxide (NO) has been recognized as a modulator in reproductive functions, but it is not clear whether NO is required for fertility. The first line of mice deficient in neuronal NO synthase (referred to herein as KN1 mice) reproduce normally. However, residual neuronal NO synthase (nNOS) activity is detected in KN1 mice due to the expression of β- and γ-nNOS splice variants. We generated a new line of nNOS knockout mice (KN2) lacking exon 6, which codes for the heme-binding domain of nNOS. KN2 mice are viable, but mated homozygotes do not produce litters, indicating that either one or both sexes are infertile. Male KN2 mice show decreased gonad weights, but sperm counts are normal. KN2 males do not display mating behavior, and consequently do not leave vaginal plugs when housed with wild-type (WT) females. KN2 females show decreased ovary weight, and histology reveals decreased corpus luteum counts. RIAs show that KN2 males have decreased plasma FSH, whereas KN2 females have increased levels of plasma LH and increased hypothalamic GnRH content. Experimental ovarian transplantation suggests that central, rather than ovarian, processes are influenced by nNOS, as KN2 ovaries ovulate at near-normal rates under WT hormonal control, whereas WT ovaries transplanted into KN2 mice have decreased ovulation rates. We observed pyloric stenosis in KN2 mice, but plasma leptin levels are normal, and no ketones are found, indicating that hypogonadism is not a result of malnutrition. We conclude that nNOS is required for normal central hormonal regulation of reproductive function.

Deletion of exon 6 of the neuronal nitric oxide synthase gene in mice results in hypogonadism and infertility.

Nitric oxide (NO) has been recognized as a modulator in reproductive functions, but it is not clear whether NO is required for fertility. The first line of mice deficient in neuronal NO synthase (referred to herein as KN1 mice) reproduce normally. However, residual neuronal NO synthase (nNOS) activity is detected in KN1 mice due to the expression of beta- and gamma-nNOS splice variants. We generated a new line of nNOS knockout mice (KN2) lacking exon 6, which codes for the heme-binding domain of nNOS. KN2 mice are viable, but mated homozygotes do not produce litters, indicating that either one or both sexes are infertile. Male KN2 mice show decreased gonad weights, but sperm counts are normal. KN2 males do not display mating behavior, and consequently do not leave vaginal plugs when housed with wild-type (WT) females. KN2 females show decreased ovary weight, and histology reveals decreased corpus luteum counts. RIAs show that KN2 males have decreased plasma FSH, whereas KN2 females have increased levels of plasma LH and increased hypothalamic GnRH content. Experimental ovarian transplantation suggests that central, rather than ovarian, processes are influenced by nNOS, as KN2 ovaries ovulate at near-normal rates under WT hormonal control, whereas WT ovaries transplanted into KN2 mice have decreased ovulation rates. We observed pyloric stenosis in KN2 mice, but plasma leptin levels are normal, and no ketones are found, indicating that hypogonadism is not a result of malnutrition. We conclude that nNOS is required for normal central hormonal regulation of reproductive function.

Effects of castration and chronic steroid treatments on hypothalamic gonadotropin-releasing hormone content and pituitary gonadotropins in male wild-type and estrogen receptor-alpha knockout mice.

Testicular androgens are integral components of the hormonal feedback loops that regulate circulating levels of LHbeta and FSH. The sites of feedback include hypothalamic areas regulating GnRH neurons and pituitary gonadotropes. To better define the roles of androgen receptor (AR), estrogen receptor-alpha (ERalpha), and estrogen receptor-beta (ERbeta) in mediating feedback effects of sex steroids on reproductive neuroendocrine function, we have determined the effects of castration and steroid replacement therapy on hypothalamic GnRH content, pituitary LHbeta and FSHbeta messenger RNA (mRNA) levels, and serum gonadotropins in male wild-type (WT) and estrogen receptor-alpha knockout (ERKO) mice. Hypothalami from intact WT and ERKO males contained similar amounts of GnRH, whereas castration significantly reduced GnRH contents in both genotypes. Replacement therapy with estradiol (E2), testosterone (T), or dihydrotestosterone (DHT) restored hypothalamic GnRH content in castrated (CAST) WT mice; only the androgens were effective in CAST ERKOs. Analyses of pituitary function revealed that LHbeta mRNA and serum LHbeta levels in intact ERKOs were 2-fold higher than those in intact WT males. Castration increased levels of LHbeta mRNA (1.5- to 2-fold) and serum LHbeta (4- to 5-fold) in both genotypes. Both E2 and T treatments significantly suppressed LHbeta mRNA and serum LH levels in CAST WT males. However, E2 was completely ineffective, and T was only partially effective in suppressing these two indexes in the CAST ERKO males. DHT treatments stimulated a 50% increase in LHbeta mRNA and serum LH levels in WT males, whereas serum LH was significantly suppressed in DHT-treated ERKO males. Although the pituitaries from intact ERKO males contained similar amounts of FSHbeta mRNA, serum FSH levels were 20% higher than those in the intact WT males. Castration increased FSHbeta mRNA levels only in WT males, but significantly increased serum FSH levels in both genotypes. Both E2 and T treatments significantly suppressed serum FSH in CAST WT males, whereas only E2 suppressed FSHbeta mRNA. DHT treatments of CAST WT mice stimulated a small increase in serum FSH, but failed to alter FSHbeta mRNA levels. None of the steroid treatments exerted any significant effect on FSHbeta mRNA or serum FSH levels in CAST ERKOs. These data suggest that hypothalamic GnRH contents can be maintained solely through AR signaling pathways. However, normal regulation of gonadotrope function requires aromatization of T and activation of ERalpha signaling pathways in the gonadotrope. In addition, serum FSH levels in male ERKOs appear to be regulated largely by nonsteroidal testicular factors such as inhibin. Finally, these data suggest that hypothalamic ERbeta may not be involved in mediating the negative feedback effects of T on serum LH and FSH in male mice.

Ovarian Follicular Cysts in Dairy Cows

Ovarian follicular cysts are anovulatory follicular structures that occur in 10 to 13% of dairy cows. This review focuses upon the dynamics of cyst growth, development, and persistence as well as on associated endocrine and cellular mechanisms. During the estrous cycle of cows, two to four waves of follicular growth occur. From a cohort of recruited follicles, one is selected for continued growth and dominance while the others undergo atresia and regress. In contrast, cysts have long been thought to be static structures that persist for extended periods. Although cysts can persist for extended periods, most regress over time and are replaced during subsequent follicular waves. The next dominant follicle either ovulates or develo6ps into a new cyst. The recruitment of a cohort of follicles from which a cyst develops and the growth rate of cysts to ovulatory size are similar to ovulatory follicular waves, but the cyst continues to grow for a longer pierod. The interval between waves of follicular growth is longer for cows with cysts than for cows with normal estrous cycles. Each wave is preceded by a transient increase in circulating FSH. Near the time of cyst development and persistence, the concentration of FSH is not different from that during normal estrous cycles. Serum concentrations of LH and estradiol-17β are higher in cows that develop cysts than in cows that do not. Conversely, hypothalamic content of GnRH is lower in cows with cysts. Thus, cysts are dynamic structures, and their development and lifespan are likely associated with altered hypothalamic-hypophysial-ovarian function.

Photoperiod and steroid-dependent adjustments in hypothalamic gonadotropic hormone-releasing hormone, dopamine, and norepinephrine content in male deer mice.

The purpose of this study was twofold: 1) to examine photoperiod-dependent and steroid-dependent adjustments in hypothalamic GnRH and catecholamine content in male deer mice, Peromyscus maniculatus, and 2) to evaluate whether the commonly used measure of GnRH accumulation serves as an accurate indicator of altered reproductive function in short-day-housed rodents. Deer mice provide a unique animal model in which to evaluate the effect of short day length on the neural substrates directing reproductive function, since individual deer mice respond to short photoperiod with a range of neuroendocrine, endocrine, metabolic, and thermoregulatory adjustments. These adjustments, in part, result in two distinct reproductive phenotypes; reproductively competent "nonresponsive" mice, and gonad-regressed "responsive" mice. Reproductively mature males were maintained on either long (16L:8D) or short (8L: 16D) photoperiod for 8 wk. Thereafter, mice were phenotyped and either remained intact, castrated, or castrated and provided with an s.c. testosterone-filled silicone elastomer capsule releasing long-day or twice long-day levels of testosterone; mice were then housed for an additional 4 wk. At autopsy, plasma levels of LH and testosterone, hypothalamic GnRH content, and hypothalamic dopamine and norepinephrine concentrations were determined. Short photoperiod caused a similar increase in GnRH content in both short-day phenotypes, and this increase was primarily localized to the medial basal hypothalamus (MBH). Castration decreased GnRH content within the MBH of mice housed on both a long and short photoperiod. However, castration also reduced GnRH content within the medial preoptic area (MPOA) only among the short-day phenotypes. Steroid replacement at levels similar to those of intact long-day males restored GnRH content in all brain regions to levels of intact mice. On the other hand, steroid replacement at twice long-day levels restored GnRH content among long-day mice but reduced GnRH content among both short-day reproductive response groups, specifically within the MBH and MPOA. There were no dramatic effects of photoperiod or steroid treatment on catecholamine concentrations. But our results do suggest that the lateral preoptic area (LPOA) and MBH may be important sites of steroid feedback among mice exposed to short photoperiods and that the LPOA and diagonal band of Broca-septal regions may specifically be associated with catecholamine-directed photoperiod-induced adjustments of reproductive responsive mice. Taken together, these data show that GnRH accumulation is not the proximate signal directing seasonal changes in pituitary-gonadal function and cast doubt on the use of this factor as an accurate indicator of altered reproductive function.

The Development of the Hypothalamic–Pituitary–Gonadal Axis in Juvenile Great Tits

The development of the hypolamic–pituitary–gonadal axis in free-living nestling great tits (Parus major) was studied from the day of hatching until about 40 days of age. The hypothalamic content of GnRH was low during the entire nestling period, although a significant increase was noted between 6 and 9 days of age. Thereafter there was no further increase. The pituitary content of LH was low in 1- to 6-day-old nestlings, but increased significantly at 9 days of age; this increase was only temporary, and levels decreased in 12-day-old nestlings. Injections of chicken GnRH increased plasma levels of LH in newly hatched great tits. The high plasma levels of LH in newly hatched great tits of both sexes decreased to basal by 9 days of age. Circulating levels of testosterone and estradiol showed more or less the same pattern as LH. Both steroids showed very high levels in newly hatched birds of both sexes. Testosterone levels were down to basal in 3-day-old nestlings, and estradiol levels were basal in 2-day-old nestlings. The only sex difference was observed in 1- and 2-day-old nestlings. During this period males had significantly higher plasma levels of testosterone than females. Gonads showed a more or less linear growth during the entire nestling period, and did not increase further after fledging. Histological and ultrastructural examinations of the testis showed that mature Leydig cells appeared for the first time in 6-day-old birds and become common in 9-day-old nestlings. The development of the seminiferous tubules, the interstitial tissue, and the tunica albuginea reached a “plateau” when the nestlings had reached an age of 9 days. At no age had the germinal epithelium developed beyond the stage of spermatogonia. GnRH treatment did not change male testosterone or female estradiol levels until the nestlings were 9 days of age. The response decreased in older nestlings. Castration of 9-day-old male nestlings showed that neither was there a negative feedback of testosterone on LH in older nestlings.

Thyroxine treatment prevents the development of photosensitivity in european starlings (Sturnus vulgaris)

In starlings, the breeding season is terminated by a state of photorefractoriness. Birds remain completely reproductively inactive as long as long days are maintained, and only exposure to short days restores photosensitivity. Two experiments investigated the role of different doses of thyroxine in the development of photosensitivity in castrated starlings. First, photorefractory castrated male starlings were moved from long (18L:6D) to short (8L:16D) days, and received in the drinking water either 1 or 10 mg · 1-1 thyroxine for the first 7 weeks of a 14-week observation period. Control birds regained photosensitivity after 5 weeks of short days, as signaled by a spontaneous increase in plasma LH, whereas the return to photosensitivity was delayed until weeks 7 and 9 in the 1- and 10-mg · 1-1 thyroxine-treated birds, respectively. In the second experiment, the effect of different doses of thyroxine was explored at the level of the hypothalamic Gn-RH neurosecretory neurones. The acquisition of photosensitivity in control birds transferred from long to short days was characterized by a marked increase in hypothalamic Gn-RH content (while long-day controls maintained low Gn-RH content). Doses of 10 and 20 mg · 1-1 of thyroxine completely prevented the return to photosensitivity, as seen through changes in either plasma LH concentrations or hypothalamic Gn-RH content, while a dose of 1 mg · 1-1 allowed a partial recovery of photosensitivity, as hypothalamic Gn-RH content increased to an intermediate level and the spontaneous rise in plasma LH occurred slowly but steadily.

Relationships among LH, FSH and prolactin secretion, storage and response to secretagogue and hypothalamic GNRH content in ovariectomized pony mares administered testosterone, dihydrotestosterone, estradiol, progesterone, dexamethasone or follicular fluid

Thirty-five ovariectomized pony mares were used to study the relationships among luteinizing hormone (LH), follicle stimulating hormone (FSH) and prolactin (PRL) concentrations in blood (secretion), in pituitary (storage) and in blood after secretagogue administration, as well as the content of gonadotropin releasing hormone (GnRH) in hypothalamic areas, under various conditions of steroidal and nonsteroidal treatment. Five mares each were treated daily for 21 d with vegetable shortening (controls), testosterone (T; 150 μg/kg of body weight, BW), dihydrotestosterone (DHT; 150 μg/kg BW), estradiol (E 2; 35 μg/kg BW), progesterone (P 4; 500 μg/kg BW), dexamethasone (DEX; 125 μg/kg BW) or charcoal-stripped equine follicular fluid (FF; 10 ml). Secretagogue injections (GnRH and thyrotropin releasing hormone, TRH, at 1 and 4 μg/kg of BW, respectively) were given one d prior to treatment and again after 15 d of treatment. Relative to controls, treatment with T, DHT and DEX reduced (P <. 05) LH secretion, storage and response to exogenous GnRH, whereas treatment with E 2 increased (P < .05) these same characteristics. Treatment with P 4 reduced (P < .05) only LH secretion. Treatment with T, DHT, E 2 and DEX reduced (P < .05) FSH secretion, whereas treatment with P 4 increased (P < .05) it and FF had no effect (P > .1). All treatments increased (P < .05) FSH storage, whereas only treatment with T and DHT increased (P < .05) the FSH response to exogenous GnRH. Other than a brief increase (P < .05) in PRL secretion in mares treated with E 2, secretion of PRL did not differ (P > .1) among groups. Only treatment with E 2 increased (P < .01) PRL storage, yet treatment with T or DHT (but not E 2) increased (P < .05) the PRL response to exogenous TRH. Content of GnRH in the body and pre-optic area of the hypothalamus was not affected (P > .1) by treatment, whereas treatment with T, E 2 and DEX increased (P < .1) GnRH content in the median eminence. For LH, secretion, storage and response to exogenous GnRH were all highly correlated (r ≥ .77; P < .01). For FSH, only storage and response to exogenous GnRH were related (r = .62; P < .01). PRL characteristics were not significantly related to one another. Moreover, the amount of GnRH in the median eminence was not related (P > .1) to any LH or FSH characteristic.

Secretory patterns of LH and FSH during development and hypothalamic and hypophysial characteristics following development of steroid-induced ovarian follicular cysts in dairy cattle

Two experiments were conducted to (1) investigate developmental endocrinology of ovarian follicular cysts (cysts) in cattle and (2) evaluate effects of cysts on hypothalamic and hypophysial characteristics. Cysts were induced with oestradiol-17 beta (15 mg) and progesterone (37.5 mg) dissolved in alcohol and injected s.c. twice daily for 7 days. Cysts were defined as the presence of follicular structures (which may or may not have been the same structure) of 2.0 cm in diameter or greater that were present for 10 days without ovulation and corpus luteum development. In Exp. 1,22 non-lactating, non-pregnant Holstein cows were allocated to 3 groups. Beginning on Day 5 (oestrus = Day 0) of the oestrous cycle, 7 cows (Controls) were treated with twice daily s.c. injections of ethanol (2 ml/injection) for 7 days. Luteolysis was then induced with PGF-2 alpha and blood samples were collected daily every 15 min for 6 h from the morning after the PGF-2 alpha injection (Day 13) until oestrus. Steroids to induce cysts were injected as previously described into the remaining cows (N = 15). Three blood samples were collected at 15-min intervals every 12 h throughout the experimental period. Additional blood samples were collected every 15 min for 6 h on a twice weekly basis. After steroid injections, follicular and luteal structures on ovaries were not detected via rectal palpation for a period of 36 +/- 4 days (static phase). Then follicles developed which ovulated within 3-7 days (non-cystic; N = 7) or increased in size with follicular structures present for 10 days (cystic; N = 8). Mean (+/- s.e.m.) concentrations of LH, FSH, oestradiol-17 beta and progesterone in serum remained low and were not different during the static phase between cows that subsequently developed cysts or ovulated. During the follicular phase, mean serum concentration of LH (ng/ml) was higher (P less than 0.1) in cows with cysts (2.9 +/- 0.2) than in cows without cysts (1.1 +/- 0.1) or control cows (1.4 +/- 0.2). In addition, LH pulse frequency (pulses/6 h) and amplitude (ng/ml) were higher (P less than 0.1) in cows with cysts (3.6 +/- 0.3 and 2.2 +/- 0.3, respectively) than in non-cystic (2.3 +/- 0.2 and 1.0 +/- 0.2, respectively) and control (1.8 +/- 0.1 and 1.1 +/- 0.2, respectively) groups during the follicular phase. There were no differences in the FSH, oestradiol-17 beta or progesterone characteristics in cows of any of the 3 groups during the follicular phase.(ABSTRACT TRUNCATED AT 400 WORDS)

Effect of supplemental lighting on puberty, follicular development, pituitary and serum concentrations of LH and hypothalamic content of GnRH in gilts

ABSTRACTOne hundred and thirty crossbred gilts (Yorkshire × Duroc × Hampshire) were exposed to either natural (40 lux) or supplemental lighting (700 lux) beginning at 8 weeks of age. Gilts received supplemental lighting from 40-W flourescent bulbs from 05·15 to 09·00 h and 16·30 to 20·15 h from 9 September to 22 April. At 4 months of age, all gilts were exposed daily for 15 min to mature boars five times weekly. Twelve gilts from each group were slaughtered at 4, 5, 6 and 7 months of age. All gilts were bled from indwelling cannulas at 20-min intervals for 4 h the day preceding slaughter. In addition, blood samples were drawn weekly from 17 gilts in each group from 7 to 9 months of age and analysed for progesterone to determine age of puberty. Twelve of 17 gilts that received supplemental lighting reached puberty at 228 (s.e. 11) days of age whereas 11 of 16 gilts that received natural lighting reached puberty at 252 (s.e. 7) days of age (P = 0·07). No significant differences occurred in anterior pituitary gland weight or pituitary concentration of LH, mean concentrations of serum LH or number of LH secretory spikes between natural and supplemental-lighted gilts at each month (all P &gt; 005). Concentration of oestradiol-17β and progesterone in follicular fluid was similar between treatment groups within months and follicular size (P &gt; 0·05). Content of GnRH in the stalk-median eminence, preoptic area and hypothalamus proper were also similar between treatment groups (P &gt; 0·05). In summary, supplemental lighting reduced the age of puberty in gilts without a detectable change in pituitary, hypothalamic or ovarian parameters.

Metabolic Interfaces between Growth and Reproduction, III. Central Mechanisms Controlling Pulsatile Luteinizing Hormone Secretion in the Nutritionally Growth-Limited Female Lamb*

Growth retardation induced by dietary restriction in the lamb results in a low frequency of episodic LH secretion and, thus, delayed puberty. Such lambs respond normally to physiological doses of GnRH, indicating that the pituitary gland can function adequately during diet-induced hypogonadotropism. The current studies investigated central mechanisms underlying diet-induced hypogonadotropism. The first aim was to determine whether the hypothalamic GnRH secretory system is capable of normal function. The initial approach was to compare hypothalamic GnRH content between lambs on a restricted diet with low LH pulse frequency (less than 1 pulse/4 h; n = 5) and lambs on an ad libitum diet with high LH pulse frequency (4.5 +/- 0.4 pulses/4 h; n = 5). RIA of extracts of preoptic area and mediobasal hypothalamus/median eminence tissue blocks revealed no differences in GnRH content between lambs on a restricted diet and those on an ad libitum diet. The second approach was to determine if LH secretion could be induced by chemical stimulation of neuronal function with N-methyl-D,L-aspartate (NMA), an excitatory amino acid agonist. Initially, a single iv bolus of NMA was given to hypogonadotropic lambs on a restricted diet. There was a dose-dependent immediate rise in serum LH concentrations. All lambs responded to the highest dose (5.0 mg/kg BW; n = 6), and four of five lambs responded to the intermediate dose (1.0 mg/kg). No lambs responded to the lowest dose (0.2 ng/kg), despite a normal response to GnRH (2.5 ng/kg BW, iv). In a second experiment, hypogonadotropic lambs on a restricted diet were treated with repeated injections of NMA (5 mg/kg BW, iv) at either hourly intervals (n = 6) or every 3 h (n = 6). Each NMA injection induced a LH pulse in both treatment regimens over the entire 7-h experimental period. Thus, the nutritionally growth-limited lamb is capable of sustained production of LH pulses, which, we presume, reflect GnRH secretion. The second aim was to test the hypothesis that endogenous opioid mechanisms inhibit LH secretion during nutritionally induced hypogonadotropism, because opioid pathways are a poor inhibitory regulator of LH secretion in the normally developing sheep, even in the absence of ovarian steroids. We were unable to detect any effects of the opiate antagonist naloxone on LH secretion in the nutritionally growth-limited lamb. We conclude that central mechanisms controlling the release, rather than synthesis, of GnRH are limiting LH secretion when sexual maturation is delayed by growth retardation. Moreover, opioid inhibition is not the primary reason for hypogonadotropism during dietary restriction.

Luteinizing hormone in nutrient-restricted ovariectomized ewes2

This study was designed to evaluate profiles of serum concentrations of LH, pituitary content of LH and GnRH receptors, and hypothalamic content of GnRH in undernourished, ovariectomized ewes. In earlier studies, pulsatile secretion of LH diminished as duration of undernutrition progressed in prepubertal or adult ovariectomized ewes. Ewes having similar body condition scores (CS) of 5 to 9 (1 = extremely thin, 5 = moderate, 9 = obese) were fed maintenance or low-energy diets (100% and 60% of NRC requirements, respectively). Blood samples for analysis of LH were collected at 15-min intervals for 4 h at initiation of the project and immediately prior to slaughter. Serum concentrations of LH did not differ (P greater than .05) among groups at the initial sampling period. At slaughter, ewes with CS less than or equal to 2 (n = 7) had lost 26.8 kg (42% of initial weight). Ewes with CS greater than or equal to 3 (n = 12) had lost an average of 13.7 kg (18% of initial weight). Concentrations of LH in ewes with final CS greater than or equal to 3 was similar (P greater than .05) to that observed during the initial sampling period. However, release of LH was reduced (P less than .01) in ewes with CS less than or equal to 2 compared with ewes with CS greater than or equal to 3 (2.6 vs 9.5 and 3.2 vs 10.5 ng/ml for basal and mean concentrations, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)