Sandra J. Legan

Neuroendocrine basis of seasonal reproduction

Publisher Summary This chapter discusses the strategy of seasonal breeding, the role of photoperiod in timing the annual reproductive cycle, the hypothalamo-pituitary mechanisms that mediate photoperiodic regulation of estrous cyclicity, and the photoperiodic pathway to luteinizing hormone (LH) pulse generator. To understand how photic input to the LH pulse generator leads to seasonal changes in gonadal activity, the sequence of endocrine events that normally leads to ovulation during the estrous cycle of the ewe must be considered. These preovulatory events occur during a 2–3 day follicular phase and include a precipitous drop in progesterone, a progressive rise in tonic LH secretion, a sustained increase in estradiol secretion, and the LH surge. The pivotal step in this sequence is the sustained increase in tonic LH secretion. A great deal of insight has been gained into the complex interplay between the neural and endocrine response systems that underlie the seasonal reproductive process in the short-day breeding ewe. Specifically, light cues activate retinal photoreceptors and are transmitted via a monosynaptic tract to the suprachiasmatic nuclei of the hypothalamus. After interacting with the circadian system, the photic information is relayed to the pineal gland that transduces the neural message into a hormonal signal in the form of a circadian rhythm of melatonin secretion. The pattern of this melatonin signal, which is interpreted as inductive or suppressive, sets the frequency of the LH pulse generator and determines the capacity of this neural oscillator to respond to the negative feedback action of estradiol. The resulting changes in the episodic pattern of gonadotropin secretion, in turn, dictate whether or not estrous cycles can occur.

Neuroendocrine Regulation of the Luteinizing Hormone-Releasing Hormone Pulse Generator in the Rat

We have analyzed the mechanisms by which several known regulators of the LHRH release process may exert their effects. For each, we have attempted to determine how and where the regulatory input is manifest and, according to our working premise, we have attempted to identify factors which specifically regulate the LHRH pulse generator. Of the five regulatory factors examined, we have identified two inputs whose primary locus of action is on the pulse-generating mechanism--one endocrine (gonadal negative feedback), and one synaptic (alpha 1-adrenergic inputs) (see Fig. 29). Other factors which regulate LHRH and LH release appear to do so in different ways. The endogenous opioid peptides, for example, primarily regulate LHRH pulse amplitude (Karahalios and Levine, 1988), a finding that is consistent with the idea that these peptides exert direct postsynaptic or presynaptic inhibition (Drouva et al., 1981). Gonadal steroids exert positive feedback actions which also result in an increase in the amplitude of LHRH release, and this action may be exerted through a combination of cellular mechanisms which culminate in the production of a unique, punctuated set of synaptic signals. Gonadal hormones and neurohormones such as NPY also exert complementary actions at the level of the pituitary gland, by modifying the responsiveness of the pituitary to the stimulatory actions of LHRH. The LHRH neurosecretory system thus appears to be regulated at many levels, and by a variety of neural and endocrine factors. We have found examples of (1) neural regulation of the pulse generator, (2) hormonal regulation of the pulse generator, (3) hormonal regulation of a neural circuit which produces a unique, punctuated synaptic signal, (4) hormonal regulation of pituitary responsiveness to LHRH, and (5) neuropeptidergic regulation of pituitary responsiveness to LHRH. While an attempt has been made to place some of these regulatory inputs into a physiological context, it is certainly recognized that the physiological significance of these mechanisms remains to be clarified. We also stress that these represent only a small subset of the neural and endocrine factors which regulate the secretion or actions of LHRH. A more comprehensive list would also include CRF, GABA, serotonin, and a variety of other important regulators. Through a combination of design and chance, however, we have been able to identify at least one major example of each type of regulatory mechanism.

The photoneuroendocrine control of seasonal breeding in the ewe

Abstract The major environmental parameter controlling seasonal breeding in ewes is photoperiod. Short days stimulate, and long days inhibit breeding activity. One of the more intriguing enigmas of neuroendocrinology is the endocrine mechanism whereby a mere change in daylength initiates or prevents estrous cycles. Recent experiments have begun to solve this problem by demonstrating that in ewes, photoperiod governs response of the hypothalamo-pituitary axis to the negative feedback action of estradiol. In long days, estradiol is a potent inhibitor of gonadotropin secretion, whereas in short days it is relatively ineffective in this regard. These photoperiod-induced changes in estradiol feedback are proposed to permit or prevent estrous cycles by controlling the occurrence of a crucial step in the sequence of events leading to ovulation, namely a sustained, preovulatory rise in LH. Thus, estrous cycles cease in long days because an increase in estradiol negative feedback prevents the sustained rise in LH. In short days, estrous cycles resume because a decrease in estradiol feedback permits the sustained rise in LH required for ovulation. Even more puzzling, at present, than the mechanism of transduction of photoperiodic information into an endocrine event controlling seasonal breeding is the problem of transmission of photoperiodic information from the environment to the hypothalamo-pituitary axis. In sheep, early investigations of the transmission pathway have elicited provocative results, only some of which are similar to those obtained in other species. Among the questions which remain to be answered are: what is the location of the photoreceptors, and what, if any, are the roles of the suprachiasmatic nuclei and the pineal in photoperiodic control of seasonal breeding? These issues promise to provide a tantalizing challenge for future investigations into the photoneuroendocrine control of seasonal breeding.

Gonadotropin-inhibitory hormone reduces sexual motivation but not lordosis behavior in female Syrian hamsters (Mesocricetus auratus)

Reproductive success is maximized when female sexual motivation and behavior coincide with the time of optimal fertility. Both processes depend upon coordinated hormonal events, beginning with signaling by the gonadotropin-releasing hormone (GnRH) neuronal system. Two neuropeptidergic systems that lie upstream of GnRH, gonadotropin-inhibitory hormone (GnIH; also known as RFamide related peptide-3) and kisspeptin, are potent inhibitory and excitatory modulators of GnRH, respectively, that participate in the timing of the preovulatory luteinizing hormone (LH) surge and ovulation. Whether these neuropeptides serve as neuromodulators to coordinate female sexual behavior with the limited window of fertility has not been thoroughly explored. In the present study, either intact or ovariectomized, hormone-treated female hamsters were implanted for fifteen days with chronic release osmotic pumps filled with GnIH or saline. The effect of GnIH on sexual motivation, vaginal scent marking, and lordosis was examined. Following mating, FOS activation was quantified in brain regions implicated in the regulation of female sexual behavior. Intracerebroventricular administration of GnIH reduced sexual motivation and vaginal scent marking, but not lordosis behavior. GnIH administration altered FOS expression in key neural loci implicated in female reproductive behavior, including the medial preoptic area, medial amygdala and bed nucleus of the stria terminalis, independent of changes in circulating gonadal steroids and kisspeptin cell activation. Together, these data point to GnIH as an important modulator of female proceptive sexual behavior and motivation, independent of downstream alterations in sex steroid production.

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.

Restricting feeding to the active phase in middle-aged mice attenuates adverse metabolic effects of a high-fat diet.

Time-restricted feeding ameliorates the deleterious effects of a high-fat diet on body weight and metabolism in young adult mice. Because obesity is highly prevalent in the middle-aged population, this study tested the hypothesis that time-restricted feeding alleviates the adverse effects of a high-fat diet in male middle-aged (12months) mice. C57BL6/J mice were fed one of three diets for 21-25weeks: 1) high-fat diet (60% total calories from fat) ad-libitum (HFD-AL), 2) HFD, time-restricted feeding (HFD-TRF), and 3) low-fat diet (10% total calories from fat) ad-libitum (LFD-AL) (n=15 each). HFD-TRF mice only had food access for 8h/day during their active period. HFD-TRF mice gained significantly less weight than HFD-AL mice (~20% vs 55% of initial weight, respectively). Caloric intake differed between these groups only during the first 8weeks and accounted for most but not all of their body weight difference during this time. TRF of a HFD lowered glucose tolerance in terms of incremental area under the curve (iAUC) (p<0.02) to that of LFD-AL mice. TRF of a HFD lowered liver weight (p<0.0001), but not retroperitoneal or epididymal fat pad weight, to that of LFD-AL mice. Neither HFD-AL nor HFD-TRF had any effect on performance in the novel object recognition or object location memory tests. Circulating corticosterone levels either before or after restraint stress were not affected by diet. In conclusion, TRF without caloric restriction is an effective strategy in middle-aged mice for alleviating the negative effects of a HFD on body weight, liver weight, and glucose tolerance.

Effects of perinatal cocaine exposure on open field behavior and the response to corticotropin releasing hormone (CRH) in rat offspring

Previous reports indicate that prenatal cocaine exposure alters specific behaviors and hypothalamic-pituitary-adrenal axis (HPA) function in the offspring. In most previous studies, cocaine was given via subcutaneous injections. However intravenous administration more closely mimics human cocaine abuse during pregnancy. Therefore, we investigated the effects of prenatal cocaine exposure via intravenous injection to the mothers on open field behavior and HPA axis function of the offspring. We hypothesized that prenatal cocaine exposure decreases immobility in a novel environment, and enhances the HPA response to stress. Dams received cocaine (COC) or vehicle (control, CON) intravenously from gestation day 8 to postnatal day (PD) 5. Behaviors were recorded in the open field on PD 28 (weanlings). As expected, perinatally cocaine-exposed offspring spent less time immobile and had a longer latency to entering the center zone. No other behavioral activities were different between the groups. On PD 43-50, adolescent male and female offspring received either corticotropin releasing hormone (CRH) or saline intravenously. Plasma adrenocorticotropic hormone (ACTH) and corticosterone (CORT) levels were determined before, and up to 60 min after injection. COC-exposed offspring of both sexes had higher basal CORT levels. Prenatal cocaine enhanced the CORT response to CRH/saline injections up to 60 min in males but not in females. These novel results show that perinatal administration of cocaine in a manner that most closely mimics human cocaine use has long-term effects on the offsprings behavioral response to stress and on HPA axis functions.

Modulation of hypoglycemia-induced increases in plasma epinephrine by estrogen in the female rat

Clinical studies have demonstrated that estrogen replacement therapy suppresses stress‐induced increases in plasma catecholamines. The present study determined whether normal circulating levels of estrogen can modulate hypoglycemia‐induced increases in plasma epinephrine (EPI). In anesthetized female rats, insulin‐induced hypoglycemia (0.25 U/kg) increased plasma EPI concentration to a significantly greater extent in 14‐day ovariectomized (OVEX) rats compared to that in sham‐operated controls. In 17β‐estradiol (E2)‐replaced OVEX rats, the hypoglycemia‐induced rise in plasma EPI was reduced significantly when compared to that in vehicle‐replaced OVEX rats. OVEX and E2 replacement had no effect on tyrosine hydroxylase or phenylethanolamine N‐methyltransferase mRNA levels in the adrenal medulla. In isolated adrenal medullary chromaffin cells, agonist‐induced increases in intracellular Ca2+ were unaffected by 48‐hr exposure to 10 nM E2. In contrast, acute (3‐min) exposure to micromolar concentrations of E2 dose‐dependently and reversibly inhibited agonist‐induced Ca2+ transients. In addition, in OVEX rats, a constant infusion of E2 significantly reduced the insulin‐induced increase in plasma EPI concentration compared to that in vehicle‐infused controls. These data demonstrate that physiologic levels of circulating E2 can modulate hypoglycemia‐induced increases in plasma EPI. This effect seems independent of steroid influence on adrenal medullary secretion or biosynthesis. In contrast, acute exposure to high levels of E2 can also suppress hypoglycemia‐induced increases in plasma epinephrine, due at least in part to inhibition of stimulus‐secretion coupling.

Chronic Elevation of Estradiol in Young Ovariectomized Rats Causes Aging-Like Loss of Steroid-Induced Luteinizing Hormone Surges

Abstract This study was designed to test the hypothesis that the loss of LH surges in response to the stimulatory actions of estradiol and progesterone in middle-aged, persistent-estrous (PE) rats may be caused by chronic elevations in circulating estradiol. Five groups of regularly cycling young rats received an s.c. estradiol implant immediately after ovariectomy (Day 0). For determination of LH surges, blood samples were collected hourly between 1200–1900 h from each of the five groups at one of the following times: 3 days, or 1, 2, 4, or 8 wk later. On the next day, either progesterone (0.5 mg/100 g BW) or corn oil was injected s.c. at 1200 h, and samples were obtained as before. Incidence and amplitude of estradiol-induced LH surges decreased during the first 2 wk of estradiol treatment, after which no surges occurred. Progesterone enhanced the incidence and amplitude of estradiol-induced LH surges thus delaying their disappearance. These results support our hypothesis and demonstrate that the stimulatory actions of estradiol and progesterone on the LH surge sequentially diminish with time after exposure to estradiol in young rats. Thus, young rats chronically treated with estradiol may be a useful model for studying the mechanisms whereby LH surges are abolished in middle age during the hyperestrogenic state of PE.

Loss of Luteinizing Hormone Surges Induced by Chronic Estradiol Is Associated with Decreased Activation of Gonadotropin-Releasing Hormone Neurons

Abstract Chronic exposure of young ovariectomized rats to elevated circulating estradiol causes loss of steroid-induced LH surges. Such LH surges are associated with cFos-induced activation of GnRH neurons; therefore, we hypothesized that chronic estradiol treatment abolishes LH surges by decreasing activation of GnRH neurons. Regularly cycling rats were ovariectomized and immediately received an estradiol implant or remained untreated. Three days or 2 or 4 wk later, the estradiol-treated rats received vehicle or progesterone at 1200 h, and 7 hourly blood samples were collected for RIA of LH. Thereafter, all rats were perfused, and the brains were examined for immunocytochemical localization of cFos and GnRH. The GnRH neurons from untreated ovariectomized rats rarely expressed cFos. As reported, LH surges induced by 3 days of estradiol treatment were associated with a 30% increase in cFos-containing GnRH neurons, and progesterone enhanced both the amplitude of LH surges and the proportion of cFos-immunopositive GnRH neurons. As hypothesized, the abolition of LH surges caused by 2 or more weeks of estradiol was paralleled by a reduction in the percentage of cFos-containing GnRH neurons, and this effect was delayed by progesterone. These results suggest that chronic estradiol abolishes steroid-induced LH surges in part by inactivating GnRH neurons.