This overview attempts to synthesise current understandings of the neuroendocrine basis of parenting. The parent–infant bond is central to the human condition, contributes to risks for mood and anxiety disorders, and provides the potential for resiliency and protection against the development of psychopathology. Animal models of parenting provide compelling evidence that biological mechanisms may be studied in humans. This has led to brain imaging and endocrine system studies of human parents using baby stimuli and concerted psychological and behavioural measures. Certain brain circuits and related hormonal systems, including subcortical regions for motivation (striatum, amygdala, hypothalamus and hippocampus) and cortical regions for social cognition (anterior cingulate, insula, medial frontal and orbitofrontal cortices), appear to be involved. These brain circuits work with a range of endocrine systems to manage stress and motivate appropriate parental caring behaviour with a flexibility appropriate to the environment. Work in this field promises to link evolving models of parental brain performance with resilience, risk and treatment toward mother–infant mental health.
parent–child relationships; brain imaging; functional magnetic resonance imaging; attachment; parenting; caregiving; cortisol; dopamine; opioids
Obesity and type 2 diabetes mellitus (T2DM) are associated with dysfunctional insulin signalling and impaired central glucose sensing. Glucose sensing neurones reside in key areas of the brain involved in glucose and energy homeostasis (e.g. ventromedial hypothalamus; VMH). We have recently shown that insulin attenuates the ability of glucose-excited (GE) neurones to sense decreased glucose. We hypothesise that this effect of insulin on VMH GE neurones is impaired during T2DM when insulin signalling is dysfunctional. To test our hypotheses, we used whole cell patch clamp recording techniques to evaluate the effects of insulin on VMH GE neurones in brain slices from wild-type and diabetic (db/db) mice. The effects of decreasing glucose from 2.5 to 0.1 mm on VMH GE neurones were similar in wild-type and db/db mice. However, decreasing glucose from 2.5 to 0.5 mm decreased the action potential frequency, membrane potential and input resistance of VMH GE neurones to a significantly greater extent in db/db versus wild-type mice. Furthermore, insulin (5 nm) blunted the effects of decreased glucose in wild-type, but not db/db mice. These differences in both glucose and insulin sensitivity between wild-type and db/db mice were completely ameliorated by the insulin sensitiser, Compound 2 (300 nm). These data are consistent with our hypothesis that impaired insulin signalling in T2DM sensitises VMH GE neurones to decreased glucose.
phosphatidylinositol-3-kinase; db/db mice; insulin; compound 2; electrophysiology
Reproductive experience (i.e. parturition and lactation) leads to persistent alterations in anxietylike behaviour that are influenced by the oestrous cycle. We recently found that repeated administration of the selective oestrogen receptors (ER)α agonist propyl-pyrazole triol (PPT) results in anxiolytic-like behaviours on the elevated plus maze (EPM) in primiparous (but not nulliparous) female rats. The present study examined the effects of the acute administration of PPT on EPM behaviour in primiparous and aged-matched, nulliparous female rats. In addition, corticosterone secretion, corticotrophin-releasing hormone (CRH) gene expression and expression of the immediate early gene product Fos in the paraventricular nucleus (PVN) and amygdala were measured either after EPM testing or in home cage controls. Acute PPT administration significantly modified EPM behaviour as a function of reproductive experience, with nulliparous females tending toward increased anxiety-like behaviours and primiparous females tending toward decreased anxiety-like behaviours. In home cage controls, PPT increased corticosterone secretion in all females; however, both vehicle- and PPT-treated, primiparous females had reduced corticosterone levels compared to their nulliparous counterparts. Significant effects of PPT on CRH mRNA within the PVN were observed after the administration of PPT but only in primiparous females tested on the EPM. PPT also increased Fos expression within the PVN of EPM-exposed females; however, both vehicle- and PPT-treated primiparous females had reduced Fos expression compared to nulliparous females. In the amygdala, PPT increased Fos immunore-activity in the central but not the medial or basolateral amygdala, although these effects were only observed in home cage females. Additionally, both vehicle- and PPT-treated home cage, primiparous females had increased Fos in the central nucleus of the amygdala compared to nullip-arous controls. Overall, these data demonstrate that reproductive experience alters the behavioural response to acute ERα activation. Moreover, the findings suggest that central regulation of the hypothalamic-adrenal-pituitary axis is modified as a consequence of reproductive experience.
Corticotrophin-releasing hormone; anxiety; stress; Fos; PVN; amygdala
Periodic ovulation in rats, mice and hamsters is the result of a surge in LH that depends on circadian gating signals emerging from the master circadian clock within the suprachiasmatic nucleus (SCN) and rising ovarian oestrogen levels. These two signals converge into the anteroventral periventricular nucleus (AVPV) and lead to the release of kisspeptin, which is responsible for surges of GnRH and, in turn, of LH release. How the AVPV integrates circadian and reproductive signals remains unclear. Here we show that the female rat AVPV itself shows circadian oscillations in the expression of the clock genes PER1 and BMAL1, which lie at the core circadian clockwork of mammals. In ovariectomized (OVX) females treated with estradiol (E2) these oscillations are in synchrony with the AVPV rhythmic expression of the KISS1 gene and the gene that codes for the arginine-vasopressin (AVP) receptor AVPr1a. Whereas clock gene oscillations are independent of oestrogen levels, circadian expression of Kiss1 and Avpr1a (also referred to as V1a) mRNA are respectively blunted and absent in ovariectomized animals without E2 replacement. Because AVP is believed to be a critical SCN transmitter to gate the LH surge, our data suggest that a there is a circadian oscillator located in the AVPV, and that such a putative oscillator could time, in an oestrogen dependent manner, the sensitivity to circadian signals emerging from the SCN and the release of kisspeptin.
kisspeptin; vasopressin; circadian oscillators; ovulation
Neuropeptide W (NPW), an endogenous ligand for the G-Protein coupled receptor GPR7 (NPBWR1), is produced in neurones in the rat hypothalamus and brain stem known to be important in the control of food intake and the neuroendocrine response to stress. In previous studies, central administration of NPW during the light phase increased food and water intake and elevated prolactin and corticosterone levels in conscious, unrestrained male rats. In the present studies, central administration of siRNA reduced NPW levels in hypothalamus and resulted in a failure of angiotensin II to stimulate water drinking or increase mean arterial pressure. In addition siRNA treated animals failed to mount a significant prolactin response to immobilisation stress, while maintaining a normal corticosterone response. These results suggest that endogenous NPW may be a physiologically relevant, downstream mediator of the central actions of angiotensin II to stimulate thirst and increase arterial pressure. In addition, NPW-producing neurones appear to participate in the hypothalamic mechanisms controlling prolactin, but not corticosterone, secretion.
Angiotensin II; Neuropeptide W; Water Drinking; Blood Pressure; Stress Hormones
Oxytocin is a major neuropeptide that modulates the brain functions involved in social behavior and interaction. Despite of the importance of oxytocin for neural control of social behavior, little is known about the molecular mechanism(s) by which oxytocin secretion in the brain is regulated. Pro-oxytocin is synthesized in the cell bodies of hypothalamic neurons in the supraoptic and paraventricular nuclei and processed to a 9-amino-acid mature form during post-Golgi transport to the secretion sites at the axon terminals and somatodendritic regions. Oxytocin secreted from the somatodendritic regions diffuses throughout the hypothalamus and its neighboring brain regions. Some oxytocin-positive axons innervate and secrete oxytocin to the brain regions distal to the hypothalamus. Brain oxytocin binds to its receptors in the brain regions involved in social behavior. Oxytocin is also secreted from the axon terminal at the posterior pituitary gland into the blood circulation. We have discovered a new molecular complex consisting of annexin A1 (ANXA1), A-kinase anchor protein 150 (AKAP150), and microtubule motor, that controls the distribution of oxytocin vesicles between the axon and the cell body in a protein kinase A (PKA)- and protein kinase C (PKC)-sensitive manner. ANXA1 showed significant co-localization with oxytocin vesicles. Activation of PKA enhanced the association of kinesin-2 with ANXA1, thus increasing the axon-localization of oxytocin vesicles. Conversely, activation of PKC decreased the binding of kinesin-2 to ANXA1, thus attenuating the axon-localization of oxytocin vesicles. Our study suggests that ANXA1 complex coordinates the actions of PKA and PKC to control the distribution of oxytocin vesicles between the axon and the cell body.
Oxytocin; Annexin A1; Kinesin-2; AKAP150; PKA; PKC
Cytochrome P450 aromatase (P450arom; aromatase) is a microsomal enzyme involved in the production of endogeneous sex steroids by converting testosterone into oestradiol. Aromatase is the product of the cyp19a1 gene and plays a crucial role in the sexual differentiation of the brain and in the regulation of reproductive functions. In the brain of mammals and birds, expression of cyp19a1 has been demonstrated in neuronal populations of the telencephalon and diencephalon. By contrast, a wealth of evidence established that, in teleost fishes, aromatase expression in the brain is restricted to radial glial cells. The present study investigated the precise neuroanatomical distribution of cyp19a1 mRNA during brain development in Xenopus laevis (late embryonic to juvenile stages). For this purpose, we used in situ hybridisation alone or combined with the detection of a proliferative (proliferating cell nuclear antigen), glial (brain lipid binding protein, Vimentin) or neuronal (acetylated tubulin; HuC/D; NeuroβTubulin) markers. We provide evidence that cyp19a1 expression in the brain is initiated from the very early larval stage and remains strongly detected until the juvenile and adult stages. At all stages analysed, we found the highest expression of cyp19a1 in the preoptic area and the hypothalamus compared to the rest of the brain. In these two brain regions, cyp19a1-positive cells were never detected in the ventricular layers. Indeed, no co-labelling could be observed with radial glial (brain lipid binding protein, Vimentin) or dividing progenitors (proliferating cell nuclear antigen) markers. By contrast, cyp19a1-positive cells perfectly matched with the distribution of post-mitotic neurones as shown by the use of specific markers (HuC/D, acetylated tubulin and NeuroβTubulin). These data suggest that, similar to that found in other tetrapods, aromatase in the brain of amphibians is found in post-mitotic neurones and not in radial glia as reported in teleosts.
Aromatase; Cyp19a1; radial glia; Xenopus; Brain
Steroid hormones act in specific regions of the brain to alter behavior and physiology. While it has been well established that the bioavailability of the steroid and the expression of its receptor is critical to understanding steroid action in brain, the importance of nuclear receptor coactivators in brain is becoming more apparent. This review will focus on the function of the p160 family of coactivators, which includes steroid receptor coactivator-1 (SRC-1), SRC-2 and SRC-3, in steroid receptor action in brain. The expression, regulation and function of these coactivators in steroid-dependent gene expression in brain and behavior will be discussed.
steroid receptor coactivator-1 (SRC-1/NcoA-1); SRC-2 (GRIP1/TIF2/NCoA-2); androgen receptor; estrogen receptor; progestin receptor; sex behavior; hypothalamus; steroid hormones
Tourette syndrome (TS) is a neurodevelopmental disorder characterized by recurring motor and phonic tics. The pathogenesis of TS is thought to reflect dysregulations in the signaling of dopamine (DA) and other neurotransmitters, which lead to excitation/inhibition imbalances in cortico-striato-thalamocortical circuits. The causes of these deficits may reflect complex gene × environment × sex (G×E×S) interactions; indeed, the disorder is markedly predominant in males, with a male-to-female prevalence ratio of ~4:1. Converging lines of evidence point to neuroactive steroids as likely molecular candidates to account for GxExS interactions in TS. Building on these premises, our group has begun examining the possibility that alterations in the steroid biosynthetic process may be directly implicated in TS pathophysiology; in particular, our research has focused on 5α-reductase (5αR), the enzyme catalyzing the key rate-limiting step in the synthesis of pregnane and androstane neurosteroids. In clinical and preclinical studies, we found that 5αR inhibitors exerted marked anti-DAergic and tic-suppressing properties, suggesting a central role for this enzyme in TS pathogenesis. Based on these data, we hypothesize that enhancements in 5αR activity in early developmental stages may lead to an inappropriate activation of the “backdoor” pathway for androgen synthesis from adrenarche until the end of puberty. We predict that the ensuing imbalances in steroid homeostasis may impair the signaling of DA and other neurotransmitters, ultimately resulting in the facilitation of tics and other behavioral abnormalities in TS.
Prenatal exposure to corticosteroids has long-term postnatal somatic and neurodevelopmental consequences. Animal studies indicate that corticosteroid exposure-associated alterations in the nervous system include hypothalamic function. Infants with infantile spasms, a devastating epileptic syndrome of infancy with characteristic spastic seizures, chaotic irregular waves on interictal electroencephalogram (EEG; hypsarhythmia) and mental deterioration, have decreased concentrations of adrenocorticotropic hormone (ACTH) and cortisol in cerebrospinal fluid strongly suggesting hypothalamic dysfunction. We have exploited this feature to develop a model of human infantile spasms by using repeated prenatal exposure to betamethasone and postnatal trigger of developmentally relevant spasms with N-methyl-D-aspartic acid (NMDA). The spasms triggered in prenatally primed rats are more severe compared to prenatally saline-injected ones and respond to ACTH, a treatment of choice for infantile spasms in humans. Using autoradiography and immunohistochemistry, we have identified a link between the spasms in our model and hypothalamus, especially the arcuate nucleus. Transcriptomic analysis of the arcuate nucleus after prenatal priming with betamethasone but before trigger of spasms indicates that prenatal betamethasone exposure down-regulates genes encoding several important proteins participating in glutamatergic and GABAergic transmission. Interestingly, there were significant sex-specific alterations after prenatal betamethasone in synapse-related gene expression but no such sex differences were found in prenatally saline-injected controls. A pair-wise relevance analysis revealed that, although the synapse gene expression in controls was independent of sex, these genes form topologically distinct gene fabrics in males and females and these fabrics are altered by betamethasone in a sex-specific manner. These findings may explain the sex differences in both normal behaviour and occurrence and severity of infantile spasms. Changes in transcript expression and their coordination may contribute to a molecular substrate of permanent neurodevelopmental changes (including infantile spasms) found after prenatal exposure to corticosteroids.
Like humans, rhesus macaques (Macaca mulatta) are large, long-lived diurnal primates, and show similar age-related changes in the secretion of many steroid hormones, including oestradiol, testosterone, cortisol, and dehydroepiandrosterone (DHEA). Consequently, they represent a pragmatic animal model in which to examine the mechanisms by which these steroidal changes contribute to perturbed sleep-wake cycles and cognitive decline in the elderly. Using remote serial blood sampling we have found the circulating levels of DHEA sulphate, as well as oestradiol and testosterone, decline markedly in old monkeys. Furthermore, using real-time PCR, we have shown that genes associated with the conversion of DHEA to oestradiol and testosterone (e.g., 3BHSD, 17BHSD, and aromatase) are highly expressed in brain areas associated with cognition and behavior, including the hippocampus, prefrontal cortex, and amygdala. Taken together, these findings suggest that administration of supplementary DHEA in the elderly may have therapeutic potential for cognitive and behavioral disorders, but with fewer negative side effects outside of the central nervous system. To test this we have developed a novel steroid supplementation paradigm for use in old animals; this involves oral administration of DHEA and testosterone at physiologically relevant times of the day to mimic the circadian hormone patterns observed in young adults. We are currently evaluating the efficacy of this steroid supplementation paradigm at reversing age-associated disorders, including perturbed sleep-wake cycles and cognitive decline as well as impaired immune response.
dehydroepiandrosterone; oestradiol; testosterone; circadian rhythms
Epigenetic alterations of histone proteins and DNA are essential for hippocampal synaptic plasticity and cognitive function, and contribute to the etiology of psychiatric disorders and neurodegenerative diseases. Hippocampal memory formation depends on histone alterations and DNA methylation, and increasing evidence suggests that regulation of these epigenetic processes by modulatory factors such as environmental enrichment, stress, and hormones substantially influences memory function. Recent work from our laboratory suggests that the ability of the sex-steroid hormone 17β-estradiol (E2) to enhance novel object recognition memory consolidation in young adult female mice is dependent on histone H3 acetylation and DNA methylation in the dorsal hippocampus. Our data also suggest that enzymes mediating DNA methylation and histone acetylation work in concert to regulate the effects of E2 on memory consolidation. These findings shed light on the epigenetic mechanisms that influence hormonal modulation of cognitive function, and may have important implications for understanding how hormones influence cognition in adulthood and aging. This review will provide a brief overview of the literature on epigenetics and memory, describe in detail our findings demonstrating that epigenetic alterations regulate E2-induced memory enhancement in female mice, and discuss future directions for research on the epigenetic regulation of E2-induced memory enhancement.
histone acetylation; DNA methylation; hippocampus; novel object recognition; estrogen
The acceptance of estradiol signaling through receptors found in the cell membrane, as well as, the nucleus has provided for a re-examination of timing and location of estradiol actions on neural circuits mediating sexual receptivity (lordosis). Estradiol membrane signaling involves the transactivation of metabotropic glutamate receptors (mGluR) that transduce steroid information through PKC signaling cascades producing rapid activation of lordosis regulating circuits. It has been known for some time that estradiol initially produces an inhibition of the medial preoptic nucleus (MPN). We have demonstrated that underlying this inhibition is estradiol acting in the arcuate nucleus to induce β-endorphin release which inhibits the MPN through a μ-opioid receptor mechanism. This transient inhibition is relieved by either subsequent progesterone treatment or longer exposure to higher doses of estradiol to facilitate lordosis behavior. We review recent findings about estradiol membrane signaling inducing dendritic spine formation in the arcuate nucleus that is critical for estradiol induction of sexual receptivity. Moreover, we discuss the evidence that in addition to ERα, several other putative membrane estrogen receptors facilitate lordosis behavior through regulation of the arcuate nucleus. These include the GRP30 and the STX activated Gq-mER. Finally, we report on the importance of GABA acting at GABAB receptors for estradiol membrane signaling that regulates lordosis circuit activation and sexual receptivity.
progesterone; MOR; β-endorphin; NPY; actuate nucleus; membrane estrogen receptor; ERα
Progestogens have actions in the midbrain ventral tegmental area (VTA) to mediate motivated behaviours, such as those involved in reproductive processes, among female rodents. In the VTA, formation and actions of one progestogen, 5α-pregnan-3α-ol-20-one (3α,5α-THP; 3α,5α-THP), are necessary and sufficient to facilitate sexual responding (measured by lordosis) of female rodents. Although 3α,5α-THP can be produced following metabolism of ovarian progesterone, 3α,5α-THP is also a neurosteroid produced de novo in brain regions, such as the VTA. There can be dynamic changes in 3α,5α-THP production associated with behavioural experience, such as mating. Questions of interest are the sources and targets of 3α,5α-THP. Regarding sources, pregnane Xenobiotic Receptor (PXR) may be a novel factor involved in 3α,5α-THP metabolism in the VTA (as well as a direct target of 3α,5α-THP). We have identified PXR in the midbrain of female rats, and manipulating PXR in this region reduces 3α,5α-THP synthesis and alters lordosis as well as affective and social behaviours. Regarding targets, recent studies have focused on the role of membrane progestin receptors (mPRs). We have analyzed expression of two of the common forms of these receptors (mPRα/paqr7 and mPRβ/paqr8) in female rats. Expression of mPRα was observed in peripheral tissues and brain areas, including hypothalamus and midbrain. Expression of mPRβ was only observed in brain tissues and was abundant in the midbrain and hypothalamus. To our knowledge, studies of these receptors in mammalian models have been limited to expression and regulation, instead of function. A question that was addressed was the functional effects of progestogens via mPRα and mPRβ in the midbrain of hormone-primed rats for lordosis. Studies to date suggest that mPRβ may be an important target of progestogens in the VTA for lordosis. Together, these studies demonstrate that PXR is involved in production of 3α,5α-THP in the midbrain VTA. Moreover, mPRs may be a target for progestogens’ actions in the VTA for lordosis.
progesterone; non-genomic; lordosis; neurosteroids; mating
Oestrogen rapidly enhances fast excitatory postsynaptic potentials, facilitates long-term potentiation (LTP), and increases spine numbers. Each effect likely contributes to the steroid’s influence on cognition and memory. In this review, we will first describe a model for the substrates of LTP that includes an outline of synaptic events that occur during induction, expression, and consolidation. Briefly, critical signaling pathways involving the small GTPases RhoA and Rac/Cdc42 are activated by theta burst-induced calcium influx and initiate actin filament assembly via phosphorylation (inactivation) of cofilin. Reorganization of the actin cytoskeleton changes spine and synapse morphology, resulting in increased concentrations of AMPA receptors at stimulated contacts. We then use the synaptic model to develop a specific hypothesis about how oestrogen affects both baseline transmission and plasticity. Brief infusions of 17β-estradiol (E2) reversibly stimulate the RhoA, cofilin phosphorylation, and actin polymerization cascade of the LTP machinery; blocking this eliminates the steroid’s effects on transmission. We accordingly propose that E2 induces a weak form of LTP and thereby increases synaptic responses, a hypothesis that also accounts for how it markedly enhances theta burst induced potentiation. While E2’s effects on the cytoskeleton could be due to direct activation of small GTPases by oestrogen receptors on the synaptic membrane, the hormone also activates TrkB receptors for Brain-Derived Neurotrophic Factor (BDNF), a neurotrophin that engages the RhoA-cofilin sequence and promotes LTP. The latter observations raise the possibility that E2 produces its effects on synaptic physiology via transactivation of neighboring receptors that have prominent roles in the management of spine actin, synaptic physiology, and plasticity.
estradiol; spines; RhoA; cofilin; pTrkB; LTP
It is now clear that estrogens are not just circulating reproductive hormones, but that they also have neurotransmitter-like properties in a wide range of brain circuits. The view of estrogens as intrinsic neuromodulators that shape behavior has been bolstered by a series of recent developments from multiple vertebrate model systems. Here, we review several recent findings from studies of songbirds showing how the identified neural circuits that govern auditory processing and sensorimotor integration are modulated by the local and acute production of estrogens. First, studies using in vivo microdialysis demonstrate that estrogens fluctuate in auditory cortex (30-min time bin resolution) when songbirds are hearing song and interacting with conspecifics. Second, estrogens rapidly boost the auditory-evoked activity of neurons in the same auditory cortical region, enhancing auditory processing. Third, local pharmacological blockade of estrogen signaling in this region impairs auditory neuronal responsiveness as well as behavioral song preferences. Fourth, the rapid estrogen actions that occur within the auditory cortex can propagate upstream (transsynaptically) to sensorimotor circuits to enhance the neural representation of song. Lastly, we present new evidence that the receptor for the rapid actions of estradiol is likely in neuronal membranes, and that traditional nuclear estrogen receptor agonists do not mimic these rapid actions. Broadly speaking, many of these findings are observed in both males and females, emphasizing the fundamental importance of estrogens in neural circuit function. Together, these and other emergent studies provide support for rapid, brain-derived estrogen signaling in regulating sensorimotor integration, learning and perception.
Neuroestrogen; neurosteroid; estradiol; electrophysiology; microdialysis
Glucagon-like peptide-1 (GLP-1) plays an important role in energy homeostasis. Injections of GLP-1 receptor (GLP-1R) agonists suppress food intake, and endogenous GLP-1 is released when nutrients enter the gut. There is also growing evidence that the GLP-1 system is involved in the regulation of body fluid homeostasis. GLP-1R agonists suppress water intake independent of their effects on food intake. It is unknown, however, whether this suppressive effect of GLP-1R agonists extends to saline intake. Accordingly, we tested the effect of the GLP-1R agonists liraglutide (0.05 µg) and exendin-4 (0.05 µg) on water and saline intakes stimulated either by angiotensin II (AngII) or by water deprivation with partial rehydration (WD-PR). Each agonist suppressed AngII-induced water intake; however, only exendin-4 suppressed saline intake. WD-PR-induced water and saline intakes were both attenuated by each agonist. Analysis of drinking microstructure after WD-PR found a reliable effect of the agonists on burst number. Furthermore, exendin-4 conditioned a robust taste avoidance to saccharine; however, there was no similar effect of liraglutide. To evaluate the relevance of the conditioned taste avoidance, we tested whether inducing visceral malaise by injection of lithium chloride (LiCl) suppressed fluid intake. Injection of LiCl did not suppress water or saline intakes. Overall, these results indicate that the fluid intake suppression by GLP-1R activation is not selective to water intake, is a function of post-ingestive feedback, and is not secondary to visceral malaise.
Fluid intake; Liraglutide; Exendin-4; thirst
Gonadotrophin-releasing hormone (GnRH) neurones of the hypothalamic-pituitary-gonadal (HPG) axis drive reproductive function and undergo age-related decreases in activation during the transition to reproductive senescence. Decreased GnRH secretion from the median eminence (ME) partially arises from attenuated glutamatergic signaling via the NMDA receptor (NMDAR), and may be due to changing NMDAR stoichiometry to favor NR2b over NR2a subunit expression with aging. We have previously shown that the systemic inhibition of NR2b-containing receptors with ifenprodil, an NR2b-specific antagonist, stimulates parameters of luteinising hormone (used as a proxy for GnRH) release in both young and middle-aged females. Here, we chronically administered ifenprodil, an NR2b-specific antagonist, at the site of GnRH terminals in the median eminence (ME) or at GnRH perikarya in the preoptic area, in reproductively senescent middle-aged female rats to determine whether NR2b antagonism could restore aspects of reproductive functionality. Effects on oestrous cyclicity, serum hormones, and protein expression of GnRH, NR2b, and phosphorylated NR2b (Tyr-1472) in the ME were measured. Chronic ifenprodil treatment in the ME, but not the preoptic area, altered oestrous cyclicity by increasing the percentage of days spent in pro-oestrus. This was accompanied by increased GnRH fluorescence intensity in the external ME zone and a greater proportion of GnRH terminals that co-labelled with pNR2b with treatment. We also observed changes in the relationships between protein immunofluorescence, serum hormone levels, and other aspects of reproductive physiology in acyclic females, as revealed by bionetwork analysis. Together, these data support the hypothesis that NMDAR-NR2b expression and phosphorylation state play a role in reproductive senescence and highlight the ME as a major player in reproductive aging.
Median Eminence; Menopause; Reproductive senescence; NMDA receptor; GnRH
Expression of the primary female sex behaviour, lordosis, in laboratory animals depends on oestrogen-induced expression of progesterone receptor (PgR) within a defined cell group in the ventrolateral portion of the ventromedial nucleus of the hypothalamus (VMH). The minimal latency from oestradiol administration to lordosis is 18 hours. During that time, ligand-bound oestrogen receptors (ER), members of a nuclear receptor superfamily, recruit transcriptional coregulators, which induce covalent modifications of histone proteins thus leading to transcriptional activation or repression of target genes. The aim of this study was to investigate early molecular epigenetic events underlying oestrogen-regulated transcriptional activation of the Pgr gene in the VMH of female mice. Oestradiol (E2) administration induced rapid and transient global histone modifications in the VMH of ovariectomised female mice. Histone H3 N-terminus phosphorylation (H3S10phK14Ac), acetylation (H3Ac) and methylation (H3K4me3) exhibited distinct temporal patterns facilitative to the induction of transcription; and a transcriptional repressive (H3K9me3) modification showed a different temporal pattern. Collectively these should create a permissive environment for the transcriptional activity necessary for lordosis, within 3-6 hours after E2-treatment. In the VMH, changes in the H3Ac and H3K4me3 levels of histone H3 were also detected at the promoter region of Pgr gene within the same time window, but were delayed in the preoptic area. Moreover, examination of histone modifications associated with the promoter of another ER-target gene, oxytocin receptor (Oxtr), revealed gene- and brain-region specific effects of E2 treatment. In the VMH of female mice, E2-treatment resulted in the recruitment of ERα to the oestrogen-response-elements-containing putative enhancer site of Pgr gene ~200kb upstream of the transcription start site (TSS), but failed to increase ERα association with the more proximal promoter region. Finally, E2 administration led to significant changes in the mRNA expression of several ER coregulators in a brain-region dependent manner. Taken together, these data indicate that in the hypothalamus and preoptic area of female mice, early responses to E2-treatment involve highly specific changes in chromatin structure, dependent on cell group, gene, histone modification studied, promoter/enhancer site and time following E2.
oestrogens; oestrogen receptor; progesterone receptor; hypothalamus; transcriptional regulation; histone modification
The neuropeptides kisspeptin (encoded by Kiss1) and RFamide-related peptide-3 (also known as GnIH; encoded by Rfrp) are potent stimulators and inhibitors, respectively, of reproduction. Whether kisspeptin or RFRP-3 might act directly on each other’s neuronal populations to indirectly modulate reproductive status is unknown. To examine possible interconnectivity of the kisspeptin and RFRP-3 systems, we performed double label in-situ hybridization (ISH) for RFRP-3’s receptors, Gpr147 and Gpr74, in hypothalamic Kiss1 neurons of adult male and female mice, as well as double-label ISH for kisspeptin’s receptor, Kiss1r, in Rfrp-expressing neurons of the hypothalamic dorsal-medial nucleus (DMN). Only a very small proportion (5–10%) of Kiss1 neurons of the anteroventral periventricular region expressed Gpr147 or Gpr74 in either sex, whereas higher co-expression (~25%) existed in Kiss1 neurons in the arcuate nucleus. Thus, RFRP-3 could signal to a small, primarily arcuate, subset of Kiss1 neurons, a conclusion supported by the finding of ~35% of arcuate kisspeptin cells receiving RFRP-3--immunoreactive fibre contacts. In contrast to the former situation, no Rfrp neurons co-expressed Kiss1r in either sex, and Tacr3, the receptor for neurokinin B (NKB; a neuropeptide co-expressed with arcuate kisspeptin neurons) was found in <10% of Rfrp neurons. Moreover, kisspeptin-immunoreactive fibres did not readily appose RFRP-3 cells in either sex, further excluding the likelihood that kisspeptin neurons directly communicate to RFRP-3 neurons. Lastly, despite abundant NKB in the DMN region where RFRP-3 soma reside, NKB was not co-expressed in the majority of Rfrp neurons. Our results suggest that RFRP-3 may modulate a small proportion of kisspeptin-producing neurons in mice, particularly in the arcuate nucleus, whereas kisspeptin neurons are unlikely to have any direct reciprocal actions on RFRP-3 neurons.
RFRP-3; GnIH; Gpr147; Gpr74; Gpr54; Kisspeptin; Kiss1; Kiss1r; Tacr3; Tac2; Neurokinin B; reproduction; hypothalamus
Orphanin FQ/nociceptin (OFQ/N) inhibits the activity of proopiomelanocortin (POMC) neurones located in the hypothalamic arcuate nucleus (ARH) that regulate female sexual behaviour and energy balance. We tested the hypothesis that gonadal steroids differentially modulate the ability of OFQ/N to inhibit these cells via presynaptic inhibition of transmitter release and postsynaptic activation of G protein-gated, inwardly-rectifying K+ (GIRK)-1 channels. Whole-cell patch clamp recordings were performed in hypothalamic slices prepared from ovariectomised rats. OFQ/N (1 μM) decreased the frequency of miniature excitatory postsynaptic currents (mEPSCs) and inhibitory postsynaptic currents (mIPSCs), and also caused a robust outward current in the presence of tetrodotoxin, in ARH neurones from vehicle- treated animals. A priming dose of oestradiol benzoate (EB; 2 μg) increased basal mEPSC frequency, markedly diminished both the OFQ/N-induced decrease in mEPSC frequency and the activation of GIRK-1 currents, and potentiated the OFQ/N-induced decrease in mIPSC frequency. Steroid treatment regimens that facilitate sexual receptivity reinstate the basal mEPSC frequency, the OFQ/N-induced decrease in mEPSC frequency and the activation of GIRK-1 currents to levels observed in vehicle-treated controls, and largely abolish the ability of OFQ/N to decrease mIPSC frequency. These effects were observed in an appreciable population of identified POMC neurones, nearly one-half of which projected to the medial preoptic nucleus. Taken together, these data reveal that gonadal steroids influence the pleiotropic actions of OFQ/N on ARH neurones, including POMC neurones, in a disparate manner. These temporal changes in OFQ/N responsiveness further implicate this neuropeptide system as a critical mediator of the gonadal steroid regulation of reproductive behaviour.
estradiol; progesterone; POMC; orphanin FQ; lordosis
Normal hypothalamic-pituitary-adrenal (HPA) axis activity leading to rhythmic and episodic release of adrenal glucocorticoids is essential for body homeostasis and survival during stress. Acting through specific intracellular receptors in the brain and periphery, glucocorticoids regulate behavior, metabolic, cardiovascular, immune, and neuroendocrine activities. In contrast to chronic elevated levels, circadian and acute stress-induced increases in glucocorticoids are necessary for hippocampal neuronal survival and memory acquisition and consolidation, through inhibiting apoptosis, facilitating glutamate transmission and inducing immediate early genes and spine formation. In addition to its metabolic actions leading to increasing energy availability, glucocorticoids have profound effects on feeding behavior, mainly through modulation of orexigenic and anorixegenic neuropeptides. Evidence is also emerging that in addition to the recognized immune suppressive actions of glucocorticoids by counteracting adrenergic proinflammatory actions, circadian elevations have priming effects in the immune system, potentiating acute defensive responses. In addition, negative feedback by glucocorticoids involves multiple mechanisms leading to limiting HPA axis activation and preventing deleterious effects of excessive glucocorticoid production. Adequate glucocorticoid secretion to meet body demands is tightly regulated by a complex neural circuitry controlling hypothalamic corticotrophin releasing hormone (CRH) and vasopressin secretion, the main regulators of pituitary adrenocorticotrophic hormone (ACTH). Rapid feedback mechanisms, likely involving non-genomic actions of glucocorticoids, mediate immediate inhibition of hypothalamic CRH and ACTH secretion, while intermediate and delayed mechanisms mediated by genomic actions involve modulation of limbic circuitry and peripheral metabolic messengers. Consistent with their key adaptive roles, HPA axis components are evolutionarily conserved, being present in the earliest vertebrates. Understanding these basic mechanisms may lead to novel approaches for the development of diagnostic and therapeutic tools for disorders related to stress and alterations of glucocorticoid secretion.
CRH; ACTH; Glucocorticoids; cortisol/corticosterone
Friendships and other rewarding affilliative bonds are associated with the actions of the nonapeptide hormone oxytocin (OT) in humans and many social mammals. We determined if OT itself is rewarding, and if that reward is dependent upon the presence of conspecifics. We evaluated the reinforcing effects of OT infusion in female mice on social (conditioned social preference, CSP), and non-social tests (conditioned place preference, CPP). Ovariectomised females received oestradiol implants and intracerebroventricular cannulas. During a pre-test, they were introduced to a 3-chamber apparatus for 10 minutes. Social and place apparatus were identical, except that each end-chamber contained a novel stimulus female for CSP, whereas they were distinguished by visual and tactile cues for CPP. For CSP, test females received OT (0, 100, 200 or 100ng) and were paired for 30 minutes with one stimulus female. On alternating days, they received saline vehicle and were paired with the opposite female, for a total of 4 pairings each. The final conditioned preference test was identical to the pre-test. OT induced CSP. Test mice that received 100ng OT increased their preference score from −67.4±22.1 seconds in pre-test to +55.7±35.1 seconds during the conditioned preference test (p<0.05). 200ng OT induced an increase in preference score from −162.7±47.3 to +74.3±23.7 seconds (p <0.001). There was no effect of 0 or 1000ng OT on CSP. An additional group of mice was tested for CPP at 200ng OT. Testing and pairings were identical to CSP. OT induced a small but significant CPP. Mice increased their preference score from −222.4±38.0 to −126.0±58.7 seconds (p<0.05). OT had no effect on anxiety or odor recognition as assessed by elevated plus maze and olfactory habituation/dishabituation tests, respectively. In conclusion, OT like other motivating stimuli (drugs, food) is rewarding when tested under solitary conditions, but is also reinforcing in a social setting.
Oxytocin; Conditioned Social Preference; Conditioned Place Preference; Social Behaviour; Dose-Response Curve