Stress activates the hypothalamic-pituitary-adrenal (HPA) axis, a neuroendocrine system that regulates responses related to feeding, reproduction, and aggression, among other homeostatic functions. Stressors significantly impact gene expression along the HPA axis and in hypothalamic nuclei that drive it, including the paraventricular nucleus (PVN). To identify genetic regulators of stress responses in the PVN, adult mice underwent 2 h of multi-modal stress before gene expression profiles were analyzed using bulk RNA sequencing. A transcription factor zinc finger and BTB domain containing 16 (Zbtb16), also known as PLZF, was identified as a stress responsive, glucocorticoid receptor (GR) target in the PVN. Zbtb16 mRNA expression was increased by two-fold in male and female mice within 2 h of restraint stress or injection of a synthetic glucocorticoid, dexamethasone (DEX). Immunohistochemistry (IHC) confirmed Zbtb16 protein expression and localization in the PVN following 20 min of restraint stress and 4 h of recovery. Cellular analyses revealed that Zbtb16 was highly expressed in CRH neurons in the PVN, neurons routinely activated post-stress as indicated by colocalization with c-FOS. Adult mice were also exposed to an immune stress by injection of tumor necrosis factor alpha (TNFα) to assess Zbtb16 regulation. Expanded analyses indicated that the cell specificity of Zbtb16 expression was region-specific, colocalizing with CRH neurons in the mid-PVN but more in astrocytes surrounding the PVN. These findings identify Zbtb16 as a glucocorticoid- and cytokine-inducible transcriptional regulator with region- and cell type-specific roles in PVN stress circuitry.

Abstract Disclosure: J.A. Sheng: None. R.J. Handa: None. S.A. Tobet: None. Background: The blood-brain barrier (BBB) protects the brain from the influx of harmful compounds in the blood. It is comprised of multiple cells, including endothelial cells sharing tight junctions, pericytes, and astrocyte endfeet. The paraventricular nucleus of the hypothalamus (PVN) is 3-5 times more vascularized than surrounding regions in the brain. The current study focuses on the BBB in the PVN as a function of stress. Recent data further suggests a high fat diet (HFD) disrupts the integrity of the BBB and leads to impairment of brain function (Li et al, 2021), suggesting a potential mechanism that may influence stress-related diseases. Methods: Adult male and female mice were placed on standard mouse chow (2918; Tekland) or a HFD (TD.06414, Tekland) for 6 weeks and further divided into control and stressed groups. Control mice were euthanized directly out of their home cage and stressed mice were euthanized 60-min after a 20-min acute restraint stress. Mice were perfused with fluorescein isothiocyanate in phosphate buffered saline followed by fixation with 4% paraformaldehyde to visualize blood vessel integrity (FITC leakage; Frahm & Tobet, 2015). Immunolabeled Glial Fibrillary Acidic Protein (GFAP; astrocytic end feet) and IBA-1 (microglia) were used to further assess BBB integrity and neuroinflammation. Results: Results showed ∼40% more FITC leakage in the PVN vasculature in adult HFD females (p<0.0001) but not males. Similar leakage was not noted in the region lateral to the PVN. IBA-1 immunoreactivity (microglia) showed a HFD-related 50% increase in raw cell counts (p<0.001) and a HFD x restraint decrease in fluorescence intensity (p<0.02) in both sexes. GFAP immunoreactivity (astrocyte end feet) additionally showed 2-fold increase in cell counts in both sexes by HFD (p<0.0001) and a female-specific HFD-induced increase in area (∼40%; p<0.001). Conclusions: Data suggest a chronic high fat diet impairs BBB integrity in PVN vasculature that can be exacerbated further by a brief exposure to stress in adult mice. This effect is being further studied at the cellular level in astrocytes and microglia. Such changes in these BBB components could indicate critical roles for the uniquely dense PVN vasculature, increasing risk of damage to neural functions that could include obesity, cardiovascular autonomic regulation, or depression-like behaviors. Supported by ORWH-NIMH U54 MH118919 SCORE. References: Li C, Shi L, Wang Y, Peng C, Wu L, Zhang Y, Du Z. (2021). High-fat diet exacerbates lead-induced blood-brain barrier disruption by disrupting tight junction integrity. Environ Toxicol. 36(7):1412-1421. doi: 10.1002/tox.23137. Frahm KA, Tobet SA. (2015). Development of the blood-brain barrier within the paraventricular nucleus of the hypothalamus: influence of fetal glucocorticoid excess. Brain Struct Funct. 220(4): 2225-2234. doi: 10.1007/s00429-014-0787-8. Presentation: Thursday, June 15, 2023

IntroductionMaternal adversity during pregnancy influences neurodevelopment in human and model animal offspring. Adversity can result from stressors coming from many different directions ranging from environmental to nutritional and physiological to immune (e.g., infection). Most stressors result in fetal overexposure to glucocorticoids that have been directly linked to long- and short-term negative impacts on neurological health of offspring. Neuropsychiatric diseases postulated to have fetal origins are diverse and include such things cardiovascular disease, obesity, affective disorders, and metabolic and immune disorders.MethodsThe experiments in the current study compare 3 stressors: prenatal exposure to dexamethasone (DEX), maternal high fat diet (HFD), and maternal caloric restriction (CR). Offspring of mothers with these treatments were examined prepubertally to evaluate stress responsiveness and stress-related behaviors in in male and female mice.ResultsPrenatal exposure to synthetic glucocorticoid, DEX, resulted in decreased neonatal body weights, reduced social interaction behavior, and hypoactive stress response offspring exposed to maternal DEX. Maternal CR resulted in decreased body weights and social interaction behavior in males and females and increased anxiety-like behavior and acute stress response only in males. HFD resulted in altered body weight gain in both sex offspring with decreased anxiety-like behavior in a female-biased manner.DiscussionThe idea that glucocorticoid responses to different stressors might serve as a common stimulus across stress paradigms is insufficient, given that different modes of prenatal stress produced differential effects. Opposite nutritional stressors produced similar outcomes for anxiety-like behavior in both sexes, social-like behavior in females, and a hyperactive adrenal stress response in males. One common theme among the three models of maternal stress (DEX, CR, and HFD) was consistent data showing their role in activating the maternal and fetal immune response. By tuning in on the more immediate immunological aspect on the developing fetus (e.g., hormones, cytokines), additional studies may tease out more direct outcomes of maternal stress in rodents and increase their translational value to human studies.

The ventromedial prefrontal cortex (vmPFC) regulates fear acquisition, fear extinction, mood, and HPA axis function. Multiple brain regions exhibit time-of-day dependent variations in learning, long term potentiation (LTP), and dendritic morphology. Glucocorticoids have been implicated in the regulation of dendritic structure in the context of stress. Glucocorticoids are also known to regulate molecular clock entrainment via upregulation of Per1 transcription. In the present study, C57BL/6 N mice were sacrificed at three distinct times of day (ZT3, ZT12, and ZT16, lights off at ZT12) and Per1 mRNA expression was measured in the infralimbic and prelimbic vmPFC subregions using droplet digital (dd) PCR after recovering from adrenalectomy or sham surgery for 10 days. Sham mice showed Per1 rhythmicity in both infralimbic (IL) and prelimbic (PL) cortex, with peak expression occurring at ZT12. Adrenalectomized mice showed reductions in Per1 amplitude at ZT12 in both IL and PL, suggesting that the vmPFC molecular clock is entrained by diurnal glucocorticoid oscillations. Thy1-eGFP mice were used to visualize and quantify dendritic spine density on deep layer pyramidal dendrites at ZT 3, 12, and 16. Spine density in both PL and IL exhibited changes between the light (inactive) and dark (active) phases, with peak spine density observed at ZT16 and trough spine density observed at ZT3. These changes in spine density were restricted to changes in long thin and stubby type spines. To determine if changes in spine density is regulated by glucocorticoid oscillations, the 11β-hydroxylase inhibitor metyrapone was administered 2 h prior to the onset of the active phase (ZT10) daily for 7 days. Metyrapone administration blocked both the diurnal peak of plasma corticosterone and peak spine densities in the IL and PL at ZT16. These results suggest that vmPFC molecular clock gene and dendritic spine diurnal rhythms depend on intact diurnal glucocorticoid oscillations.

Prenatal insults leading to increased fetal glucocorticoid exposure can sex‐selectively impact cardiovascular function in adulthood. In support of this, we have shown that when pregnant rat dams are treated with the glucocorticoid, dexamethasone (DEX), for the last 4 days of gestation, female‐specific changes resulting in enhanced pressor and heart rate (HR) responses to stress occur in adult offspring. Moreover, we demonstrated that females exposed to DEX in uteroshow altered autonomic function compared to vehicle‐exposed females. This is characterized by a reduced high frequency (HF) power component of heart rate variability (HRV) in DEX‐exposed rats, suggesting a lower reliance on parasympathetic drive. The present study evaluated the degree to which the sex‐specific changes in autonomically‐driven cardiovascular responses are due to activational effects of gonadal steroid hormones. Pregnant dams were administered DEX (0.4mg/kg per day, s.c.) or vehicle on gestation days 18‐21. This resulted in a significant reduction in birthweight in DEX‐exposed males and females. At 8 weeks, rats underwent a gonadectomy (GDX) or sham surgery, or remained intact, and at 10 weeks rats were instrumented with radiotelemetric transmitters for direct recording of arterial pressure, HR, and HRV in conscious, freely moving male and female rats. At 11‐12 weeks rats were placed in a restraint tube for 20 minutes, followed by a 3‐hour recovery period, to assess whether GDX alters the sex‐specific stress responses in DEX‐exposed offspring. Restraint‐stress testing was performed on diestrus in intact and sham females, and absence of cycling in GDX females was confirmed via cytological analysis. We demonstrate that intact females, but not males, that were exposed to DEX in utero exhibit an exaggerated pressor response to restraint, as compared to vehicle‐exposed females. We found that GDX did not alter stress‐responsive MAP in males regardless of prenatal treatment, suggesting testosterone does not play a role in acute cardiovascular stress responses in adult rats. In vehicle‐exposed females, when compared to intact, both sham and GDX surgery resulted in an exaggerated pressor response to restraint. However, in females that were prenatally exposed to DEX, there was no difference in the pressor response to restraint between intact, sham, and GDX rats. This suggests that the exaggerated pressor response observed in DEX females compared to males is not due to activational effects of estradiol. GDX was shown to have opposing effects on HRV in males and females, with no influence of prior DEX. In females, GDX resulted in a tendency toward reduced HF (p=0.065) and low frequency (LF, p=0.074) power component of HRV, whereas males showed a significant increase in HF (p=0.044) and a tendency toward increased LF (0.051). Taken together, activational effects of gonadal steroids appear to impact HRV, but not acute cardiovascular responses to stress. It may be that gonadal steroids act at an organizational level to mediate the sex difference observed in rats exposed to DEX in utero. Future studies to identify the mechanisms by which prenatal dexamethasone produce long‐term changes in cardiovascular function will be important for better understanding the sex‐specific consequences of prenatal programming over the lifespan.

Chronic stress is a significant risk factor for negative health outcomes. Furthermore, imbalance of autonomic nervous system control leads to dysregulation of physiological responses to stress and contributes to the pathogenesis of cardiometabolic and psychiatric disorders. However, research on autonomic stress responses has historically focused on males, despite evidence that females are disproportionality affected by stress-related disorders. Accordingly, this mini-review focuses on the influence of biological sex on autonomic responses to stress in humans and rodent models. The reviewed literature points to sex differences in the consequences of chronic stress, including cardiovascular and metabolic disease. We also explore basic rodent studies of sex-specific autonomic responses to stress with a focus on sex hormones and hypothalamic-pituitary-adrenal axis regulation of cardiovascular and metabolic physiology. Ultimately, emerging evidence of sex differences in autonomic-endocrine integration highlights the importance of sex-specific studies to understand and treat cardiometabolic dysfunction.

In utero exposure to glucocorticoids in late gestation programs changes in cardiovascular function. The objective of this study was to determine the degree to which angiotensin II mediates sex-biased changes in autonomic function as well as basal and stress-responsive cardiovascular function following in utero glucocorticoid exposure. Pregnant rats were administered the synthetic glucocorticoid dexamethasone (Dex; 0.4 mg/kg/day sc) or vehicle on gestation days 18-21. Mean arterial pressure, heart rate, and heart rate variability (HRV) were measured via radiotelemetry in freely moving, conscious adult rats. To evaluate the impact of stress, rats were placed in a restraint tube for 20 min. In a separate cohort of rats, restraint stress was performed before and after chronic treatment with the angiotensin type 1 receptor antagonist, losartan (30 mg/kg/day ip). Frequency domain analysis of HRV was evaluated, and data were integrated into low-frequency (LF, 0.20-0.75 Hz) and high-frequency (HF, 0.75-2.00 Hz) bands. Prenatal Dex resulted in an exaggerated pressor and heart rate response to restraint in female offspring that was attenuated by prior losartan treatment. HF power was higher in vehicle-exposed female rats compared with Dex females. Following losartan, HF power was equivalent between female vehicle and Dex-exposed rats. In utero exposure to Dex produced female-biased alterations in stress-responsive cardiovascular function, which may be indicative of a reduction in parasympathetic activity. Moreover, these findings suggest this autonomic dysregulation may be mediated, in part, by long-term changes in renin-angiotensin signaling.NEW & NOTEWORTHY Our findings reveal the involvement of angiotensin II on sex-selective cardiovascular function and autonomic changes in adult offspring exposed to dexamethasone during the last 4 days of gestation. We show that angiotensin II receptor blockade reverses the exaggerated pressor and heart rate response to acute restraint stress and the autonomic dysregulation observed in female, but not male, offspring exposed to dexamethasone in utero.

hypothalamic-pituitary-adrenal (HPA) axis mediates the physiological response to stressors and also synchronizes different physiological systems to environmental cues. Changes in day length (i.e., photoperiod) as well as chronic exposure to stressors are known to impact the HPA axis activity regulating the levels of glucocorticoid hormones. Over-exposure to inappropriate levels of glucocorticoids has been implicated in increased disease risk. In the present study, we examined the impact of chronic stress, using a chronic variable stress (CVS) paradigm, in combination with changes in photoperiod on physiological and behavioral measures, as well as on the reactivity and regulation of the HPA axis, in male and female mice. Six weeks of CVS, regardless of the photoperiod condition, decreased the body weight and attenuated the HPA axis reactivity to an acute stressor in both sexes. The attenuated HPA axis reactivity observed in stressed animals was related to reduced Proopiomelanocortin (POMC) mRNA levels in the pituitary of females. The gene expression analyses of key regulators of the HPA axis also indicated a sex-dependent effect with opposite patterns in the pituitary and adrenal glands. CVS effects on behavior were limited and related to an anxiety-like phenotype in both sexes, regardless of photoperiod condition. Our findings highlight sex-specific differences in the HPA axis and also sex-dependent effects of CVS on physiological parameters.(c) 2022 IBRO. Published by Elsevier Ltd. All rights reserved.

Sex differences in the neuroendocrine response to acute stress occur in both animals and humans. In rodents, stressors such as restraint and novelty induce a greater activation of the hypothalamic-pituitary-adrenal axis (HPA) in females compared to males. The nature of this difference arises from steroid actions during development (organizational effects) and adulthood (activational effects). Androgens decrease HPA stress responsivity to acute stress, while estradiol increases it. Androgenic down-regulation of HPA responsiveness is mediated by the binding of testosterone (T) and dihydrotestosterone (DHT) to the androgen receptor, as well as the binding of the DHT metabolite, 3β-diol, to the β form of the estrogen receptor (ERβ). Estradiol binding to the α form of the estrogen receptor (ERα) increases HPA responsivity. Studies of human sex differences are relatively few and generally employ a psychosocial paradigm to measure stress-related HPA activation. Men consistently show greater HPA reactivity than women when being evaluated for achievement. Some studies have found greater reactivity in women when being evaluated for social performance. The pattern is inconsistent with rodent studies but may involve the differential nature of the stressors employed. Psychosocial stress is nonphysical and invokes a significant degree of top-down processing that is not easily comparable to the types of stressors employed in rodents. Gender identity may also be a factor based on recent work showing that it influences the neural processing of positive and negative emotional stimuli independent of genetic sex. Comparing different types of stressors and how they interact with gender identity and genetic sex will provide a better understanding of sex steroid influences on stress-related HPA reactivity.

Androgens play a pivotal role during development. These gonadal hormones and their receptors exert organizational actions that shape brain morphology in regions controlling the stress regulatory systems in a male-specific manner. Specifically, androgens drive sex differences in the hypothalamic/pituitary/adrenal (HPA) axis and corresponding hypothalamic neuropeptides. While studies have examined the role of estradiol and its receptors in sex differences in the HPA axis and associated behaviors, the role of androgens remains far less studied. Androgens are generally thought to modulate the HPA axis through the activation of androgen receptors (ARs). They can also impact the HPA axis through reduction to estrogenic metabolites that can bind estrogen receptors in the brain and periphery. Such regulation of the HPA axis stress response by androgens can often result in sex-biased risk factors for stress-related disorders, such as anxiety and depression. This review focuses on the biosynthesis pathways and molecular actions of androgens and their nuclear receptors. The impact of androgens on hypothalamic neuropeptide systems (corticotropin-releasing hormone, arginine vasopressin, oxytocin, dopamine, and serotonin) that control the stress response and stress-related disorders is discussed. Finally, this review discusses potential therapeutics involving androgens (androgen replacement therapies, selective AR modulator therapies) and ongoing clinical trials.

BACKGROUND: Sex differences in incidence and/or presentation of schizophrenia (SCZ), major depressive disorder (MDD), and bipolar disorder (BIP) are pervasive. Previous evidence for shared genetic risk and sex differences in brain abnormalities across disorders suggest possible shared sex-dependent genetic risk. METHODS: We conducted the largest to date genome-wide genotype-by-sex (GxS) interaction of risk for these disorders using 85,735 cases (33,403 SCZ, 19,924 BIP, and 32,408 MDD) and 109,946 controls from the PGC (Psychiatric Genomics Consortium) and iPSYCH. RESULTS: Across disorders, genome-wide significant single nucleotide polymorphism-by-sex interaction was detected for a locus encompassing NKAIN2 (rs117780815, p = 3.2 x 10(-8)), which interacts with sodium/potassium-transporting ATPase (adenosine triphosphatase) enzymes, implicating neuronal excitability. Three additional loci showed evidence (p < 1 x 10(-8)) for cross-disorder GxS interaction (rs7302529, p = 1.6 x 10(-7); rs73033497, p = 8.8 x 10(-7); rs7914279, p = 6.4 x 10(-7)), implicating various functions. Gene-based analyses identified GxS interaction across disorders (p = 8.97 x 10(-7)) with transcriptional inhibitor SLTM. Most significant in SCZ was a MOCOS gene locus (rs11665282, p = 1.5 x 10(-7)), implicating vascular endothelial cells. Secondary analysis of the PGC-SCZ dataset detected an interaction (rs13265509, p = 1.1 x 10(-7)) in a locus containing IDO2, a kynurenine pathway enzyme with immunoregulatory functions implicated in SCZ, BIP, and MDD. Pathway enrichment analysis detected significant GxS interaction of genes regulating vascular endothelial growth factor receptor signaling in MDD (false discovery rate-corrected p < .05). CONCLUSIONS: In the largest genome-wide GXS analysis of mood and psychotic disorders to date, there was substantial genetic overlap between the sexes. However, significant sex-dependent effects were enriched for genes related to neuronal development and immune and vascular functions across and within SCZ, BIP, and MDD at the variant, gene, and pathway levels.

Background: Obesity (OB) and major depressive disorder (MDD) are chronic conditions associated with disease burden, and their comorbidity appears more common among women. Mechanisms linking these conditions may involve inflammatory and metabolic pathways. The goal of this study was to evaluate the impact of MDD on relationships between OB and cardiometabolic function, and sex differences therein. Materials and Methods: Adult offspring from the New England Family Studies (NEFS) were assessed at ages 39-50, including anthropometry, cardiometabolic profile assays, and metabolic syndrome. Individuals were grouped by body mass index (BMI) and MDD status: healthy weight with (n = 50) or without MDD (n = 95) and obese with (n = 79) or without MDD (n = 131). The interaction of (recurrent) MDD and BMI on cardiometabolic markers was tested using quantile regression models. Results: Participants with MDD exhibited significantly higher hemoglobin A1c (HbA1c) than those without MDD (5.60% vs. 5.35%, p < 0.05). Women with comorbid recurrent MDD and OB had higher HbA1c levels compared to obese women without MDD (5.75% vs. 5.44%, p < 0.05); an interaction between MDD and BMI status was not observed among men. Conclusions: We demonstrated sex differences in the interaction between BMI and recurrent MDD status on a primary biomarker for diabetes risk, suggesting a common metabolic pathway predisposing women to these comorbid conditions. Further investigation is needed to identify mechanisms that may lead to more effective, sex-dependent screening and therapies.

Background The Period Circadian Regulator 2 ( Per2 ) gene is important for the modulation of circadian rhythms that influence biological processes. Circadian control of the hypothalamus-pituitary-adrenal (HPA) axis is critical for regulation of hormones involved in the stress response. Dysregulation of the HPA axis is associated with neuropsychiatric disorders. Therefore, it is important to understand how disruption of the circadian rhythm alters the HPA axis. One way to address this question is to delete a gene involved in regulating a central circadian gene such as Per2 in an animal model and to determine how this deletion may affect the HPA axis and behaviors that are altered when the HPA axis is dysregulated. To study this, corticosterone (CORT) levels were measured through the transition from light (inactive phase) to dark (active phase). Additionally, CORT levels as well as pituitary and adrenal mRNA expression were measured following a mild restraint stress. Mice were tested for depressive-like behaviors (forced swim test (FST)), acoustic startle response (ASR), and pre-pulse inhibition (PPI). Results The present results showed that Per2 knockout impacted CORT levels, mRNA expression, depressive-like behaviors, ASR and PPI. Unlike wild-type (WT) mice, Per2 knockout ( Per2 ) mice showed no diurnal rise in CORT levels at the onset of the dark cycle. Per2−/− mice had enhanced CORT levels and adrenal melanocortin receptor 2 ( Mc2R ) mRNA expression following restraint. There were no changes in expression of any other pituitary or adrenal gene. In the FST, Per2−/− mice spent more time floating (less time struggling) than WT mice, suggesting increased depressive-like behaviors. Per2−/− mice had deficits in ASR and PPI startle responses compared to WT mice. Conclusions In summary, these findings showed that disruption of the circadian system via Per2 gene deletion dysregulated the HPA stress axis and is subsequently correlated with increased depressive-like behaviors and deficits in startle response.

Abstract The hypothalamic pituitary adrenal (HPA) axis responds to environmental perturbations to maintain homeostasis. Pregnancy demands extensive modifications in HPA axis function to prepare for increased energy and metabolic demands required to meet the needs of mother and offspring. Short-term effects of pregnancy on the HPA axis have been shown, but data is lacking regarding the long-term effects in middle-aged female mice no longer breeding. Since changes of the HPA axis are further found with age, in this study, we examined both parity- and age-related interactions on the HPA axis in female mice. Wildtype C57bl/6N females were divided into nulliparous young (NY) (3-6 mo) and nulliparous middle-aged (NM) or multiparous retired-breeder middle-aged (RBM) (8-10 mo) groups. RBM mice were killed at least 4 weeks after their last litter was weaned. Control mice were euthanized directly out of the home cage and experimental groups were euthanized at 0 min, 30 min, or 90 min recovery (n=8-10/ group) after 2h of multi-modal stress (MMS; restraint, noise, shaker, light). (Paraventricular nucleus of the hypothalamus (PVN) neuronal activity was quantified by c-FOS immunoreactivity (-ir), and plasma corticosterone (CORT) levels were measured by radioimmunoassay. Corticotropin releasing hormone (CRH) mRNA was assayed by in situ hybridization. Two-way ANOVA showed effects of age (p<0.0248), parity (p<0.0021), time (p<0.0001), and interaction (p<0.0009) on CORT levels. Specifically, basal CORT levels were reduced in NM/RBM versus NY mice. In all groups, CORT levels were significantly elevated by MMS. There was no difference between CORT levels immediately after MMS in NY or NM groups, but CORT levels after 30- and 90- min recovery from MMS remained elevated in NM, indicating reduced negative feedback with age. Additionally, RBM plasma CORT was further reduced in all time groups versus NM, accompanied by a return to baseline CORT after 90 min recovery, suggesting a parity-dependent effect on the HPA axis. Changes in CORT levels were correlated with c-FOS-ir. MMS increased PVN c-FOS-ir in all groups compared to controls and c-FOS-ir in NM was significantly greater than PVN c-FOS of RBM. Further, while c-FOS-ir in the NY females was reduced to baseline 30 min after MMS, the return to baseline was more gradual in NM. No effect of parity or age was seen in Crh mRNA. Collectively, our findings show that activation of the HPA axis in females involves interactions between age- and parity- dependent function. Our findings further show activation and inhibition of the HPA axis in females involving long-term changes that occur after pregnancy, which may increase risk for stress- or postpartum- related disorders. Supported by NIDDK 1-R01 DK105826

The hypothalamic-pituitary-adrenal axis is a complex system of neuroendocrine pathways and feedback loops that function to maintain physiological homeostasis. Abnormal development of the hypothalamic-pituitary-adrenal (HPA) axis can further result in long-term alterations in neuropeptide and neurotransmitter synthesis in the central nervous system, as well as glucocorticoid hormone synthesis in the periphery. Together, these changes can potentially lead to a disruption in neuroendocrine, behavioral, autonomic, and metabolic functions in adulthood. In this review, we will discuss the regulation of the HPA axis and its development. We will also examine the maternal-fetal hypothalamic-pituitary-adrenal axis and disruption of the normal fetal environment which becomes a major risk factor for many neurodevelopmental pathologies in adulthood, such as major depressive disorder, anxiety, schizophrenia, and others.

The pituitary gland is composed of two functionally unique lobes known as the anterior and posterior pituitary. The primary purpose of the posterior lobe of the pituitary, also known as the neurohypophysis, is to act as a conduit whereby hormones made within the hypothalamus can be secreted directly into the peripheral circulation. The major hormones of the posterior pituitary are oxytocin (OT) and vasopressin (AVP). These two related neuropeptides are made by neurons in the paraventricular and supraoptic nucleus of the hypothalamus. They are structurally similar and are transcribed in opposite directions off the same chromosomal region where their genes are arranged in a tail-to-tail fashion. OT and AVP are phylogenetically ancient hormones and are conserved through evolution. Moreover, they operate by binding a distinct family of G protein–coupled receptors. Activation of the OT receptor induces changes in intracellular calcium to cause myometrial contraction necessary for parturition, whereas activation of AVP receptors (V1aR, V1bR, and V2R) is responsible for the effects of AVP on the regulation of blood pressure and water balance, both of which are essential for homeostasis. OT and AVP receptors are also found in brain, where OT and AVP regulate a number of behaviors as well as autonomic responses.

The interaction between isoflavones and the gut microbiota has been highlighted as a potential regulator of obesity and diabetes. In this study, we examined the interaction between isoflavones and a shortened activity photoperiod on the gut microbiome. Male mice were exposed to a diet containing no isoflavones (NIF) or a regular diet (RD) containing the usual isoflavones level found in a standard vivarium chow. These groups were further divided into regular (12L:12D) or short active (16L:8D) photoperiod, which mimics seasonal changes observed at high latitudes. White adipose tissue and genes involved in lipid metabolism and adipogenesis processes were analysed. Bacterial genomic DNA was isolated from fecal boli, and 16S ribosomal RNA sequencing was performed. NIF diet increased body weight and adipocyte size when compared to mice on RD. The lack of isoflavones and photoperiod alteration also caused dysregulation of lipoprotein lipase ( Lpl ), glucose transporter type 4 ( Glut-4 ) and peroxisome proliferator-activated receptor gamma ( Pparg ) genes. Using 16S ribosomal RNA sequencing, we found that mice fed the NIF diet had a greater proportion of Firmicutes than Bacteroidetes when compared to animals on the RD. These alterations were accompanied by changes in the endocrine profile, with lower thyroid-stimulating hormone levels in the NIF group compared to the RD. Interestingly, the NIF group displayed increased locomotion as compared to the RD group. Together, these data show an interaction between the gut bacterial communities, photoperiod length and isoflavone compounds, which may be essential for understanding and improving metabolic health.

The chronic variable stress (CVS) paradigm is frequently used to model the changes in hypothalamic pituitary adrenal (HPA) axis function characteristic of many stress-related diseases. However, male C57BL/6 mice are typically resistant to CVS's effects, making it difficult to determine how chronic stress exposure may alter acute HPA function and regulation in these mice. As social support in rodents can profoundly influence physiological and behavioral processes, including the HPA axis, we sought to characterize the effects of CVS exposure on basal and acute stress-induced HPA axis function in pair- and single-housed adult male mice. Despite all subjects exhibiting decreased body weight gain after six weeks of CVS, the corticosterone response to a novel, acute restraint stressor was enhanced by CVS exclusively in single-housed males. CVS also significantly increased arginine vasopressin (AVP) mRNA in the hypothalamic paraventricular nucleus (PVN) in single-housed males only. Moreover, in single-, but not pair-housed mice, CVS attenuated decreases in circulating OT found following acute restraint. Only the effect of CVS to elevate PVN corticotropin releasing hormone (CRH) mRNA levels after an acute stressor was restricted to pair-housed mice. Collectively, our findings suggest that social isolation reveals effects of CVS on the HPA axis in male C57BL/6 mice.

It is well established that even transient prenatal insults can impact cardiovascular (CV) function in adulthood. We hypothesized that adult CV disease may have its origins in utero as a result of exposure to elevated levels of glucocorticoids arising as a result of prenatal stress or inflammation. In support of this, we showed that when pregnant rat dams are treated with the synthetic glucocorticoid, dexamethasone (DEX), for 4 days in late gestation, there are female‐specific changes resulting in enhanced pressor and tachycardic responses to stress in adult offspring. We further showed that these effects in DEX exposed females can be reversed following treatment with the angiotensin II type 1 receptor antagonist, losartan. To better understand whether autonomic imbalance drives the sex‐specific CV effects of adult offspring prenatally exposed to DEX, we investigated the impact of DEX on heart rate variability (HRV). Pregnant dams were administered DEX (0.4mg/kg per day, s.c.) or vehicle on gestation days 18–21. This resulted in a significant reduction in birthweight in DEX‐exposed males and females. At 2–3 months of age, rats were instrumented with radiotelemetric transmitters for direct recording of arterial pressure, heart rate, and HRV. In a separate cohort, female rats were re‐evaluated following 9 days of losartan (30mg/kg per day, i.p.). HRV was analyzed in the frequency domain by obtaining arterial pressure waveforms recorded over 2–3 days. Interbeat intervals were analyzed (Kubios Software), and a fast fourier transformation was performed and data integrated into two frequency bands: low frequency (LF: 0.20–0.75 Hz) and high frequency (0.75–2.00 Hz). The LF component represents both the sympathetic (SYM) and parasympathetic (PS) input, while the HF component corresponds to PS nervous system activity. The LF/HF ratio is used to assess relative SYM activity. Data were analyzed by Two Way ANOVA (sex x prenatal treatment). There was a significant effect of sex with females having greater HF power, and a significant interaction whereby HF power increased in males and decreased in females prenatally exposed to DEX. Neither sex nor prenatal DEX had any impact on LF power; however, there was a significant sex effect and interaction when assessing LF/HF. DEX decreased the ratio in males, but tended to increase it in females exposed to DEX, in utero. Losartan treatment in DEX exposed females significantly increased HF to a level that was not different from vehicle rats. The increased HF power in females suggests a greater reliance on PS drive than males. However, prenatal DEX exposure produces sex‐specific reductions PS drive in females, that is reversible by losartan treatment. This decrease in PS activity may underlie the altered pressor and tachycardic stress responses that we have previously observed in these rats. Additionally, these findings suggest that autonomic dysregulation following prenatal DEX may be mediated in part due to long‐term changes in central renin‐angiotensin signaling. Further studies will use SYM and PS antagonists to further elucidate the impact of DEX and sex on cardiac autonomic imbalance in the pursuit of examining the long‐term consequence of age‐ and sex‐related changes of DEX on CV function.Support or Funding InformationSpringboard, ABRC

Estradiol and testosterone are powerful steroid hormones that impact brain function in numerous ways. During development, these hormones can act to program the adult brain in a male or female direction. During adulthood, gonadal steroid hormones can activate or inhibit brain regions to modulate adult functions. Sex differences in behavioral and neuroendocrine (i.e., hypothalamic pituitary adrenal (HPA) axis) responses to stress arise as a result of these organizational and activational actions. The sex differences that are present in the HPA and behavioral responses to stress are particularly important considering their role in maintaining homeostasis. Furthermore, dysregulation of these systems can underlie the sex biases in risk for complex, stress-related diseases that are found in humans. Although many studies have explored the role of estrogen and estrogen receptors in mediating sex differences in stress-related behaviors and HPA function, much less consideration has been given to the role of androgens. While circulating androgens can act by binding and activating androgen receptors, they can also act by metabolism to estrogenic molecules to impact estrogen signaling in the brain and periphery. This review focuses on androgens as an important hormone for modulating the HPA axis and behaviors throughout life and for setting up sex differences in key stress regulatory systems that could impact risk for disease in adulthood. In particular, impacts of androgens on neuropeptide systems known to play key roles in HPA and behavioral responses to stress (corticotropin-releasing factor, vasopressin, and oxytocin) are discussed. A greater knowledge of androgen action in the brain is key to understanding the neurobiology of stress in both sexes.