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Chronic immobilization stress (CIS) results in sex-dependent changes in opioid peptide levels and receptor subcellular distributions within the rat dorsal hippocampus, which are paralleled with an inability for males to acquire conditioned place preference (CPP) to oxycodone. Here, RNAScope in situ hybridization was used to determine the expression of hippocampal opioid peptides and receptors in unstressed (US) and CIS estrus female and male adult (∼2.5 months old ) Sprague Dawley rats. In all groups, dentate granule cells expressed PENK and PDYN; additionally, numerous interneurons expressed PENK. OPRD1 and OPRM1 were primarily expressed in interneurons, and to a lesser extent, in pyramidal and granule cells. OPRK1-was expressed in sparsely distributed interneurons. There were few baseline sex differences: US females compared to US males had more PENK-expressing and fewer OPRD1-expressing granule cells and more OPRM1-expressing CA3b interneurons. Several expression differences emerged after CIS. Both CIS females and males compared to their US counterparts had elevated: (1) PENK-expressing dentate granule cells and interneurons in CA1 and CA2/3a; (2) OPRD1 probe number and cell expression in CA1, CA2/3a and CA3b and the dentate gyrus; and (3) OPRK1-expressing interneurons in the dentate hilus. Also, CIS males compared to US males had elevated: (1) PDYN expression in granule cells; (2) OPRD1 probe and interneuron expression in CA2/3a; (3) OPRM1 in granule cells; and (4) OPRK1 interneuron expression in CA2/3a. The sex-specific changes in hippocampal opioid gene expression may impact network properties and synaptic plasticity processes that may contribute to the attenuation of oxycodone CPP in CIS males., (© 2021 Wiley Periodicals LLC.)

Oxycodone (Oxy) conditioned place preference (CPP) in Sprague Dawley rats results in sex-specific alterations in hippocampal opioid circuits in a manner that facilitates opioid-associative learning processes, particularly in females. Here, we examined if Oxy (3 mg/kg, I.P.) or saline (Sal) injections not paired with behavioral testing similarly affect the hippocampal opioid system. Sal-injected females compared to Sal-injected males had: (1) higher densities of cytoplasmic delta opioid receptors (DOR) in GABAergic hilar dendrites suggesting higher baseline reserve DOR pools and (2) elevated phosphorylated DOR levels, but lower phosphorylated mu opioid receptor (MOR) levels in CA3a suggesting that the baseline pools of activated opioid receptors vary in females and males. In contrast to CPP studies, Oxy-injections in the absence of behavioral tests resulted in few changes in the hippocampal opioid system in either females or males. Specifically, Oxy-injected males compared to Sal-injected males had fewer DORs near the plasma membrane of CA3 pyramidal cell dendrites and in CA3 dendritic spines contacted by mossy fibers, and lower pMOR levels in CA3a. Oxy-injected females compared to Sal-injected females had higher total DORs in GABAergic dendrites and lower total MORs in parvalbumin-containing dendrites. Thus, unlike Oxy CPP, Oxy-injections redistributed opioid receptors in hippocampal neurons in a manner that would either decrease (males) or not alter (females) excitability and plasticity processes. These results indicate that the majority of changes within hippocampal opioid circuits that would promote opioid-associative learning processes in both females and males do not occur with Oxy administration alone, and instead must be paired with CPP., (© 2020 Wiley Periodicals LLC.)

Obstructive sleep apnea (OSA), a chronic sleep disorder characterized by repetitive reduction or cessation of airflow during sleep, is widely prevalent and is associated with adverse neurocognitive sequelae including increased risk of Alzheimer's disease (AD). In humans, OSA is more common in elderly males. OSA is characterized by sleep fragmentation and chronic intermittent hypoxia (CIH), and recent epidemiological studies point to CIH as the best predictor of neurocognitive sequelae associated with OSA. The sex- and age- specific effects of OSA-associated CIH on specific cell populations such as γ-aminobutyric acid (GABA)-ergic neurons in the hippocampus and the medial prefrontal cortex (mPFC), regions important for cognitive function, remain largely unknown. The present study examined the effect of 35 days of either moderate (10% oxygen) or severe (5% oxygen) CIH on GABAergic neurons in the mPFC and hippocampus of young and aged male and female mice as well as post-accelerated ovarian failure (AOF) female mice. In the mPFC and hippocampus, the number of GABA-labeled neurons increased in aged and young severe CIH males compared to controls but not in young moderate CIH males. This change was not representative of the individual GABAergic cell subpopulations, as the number of parvalbumin-labeled neurons decreased while the number of somatostatin-labeled neurons increased in the hippocampus of severe CIH young males only. In all female groups, the number of GABA-labeled cells was not different between CIH and controls. However, in the mPFC, CIH increased the number of parvalbumin-labeled neurons in young females and the number of somatostatin-labeled cells in AOF females but decreased the number of somatostatin-labeled cells in aged females. In the hippocampus, CIH decreased the number of somatostatin-labeled neurons in young females. CIH decreased the density of vesicular GABA transporter in the mPFC of AOF females only. These findings suggest sex-specific changes in GABAergic neurons in the hippocampus and mPFC with males showing an increase of this cell population as compared to their female counterparts following CIH. Age at exposure and severity of CIH also differentially affect the GABAergic cell population in mice., Competing Interests: Declaration of Competing Interest The authors declare no competing financial interest., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Following oxycodone conditioned place preference (CPP) in naïve female and male Sprague Dawley rats, delta- and mu-opioid receptors (DORs and MORs) redistribute in hippocampal CA3 pyramidal cells and GABAergic interneurons in a manner that would promote opioid-associative learning processes, particularly in females. MORs and DORs similarly redistribute in CA3 and hilar neurons following chronic immobilization stress (CIS) in females, but not males, essentially "priming" the opioid system for oxycodone-associative learning. Following CIS, only females acquire oxycodone CPP. The present study determined whether sex and CIS differentially affect the levels of phosphorylated MORs and DORs (pMORs and pDORs) in the hippocampus following oxycodone CPP as phosphorylation is important for opioid receptor internationalization and trafficking. In naïve oxycodone-injected (Oxy) female rats, the density of pMOR-immunoreactivity (ir) was increased in CA1 stratum oriens and CA3a,b strata lucidum and radiatum compared to saline-injected (Sal)-females. Additionally, the density of pDOR-ir increased in the pyramidal cell layer and stratum radiatum of CA2/3a in Oxy-males compared to Sal-males. In CIS females that acquire CPP, pDOR-ir levels were increased in the CA2/3a. These findings indicate only rats that acquire oxycodone CPP have activated MORs and DORs in the hippocampus but that the subregion containing activated opioid receptors differs in females and males. These results are consistent with previously observed sex differences in the hippocampal opioid system following Oxy-CPP., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Infant maltreatment increases vulnerability to physical and mental disorders, yet specific mechanisms embedded within this complex infant experience that induce this vulnerability remain elusive. To define critical features of maltreatment-induced vulnerability, rat pups were reared from postnatal day 8 (PN8) with a maltreating mother, which produced amygdala and hippocampal deficits and decreased social behavior at PN13. Next, we deconstructed the maltreatment experience to reveal sufficient and necessary conditions to induce this phenotype. Social behavior and amygdala deficits (volume, neurogenesis, c-Fos, local field potential) required combined chronic high corticosterone and maternal presence (not maternal behavior). Hippocampal deficits were induced by chronic high corticosterone regardless of social context. Causation was shown by blocking corticosterone during maltreatment and suppressing amygdala activity during social behavior testing. These results highlight (1) that early life maltreatment initiates multiple pathways to pathology, each with distinct causal mechanisms and outcomes, and (2) the importance of social presence on brain development., Competing Interests: The authors declare no competing interest.

Background: Previous studies identified several separate risk factors for stress-induced disorders. However, an integrative model of susceptibility versus resilience to stress including measures from brain-body domains is likely to yield a range of multiple phenotypic information to promote successful adaptation to stress., Methods: We used computational and molecular approaches to test whether 1) integrative brain-body behavioral, immunological, and structural domains characterized and predicted susceptibility or resilience to social defeat stress (SDS) in mice and 2) administration of acetyl-L-carnitine promoted resilience at the SDS paradigm., Results: Our findings identified multidimensional brain-body predictors of susceptibility versus resilience to SDS. The copresence of anxiety, decreased hippocampal volume, and elevated systemic interleukin-6 characterized a susceptible phenotype that developed behavioral and neurobiological deficits after exposure to SDS. The susceptible phenotype showed social withdrawal and impaired transcriptomic-wide changes in the ventral dentate gyrus after SDS. At the individual level, a computational approach predicted whether a given animal developed SDS-induced social withdrawal, or remained resilient, based on the integrative in vivo measures of anxiety and immune system function. Finally, we provide initial evidence that administration of acetyl-L-carnitine promoted behavioral resilience at the SDS paradigm., Conclusions: The current findings of multidimensional brain-body predictors of susceptibility versus resilience to stress provide a starting point for in vivo models of mechanisms predisposing apparently healthy individuals to develop the neurobiological and behavioral deficits resulting from stress exposure. This framework can lead to novel therapeutic strategies to promote resilience in susceptible phenotypes., (Copyright © 2019 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.)

Background: In humans, accumulated adverse experiences during childhood increase the risk of anxiety disorders and attention-deficit/hyperactivity disorder. In rodents, the ventral hippocampus (vHIP) is associated with anxiety regulation, and lesions in this region alter both anxiety-like behavior and activity levels. Neuronal oscillations in the vHIP of the theta frequency range (4-12 Hz) have been implicated in anxious states and derive in part from the activity of inhibitory interneurons in the hippocampus, some of which are enwrapped with perineuronal nets (PNNs), extracellular matrix structures known to regulate plasticity. We sought to investigate the associations among early life stress-induced anxiety and hyperactivity with vHIP neuronal oscillations, inhibitory interneurons, and PNNs in mice., Methods: We used repeated maternal separation with early weaning (MSEW) to model accumulated early life adversity in mouse offspring and studied the underlying cellular and electrophysiological changes in the vHIP that are associated with excessive anxiety and hyperactivity., Results: We found increased anxiety-like behavior and activity levels in MSEW adult males, along with increased theta power and enhanced theta-gamma coupling in the vHIP. MSEW mice showed reduced intensity of parvalbumin as well as increased PNN intensity around parvalbumin-positive interneurons in the vHIP. We further observed that MSEW increased orthodenticle homeobox protein 2, a transcription factor promoting PNN development, in the choroid plexus, where it is produced, as well as in parvalbumin-positive interneurons, where it is sequestered., Conclusions: These findings raise the possibility of causal links among parvalbumin-positive interneurons, PNNs, orthodenticle homeobox protein 2, and MSEW-induced anxiety and hyperactivity., (Copyright © 2019 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.)

Regular exercise and dietary supplements with antioxidants each have the potential to improve cognitive function and attenuate cognitive decline, and, in some cases, they enhance each other. Our current results reveal that low-intensity exercise (mild exercise, ME) and the natural antioxidant carotenoid astaxanthin (AX) each have equivalent beneficial effects on hippocampal neurogenesis and memory function. We found that the enhancement by ME combined with AX in potentiating hippocampus-based plasticity and cognition is mediated by leptin (LEP) made and acting in the hippocampus. In assessing the combined effects upon wild-type (WT) mice undergoing ME with or without an AX diet for four weeks, we found that, when administrated alone, ME and AX separately enhanced neurogenesis and spatial memory, and when combined they were at least additive in their effects. DNA microarray and bioinformatics analyses revealed not only the up-regulation of an antioxidant gene, ABHD3 , but also that the up-regulation of LEP gene expression in the hippocampus of WT mice with ME alone is further enhanced by AX. Together, they also increased hippocampal LEP (h-LEP) protein levels and enhanced spatial memory mediated through AKT/STAT3 signaling. AX treatment also has direct action on human neuroblastoma cell lines to increase cell viability associated with increased LEP expression. In LEP-deficient mice ( ob/ob ), chronic infusion of LEP into the lateral ventricles restored the synergy. Collectively, our findings suggest that not only h-LEP but also exogenous LEP mediates effects of ME on neural functions underlying memory, which is further enhanced by the antioxidant AX., Competing Interests: Conflict of interest statement: J.S.Y. and H.S. are listed as inventors on patent application WO2017213197A1. H.S. has received a research grant and served as a scientific advisor for AstaReal Co. Ltd. All other authors declare that they have no competing interests.

In animals, stress and corticosteroid excess are associated with decreases in memory performance and hippocampal volume that may be prevented with agents that decrease glutamate release. Humans also demonstrate changes in memory and hippocampus with corticosteroids. In this report the effects of glutamate-release inhibitor lamotrigine on hippocampal structure and memory were examined in people receiving medically needed prescription corticosteroid therapy. A total of 54 outpatient adults (n = 28 women) receiving chronic (≥ 6 months) oral corticosteroid therapy were randomized to lamotrigine or placebo for 48 weeks. Declarative memory was assessed using the Rey Auditory Verbal Learning Test (RAVLT); structural magnetic resonance imaging (MRI) as well as single-voxel proton MR spectroscopy ( 1 HMRS) focused on hippocampus were obtained at baseline and week 48. Utilizing a mixed-model approach, structural and biochemical data were examined by separate ANOVAs, and memory was assessed with a multi-level longitudinal model. RAVLT total scores demonstrated significantly better declarative memory performance with lamotrigine than placebo (p = 0.047). Hippocampal subfield volumes were not significantly different between the treatment groups. In summary, lamotrigine was associated with less decline in declarative memory performance than placebo in corticosteroid-treated patients. Findings suggest that, in humans as well as in animal models, glutamate release inhibitors may attenuate some of the effects on the human memory associated with corticosteroids., (Copyright © 2018. Published by Elsevier B.V.)

Preclinical studies implicate insulin signaling as a modulator of dopamine transmission, but human data is currently limited. We hypothesize that changes in the expression of insulin receptor-related genes in the post-mortem brain tissue of patients with mood and psychotic disorders mediate the expression of dopamine regulation-related genes. From a database containing microarray data from the post-mortem dorsolateral prefrontal cortex (dlPFC) (healthy controls [HC]: n = 209; patients: n = 321) and hippocampus (HC: n = 180; patients: n = 196), we conducted a hypothesis-driven analysis through the a priori selection of 12 dopamine- and 3 insulin-related genes. Mediation and moderated mediation models, accounting for the role of body mass index (BMI), were used. In the dlPFC, expressions of insulin receptor- and dopamine regulation-related genes were moderated by BMI, with significantly lower expression in high BMI patients. In the hippocampus, there were significantly lower expressions of these genes, which were not moderated by BMI. Illnesses by BMI effects on expression of dopamine genes were fully mediated by expression of insulin receptor gene (INSR). Analysis of conditional indirect effects showed interactions between INSR and BMI, indicating significantly stronger indirect effects at higher BMI values. In the hippocampus we observed that expression of insulin receptor substrate 1 and 2 fully mediated the effects of illnesses on expression of dopamine genes. In conclusion, differential expression of dopamine-related genes was related to altered expression of insulin signaling genes. BMI had region-specific effects, supporting the hypothesis that metabolic systems are critical mediators of dopaminergic function., (Copyright © 2018. Published by Elsevier Ltd.)

Although opioid addiction has risen dramatically, the role of gender in addiction has been difficult to elucidate. We previously found sex-dependent differences in the hippocampal opioid system of Sprague-Dawley rats that may promote associative learning relevant to drug abuse. The present studies show that although female and male rats acquired conditioned place preference (CPP) to the mu-opioid receptor (MOR) agonist oxycodone (3 mg/kg, I.P.), hippocampal opioid circuits were differentially altered. In CA3, Leu-Enkephalin-containing mossy fibers had elevated levels in oxycodone CPP (Oxy) males comparable to those in females and sprouted in Oxy-females, suggesting different mechanisms for enhancing opioid sensitivity. Electron microscopy revealed that in Oxy-males delta opioid receptors (DORs) redistributed to mossy fiber-CA3 synapses in a manner resembling females that we previously showed is important for opioid-mediated long-term potentiation. Moreover, in Oxy-females DORs redistributed to CA3 pyramidal cell spines, suggesting the potential for enhanced plasticity processes. In Saline-injected (Sal) females, dentate hilar parvalbumin-containing basket interneuron dendrites had fewer MORs, however plasmalemmal and total MORs increased in Oxy-females. In dentate hilar GABAergic dendrites that contain neuropeptide Y, Sal-females compared to Sal-males had higher plasmalemmal DORs, and near-plasmalemmal DORs increased in Oxy-females. This redistribution of MORs and DORs within hilar interneurons in Oxy-females would potentially enhance disinhibition of granule cells via two different circuits. Together, these results indicate that oxycodone CPP induces sex-dependent redistributions of opioid receptors in hippocampal circuits in a manner facilitating opioid-associative learning processes and may help explain the increased susceptibility of females to opioid addiction acquisition and relapse., (Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.)

The development of medial temporal lobe circuits is critical for subsequent learning and memory functions later in life. The present study reports the expression of progesterone receptor (PR), a powerful transcription factor of the nuclear steroid receptor superfamily, in Cajal-Retzius cells of the molecular layer of the dentate gyrus of rats. PR was transiently expressed from the day of birth through postnatal day 21, but was absent thereafter. Although PR immunoreactive (PR-ir) cells did not clearly express typical markers of mature neurons, they possessed an ultrastructural morphology consistent with neurons. PRir cells did not express markers for GABAergic neurons, neuronal precursor cells, nor radial glia. However, virtually all PR cells co-expressed the calcium binding protein, calretinin, and the glycoprotein, reelin, both reliable markers for Cajal-Retzius neurons, a transient population of developmentally critical pioneer neurons that guide synaptogenesis of perforant path afferents and histogenesis of the dentate gyrus. Indeed, inhibition of PR activity during the first two weeks of life impaired adult performance on both the novel object recognition and object placement memory tasks, two behavioral tasks hypothesized to describe facets of episodic-like memory in rodents. These findings suggest that PR plays an unexplored and important role in the development of hippocampal circuitry and adult memory function., (© 2018 Wiley Periodicals, Inc.)

Corticotropin-releasing factor receptors (CRFR1) contribute to stress-induced adaptations in hippocampal structure and function that can affect learning and memory processes. Our prior studies showed that female rats with elevated estrogens compared to males have more plasmalemmal CRFR1 in CA1 pyramidal cells, suggesting a greater sensitivity to stress. Here, we examined the distribution of hippocampal CRFR1 following chronic immobilization stress (CIS) in female and male rats using immuno-electron microscopy. Without stress, total CRFR1 dendritic levels were higher in females in CA1 and in males in the hilus; moreover, plasmalemmal CRFR1 was elevated in pyramidal cell dendrites in CA1 in females and in CA3 in males. Following CIS, near-plasmalemmal CRFR1 increased in CA1 pyramidal cell dendrites in males but not to levels of control or CIS females. In CA3 and the hilus, CIS decreased cytoplasmic and total CRFR1 in dendrites in males only. These results suggest that in naive rats, CRF could induce a greater activation of CA1 pyramidal cells in females than males. Moreover, after CIS, which leads to even greater sex differences in CRFR1 by trafficking it to different subcellular compartments, CRF could enhance activation of CA1 pyramidal cells in males but to a lesser extent than either unstressed or CIS females. Additionally, CA3 pyramidal cells and inhibitory interneurons in males have heightened sensitivity to CRF, regardless of stress state. These sex differences in CRFR1 distribution and trafficking in the hippocampus may contribute to reported sex differences in hippocampus-dependent learning processes in baseline conditions and following chronic stress., (Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.)

We demonstrate that stress differentially regulates glutamate homeostasis in the dorsal and ventral hippocampus and identify a role for the astroglial xCT in ventral dentate gyrus (vDG) in stress and antidepressant responses. We provide an RNA-seq roadmap for the stress-sensitive vDG. The transcription factor REST binds to xCT promoter in co-occupancy with the epigenetic marker H3K27ac to regulate expression of xCT, which is also reduced in a genetic mouse model of inherent susceptibility to depressive-like behavior. Pharmacologically, modulating histone acetylation with acetyl-L-carnitine (LAC) or acetyl-N-cysteine (NAC) rapidly increases xCT and activates a network with mGlu2 receptors to prime an enhanced glutamate homeostasis that promotes both pro-resilient and antidepressant-like responses. Pharmacological xCT blockage counteracts NAC prophylactic effects. GFAP + -Cre-dependent overexpression of xCT in vDG mimics pharmacological actions in promoting resilience. This work establishes a mechanism by which vDG protection leads to stress resilience and antidepressant responses via epigenetic programming of an xCT-mGlu2 network., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Sleep-disordered breathing (SDB) is a common disorder in aging that is associated with cognitive decline, including significant executive dysfunction, for which the neurobiological underpinnings remain poorly understood. Using proton magnetic resonance spectroscopy (1H MRS), this study assessed whether dysregulation of the homeostatic balance of the major inhibitory and excitatory amino acid neurotransmitter systems of γ-aminobutyric acid (GABA) and glutamate, respectively, play a role in SDB. Levels of GABA and those of the combined resonances of glutamate and glutamine (Glx), were measured by 1H MRS in the left dorsolateral prefrontal cortex (l-DLPFC) and bilateral hippocampal regions of 19 older adults (age ± SD: 66.1 ± 1.9 years) with moderate to severe SDB, defined as having an Apnea-Hypopnea Index (AHI) greater than 15 as assessed by polysomnography, and in 14 older adults (age ± SD: 62.3 ± 1.3 years) without SDB (AHI < 5). In subjects with SDB, levels of l-DLPFC GABA, but not Glx, were significantly lower than in control subjects ( P < 0.0002). Additionally, there was a negative correlation between l-DLPFC GABA levels, but not Glx, and SDB severity by AHI ( r = -0.68, P < 0.0001), and a positive correlation between l-DLPFC GABA levels, but not Glx, and minimal oxygen saturation during sleep ( r = 0.62, P = 0.0005). By contrast, no group differences or oxygenation associations were found for levels of GABA or Glx in right or left hippocampal region. These findings are interpreted in terms of a pathophysiological model of SDB in which hypoxia-mediated inhibitory neurotransmission deficit in DLPFC could lead to hyperexcitability and, potentially neuronal dysfunction and cognitive decline., Competing Interests: The authors declare no conflict of interest.

Aims/hypothesis: Type 2 diabetes is likely to be an independent risk factor for hippocampal-based memory dysfunction, although this complication has yet to be investigated in detail. As dysregulated glycometabolism in peripheral tissues is a key symptom of type 2 diabetes, it is hypothesised that diabetes-mediated memory dysfunction is also caused by hippocampal glycometabolic dysfunction. If so, such dysfunction should also be ameliorated with moderate exercise by normalising hippocampal glycometabolism, since 4 weeks of moderate exercise enhances memory function and local hippocampal glycogen levels in normal animals., Methods: The hippocampal glycometabolism in OLETF rats (model of human type 2 diabetes) was assessed and, subsequently, the effects of exercise on memory function and hippocampal glycometabolism were investigated., Results: OLETF rats, which have memory dysfunction, exhibited higher levels of glycogen in the hippocampus than did control rats, and breakdown of hippocampal glycogen with a single bout of exercise remained unimpaired. However, OLETF rats expressed lower levels of hippocampal monocarboxylate transporter 2 (MCT2, a transporter for lactate to neurons). Four weeks of moderate exercise improved spatial memory accompanied by further increase in hippocampal glycogen levels and restoration of MCT2 expression independent of neurotrophic factor and clinical symptoms in OLETF rats., Conclusions/interpretation: Our findings are the first to describe detailed profiles of glycometabolism in the type 2 diabetic hippocampus and to show that 4 weeks of moderate exercise improves memory dysfunction in type 2 diabetes via amelioration of dysregulated hippocampal glycometabolism. Dysregulated hippocampal lactate-transport-related glycometabolism is a possible aetiology of type-2-diabetes-mediated memory dysfunction.

Glucocorticoids (GCs) are involved in stress and circadian regulation, and produce many actions via the GC receptor (GR), which is classically understood to function as a nuclear transcription factor. However, the nuclear genome is not the only genome in eukaryotic cells. The mitochondria also contain a small circular genome, the mitochondrial DNA (mtDNA), that encodes 13 polypeptides. Recent work has established that, in the brain and other systems, the GR is translocated from the cytosol to the mitochondria and that stress and corticosteroids have a direct influence on mtDNA transcription and mitochondrial physiology. To determine if stress affects mitochondrially transcribed mRNA (mtRNA) expression, we exposed adult male rats to both acute and chronic immobilization stress and examined mtRNA expression using quantitative RT-PCR. We found that acute stress had a main effect on mtRNA expression and that expression of NADH dehydrogenase 1, 3, and 6 (ND-1, ND-3, ND-6) and ATP synthase 6 (ATP-6) genes was significantly down-regulated. Chronic stress induced a significant up-regulation of ND-6 expression. Adrenalectomy abolished acute stress-induced mtRNA regulation, demonstrating GC dependence. ChIP sequencing of GR showed that corticosterone treatment induced a dose-dependent association of the GR with the control region of the mitochondrial genome. These findings demonstrate GR and stress-dependent transcriptional regulation of the mitochondrial genome in vivo and are consistent with previous work linking stress and GCs with changes in the function of brain mitochondria.

The hippocampus provided the gateway into much of what we have learned about stress and brain structural and functional plasticity, and this initial focus has expanded to other interconnected brain regions, such as the amygdala and prefrontal cortex. Starting with the discovery of adrenal steroid, and later, estrogen receptors in the hippocampal formation, and subsequent discovery of dendritic and spine synapse remodeling and neurogenesis in the dentate gyrus, mechanistic studies have revealed both genomic and rapid non-genomic actions of circulating steroid hormones in the brain. Many of these actions occur epigenetically and result in ever-changing patterns of gene expression, in which there are important sex differences that need further exploration. Moreover, glucocorticoid and estrogen actions occur synergistically with an increasing number of cellular mediators that help determine the qualitative nature of the response. The hippocampus has also been a gateway to understanding lasting epigenetic effects of early-life experiences. These findings in animal models have resulted in translation to the human brain and have helped change thinking about the nature of brain malfunction in psychiatric disorders and during aging, as well as the mechanisms of the effects of early-life adversity on the brain and the body.

Stress plays a substantial role in shaping behavior and brain function, often with lasting effects. How these lasting effects occur in the context of a fixed postmitotic neuronal genome has been an enduring question for the field. Synaptic plasticity and neurogenesis have provided some of the answers to this question, and more recently epigenetic mechanisms have come to the fore. The exploration of epigenetic mechanisms recently led us to discover that a single acute stress can regulate the expression of retrotransposons in the rat hippocampus via an epigenetic mechanism. We propose that this response may represent a genomic stress response aimed at maintaining genomic and transcriptional stability in vulnerable brain regions such as the hippocampus. This finding and those of other researchers have made clear that retrotransposons and the genomic plasticity they permit play a significant role in brain function during stress and disease. These observations also raise the possibility that the transposome might have adaptive functions at the level of both evolution and the individual organism.

Both estrous cycle and sex affect the numbers and types of neuronal and glial profiles containing the classical estrogen receptors α and β, and synaptic levels in the rodent dorsal hippocampus. Here, we examined whether the membrane estrogen receptor, G-protein-coupled estrogen receptor 1 (GPER1), is anatomically positioned in the dorsal hippocampus of mice to regulate synaptic plasticity. By light microscopy, GPER1-immunoreactivity (IR) was most noticeable in the pyramidal cell layer and interspersed interneurons, especially those in the hilus of the dentate gyrus. Diffuse GPER1-IR was found in all lamina but was most dense in stratum lucidum of CA3. Ultrastructural analysis revealed discrete extranuclear GPER1-IR affiliated with the plasma membrane and endoplasmic reticulum of neuronal perikarya and dendritic shafts, synaptic specializations in dendritic spines, and clusters of vesicles in axon terminals. Moreover, GPER1-IR was found in unmyelinated axons and glial profiles. Overall, the types and amounts of GPER1-labeled profiles were similar between males and females; however, in females elevated estrogen levels generally increased axonal labeling. Some estradiol-induced changes observed in previous studies were replicated by the GPER agonist G1: G1 increased PSD95-IR in strata oriens, lucidum, and radiatum of CA3 in ovariectomized mice 6 h after administration. In contrast, estradiol but not G1 increased Akt phosphorylation levels. Instead, GPER1 actions in the synapse may be due to interactions with synaptic scaffolding proteins, such as SAP97. These results suggest that although estrogen's actions via GPER1 may converge on the same synaptic elements, different pathways are used to achieve these actions., (Copyright © 2015 the authors 0270-6474/15/352384-14$15.00/0.)

The hippocampus is classically involved in memory consolidation, spatial navigation and is involved in the stress response. Migraine is an episodic disorder characterized by intermittent attacks with a number of physiological and emotional stressors associated with or provoking each attack. Given that migraine attacks can be viewed as repeated stressors, alterations in hippocampal function and structure may play an important role in migraine pathophysiology. Using high-resolution magnetic resonance imaging, hippocampal morphometric and functional differences (in response to noxious heat stimulation) were compared in age and gender-matched acute episodic migraineurs with high (HF) versus low (LF) frequency of migraine attacks. Morphometric results were compared with age and gender-matched healthy control (HC) cohort. Significant larger bilateral hippocampal volume was found in LF group relative to the HF and HC groups suggestive of an initial adaptive plasticity that may then become dysfunctional with increased frequency. Functional correlates of greater deactivation (LF > HF) in the same hippocampal regions in response to noxious stimulation was also accompanied by overall reduction in functional connectivity of the hippocampus with other brain regions involved in pain processing in the HF group. The results implicate involvement of hippocampus in the pathophysiology of the migraine.

In the hippocampus, ovarian hormones and sex can alter the trafficking of delta opioid receptors (DORs) and the proportion of DORs that colocalize with the stress hormone, corticotropin releasing factor. Here, we assessed the effects of acute immobilization stress (AIS) and sex on the phosphorylation of DORs in the rat hippocampus. We first localized an antibody to phosphorylated DOR (pDOR) at the SER363 carboxy-terminal residue, and demonstrated its response to an opioid agonist. By light microscopy, pDOR-immunoreactivity (ir) was located predominantly in CA2/CA3a pyramidal cell apical dendrites and in interneurons in CA1-3 stratum oriens and the dentate hilus. By electron microscopy, pDOR-ir primarily was located in somata and dendrites, associated with endomembranes, or in dendritic spines. pDOR-ir was less frequently found in mossy fibers terminals. Quantitative light microscopy revealed a significant increase in pDOR-ir in the CA2/CA3a region of male rats 1h following an injection of the opioid agonist morphine (20mg/kg, I.P). To look at the effects of stress on pDOR, we compared pDOR-ir in males and cycling females after AIS. The level of pDOR-ir in stratum radiatum of CA2/CA3a was increased in control estrus (elevated estrogen and progesterone) females compared to proestrus and diestrus females and males. However, immediately following 30min of AIS, no significant differences in pDOR levels were seen across estrous cycle phase or sex. These findings suggest that hippocampal levels of phosphorylated DORs vary with estrous cycle phase and that acute stress may dampen the differential effects of hormones on DOR activation in females., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The hippocampus is a highly plastic brain region particularly susceptible to the effects of environmental stress; it also shows dynamic changes in epigenetic marks in response to stress and learning. We have previously shown that, in the rat, acute (30 min) restraint stress induces a substantial, regionally specific, increase in hippocampal levels of the repressive histone H3 lysine 9 trimethylation (H3K9me3). Because of the large magnitude of this effect and the fact that stress can induce the expression of endogenous retroviruses and transposable elements in many systems, we hypothesized that the H3K9me3 response was targeted to these elements as a means of containing potential genomic instability. We used ChIP coupled with next generation sequencing (ChIP-Seq) to determine the genomic localization of the H3K9me3 response. Although there was a general increase in this response across the genome, our results validated this hypothesis by demonstrating that stress increases H3K9me3 enrichment at transposable element loci and, using RT-PCR, we demonstrate that this effect represses expression of intracisternal-A particle endogenous retrovirus elements and B2 short interspersed elements, but it does not appear to have a repressive effect on long interspersed element RNA. In addition, we present data showing that the histone H3K9-specific methyltransferases Suv39h2 is up-regulated by acute stress in the hippocampus, and that this may explain the hippocampal specificity we observe. These results are a unique demonstration of the regulatory effect of environmental stress, via an epigenetic mark, on the vast genomic terra incognita represented by transposable elements.

Glucocorticoid (GC) hormones, released by the adrenals in response to stress, are key regulators of neuronal plasticity. In the brain, the hippocampus is a major target of GC, with abundant expression of the GC receptor. GC differentially affect the hippocampal transcriptome and consequently neuronal plasticity in a subregion-specific manner, with consequences for hippocampal information flow and memory formation. Here, we show that GC directly affect the mammalian target of rapamycin (mTOR) signaling pathway, which plays a central role in translational control and has long-lasting effects on the plasticity of specific brain circuits. We demonstrate that regulators of the mTOR pathway, DNA damage-induced transcript (DDIT)4 and FK506-binding protein 51 are transcriptionally up-regulated by an acute GC challenge in the dentate gyrus (DG) subregion of the rat hippocampus, most likely via a GC-response element-driven mechanism. Furthermore, two other mTOR pathway members, the mTOR regulator DDIT4-like and the mTOR target DDIT3, are down-regulated by GC in the rat DG. Interestingly, the GC responsiveness of DDIT4 and DDIT3 was lost in animals with a recent history of chronic stress. Basal hippocampal mTOR protein levels were higher in animals exposed to chronic stress than in controls. Moreover, an acute GC challenge significantly reduced mTOR protein levels in the hippocampus of animals with a chronic stress history but not in unstressed controls. Based on these findings, we propose that direct regulation of the mTOR pathway by GC represents an important mechanism regulating neuronal plasticity in the rat DG, which changes after exposure to chronic stress.

Mild exercise activates hippocampal neurons through the glutamatergic pathway and also promotes adult hippocampal neurogenesis (AHN). We hypothesized that such exercise could enhance local androgen synthesis and cause AHN because hippocampal steroid synthesis is facilitated by activated neurons via N-methyl-D-aspartate receptors. Here we addressed this question using a mild-intense treadmill running model that has been shown to be a potent AHN stimulator. A mass-spectrometric analysis demonstrated that hippocampal dihydrotestosterone increased significantly, whereas testosterone levels did not increase significantly after 2 wk of treadmill running in both orchidectomized (ORX) and sham castrated (Sham) male rats. Furthermore, analysis of mRNA expression for the two isoforms of 5α-reductases (srd5a1, srd5a2) and for androgen receptor (AR) revealed that both increased in the hippocampus after exercise, even in ORX rats. All rats were injected twice with 5'-bromo-2'deoxyuridine (50 mg/kg body weight, i.p.) on the day before training. Mild exercise significantly increased AHN in both ORX and Sham rats. Moreover, the increase of doublecortin or 5'-bromo-2'deoxyuridine/NeuN-positive cells in ORX rats was blocked by s.c. flutamide, an AR antagonist. It was also found that application of an estrogen receptor antagonist, tamoxifen, did not suppress exercise-induced AHN. These results support the hypothesis that, in male animals, mild exercise enhances hippocampal synthesis of dihydrotestosterone and increases AHN via androgenenic mediation.

Nearly 12% of US children are exposed to intense adverse experiences. Research has demonstrated that these experiences can negatively impact adult health, often resulting in psychopathology. Less attention, however, is given to the impact of childhood adverse experiences on childhood health and wellbeing. Using a rodent model of chronic juvenile stress (restraint 6 h daily from postnatal day 20 to 41), we report that chronic stress has significant immediate morbidities in both males and females during this developmental window. Specifically, we demonstrate that chronic juvenile stress produces depressive-like behavior and significant neuronal remodeling of brain regions likely involved in these behavioral alterations: the hippocampus, prefrontal cortex and amygdala. Chronically stressed males and females exhibit anhedonia, increased locomotion when exposed to novelty, and altered coping strategies when exposed to acute stress. Coincident with these behavioral changes, we report simplification of dendrites in the hippocampus and prefrontal cortex and concurrent hypertrophy of dendrites in the amygdala. Taken together, these results demonstrate that chronically stressed juveniles exhibit aberrant behavioral responses to acute challenges that occur in conjunction with stress-induced remodeling of brain regions intimately involved in regulating emotionality and stress reactivity. Further, the absence of sex differences in our reported stress responses, likely speaks to the decreased sensitivity of immature HPA regulating brain regions to sex hormones., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Rationale and Objectives: The rat model of prenatal restraint stress (PRS) replicates factors that are implicated in the etiology of anxious/depressive disorders. We used this model to test the therapeutic efficacy of agomelatine, a novel antidepressant that behaves as a mixed MT1/MT2 melatonin receptor agonist/5-HT(2c) serotonin receptor antagonist., Results: Adult PRS rats showed behavioral, cellular, and biochemical abnormalities that were consistent with an anxious/depressive phenotype. These included an increased immobility in the forced swim test, an anxiety-like behavior in the elevated plus maze, reduced hippocampal levels of phosphorylated cAMP-responsive element binding protein (p-CREB), reduced hippocampal levels of mGlu2/3 and mGlu5 metabotropic glutamate receptors, and reduced neurogenesis in the ventral hippocampus, the specific portion of the hippocampus that encodes memories related to stress and emotions. All of these changes were reversed by a 3- or 6-week treatment with agomelatine (40-50 mg/kg, i.p., once a day). Remarkably, agomelatine had no effect in age-matched control rats, thereby behaving as a "disease-dependent" drug., Conclusions: These data indicate that agomelatine did not act on individual symptoms but corrected all aspects of the pathological epigenetic programming triggered by PRS. Our findings strongly support the antidepressant activity of agomelatine and suggest that the drug impacts mechanisms that lie at the core of anxious/depressive disorders.

Tropomyosin-related kinase B receptor (TrkB) is a neurotrophin receptor important for the synaptic plasticity underlying hippocampal-dependent learning and memory. Because this receptor is widely expressed in hippocampal neurons, the precise location of TrkB activation is likely important for its specific actions. The goal of this study was to identify the precise sites of TrkB activation in the mouse hippocampal formation and to determine any changes in the distribution of activated TrkB under conditions of enhanced brain-derived neurotrophic factor (BDNF) expression and hippocampal excitability. Using electron microscopy, we localized TrkB phosphorylated at tyrosine 816 (pTrkB) in the hippocampal formation of male and female mice under conditions of naturally low circulating estradiol and naturally high circulating estradiol, when BDNF expression, TrkB signaling, and synaptic plasticity are enhanced. To compare relative amounts of pTrkB in each group, we counted profiles containing pTrkB-immunoreactivity (pTrkB-ir) in all hippocampal subregions. pTrkB-ir was in axons, axon terminals, dendrites, and dendritic spines of neurons in the hippocampal formation, but the majority of pTrkB-ir localized to presynaptic profiles. pTrkB-ir also was abundant in glial profiles, which were further identified as microglia using immunofluorescence and confocal microscopy. Axonal and glial pTrkB-ir and pTrkB-ir in the CA1 stratum radiatum were more abundant in high-estradiol states (proestrus females) than low-estradiol states (estrus and diestrus females and males). These findings suggest that presynaptic TrkB is positioned to modulate estradiol-mediated and BDNF-dependent synaptic plasticity. Furthermore, they suggest a novel role for TrkB in microglial function in the neuroimmune system.

Circulating estrogen levels and hippocampal-dependent cognitive functions decline with aging. Moreover, the responses of hippocampal synaptic structure to estrogens differ between aged and young rats. We recently reported that estrogens increase levels of post-synaptic proteins, including PSD-95, and opioid peptides leu-enkephalin and dynorphin in the hippocampus of young animals. However, the influence of ovarian hormones on synaptic protein and opioid peptide levels in the aging hippocampus is understudied. Here, young (3- to 5-month-old), middle-aged (9- to 12-month-old), and aged (about 22-month-old) female rats were ovariectomized and then, 4 weeks later, subcutaneously implanted with a silastic capsule containing vehicle or 17β-estradiol. After 48 h, rats were subcutaneously injected with progesterone or vehicle and sacrificed 1 day later. Coronal sections through the dorsal hippocampus were processed for quantitative peroxidase immunohistochemistry of leu-enkephalin, dynorphin, synaptophysin, and PSD-95. With age, females showed opposing changes in leu-enkephalin and dynorphin levels in the mossy fiber pathway, particularly within the hilus, and regionally specific changes in synaptic protein levels. 17β-estradiol, with or without progesterone, altered leu-enkephalin levels in the dentate gyrus and synaptophysin levels in the CA1 of young but not middle-aged or aged females. Additionally, 17β-estradiol decreased synaptophysin levels in the CA3 of middle-aged females. Our results support and extend previous findings indicating 17β-estradiol modulation of hippocampal opioid peptides and synaptic proteins while demonstrating regional and age-specific effects. Moreover, they lend credence to the "window of opportunity" hypothesis during which hormone replacement can modulate hippocampal structure and circuitry to improve cognitive outcomes., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Inappropriate recollections and responses in stressful conditions are hallmarks of post-traumatic stress disorder and other anxiety and mood disorders, but how stress contributes to the disorders is unclear. Here we show that stress itself reactivates memories even if the memory is unrelated to the stressful experience. Forced-swim stress one day after learning enhanced memory recall. One-day post-learning amnestic treatments were ineffective unless administered soon after the swim, indicating that a stressful experience itself can reactivate unrelated consolidated memories. The swim also triggered inter-hemispheric transfer of a lateralized memory, confirming stress reactivates stable memories. These novel effects of stress on memory required the hippocampus although the memories themselves did not, indicating hippocampus-dependent modulation of extra-hippocampal memories. These findings that a stressful experience itself can activate memory suggest the novel hypothesis that traumatic stress reactivates pre-trauma memories, linking them to memory for the trauma and pathological facilitation of post-traumatic recall., Competing Interests: The authors have declared that no competing interests exist.

Stress of social defeat in rodents is known to have a strong and long-lasting effect on brain, physiology and behavior, which bears similarities with certain human stress related psychopathologies. Previous experiments in this lab showed that social defeat stress suppresses testosterone secretion and causes a lasting desensitization of the serotonergic 5-HT(1A) receptors. Testosterone supplementation in socially stressed tree shrews prevented a decrease in hippocampal 5-HT(1A) receptor binding. These receptors are hypothesized to play an important role in neurogenesis in this brain structure. We designed the present experiment to test if social defeat reduces hippocampal cell proliferation and neurogenesis in rats and if testosterone supplementation can prevent this reduction. The results indicate that repeated social defeat stress on 5 successive days induces a significant drop in plasma testosterone levels in male rats and suppresses hippocampal cell proliferation 24h and 3weeks after the end of the stress period. Testosterone supplementation prevented the social stress induced drop in plasma testosterone levels. The hormone supplementation also reduced the negative effect of stress on hippocampal BrdU labeling at 3weeks post-defeat. This effect was, however, rather weak and was caused by the tendency of the hormone in itself to suppress proliferation and the failure to fully recover the proliferation rate. Survival of dentate gyrus cells that either proliferated prior to the stress period or 24h after the last defeat was not affected by the social defeats. Thus the stress-induced lowering of hippocampal cell proliferation is not likely to be caused by transient inhibition of testosterone secretion during social stress., (Copyright © 2010 Elsevier Inc. All rights reserved.)

Estrogen receptor-alpha (ERalpha), estrogen receptor-beta (ERbeta), and progestin receptor (PR) immunoreactivities are localized to extranuclear sites in the rat hippocampal formation. Because rats and mice respond differently to estradiol treatment at a cellular level, the present study examined the distribution of ovarian hormone receptors in the dorsal hippocampal formation of mice. For this, antibodies to ERalpha, ERbeta, and PR were localized by light and electron immunomicroscopy in male and female mice across the estrous cycle. Light microscopic examination of the mouse hippocampal formation showed sparse nuclear ERalpha and PR immunoreactivity (-ir) most prominently in the CA1 region and diffuse ERbeta-ir primarily in the CA1 pyramidal cell layer as well as in a few interneurons. Ultrastructural analysis additionally revealed discrete extranuclear ERalpha-, ERbeta-, and PR-ir in neuronal and glial profiles throughout the hippocampal formation. Although extranuclear profiles were detected in all animal groups examined, the amount and types of profiles varied with sex and estrous cycle phase. ERalpha-ir was highest in diestrus females, particularly in dendritic spines, axons, and glia. Similarly, ERbeta-ir was highest in estrus and diestrus females, mainly in dendritic spines and glia. Conversely, PR-ir was highest during proestrus, mostly in axons. Except for very low levels of extranuclear ERbeta-ir in mossy fiber terminals in mice, the labeling patterns in the mice for all three antibodies were similar to the ultrastructural labeling found previously in rats, suggesting that regulation of these receptors is well conserved across the two species.

The alpha7 subunit of the nicotinic acetylcholine receptor (NAchRalpha7) is one of the principal brain receptors for nicotine and is thought to be a mediator of nicotine's pro-cognitive effects. While nicotine is known to interact with the stress axis, little is known about the effect of stress or corticosteroids on the expression in the hippocampus, a brain region important to both cognition and stress reactivity. We examined the effects of chronic (21 day) restraint stress (CRS) and adrenalectomy with hormone replacement with the selective mineralocorticoid receptor (MR) agonist aldosterone, the selective glucocorticoid receptor (GR) agonist RU28,362 or corticosterone for 7 days, on the hippocampal expression of NAchRalpha7 mRNA and protein, as measured by (125)I alpha-Bungarotoxin autoradiography. We found that CRS increased the levels of NAchRalpha7 mRNA in the CA1, CA3 and dentate gyrus while levels of the protein were lowered by the same treatment. Corticosteroid replacement showed a GR specific increase in NAchRalpha7 mRNA, consistent with a corticosteroid mediated effect of CRS. While the mechanism behind these observations is as yet unclear, they may be neuroprotective against the damaging effects of CRS or an example of adaptation to the allostatic load produced by CRS., (Copyright 2010 Elsevier B.V. All rights reserved.)

Natural fluctuations in circulating estradiol are associated with behavioral changes, including severe disturbances in mood and cognition in some women. Common genetic variation in some of the molecular mediators of estradiol effects on these behaviors, in brain regions such as the hippocampus, may explain individual variation in estradiol effects on behavior. We tested whether the common human variant BDNF Val66Met interacts with estradiol in the control of hippocampal function in cycling female mice homozygous for the wild-type Val or BDNF Met variant. BDNF Met increased anxiety behavior, impaired memory, and increased expression of BDNF and its receptor TrkB in the hippocampal formation. BDNF Met also dramatically altered the fluctuation of spatial memory, hippocampal Akt phosphorylation, and PSD-95 protein expression across the estrous cycle. The variant BDNF Val66Met should therefore be considered as a strong candidate for mediating genetic differences in ovarian steroid-related behavioral changes and disorders.

The hippocampal formation is a brain region noted for its plasticity in response to stressful events and adrenal steroid hormones. Recent work has shown that chromatin remodeling in various brain regions, including the hippocampus, is associated with the effects of stress in a variety of models. We chose to examine the effects of stress, stress duration, corticosterone administration, and fluoxetine treatment on the levels of hippocampal histone H3 methylation at lysines 4, 9, and 27, marks associated, respectively, with active transcription, heterochromatin formation, and transcriptional repression. We found that acute stress increased the levels of H3K9 tri-methylation (H3K9me3) in the dentate gyrus (DG) and CA1, while it reduced levels of H3K9 mono-methylation (H3K9me1) and H3K27 tri-methylation (H3K27me3) in the same regions, and had no effect on levels of H3K4 tri-methylation (H3K4me3). Seven days of restraint stress reduced levels of H3K4me3 in the CA1 and H3K27me3 in the DG and CA1, while increasing basal levels of H3K9me3. Chronic restraint stress (CRS) for 21 days mildly increased levels of H3K4me3 and reduced H3K9me3 levels in the DG. Treatment with fluoxetine during CRS reversed the decrease in DG H3K9me3, but had no effect on the other marks. These results show a complex, surprisingly rapid, and regionally specific pattern of chromatin remodeling within hippocampus produced by stress and anti-depressant treatment that may open an avenue of understanding the interplay of stress and hippocampal gene expression, and reveal the outlines of a potential chromatin stress response that may be diminished or degraded by chronic stress.

Changes in hippocampal CA1 dendritic spine density and synaptic number across the estrous cycle in female rats correlate with increased hippocampal-dependent cognitive performance in a manner that is dependent on estrogen receptors (ERs). Two isoforms of the estrogen receptor, alpha and beta are present in the rat hippocampus and distinct effects on cognitive behavior have been described for each receptor. The present study generated a profile of synaptic proteins altered by administration of estradiol benzoate, the ERalpha selective agonist PPT (1,3,5-tris (4-hydroxyphenyl)-4-propyl-1H-pyrazole) and the ERbeta selective agonist DPN (2,3-bis (4-hydroxyphenyl) propionitrile) alone and in combination in comparison to vehicle in the CA1 region of the dorsal hippocampus. In the stratum radiatum, estradiol, DPN, and PPT increased PSD-95 and AMPA-type glutamate receptor subunit GluR1. Only DPN administration regulated expression of AMPA receptor subunits GluR2 and GluR3, increasing and decreasing levels respectively. DPN also increased GluR2 expression in the other lamina of the CA1. These results support previous reports that estradiol and isoform specific agonists differentially activate ERalpha and ERbeta to regulate protein expression. The distinct effects of DPN and PPT administration on synaptic proteins suggest that the desired therapeutic outcome of estrogen may be accomplished by using specific estrogen receptor agonists. Moreover, the effects of estradiol treatment on PSD-95 expression are consistent with a growing body of evidence that this postsynaptic protein is a key marker of estrogen action related to spine synapse formation.

Exposure to chronic stress has been found to decrease cannabinoid CB(1) receptor expression in the hippocampus; however, the specificity of this phenomenon to specific subfields of the hippocampus has not been characterized. To this extent, male Sprague-Dawley rats were exposed to 21 days of restraint stress (6 h/day), after which autoradiographical analysis of cannabinoid CB(1) receptor binding site densities were examined in the CA1, CA3 and dentate gyrus subfields of the hippocampus. Chronic stress was found to produce a significant reduction in cannabinoid CB(1) receptor binding in the dentate gyrus, while increasing cannabinoid CB(1) receptor binding in the CA3. There was no effect of chronic stress on cannabinoid CB(1) receptor binding in the CA1, or two other proximal regions, the retrosplenial cortical gyrus and the laterodorsal thalamus. Given the role of hippocampal cannabinoid CB(1) receptor activity in the maintenance of synaptic integrity and neuroplasticity in the hippocampus, these data suggest that changes in cannabinoid CB(1) receptor activity following stress may contribute to stress-induced modulation of these processes.

Progesterone's effects on hippocampus-dependent behavior and synaptic connectivity maybe mediated through the progestin receptor (PR). Although estrogen induces PR mRNA and cytosolic PR in the hippocampus, nuclear PR immunoreactivity is undetectable by light microscopy, suggesting that PR is present at extranuclear sites. To determine whether this is the case, we used immunoelectron microscopy to examine PR distribution in the hippocampal formation of proestrus rats. Ultrastructural analysis revealed that PR labeling is present in extranuclear profiles throughout the CA1 and CA3 regions and dentate gyrus, and, in contrast to light microscopic findings, in nuclei of a few pyramidal and subgranular zone cells. Most neuronal PR labeling is extranuclear and is divided between pre- and postsynaptic compartments; approximately 30% of labeled profiles were axon terminals and 30% were dendrites and dendritic spines. In most laminae, except in CA3 stratum lucidum, about 15% of PR-immunoreactive profiles were unmyelinated axons. In stratum lucidum, where the mossy fiber axons course, more than 50% of PR-labeled profiles were axonal. The remaining 25% of PR-labeled profiles were glia, some resembling astrocytes. PR labeling is strongly dependent on estrogen priming, insofar as few PR-labeled profiles were detected in ovariectomized, oil-replaced females. Synapses formed by PR-labeled terminals were predominantly asymmetric, consistent with a role for progesterone in directly regulating excitatory transmission. These findings suggest that some of progesterone's actions in the hippocampal formation may be mediated by direct and rapid actions on extranuclear PRs and that PRs are well positioned to regulate progesterone-induced changes at synapses.

Estrogens have direct effects on the brain areas controlling cognition. One of the most studied of these regions is the dorsal hippocampal formation, which governs the formation of spatial and episodic memories. In laboratory animals, most investigators report that estrogen enhances synaptic plasticity and improves performance on hippocampal-dependent cognitive behaviors. This review summarizes work conducted in our laboratory and others toward identifying estrogen's actions in the hippocampal formation, and the mechanisms for these actions. Physiologic and pharmacologic estrogen affects cognitive behavior in mammals, which may be applicable to human health and disease. The effects of estrogen in the hippocampal formation that lead to modulation of hippocampal function include effects on cell morphology, synapse formation, signaling, and excitability that have been studied in laboratory mice, rats, and primates. Finally, estrogen may signal through both nuclear and extranuclear hippocampal estrogen receptors to achieve its downstream effects.

Numerous studies have established that adolescence is marked by substantial changes in stress reactivity and hippocampal function. Glucocorticoid receptors (GRs) in the hippocampus are imperative in corticosterone-dependent gene transcription when glucocorticoid levels are relatively high, such as during periods of stress. As reported previously, in reaction to acute stress, prepubertal animals show a significantly more protracted corticosterone response compared to adults. Chronic stress, however, results in a higher peak response, but a faster return to baseline in prepubertal compared to adult animals. Thus, depending on the developmental stage and experience of the animal, the hippocampus is exposed to different concentrations and durations of corticosterone. The present set of experiments assessed the effects of acute or repeated stress on GR mRNA expression in the dorsal and ventral hippocampal formation either before or after pubertal maturation in male rats. We found that acute stress results in a significant decrease in GR mRNA in the CA1 pyramidal cell layer and dentate gyrus in the dorsal and ventral hippocampal formation of both prepubertal and adult males. In response to repeated stress, we found no differences in GR expression in either the dorsal or ventral hippocampus. Thus, despite the dramatic differences in corticosterone concentration following stress at these two developmental stages, the stress-induced changes in GR expression in the hippocampus before and after pubertal maturation were more similar than different. These data point to a dissociation between differential stress-induced corticosterone responses and regulation of hippocampal GR levels in prepubertal and adult animals., ((c) 2007 S. Karger AG, Basel)

Magnetic resonance imaging (MRI) in association with Jacobian-modulated voxel-based morphometry (VBM) was used to test for regional variation in gray matter over the menstrual cycle. T1-weighted anatomical images were acquired using a spoiled gradient recalled acquisition sequence in 21 women. Each subject was scanned twice: once during the postmenstrual late-follicular phase (Days 10-12 after onset of menses), and once during the premenstrual late-luteal phase (1-5 days before the onset of menses). Gray matter was relatively increased in the right anterior hippocampus and relatively decreased in the right dorsal basal ganglia (globus pallidus/putamen) in the postmenstrual phase. Verbal declarative memory was increased in the postmenstrual vs. premenstrual phase. This first report of human brain structural plasticity associated with the endogenous menstrual cycle extends well-established animal findings of hormone-mediated hippocampal plasticity to humans, and has implications for understanding alterations in cognition and behavior across the menstrual cycle.

Depression is associated with abnormal neuronal plasticity. AMPA receptors mediate transmission and plasticity at excitatory synapses in a manner which is positively regulated by phosphorylation at Ser831-GluR1, a CaMKII/PKC site, and Ser845-GluR1, a PKA site. Treatment with the selective serotonin (5-hydroxytryptamine; 5-HT) reuptake inhibitor fluoxetine increases P-Ser845-GluR1 but not P-Ser831-GluR1. Here, it was found that treatment with another antidepressant, tianeptine, increased P-Ser831-GluR1 in the frontal cortex and the CA3 region of hippocampus and P-Ser845-GluR1 in the CA3 region of hippocampus. A receptorome profile detected no affinity for tianeptine at any monaminergic receptors or transporters, confirming an atypical profile for this compound. Behavioural analyses showed that mice bearing point mutations at both Ser831- and Ser845-GluR1, treated with saline, exhibited increased latency to enter the centre of an open field and increased immobility in the tail-suspension test compared to their wild-type counterparts. Chronic tianeptine treatment increased open-field locomotion and reduced immobility in wild-type mice but not in phosphomutant GluR1 mice. P-Ser133-CREB was reduced in the CA3 region of hippocampus in phosphomutant mice, and tianeptine decreased P-Ser133-CREB in this region in wild-type, but not in phosphomutant, mice. Tianeptine increased P-Ser133-CREB in the CA1 region in wild-type mice but not in phosphomutant GluR1 mice. There were higher basal P-Ser133-CREB and c-fos levels in frontal and cingulate cortex in phosphomutant GluR1 mice; these changes in level were counteracted by tianeptine in a GluR1-independent manner. Using phosphorylation assays and phosphomutant GluR1 mice, this study provides evidence that AMPA receptor phosphorylation mediates certain explorative and antidepressant-like actions under basal conditions and following tianeptine treatment.

The hippocampus is a malleable brain region that responds to external agents such as hormones and stressors. Investigations that began in our laboratories with the Golgi technique and an appreciation of hippocampal neuroanatomy at the light and electron microscopic levels have led us down a path that has uncovered unexpected structural plasticity in the adult brain along with unanticipated cellular and molecular mechanisms of this plasticity and of hormone mediation of these effects. This chapter reviews the history of discoveries in our two laboratories involving the actions of estradiol and stress hormones on neuronal structure and function and then discusses the insight to hormone-brain interactions that this has engendered. These discoveries have led us to a new view of brain structural plasticity and the role and mechanism of steroid hormone action involving both genomic and non-genomic pathways. This new view is consistent with the predictions of Cajal in his book "The Structure of Ammon's horn", 1892.

Although chronic voluntary physical activity has been shown to enhance hippocampal brain-derived neurotrophic factor (BDNF) expression in animals, the effects of forced exercise on a treadmill have not been fully investigated. We assessed induction of c-fos and BDNF expression with acute exercise at different running intensities. The mRNA for c-fos, a marker for neuronal activation, was up-regulated even under low-intensity running (15 m/min), although its induction appeared to be intensity dependent. On the other hand, increases in BDNF mRNA and protein were seen only at low-intensity running. At moderate-intensity running (25 m/min) which elevated blood lactate and corticosterone levels, induction of BDNF mRNA, but not its protein, was even depressed. Our study shows the first evidence that with an acute low-intensity exercise that is minimally stressful, hippocampal activation and BDNF expression can be achieved lending support to the idea that mild exercise could yield to greater benefits in hippocampal functions compared to the more strenuous forms.

CART (Cocaine-Amphetamine-Regulated Transcript) has been shown to be regulated by corticosteroids in the hypothalamus, but its regulation by corticosteroids and stress has not been well examined in the hippocampus or the amygdala. Further, CART has been implicated in the transition to puberty. In this study we examine the effects of acute (30 min) stress on CART mRNA in prepubescent and adult rats. In addition, we examined chronic (21 day x 6 h) restraint stress upon the expression of CART mRNA in the hippocampus and the amygdala and the effects of 7 days of adrenalectomy and corticosteroid replacement upon CART expression in these regions of the adult rat brain. We found an up-regulation of CART mRNA in the central amygdala induced by acute but not chronic stress and an up-regulation in the dentate gyrus induced by chronic but not acute stress. Adrenalectomy reduced CART expression in the dentate gyrus but not the amygdala and this effect was blocked by corticosterone but not RU28,362 or aldosterone replacement, suggesting a synergism of mineralocorticoid and glucocorticoid receptors. Our data establish that CART expression is regulated by stress in a regionally and time specific manner and that CART is regulated by corticosteroid actions in the hippocampus.

Since the first finding that 17beta-estradiol (E) can regulate CA1 pyramidal cell synapse formation, subsequent studies have explored many potential E-dependent mechanisms occurring within CA1 pyramidal cells. Fewer studies have focused on E-dependent processes outside of the pyramidal cell that may influence events activity of the pyramidal cells. This review considers hippocampal interneurons, which can potently regulate the excitability of simultaneously firing pyramidal cells. In particular, we discuss neuropeptide Y (NPY) expression by these interneurons because our published findings show that NPY expression is increased by E in a subset of interneurons which coincidentally exhibit E-regulated increase in GABA synthesis and are uniquely situated anatomically such that they may regulate synaptic activity. Here we review the role of different phenotypes of CA1 interneurons, and we propose a model in which E-stimulated NPY gene expression and the release of NPY by interneurons inhibits glutamate release presynaptically and alters glutamate-dependent synaptic events in the rat hippocampus during adulthood.

Background: Exposure to stress levels of glucocorticoids produces physiological responses that are characteristic of type 2 diabetes, such as peripheral insulin resistance and impairment in insulin-stimulated trafficking of glucose transporter 4 (GLUT4) in muscle and fat. In the central nervous system, stress produces neuroanatomical and neurochemical changes in the hippocampus that are associated with cognitive impairments., Methods: In view of these observations, the current studies examined the effects of short-term (1 week) exposure of stress levels of glucocorticoids upon insulin receptor (IR) expression and signaling, including GLUT4 translocation, in the rat hippocampus., Results: One week of corticosterone (CORT) treatment produced insulin resistance in response to peripheral glucose challenge. In the hippocampus, IR expression was unchanged in CORT-treated rats as compared with vehicle-treated rats. However, insulin-stimulated phosphorylation of the IR, total Akt levels and total GLUT4 levels were reduced in CORT-treated rats when compared to controls. In addition, insulin-stimulated translocation of hippocampal GLUT4 to the plasma membrane was completely abolished in CORT-treated rats., Conclusions: These results demonstrate that in addition to eliciting peripheral insulin resistance, short-term CORT administration impairs insulin signaling in the rat hippocampus, effects that may contribute to the deleterious consequences of hypercortisolemic/hyperglycemic states observed in type 2 diabetes.

The hippocampal formation is a highly plastic brain structure that undergoes structural remodeling in response to internal and external challenges such as metabolic imbalance and repeated stress. We investigated whether the extreme alterations in metabolic status that occur during the course of hibernation in European hamsters cause structural changes in the dendritic arborizations of the CA3 pyramidal neurons and their main excitatory afferents, the mossy fiber terminals (MFT), that originate in the dentate gyrus. We report that apical, but not basal, dendritic trees of Golgi-impregnated CA3 principal neurons are significantly shorter, less branched, and less spiny in hypothermic hamsters compared with active animals. After the induction of arousal from torpor, within 2 h, the apical dendritic lengths, branching patterns, and spine density estimations returned to levels found in active, euthermic hamsters. The ultrastructure of MFT in hibernating hamsters showed a significant reduction in synaptic vesicle density and in the percentage of MFT area covered by spine profiles. Awakened hamsters showed restoration of MFT morphology to that seen in active animals. MFT of torpid animals also showed a significant increase in the percentage area of mitochondrial profiles that remained higher 3 h after induced arousal from hibernation compared with euthermic controls. Thus, the torpid/awakening cycle of the hibernating European hamster causes a rapid and reversible morphological reorganization of intrahippocampal subregions involved in information processing. The reported reductions in morphological connectivity between the dentate gyrus and the CA3 subregions could underlie the cessation of exploratory activity and spatial navigation skills during hibernation.

Early life experience can induce long-lasting changes in brain and behaviour that are opposite in direction, such as enhancement or impairment in regulation of stress response, structural and functional integrity of the hippocampus, and learning and memory. To explore how multiple early life events jointly determine developmental outcome, we investigated the combined effects of neonatal trauma (anoxia on postnatal day 1, P1) and neonatal novelty exposure (P2-21) on adult social recognition memory (3 months of age) and synaptic plasticity in the CA1 of the rat hippocampus (4.5-8 months of age). While neonatal anoxia selectively reduced post-tetanic potentiation (PTP), neonatal novel exposure selectively increased long-term potentiation (LTP). No interaction between anoxia and novelty exposure was found on either PTP or LTP. These findings suggest that the two contrasting neonatal events have selective and distinct effects on two different forms of synaptic plasticity. At the level of behaviour, the effect of novelty exposure on LTP was associated with increased social memory, and the effect of anoxia on PTP was not accompanied by changes in social memory. Such a finding suggests a bias toward the involvement of LTP over PTP in social memory. Finally, we report a surprising finding that an early behavioural measure of emotional response to a novel environment obtained at 25 days of age can predict adult LTP measured several months later. Therefore, individual differences in emotional responses present during the juvenile stage may contribute to adult individual differences in cellular mechanisms that underlie learning and memory.

Chronic restraint stress in mice affects hippocampal structure and function. Mice were subjected to daily restraint for 3 weeks, and gene expression in hippocampus was compared to controls using large-scale cDNA microarrays. We found that 444 genes were differentially expressed, and further analysis of 6 genes by real-time reverse transcription PCR confirmed that 3 of them were downregulated by stress. These 3 genes, growth factor receptor-bound protein 2 (Grb2), phosphatidylinositol-4-phosphate 5-kinase, type 1 beta (Pip5k1b), and glutathione S-transferase, pi2 (Gstp2), were also analyzed by in situ hybridization. The downregulation of Gstp2 may induce an increase of oxidative damage in the pyramidal cells of the CA1 and CA3 regions and granular layer of the dentate gyrus, leading to structural and functional damage. Those regions are affected by stress, and our results could help understand further the mechanisms involved in the occurrence of stress-related disorders.