Characterisation of the consequences of maternal immune activation on distinct cell populations in the developing rat spinal cord.

Maternal immune activation (MIA) during gestation has been implicated in the development of neurological disorders such as schizophrenia and autism. Epidemiological studies have suggested that the effect of MIA may depend on the gestational timing of the immune challenge and the region of the central nervous system (CNS) in question. This study investigated the effects of MIA with 100 µg/kg lipopolysaccharide at either Embryonic days (E)12 or E16 on the oligodendrocytes, microglia and astrocytes of the offspring spinal cord. At E16, MIA decreased the number of olig2+ and Iba-1+ cells in multiple grey and white matter regions of the developing spinal cord 5 h after injection. These decreases were not observed at postnatal day 14. In contrast, MIA at E12 did not alter Olig2+ or Iba-1+ cell number in the developing spinal cord 5 h after injection, however, Olig2+ cell number was decreased in the ventral grey matter of the P14 spinal cord. No changes were observed in glial fibrillary acidic protein (GFAP) expression at P14 following MIA at either E12 or E16. These data suggest that E16 may be a window of immediate vulnerability to MIA during spinal cord development, however, the findings also suggest that the developmental process may be capable of compensation over time. Potential changes in P14 animals following the challenge at E12 are indicative of the complexity of the effects of MIA during the developmental process.

Development of mesenteric tissues.

Mesothelial, neurovascular, lymphatic, adipose and mesenchymal tissues make up the mesentery. These tissues are pathobiologically important for numerous reasons. Collectively, they form a continuous, discrete and substantive organ. Additionally, they maintain abdominal digestive organs in position and in continuity with other systems. Furthermore, as they occupy a central position, they mediate transmission of signals between the abdominal digestive system and the remainder of the body. Despite this physiologic centrality, mesenteric tissue development has received little investigatory focus. However, recent advances in our understanding of anatomy demonstrate continuity between all mesenteric tissues, thereby linking previously unrelated studies. In this review, we examine the development of mesenteric tissue in normality and in the setting of congenital abnormalities.

Anatomy of the mesentery: Historical development and recent advances.

Recent appraisals of mesenteric anatomy clarify its structure and show a continuous and helical-shaped organ. This new model signifies a departure from the conventional model which described multiple, separate "mesenteries". Renaissance anatomists depicted the mesentery as a continuous structure. Events that led to replacement of a continuous with a fragmented model span several centuries. In effect, the scientific and clinical community has come full circle and back to the Renaissance model. Here we review the historical development of our understanding of the mesentery. We discuss how the fragmented model replaced the continuous model. Additionally, we examine factors that contributed to recent advances in mesenteric anatomy as these present new opportunities for systematic investigation.

Anatomy of the mesentery: Current understanding and mechanisms of attachment.

An understanding of the anatomy of the mesentery is necessary to undertake any appraisal of the literature on its development. The mesentery is the collection of tissues that maintain all abdominal digestive organs in position and connection with the rest of the body. Therefore, it is also necessary to detail the exact mechanisms that maintain the mesentery in position. We explore these mechanisms, including the supportive functions of structures such as Toldt's fascia, the peritoneal reflection, and vascular connections, in this article.

Communication, collaboration and contagion: "Virtualisation" of anatomy during COVID-19.

COVID-19 has generated a global need for technologies that enable communication, collaboration, education and scientific discourse whilst maintaining physical distance. University closures due to COVID-19 and physical distancing measures disrupt academic activities that previously occurred face-to-face. Restrictions placed on universities due to COVID-19 have precluded most conventional forms of education, assessment, research and scientific discourse. Anatomists now require valid, robust and easy-to-use communication tools to facilitate remote teaching, learning and research. Recent advances in communication, video conferencing and digital technologies may facilitate continuity of teaching and research activities. Examples include highly-interactive video conferencing technology, collaborative tools, social media and networking platforms. In this narrative review, we examine the utility of these technologies in supporting effective communication and professional activities of anatomists during COVID-19 and after.

An ex vivo model to quantitatively analyze cell migration in tissue.

BACKGROUND: Within the developing central nervous system, the ability of cells to migrate throughout the tissue parenchyma to reach their target destination and undergo terminal differentiation is vital to normal central nervous system (CNS) development. To develop novel therapies to treat the injured CNS, it is essential that the migratory behavior of cell populations is understood. Many studies have examined the ability of individual neurons to migrate through the developing CNS, describing specific modes of migration including locomotion and somal translocation. Few studies have investigated the mass migration of large populations of neural progenitors, particularly in the developing the spinal cord. Here, we describe a method to robustly analyze large numbers of migrating cells using a co-culture assay. RESULTS: The ex vivo tissue model promotes the survival and differentiation of co-cultured progenitor cells. Using this assay, we demonstrate that migrating neuroepithelial progenitor cells display region specific migration patterns within the dorsal and ventral spinal cord at defined developmental time points. CONCLUSIONS: The technique described here is a viable ex vivo model to quantitatively analyze cell migration and differentiation. We demonstrate the ability to detect changes in cell migration within distinct tissue region across tissue samples using the technique described here. Developmental Dynamics 247:201-211, 2018. © 2017 Wiley Periodicals, Inc.

Acute in utero exposure to lipopolysaccharide induces inflammation in the pre- and postnatal brain and alters the glial cytoarchitecture in the developing amygdala.

BACKGROUND: Maternal immune activation (MIA) is a risk factor for neurodevelopmental disorders such as autism and schizophrenia, as well as seizure development. The amygdala is a brain region involved in the regulation of emotions, and amygdalar maldevelopment due to infection-induced MIA may lead to amygdala-related disorders. MIA priming of glial cells during development has been linked to abnormalities seen in later life; however, little is known about its effects on amygdalar biochemical and cytoarchitecture integrity. METHODS: Time-mated C57BL6J mice were administered a single intraperitoneal injection of 50 µg/kg lipopolysaccharide (LPS) on embryonic day 12 (E12), and the effects of MIA were examined at prenatal, neonatal, and postnatal developmental stages using immunohistochemistry, real-time quantitative PCR, and stereological quantification of cytoarchitecture changes. RESULTS: Fetal brain expression of pro-inflammatory cytokines (IL-1ß, TNFα, and IL-6) was significantly upregulated at 4 h postinjection (E12) and remained elevated until the day of birth (P0). In offspring from LPS-treated dams, amygdalar expression of pro-inflammatory cytokines was also increased on day 7 (P7) and expression was sustained on day 40 (P40). Toll-like receptor (TLR-2, TLR-4) expression was also upregulated in fetal brains and in the postnatal amygdala in LPS-injected animals. Morphological examination of cells expressing ionized calcium-binding adaptor molecule 1 (Iba-1) and glial fibrillary acidic protein (GFAP) suggested long-term microglial activation and astrogliosis in postnatal amygdalar regions. CONCLUSIONS: Our results showed that LPS-induced MIA at E12 induces a pro-inflammatory cytokine profile in the developing fetal brain that continues up to early adulthood in the amygdala. Inflammation elicited by MIA may activate cells in the fetal brain and lead to alterations in glial (microglia and astrocyte) cells observed in the postnatal amygdala. Moreover, increased pro-inflammatory cytokines and their effects on glial subpopulations may in turn have deleterious consequences for neuronal viability. These MIA-induced changes may predispose offspring to amygdala-related disorders such as heightened anxiety and depression and also neurodevelopmental disorders, such as autism spectrum disorders.

Contribution of amygdala pathology to comorbid emotional disturbances in temporal lobe epilepsy.

The amygdala contributes to the generation and propagation of epileptiform activity in temporal lobe epilepsy (TLE). Ictal symptoms such as fear, dreamy states (déjà vu, memory flashbacks, experiential hallucinations), epigastric auras, or sympathetic outflow with cardiovascular changes are often linked to a seizure focus in the amygdala. However, the amygdala may also play a role in comorbid anxiety, depression, and other psychiatric symptoms experienced in the interictal phase, especially in pharmacoresistant TLE. The few studies available on TLE-related alterations in surgical amygdala specimens indicate loss of both excitatory spiny projection neurons as well as interneurons in nuclei with a cortex-like architecture, which may influence mechanisms of feedforward and feedback inhibition. Studies of the human amygdala indicate global alterations in the density of AMPA/kainate, metabotropic glutamate, γ-aminobutyric acid type A (GABAA ), muscarinic M2 and M3, serotonergic 5-HT1A, and adrenergic α1 receptors. Also, amygdala GABAergic and neuropeptide Y (NPY) systems affected in human TLE are both involved in antiepileptic and anxiolytic effects. Experimental and human positron emission tomography studies indicate changes in amygdala serotonergic, NPY Y1 receptor, neurokinin, and opioid systems in emotional disturbances in TLE. Of particular interest is the reduction in amygdala volume in conjunction with ictal fear, seizure focus in the amygdala, and amygdala and hippocampal sclerosis in TLE patients. In contrast, patients with interictal depression often have an intact or even enlarged amygdala and a negative MRI associated with amygdala hypometabolism, which can be associated with limbic autoimmune encephalitis. These findings suggest a differential role of TLE-related amygdala changes in ictal and interictal emotional disturbances.

Imaging oxygen in neural cell and tissue models by means of anionic cell-permeable phosphorescent nanoparticles.

Cell-permeable phosphorescent probes enable the study of cell and tissue oxygenation, bioenergetics, metabolism, and pathological states such as stroke and hypoxia. A number of such probes have been described in recent years, the majority consisting of cationic small molecule and nanoparticle structures. While these probes continue to advance, adequate staining for the study of certain cell types using live imaging techniques remains elusive; this is particularly true for neural cells. Here we introduce novel probes for the analysis of neural cells and tissues: negatively charged poly(methyl methacrylate-co-methacrylic acid)-based nanoparticles impregnated with a phosphorescent Pt(II)-tetrakis(pentafluorophenyl)porphyrin (PtPFPP) dye (this form is referred to as PA1), and with an additional reference/antennae dye poly(9,9-diheptylfluorene-alt-9,9-di-p-tolyl-9H-fluorene) (this form is referred to as PA2). PA1 and PA2 are internalised by endocytosis, result in efficient staining in primary neurons, astrocytes, and PC12 cells and multi-cellular aggregates, and allow for the monitoring of local O(2) levels on a time-resolved fluorescence plate reader and PLIM microscope. PA2 also efficiently stains rat brain slices and permits detailed O(2) imaging experiments using both one and two-photon intensity-based modes and PLIM modes. Multiplexed analysis of embryonic rat brain slices reveals age-dependent staining patterns for PA2 and a highly heterogeneous distribution of O(2) in tissues, which we relate to the localisation of specific progenitor cell populations. Overall, these anionic probes are useful for sensing O(2) levels in various cells and tissues, particularly in neural cells, and facilitate high-resolution imaging of O(2) in 3D tissue models.

Expression of neuropeptide Y1 receptors in the amygdala and hippocampus and anxiety-like behavior associated with Ammon's horn sclerosis following intrahippocampal kainate injection in C57BL/6J mice.

Damage to the amygdala is often linked to Ammon's horn sclerosis (AHS) in surgical specimens of patients suffering from temporal lobe epilepsy (TLE). Moreover, amygdalar pathology is thought to contribute to the development of anxiety symptoms frequently found in TLE. The neuropeptide Y (NPY) Y1 receptor is critical in the regulation of anxiety-related behavior and epileptiform activity in TLE. Therefore, intrahippocampal kainate (KA) injection was performed to induce AHS-associated TLE and to investigate behavioral and cytoarchitectural changes that occur in the amygdala related to Y1 receptor expression. Status epilepticus was induced by intrahippocampal KA injection in C57BL/6J mice. Anxiety-like behavior was assessed using the elevated plus maze (EPM). Pathology of hippocampus and amygdala (volume loss and gliosis) was examined in KA-injected and saline-injected controls. Y1 receptor expression was measured using immunohistochemistry and ELISA. Animal injected with KA showed increased anxiety-like behaviors and reduced risk assessment in the EPM test compared with saline-injected controls. In the ipsilateral hippocampus of KA-injected animals, CA1 ablation, granule cell dispersion, and volume reduction were accompanied by astrogliosis indicating the development of AHS. In the amygdala, a significant decrease in the volume of nuclei and numbers of neurons was observed in the ipsilateral lateral, basolateral, and central amygdalar nuclei, which was accompanied by astrogliosis. In addition, a decrease in Y1 receptor-expressing cells in the ipsilateral CA1 and CA3 sectors of the hippocampus, ipsilateral and contralateral granule cell layer of the dentate gyrus, and ipsilateral central nucleus of the amygdala was found, consistent with a reduction in Y1 receptor protein levels. Our results suggest that plastic changes in hippocampal and/or amygdalar Y1 receptor expression may negatively impact anxiety levels. Moreover, intrahippocampal KA injection can induce amygdalar damage suggesting that AHS-associated amygdala damage may contribute to behavioral alterations seen in patients with TLE.

The spatial and temporal arrangement of the radial glial scaffold suggests a role in axon tract formation in the developing spinal cord.

Radial glial cells serve diverse roles during the development of the central nervous system (CNS). In the embryonic brain, they are recognised as guidance conduits for migrating neuroblasts and as multipotent stem cells, generating both neurons and glia. While their stem cell capacities in the developing spinal cord are as yet not fully clarified, they are classically seen as a population of astrocytes precursors, before gradually disappearing as the spinal cord matures. Although the origins and lineages of CNS radial glial cells are being more clearly understood, the relationships between radial glial cells and growing white matter (WM) tracts are largely unknown. Here, we provide an in-depth description of the distribution and organisation of radial glial cell processes during the peak periods of axonogenesis in the rat spinal cord. We show that radial glial cell distribution is highly ordered in the WM from E14 to E18, when the initial patterning of axon tracts is taking place. We report that the density of radial glial cell processes is tightly conserved throughout development in the dorsal, lateral and ventral WM funiculi along the rostrocaudal axis of the spinal cord. We provide evidence that from E16 the dorsal funiculi grow within and are segregated by fascicles of processes emanating from the dorsomedial septum. The density of radial glial cells declines with the maturation of axon tracts and coincides with the onset of the radial glial cell-astrocyte transformation. As such, we propose that radial glial cells act as structural scaffolds by compartmentalising and supporting WM patterning in the spinal cord during embryonic development.

Spinal cord neuroepithelial progenitor cells display developmental plasticity when co-cultured with embryonic spinal cord slices at different stages of development.

All neurons and glial cells of the vertebrate CNS are derived from embryonic neuroepithelial progenitor cells (NEP). Distinct modes of radial neuronal migration, locomotion, and somal translocation have been described in the cerebral cortex, but less is known about the migratory behavior of neuroepithelial cells and their neuronal and glial descendants in the developing spinal cord. Here a novel spinal cord slice co-culture was developed to investigate the migration and differentiation potential of NEPs in the developing spinal cord. E12 NEPs from eGFP transgenic mouse cells were co-cultured with E12, E14, E16, and E18 organotypic spinal cord slices. Time-lapse confocal microscopy and quantitative 3D image analysis revealed that the co-cultured E12 eGFP NEP cells differentiated at a faster rate with increasing age of embryonic spinal cord slice but migrated further in younger slices. Furthermore, it revealed fast tangentially migrating cells and slower radially migrating cells undergoing locomotion and somal translocation. The ability of NEP cells to alter their migration and differentiation within embryonic microenvironments of different ages highlights their developmental plasticity and ability to respond to temporally expressed extrinsic signals.

Hypothesis testing for neural cell growth experiments using a hybrid branching process model.

Neuron branching patterns can characterize neural cell types and act as markers for neurodegenerative disease and neural development. We develop a hybrid Markovian model for neural branching that extends previously published models by (i) using a discretized gamma model to account for underdispersion in primary branching, (ii) incorporating both bifurcation and trifurcation branching events to accommodate observed data, and (iii) only requiring branch counts and not branching topology as observations, allowing larger numbers of neurons to be sampled than in previous literature. Inference for primary branching is achieved through a gamma generalized linear model. Due to incomplete data, bifurcation and trifurcation probabilities are estimated using an expectation-maximization algorithm, which is shown to give consistent estimates using simulation studies and theoretical arguments. In simulation studies, comparison of standard errors shows no significant loss of accuracy relative to when topological information is available. A unified methodology for testing hypotheses using likelihood ratio tests (LRTs) is developed. The methodology is applied to an experiment where neurons are cocultured with different treatments: growth factor (GF), hypothalamic-astroglial conditioned medium (HY), and combination. The model provides statistically adequate fit at all branching orders. All treatments cause significantly higher branching at primary and secondary orders relative to control (p-value < 0.01), but not at higher branching orders, suggesting genetic regulation by the treatments. Using a computationally feasible lower bound on the LRT, bifurcation probabilities are shown to decrease exponentially with branching order for all treatments except HY (p-value 0.03).

The development and structure of the mesentery.

The position of abdominal organs, and mechanisms by which these are centrally connected, are currently described in peritoneal terms. As part of the peritoneal model of abdominal anatomy, there are multiple mesenteries. Recent findings point to an alternative model in which digestive organs are connected to a single mesentery. Given that direct evidence of this is currently lacking, we investigated the development and shape of the entire mesentery. Here we confirm that, within the abdomen, there is one mesentery in which all abdominal digestive organs develop and remain connected to. We show that all abdominopelvic organs are organised into two, discrete anatomical domains, the mesenteric and non-mesenteric domain. A similar organisation occurs across a range of animal species. The findings clarify the anatomical foundation of the abdomen; at the foundation level, the abdomen comprises a visceral (i.e. mesenteric) and somatic (i.e. musculoskeletal) frame. The organisation at that level is a fundamental order that explains the positional anatomy of all abdominopelvic organs, vasculature and peritoneum. Collectively, the findings provide a novel start point from which to systemically characterise the abdomen and its contents.

Rod and cone photoreceptor cells produce ROS in response to stress in a live retinal explant system.

PURPOSE: The production of reactive oxygen species (ROS) can lead to oxidative stress, which is a strong contributory factor to many ocular diseases. In this study, the removal of trophic factors is used as a model system to investigate the effects of stress in the retina. The aims were to determine if both rod and cone photoreceptor cells produce ROS when they are deprived of trophic factor support and to demonstrate if the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox) enzymes are responsible for this ROS production. METHODS: Retinas were explanted from mice aged between postnatal days 8-10 and cultured overnight. The following morning, confocal microscopy combined with various fluorescent probes was used to detect the production of ROS. Each time peanut agglutinin (PNA), a cone photoreceptor marker, was used to facilitate orientation of the retina. Dihydroethidium and dihydrorhodamine 123 (DHR123) were used to determine which cells produce ROS. Subsequently, western blots of retinal serial sections were used to detect the presence of Noxs in the different retinal layers. The Nox inhibitor apocynin was then tested to determine if it altered the production of ROS within these cells. RESULTS: Live retinal explants, viewed at high magnifications using confocal microscopy, displayed an increase in the fluorescent products of dihydroethidium and DHR123 upon serum removal when compared to controls. DHR123 fluorescence, once oxidized, localized to mitochondria and was found in the same focal plane as the PNA staining. This showed that cones and rods produced ROS when stressed. Retinal serial sectioning established that the photoreceptor layer expressed Nox4, dual oxidase (Duox) 1, and Duox2 at varying levels. Finally, the Nox inhibitor apocynin decreased the burst stimulated by the stress of serum removal. CONCLUSIONS: Confocal microscopy and PNA staining allowed differentiation of cell types within the outermost layers of the retina, demonstrating that both rods and cones generated ROS in response to the stress of serum deprivation. Nox4 was the most abundantly expressed Nox in the photoreceptor layer, but Duox1 and Duox2 were also present at detectable levels, and as apocynin reduced the levels of ROS produced, this implied that these proteins may play some role in this production.

Sonic hedgehog promotes the generation of myelin proteins by transplanted oligosphere-derived cells.

The generation of large numbers of functionally relevant cells for transplantation remains central to the use of cell replacement as a therapeutic strategy for neurodegenerative diseases. In this study we have analyzed the effect of sonic hedgehog (Shh) pretreatment on the myelinating potential of transplanted oligosphere-derived cells. The retina was chosen as a model for assessing this myelinating capability because 1) there is a lack of endogenous myelin in the normal rodent retina and 2) the retinal ganglion cell (RGC) axons are receptive to myelination, once myelinating cells have access to the retinal nerve fiber layer. Initially, oligospheres were generated in the presence of B104 CM but without the addition of Shh. After transplantation, 60% of the animals developed tumors in the eye that had received the transplant and were not analyzed for the presence of myelin. In the remaining retinas, the transplanted oligosphere-derived cells were not myelin competent, as indicated by the absence of myelin proteins in the retinal nerve fiber layer. In contrast, when B104 CM oligospheres were generated in the presence of Shh, myelin proteins were found in the nerve fiber layer after transplantation. In addition, the amount of myelin proteins synthesized increased with time posttransplantation, with the majority of the nerve fiber layer immunoreactive for these proteins in some retinas after 2 months. This study has demonstrated that growth as oligospheres and endogenously derived growth/differentiation factors alone are not sufficient to induce the differentiation of B104-treated oligosphere-derived cells and that pretreating the oligospheres by growth in the presence of Shh before transplantation is essential to induce their myelinating competence.

Reelin: Diverse roles in central nervous system development, health and disease.

Over the past 20 years the structure and function of Reelin, an extracellular glycoprotein with a role in cell migration and positioning during development has been elucidated. Originally discovered in mice exhibiting a peculiar gait and hypoplastic cerebellar tissue, Reelin is secreted from Cajal-Retzius neurons during embryonic life and has been shown to act as a stop signal, guiding migrating radial neurons in a gradient-dependent manner. Reelin carries out its function by binding to the receptors, very low-density lipoprotein receptor (VLDLR) and apolipoprotein E receptor 2 (ApoER2) resulting in the phosphorylation of the intracellular protein Disabled-1 (Dab-1) which is essential for effective Reelin signaling. Abnormalities in the RELN gene can result in multiple unusual structural outcomes including disruption of cortical layers, heterotopia, polymicrogyria and lissencephaly. Recent research has suggested a potential role for Reelin in the pathogenesis of neurological diseases such as schizophrenia, autism and Alzheimer's disease. This short review will address the current understanding of the structure and function of this protein and its emerging role in the development of neurological disorders.

The Human Mesenteric Lymph Node Microbiome Differentiates Between Crohn's Disease and Ulcerative Colitis.

BACKGROUND AND AIMS: Mesenteric lymph nodes are sites in which translocated bacteria incite and progress immunological responses. For this reason, understanding the microbiome of mesenteric lymph nodes in inflammatory bowel disease is important. The bacterial profile of Crohn's disease mesenteric lymph nodes has been analysed using culture-independent methods in only one previous study. This study aimed to investigate the mesenteric lymph node microbiota from both Crohn's disease and ulcerative colitis patients. METHODS: Mesenteric lymph nodes were collected from Crohn's disease and ulcerative colitis patients undergoing resection. Total DNA was extracted from mesenteric lymph nodes and assessed for the presence of bacterial DNA [16S]. All work was completed in a sterile environment using aseptic techniques. Samples positive for 16S DNA underwent next-generation sequencing, and the identity of bacterial phyla and species were determined. RESULTS: Crohn's disease mesenteric lymph nodes had a distinctly different microbial profile to that observed in ulcerative colitis. The relative abundance of Firmicutes was greater in nodes from ulcerative colitis patients, whereas Proteobacteria were more abundant in Crohn's disease. Although species diversity was reduced in the mesenteric lymph nodes of patients with Crohn's disease, these lymph nodes contained greater numbers of less dominant phyla, mainly Fusobacteria. CONCLUSION: This study confirms that there are distinct differences between the Crohn's disease and ulcerative colitis mesenteric lymph node microbiomes. Such microbial differences could aid in the diagnosis of Crohn's disease or ulcerative colitis, particularly in cases of indeterminate colitis at time of resection, or help explain their mechanisms of development and progression.

Sexually dimorphic effects of maternal separation stress on corticotrophin-releasing factor and vasopressin systems in the adult rat brain.

Neonatal maternal separation has been widely used to model the well-established causal relationship between stress in early life and the later development of depression. As corticotrophin-releasing factor (CRF) and vasopressin (AVP) have been implicated in depression, we aimed to determine the long-term effects of maternal separation stress on these neuropeptide systems, and also to explore whether these effects are gender-dependent. Immunohistochemical staining of CRF, AVP and c-Fos was used to assess whether these neuropeptide systems were affected following an acute swim stress in male and female maternally separated rats. There was an increase in CRF-immunoreactivity (IR) (p<0.05), and an increased co-localisation of c-Fos and CRF (p<0.05) following stress, in the paraventricular nucleus of the hypothalamus (PVN) of maternally separated female rats only. We found no differences in CRF in the hypothalamus of maternally separated and control male rats. However, male maternally separated rats exhibited decreases in AVP-IR in both the non-stressed and stressed groups relative to controls (p<0.001). These data provide further evidence of the involvement of the neuropeptides CRF and AVP in the long-term maladaptive effects of maternal separation stress in early life. The enhanced CRF response to stress in MS females relative to males suggests that maternal separation stress results in a more reactive neuroendocrinological stress system in females, than in males. Furthermore, the sexually dimorphic effects of maternal separation on these neuropeptides indicate that gender is an important factor influencing the trajectory of early life stress effects on CRF and AVP systems in the brain.