In the central nervous system, angiotensin II (AngII) binds to angiotensin type 1 receptors (AT1R) to affect autonomic and endocrine functions as well as learning and memory. However, understanding the function of cells containing AT1Rs has been restricted by limited availability of specific antisera, difficulties discriminating AT1 receptor-immunoreactive cells in many brain regions and, the identification of AT1R-containing neurons for physiological and molecular studies. Here, we demonstrate that an Agtr1a bacterial artificial chromosome (BAC) transgenic mouse line that expresses type A AT1Rs (AT1aRs) identified by enhanced green fluorescent protein (EGFP) overcomes these shortcomings. Throughout the brain, AT1aR-EGFP was detected in the nuclei and cytoplasm of cells, most of which were neurons. EGFP often extended into dendritic processes and could be identified either natively or with immunolabeling of EGFP. The distribution of AT1aR-EGFP cells in brain closely corresponded to that reported for AngII binding and AT1aR protein and mRNA. In particular, AT1aR-EGFP cells were in autonomic regions (e.g., hypothalamic paraventricular nucleus, central nucleus of the amygdala, parabrachial nucleus, nuclei of the solitary tract and rostral ventrolateral medulla) and in regions involved in electrolyte and fluid balance (i.e., subfornical organ) and learning and memory (i.e., cerebral cortex and hippocampus). Additionally, dual label electron microscopic studies in select brain areas demonstrate that cells containing AT1aR-EGFP colocalize with AT1R-immunoreactivity. Assessment of AngII-induced free radical production in isolated EGFP cells demonstrated feasibility of studies investigating AT1aR signaling ex vivo. These findings support the utility of Agtr1a BAC transgenic reporter mice for future studies understanding the role of AT1 receptor containing cells in brain function.
autonomic nuclei; hypothalamus; subfornical organ; amygdala; nucleus of the solitary tract; rostral ventrolateral medulla
Natural disasters represent an increasing threat both in terms of incidence and severity as a result of climate change. Although much is known about individual responses to disasters, much less is known about the social and contextual response and how this interacts with individual trajectories in terms of mental health, wellbeing and social connectedness. The 2009 bushfires in Victoria, Australia caused much loss of life, property destruction, and community disturbance. In order to progress future preparedness, response and recovery, it is crucial to measure and understand the impact of disasters at both individual and community levels.
This study aims to profile the range of mental health, wellbeing and social impacts of the Victorian 2009 bushfires over time using multiple methodologies and involving multiple community partners. A diversity of communities including bushfire affected and unaffected will be involved in the study and will include current and former residents (at the time of the Feb 2009 fires). Participants will be surveyed in 2012, 2014 and, funding permitting, in 2016 to map the predictors and outcomes of mental health, wellbeing and social functioning. Ongoing community visits, as well as interviews and focus group discussions in 2013 and 2014, will provide both contextual information and evidence of changing individual and community experiences in the medium to long term post disaster. The study will include adults, adolescents and children over the age of 5.
Conducting the study over five years and focussing on the role of social networks will provide new insights into the interplay between individual and community factors and their influence on recovery from natural disaster over time. The study findings will thereby expand understanding of long term disaster recovery needs for individuals and communities.
Disasters; Social networks; Mental health; Epidemiologic methods; Qualitative research; Community-based participatory research
Reporting guidelines can be used to encourage standardised and comprehensive reporting of health research. In light of the global commitment to health equity, we have previously developed and published a reporting guideline for equity-focused systematic reviews (PRISMA-E 2012). The objectives of this study were to explore the utility of the equity extension items included in PRISMA-E 2012 from a systematic review author perspective, including facilitators and barriers to its use. This will assist in designing dissemination and knowledge translation strategies. We conducted a survey of systematic review authors to expose them to the new items in PRISMA-E 2012, establish the extent to which they had historically addressed those items in their own reviews, and gather feedback on the usefulness of the new items. Data were analysed using Microsoft Excel 2008 and Stata (version 11.2 for Mac). Of 151 respondents completing the survey, 18.5% (95% CI: 12.7% to 25.7%) had not heard of the PRISMA statement before, although 83.4% (95% CI: 77.5% to 89.3%) indicated that they plan to use PRISMA-E 2012 in the future, depending on the focus of their review. Most (68.9%; 95% CI: 60.8% to 76.2%) thought that using PRISMA-E 2012 would lead them to conduct their reviews differently. Important facilitators to using PRISMA-E 2012 identified by respondents were journal endorsement and incorporation of the elements of the guideline into systematic review software. Barriers identified were lack of time, word limits and the availability of equity data in primary research. This study has been the first to ‘road-test’ the new PRISMA-E 2012 reporting guideline and the findings are encouraging. They confirm the acceptability and potential utility of the guideline to assist review authors in reporting on equity in their reviews. The uptake and impact of PRISMA-E 2012 over time on design, conduct and reporting of primary research and systematic reviews should continue to be examined.
Knowledge translation strategies are an approach to increase the use of evidence within policy and practice decision-making contexts. In clinical and health service contexts, knowledge translation strategies have focused on individual behavior change, however the multi-system context of public health requires a multi-level, multi-strategy approach. This paper describes the design of and implementation plan for a knowledge translation intervention for public health decision making in local government.
Four preliminary research studies contributed findings to the design of the intervention: a systematic review of knowledge translation intervention effectiveness research, a scoping study of knowledge translation perspectives and relevant theory literature, a survey of the local government public health workforce, and a study of the use of evidence-informed decision-making for public health in local government. A logic model was then developed to represent the putative pathways between intervention inputs, processes, and outcomes operating between individual-, organizational-, and system-level strategies. This formed the basis of the intervention plan.
The systematic and scoping reviews identified that effective and promising strategies to increase access to research evidence require an integrated intervention of skill development, access to a knowledge broker, resources and tools for evidence-informed decision making, and networking for information sharing. Interviews and survey analysis suggested that the intervention needs to operate at individual and organizational levels, comprising workforce development, access to evidence, and regular contact with a knowledge broker to increase access to intervention evidence; develop skills in appraisal and integration of evidence; strengthen networks; and explore organizational factors to build organizational cultures receptive to embedding evidence in practice. The logic model incorporated these inputs and strategies with a set of outcomes to measure the intervention’s effectiveness based on the theoretical frameworks, evaluation studies, and decision-maker experiences.
Documenting the design of and implementation plan for this knowledge translation intervention provides a transparent, theoretical, and practical approach to a complex intervention. It provides significant insights into how practitioners might engage with evidence in public health decision making. While this intervention model was designed for the local government context, it is likely to be applicable and generalizable across sectors and settings.
Australia New Zealand Clinical Trials Register ACTRN12609000953235.
Knowledge translation; Evidence; Public health; Decision-making
Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-generated reactive oxygen species (ROS) are highly implicated in the development of angiotensin II (AngII)-dependent hypertension mediated in part through the hypothalamic paraventricular nucleus (PVN). This region contains vasopressin and non-vasopressin neurons that are responsive to cardiovascular dysregulation, however it is not known if ROS is generated by one or both cell-types in response to “slow pressor” infusion of AngII. We addressed this question using ROS imaging and electron microscopic dual labeling for vasopressin and p47phox, a cytoplasmic NADPH oxidase subunit requiring mobilization to membranes for the initiation of ROS production. C57BL/6 mice or vasopressin-enhanced green fluorescent protein (VP-eGFP) mice were infused systemically with saline or AngII (600 ng/kg/min; s.c.) for two weeks during which they slowly developed hypertension. Ultrastructural analysis of the PVN demonstrated p47phox immunolabeling in many glial and neuronal profiles, most of which were postsynaptic dendrites. Compared with saline, AngII recipient mice had a significant increase in p47phox immunolabeling on endomembranes just beneath the plasmalemmal surface (+42.1±11.3%; p<0.05) in non-vasopressin dendrites. In contrast, AngII infusion decreased p47phox immunolabeling on the plasma membrane (−35.5±16.5%; p<0.05) in vasopressin dendrites. Isolated non-VP-eGFP neurons from the PVN of AngII-infused mice also showed an increase in baseline ROS production not seen in VP-eGFP neurons. Our results suggest that chronic low dose AngII may offset the homeostatic control of blood pressure by differentially affecting membrane assembly of NADPH oxidase and ROS production in vasopressin and non-vasopressin neurons located within the PVN.
cardiovascular regulation; vasopressin; reactive oxygen species; electron microscopy
At the Rio Summit in 2011 on Social Determinants of Health, the global community recognized a pressing need to take action on reducing health inequities. This requires an improved evidence base on the effects of national and international policies on health inequities. Although systematic reviews are recognized as an important source for evidence-informed policy, they have been criticized for failing to assess effects on health equity.
This article summarizes guidance on both conducting systematic reviews with a focus on health equity and on methods to translate their findings to different audiences. This guidance was developed based on a series of methodology meetings, previous guidance, a recently developed reporting guideline for equity-focused systematic reviews (PRISMA-Equity 2012) and a systematic review of methods to assess health equity in systematic reviews.
We make ten recommendations for conducting equity-focused systematic reviews; and five considerations for knowledge translation. Illustrative examples of equity-focused reviews are provided where these methods have been used.
Implementation of the recommendations in this article is one step toward monitoring the impact of national and international policies and programs on health equity, as recommended by the 2011 World Conference on Social Determinants of Health.
Health Equity; Evidence Synthesis; Knowledge Translation; Systematic Reviews
Neighboring genes are often coordinately expressed within cis-regulatory modules, but evidence that nonparalogous genes share functions in mammals is lacking. Here, we report that mutation of either TMEM138 or TMEM216 causes a phenotypically indistinguishable human ciliopathy, Joubert syndrome. Despite a lack of sequence homology, the genes are aligned in a head-to-tail configuration and joined by chromosomal rearrangement at the amphibian-to-reptile evolutionary transition. Expression of the two genes is mediated by a conserved regulatory element in the noncoding intergenic region. Coordinated expression is important for their interdependent cellular role in vesicular transport to primary cilia. Hence, during vertebrate evolution of genes involved in ciliogenesis, nonparalogous genes were arranged to a functional gene cluster with shared regulatory elements.
Community-based programs aimed at improving cooking skills, cooking confidence and individual eating behaviours have grown in number over the past two decades. Whilst some evidence exists to support their effectiveness, only small behavioural changes have been reported and limitations in study design may have impacted on results.
This paper describes the first evaluation of the Jamie Oliver Ministry of Food Program (JMoF) Australia, in Ipswich, Queensland. JMoF Australia is a community-based cooking skills program open to the general public consisting of 1.5 hour classes weekly over a 10 week period, based on the program of the same name originating in the United Kingdom.
A mixed methods study design is proposed. Given the programmatic implementation of JMoF in Ipswich, the quantitative study is a non-randomised, pre-post design comparing participants undergoing the program with a wait-list control group. There will be two primary outcome measures: (i) change in cooking confidence (self-efficacy) and (ii) change in self-reported mean vegetable intake (serves per day). Secondary outcome measures will include change in individual cooking and eating behaviours and psycho-social measures such as social connectedness and self-esteem. Repeated measures will be collected at baseline, program completion (10 weeks) and 6 months follow up from program completion. A sample of 250 participants per group will be recruited for the evaluation to detect a mean change of 0.5 serves a day of vegetables at 80% power (0.5% significance level). Data analysis will assess the magnitude of change of these variables both within and between groups and use sub group analysis to explore the relationships between socio-demographic characteristics and outcomes.
The qualitative study will be a longitudinal design consisting of semi-structured interviews with approximately 10-15 participants conducted at successive time points. An inductive thematic analysis will be conducted to explore social, attitudinal and behavioural changes experienced by program participants.
This evaluation will contribute to the evidence of whether cooking programs work in terms of improving health and wellbeing and the underlying mechanisms which may lead to positive behaviour change.
Australian and New Zealand Trial registration number: ACTRN12611001209987.
The endogenous hippocampal opioid systems are implicated in learning associated with drug use. Recently, we showed that ovarian hormones regulate enkephalin levels in the mossy fiber pathway. This pathway overlaps with parvalbumin (PARV)-basket interneurons that contain the enkephalin-activated mu opioid receptors (MORs) and are important for controlling the “temporal timing” of granule cells. Here, we evaluated the influence of ovarian steroid on the trafficking of MORs in PARV interneurons. Two groups of female rats were analyzed: cycling rats in proestrus (relatively high estrogens) or diestrus; and ovariectomized rats euthanized 6, 24 or 72 hr after estradiol benzoate (10μg, s.c.) administration. Dorsal hippocampal sections were dually immunolabeled for MORs and PARV and examined by light and electron microscopy. As in males, in females MOR-immunoreactivity (-ir) was in numerous PARV-labeled perikarya, dendrites and terminals in the dentate hilar region. Variation in ovarian steroid levels altered the subcellular distribution of MORs in PARV-labeled dendrites but not terminals. In normal cycling rats, MOR-gold particles on the plasma membrane of small PARV-labeled dendrites (area < 1μm2) had higher density in proestrus rats than in diestrus rats. Likewise, in ovariectomized rats MORs showed higher density on the plasma membrane of small PARV-labeled dendrites 72 hrs after estradiol exposure. The number of PARV-labeled cells was not affected by estrous cycle phase or estrogen levels. These results demonstrate that estrogen levels positively regulate the availability of MORs on GABAergic interneurons in the dentate gyrus, suggesting cooperative interaction between opioids and estrogens in modulating principal cell excitability.
estrogen; estrous cycle; ovariectomy; hippocampus; endogenous opioids
From its origins in how the brain controls the endocrine system via the hypothalamus and pituitary gland, neuroendocrinology has evolved into a science that now includes hormone action on many aspects of brain function. These actions involve the whole central nervous system and not just the hypothalamus. Advances in our understanding of cellular and molecular actions of steroid hormones have gone beyond the important cell nuclear actions of steroid hormone receptors to include signaling pathways that intersect with other mediators such as neurotransmitters and neuromodulators. This has, in turn, broadened the search for and identification of steroid receptors to include non-nuclear sites in synapses, dendrites, mitochondria and glial cells, as well as cell nuclei. The study of estrogen receptors and estrogen actions on processes related to cognition, mood, autonomic regulation, pain and neuroprotection, among other functions, has led the way in this new view of hormone actions on the brain. In this review we summarize past and current work in our laboratory on this topic. This exciting and growing field involving many laboratories continues to reshape our ideas and approaches to neuroendocrinology both at the bench and the bedside.
estrogens; progesterone; rapid non-genomic actions; hippocampus; cognition; mood; autonomic regulation
Estradiol affects hippocampal-dependent spatial memory and underlying structural and electrical synaptic plasticity in female mice and rats. Using estrogen receptor (ER) alpha and beta knockout mice and wild-type littermates, we investigated the role of ERs in estradiol effects on multiple pathways important for hippocampal plasticity and learning. Six hours of estradiol administration increased immunoreactivity for phosphorylated Akt throughout the hippocampal formation, while 48 hours of estradiol increased immunoreactivity for phosphorylated TrkB receptor. Estradiol effects on phosphorylated Akt and TrkB immunoreactivities were abolished in ER alpha and ER beta knockout mice. Estradiol also had distinct effects on immunoreactivity for PSD-95 and BDNF mRNA in ER alpha and beta knockout mice. Thus, estradiol acts through both ERs alpha and beta in several subregions of the hippocampal formation. The different effects of estradiol at 6 and 48 hours indicate that several mechanisms of estrogen receptor signaling contribute to this female hormone’s influence on hippocampal synaptic plasticity. By further delineating these mechanisms, we will better understand and predict the effects of endogenous and exogenous ovarian steroids on mood, cognition, and other hippocampal-dependent behaviors.
estrogen receptor; estrogen; synaptic plasticity; hippocampus
Alzheimer’s disease (AD) is characterised, not only by cognitive deficits and neuropathological changes, but also by several non-cognitive behavioural symptoms that can lead to a poorer quality of life. Circadian disturbances in core body temperature and physical activity are reported in AD patients, although the cause and consequences of these changes are unknown. We therefore characterised circadian patterns of body temperature and activity in male triple transgenic AD mice (3xTgAD) and non-transgenic (Non-Tg) control mice by remote radiotelemetry. At 4 months of age, daily temperature rhythms were phase advanced and by 6 months of age an increase in mean core body temperature and amplitude of temperature rhythms were observed in 3xTgAD mice. No differences in daily activity rhythms were seen in 4- to 9-month-old 3xTgAD mice, but by 10 months of age an increase in mean daily activity and the amplitude of activity profiles for 3xTgAD mice were detected. At all ages (4–10 months), 3xTgAD mice exhibited greater food intake compared with Non-Tg mice. The changes in temperature did not appear to be solely due to increased food intake and were not cyclooxygenase dependent because the temperature rise was not abolished by chronic ibuprofen treatment. No β-amyloid (Aβ) plaques or neurofibrillary tangles were noted in the hypothalamus of 3xTgAD mice, a key area involved in temperature regulation, although these pathological features were observed in the hippocampus and amygdala of 3xTgAD mice from 10 months of age. These data demonstrate age-dependent changes in core body temperature and activity in 3xTgAD mice that are present before significant AD-related neuropathology and are analogous to those observed in AD patients. The 3xTgAD mouse might therefore be an appropriate model for studying the underlying mechanisms involved in non-cognitive behavioural changes in AD.
Vivian Welch and colleagues present consensus-based guidelines for reporting equity-focused systematic reviews, the PRISMA-Equity extension.
Alzheimer's disease (AD) is associated with non-cognitive symptoms such as changes in feeding behaviour that are often characterised by an increase in appetite. Increased food intake is observed in several mouse models of AD including the triple transgenic (3×TgAD) mouse, but the mechanisms underlying this hyperphagia are unknown. We therefore examined feeding behaviour in 3×TgAD mice and tested their sensitivity to exogenous and endogenous satiety factors by assessing food intake and activation of key brain regions. In the behavioural satiety sequence (BSS), 3×TgAD mice consumed more food after a fast compared to Non-Tg controls. Feeding and drinking behaviours were increased and rest decreased in 3×TgAD mice, but the overall sequence of behaviours in the BSS was maintained. Exogenous administration of the satiety factor cholecystokinin (CCK; 8–30 µg/kg, i.p.) dose-dependently reduced food intake in Non-Tg controls and increased inactive behaviour, but had no effect on food intake or behaviour in 3×TgAD mice. CCK (15 µg/kg, i.p.) increased c-Fos protein expression in the supraoptic nucleus of the hypothalamus, and the nucleus tractus solitarius (NTS) and area postrema of the brainstem to the same extent in Non-Tg and 3×TgAD mice, but less c-Fos positive cells were detected in the paraventricular hypothalamic nucleus of CCK-treated 3×TgAD compared to Non-Tg mice. In response to a fast or a period of re-feeding, there was no difference in the number of c-Fos-positive cells detected in the arcuate nucleus of the hypothalamus, NTS and area postrema of 3×TgAD compared to Non-Tg mice. The degree of c-Fos expression in the NTS was positively correlated to food intake in Non-Tg mice, however, this relationship was absent in 3×TgAD mice. These data demonstrate that 3×TgAD mice show increased feeding behaviour and insensitivity to satiation, which is possibly due to defective gut-brain signalling in response to endogenous satiety factors released by food ingestion.
There are an increasing number of children in Australia growing up with same-sex attracted parents. Although children from same-sex parent families do in general perform well on many psychosocial measures recent research is beginning to consider some small but significant differences when these children are compared with children from other family backgrounds. In particular studies suggest that there is an association between the stigma that same-sex parent families experience and child wellbeing. Research to date lacks a holistic view with the complete physical, mental and social wellbeing of children not yet addressed. In addition, most studies have focused only on families with lesbian parents and have studied only small numbers of children.
The Australian Study of Child Health in Same-Sex Families (ACHESS) is a national study that aims to determine the complete physical, mental and social wellbeing of Australian children under the age 18 years with at least one parent who self identifies as being same-sex attracted. There will be a particular focus on the impact that stigma and discrimination has on these families. Parent and child surveys will be used to collect data and will be available both online and in paper form. Measures have been chosen whenever possible that have sound conceptual underpinnings, robust psychometric properties and Australian normative data, and include the Child Health Questionnaire (CHQ), the Strengths and Difficulties Questionnaire (SDQ) and the Kessler Psychological Distress Scale (K10).
ACHESS aims to be the largest study of its kind and will for the first time produce a detailed quantitative analysis of Australian children with same-sex attracted parents. By inviting participants to take part in further research it will also establish a valuable cohort of children, and their families, to launch future waves of research that will help us better understand the health and wellbeing of children with same-sex attracted parents.
Child health; Gay parents; Lesbian parents; Same-sex parents; Homosexuality; Sexual orientation
Often new arrivals from refugee backgrounds have experienced poor health and limited access to healthcare services. The maternal and child health (MCH) service in Victoria, Australia, is a joint local and state government operated, cost-free service available to all mothers of children aged 0–6 years. Although well-child healthcare visits are useful in identifying health issues early, there has been limited investigation in the use of these services for families from refugee backgrounds. This study aims to explore experiences of using MCH services, from the perspective of families from refugee backgrounds and service providers.
We used a qualitative study design informed by the socioecological model of health and a cultural competence approach. Two geographical areas of Melbourne were selected to invite participants. Seven focus groups were conducted with 87 mothers from Karen, Iraqi, Assyrian Chaldean, Lebanese, South Sudanese and Bhutanese backgrounds, who had lived an average of 4.7 years in Australia (range one month-18 years). Participants had a total of 249 children, of these 150 were born in Australia. Four focus groups and five interviews were conducted with MCH nurses, other healthcare providers and bicultural workers.
Four themes were identified: facilitating access to MCH services; promoting continued engagement with the MCH service; language challenges; and what is working well and could be done better. Several processes were identified that facilitated initial access to the MCH service but there were implications for continued use of the service. The MCH service was not formally notified of new parents arriving with young children. Pre-arranged group appointments by MCH nurses for parents who attended playgroups worked well to increase ongoing service engagement. Barriers for parents in using MCH services included access to transportation, lack of confidence in speaking English and making phone bookings. Service users and providers reported that continuity of nurse and interpreter is preferred for increasing client-provider trust and ongoing engagement.
Although participants who had children born in Melbourne had good initial access to, and experience of, using MCH services, significant barriers remain. A systems-oriented, culturally competent approach to service provision would improve the service utilisation experience for parents and providers, including formalising links and notifications between settlement services and MCH services.
Refugee; Maternal and child health; Access to health services; Cultural competence
Rapid actions of estrogens were first described over 40 years ago. However, the importance of rapid estrogen-mediated actions in the central nervous system (CNS) has only now becoming apparent. Several lines of evidence demonstrate that rapid estrogen-mediated signaling elicits potent effects on molecular and cellular events, resulting in the fine-tuning of neuronal circuitry. At an ultrastructural level, the details of estrogen receptor localization and how these are regulated by the circulating hormone and age, are now becoming evident. Furthermore, the mechanisms that allow membrane-associated estrogen receptors to couple with intracellular signaling pathways are also now being revealed. Elucidation of complex actions of rapid estrogen-mediated signaling on synaptic proteins, connectivity and synaptic function in pyramidal neurons has demonstrated that this neurosteroid engage specific mechanisms in different areas of the brain. The regulation of synaptic properties most likely underlies the ‘fine-tuning’ of neuronal circuitry. This in turn may influence how learned behaviors are encoded by different circuitry in male and female subjects. Importantly, as estrogens have been suggested as potential treatments of a number of disorders of the CNS, advancements in our understanding of rapid estrogen signaling in the brain will serve to aid in the development of potential novel estrogen-based treatments.
Opioids play a critical role in hippocampally dependent behavior and plasticity. In the hippocampal formation, mu opioid receptors (MOR) are prominent in parvalbumin (PARV) containing interneurons. Previously we found that gonadal hormones modulate the trafficking of MORs in PARV interneurons. Although sex differences in response to stress are well documented, the point at which opioids, sex and stress interact to influence hippocampal function remains elusive. Thus, we used quantitative immunocytochemistry in combination with light and electron microscopy for the phosphorylated MOR at the SER375 carboxy-terminal residue (pMOR) in male and female rats to assess these interactions. In both sexes, pMOR-immunoreactivity (ir) was prominent in axons and terminals and in a few neuronal somata and dendrites, some of which contained PARV in the mossy fiber pathway region of the dentate gyrus (DG) hilus and CA3 stratum lucidum. In unstressed rats, the levels of pMOR-ir in the DG or CA3 were not affected by sex or estrous cycle stage. However, immediately following 30 minutes of acute immobilization stress (AIS), males had higher levels of pMOR-ir whereas females at proestrus and estrus (high estrogen stages) had lower levels of pMOR-ir within the DG. In contrast, the number and types of neuronal profiles with pMOR-ir were not altered by AIS in either males or proestrus females. These data demonstrate that although gonadal steroids do not affect pMOR levels at resting conditions, they are differentially activated both pre- and post-synaptic MORs following stress. These interactions may contribute to the reported sex differences in hippocampally dependent behaviors in stressed animals.
opioids; sex differences; estrogens; mossy fiber pathway
Current rodent models of menopause fail to adequately recapitulate the menopause transition. The intact aging model fails to achieve very low estrogen levels, and the ovariectomy model lacks a perimenopause phase. A new rodent model of Accelerated Ovarian Failure (AOF) successfully replicates human perimenopause and postmenopause, including estrous acyclicity and fluctuating, followed by undetectable, estrogen levels, and allows for the dissociation of the effects of hormone levels from the effects of aging. In this model, an ovotoxic chemical, 4-vinylcyclohexene diepoxide (VCD), selective for primary and primordial follicles, is injected intraperitonelly in animals for 15 days. As the mature follicle population is depleted through natural cycling, ovarian failure follows increasing periods of acyclity. Administered at low doses, VCD specifically causes apoptotic cell death of primordial follicles but does not affect other peripheral tissues, including the liver and spleen, nor does it cross the blood-brain barrier. In addition to reducing confounds associated with genetic and surgical manipulations, the AOF model maintains the presence of ovarian tissue which importantly parallels to the menopause transition in humans. The VCD injection procedure can be applied to studies using transgenic or knock-out mice strains, or in other disease-state models (e.g., ischemia, atherosclerosis, or diabetes). This AOF model of menopause will generate new insights into women's health particularly in determining the critical periods (i.e., a window of opportunity) during perimenopause for restoring ovarian hormones for the most efficacious effect on memory and mood disorders as well as other menopausal symptoms.
estrogen; hormone replacement therapy; perimenopause
Estradiol (E) mediates increased synaptogenesis in the hippocampal CA1 stratum radiatum (sr) and enhances memory in young and some aged female rats, depending on dose and age. Young females rats express more estrogen receptor α (ERα) immunolabeling in CA1sr spine synapse complexes than aged rats and ERα regulation is E sensitive in young but not aged rats. The current study examined whether estrogen receptor β (ERβ) expression in spine synapse complexes may be altered by age or E treatment. Young (3–4 months) and aged (22–23 months) female rats were ovariectomized 7 days prior to implantation of silastic capsules containing either vehicle (cholesterol) or E (10% in cholesterol) for 2 days. ERβ immunoreactivity (ir) in CA1sr was quantitatively analyzed using post-embedding electron microscopy. ERβ-ir was more prominent postsynaptically than presynaptically and both age and E treatment affected its synaptic distribution. While age decreased the spine synaptic complex localization of ERβ-ir (i.e., within 60 nm of the pre- and post-synaptic membranes), E treatment increased synaptic ERβ in both young and aged rats. In addition, the E treatment, but not age, increased dendritic shaft labeling. This data demonstrates that like ERα the levels of ERβ-ir decrease in CA1 axospinous synapses with age, however, unlike ERα the levels of ERβ-ir increase in these synapses in both young and aged rats in response to E. This suggests that synaptic ERβ may be a more responsive target to E, particularly in aged females.
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 (3–5 mo old), middle-aged (9–12 mo old), and aged (about 22 mo old) female rats were ovariectomized for 4 weeks and then subcutaneously implanted with a silastic capsule containing vehicle or 17β-estradiol. After 48 hours, rats were subcutaneously injected with progesterone or vehicle and sacrificed one 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.
aging; estrogen; hippocampus; opioids; synaptic protein
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 BDNF expression and hippocampal excitability. Using electron microscopy, we localized TrkB phosphorylated at tyrosine 816 in the hippocampal formation of male mice 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 and 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.
Neurotrophin; Estrogen [Estradiol]; Estrous; Hippocampus; Receptor; Synaptic plasticity; BDNF
Childhood mental health problems are highly prevalent, experienced by one in five children living in socioeconomically disadvantaged families. Although childcare settings, including family day care are ideal to promote children's social and emotional wellbeing at a population level in a sustainable way, family day care educators receive limited training in promoting children's mental health. This study is an exploratory wait-list control cluster randomised controlled trial to test the appropriateness, acceptability, cost, and effectiveness of "Thrive," an intervention program to build the capacity of family day care educators to promote children's social and emotional wellbeing. Thrive aims to increase educators' knowledge, confidence and skills in promoting children's social and emotional wellbeing.
This study involves one family day care organisation based in a low socioeconomic area of Melbourne. All family day care educators (term used for registered carers who provide care for children for financial reimbursement in the carers own home) are eligible to participate in the study. The clusters for randomisation will be the fieldworkers (n = 5) who each supervise 10-15 educators. The intervention group (field workers and educators) will participate in a variety of intervention activities over 12 months, including workshops; activity exchanges with other educators; and focused discussion about children's social and emotional wellbeing during field worker visits. The control group will continue with their normal work practice. The intervention will be delivered to the intervention group and then to the control group after a time delay of 15 months post intervention commencement. A baseline survey will be conducted with all consenting educators and field workers (n = ~70) assessing outcomes at the cluster and individual level. The survey will also be administered at one month, six months and 12 months post-intervention commencement. The survey consists of questions measuring perceived levels of knowledge, confidence and skills in promoting children's social and emotional wellbeing. As much of this intervention will be delivered by field workers, field worker-family day care educator relationships are key to its success and thus supervisor support will also be measured. All educators will also have an in-home quality of care assessment at baseline, one month, six months and 12 months post-intervention commencement. Process evaluation will occur at one month, six months and 12 months post-intervention commencement. Information regarding intervention fidelity and economics will also be assessed in the survey.
A capacity building intervention in child mental health promotion for family day care is an essential contribution to research, policy and practice. This initiative is the first internationally, and essential in building an evidence base of interventions in this extremely policy-timely setting.
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In the brain, estrogen receptor β (ERβ) plays important roles in autonomic functions, stress reactivity and learning and memory processes. However, understanding the function of ERβ has been restricted by the limited availability of specific antisera, by difficulties discriminating the discrete localization of ERβ-immunoreactivity (ir) at the light microscopic level in many brain regions and the identification of ERβ-containing neurons in neurophysiological and molecular studies. Here, we demonstrate that a Esr2 bacterial artificial chromosome (BAC) transgenic mouse line that expresses ERβ identified by enhanced green fluorescent protein (EGFP) overcomes these shortcomings. Throughout the brain, ERβ-EGFP was detected in the nuclei and cytoplasm of cells, the majority of which resembled neurons. EGFP often extended into dendritic processes and could be identified either natively or following intensification of EGFP using immunolabeling. The distribution of ERβ-EGFP cells in brain closely corresponded to that reported for ERβ protein and mRNA. In particular, ERβ-EGFP cells were found in autonomic brain regions (i.e., hypothalamic paraventricular nucleus, rostral ventrolateral medulla and nucleus of the solitary tract), in regions associated with anxiety and stress behaviors (i.e., bed nucleus of the stria terminalis, amygdala, periaqueductal gray, raphe and parabrachial nuclei) and in regions involved in learning and memory processes (i.e., basal forebrain, cerebral cortex and hippocampus). Additionally, dual label light and electron microscopic studies in select brain areas demonstrate that cell containing ERβ-EGFP colocalize with both nuclear and extranuclear ERβ-immunoreactivity. These findings support the utility of Esr2 BAC transgenic reporter mice for future studies understanding the role of ERβ in CNS function.
autonomic nuclei; amygdala; raphe nuclei; hippocampus; cerebral cortex; hypothalamus
Estrogen receptor-α (ERα), -β (ERβ) and progestin receptor (PR) immunoreactivities are localized to extranuclear sites in the rat hippocampal formation. Since 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 ERα, ERβ, 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 ERα–, and PR-immunoreactivity (-ir) most prominent in the CA1 region and diffuse ERβ-ir primarily in the CA1 pyramidal cell layer as well as in a few interneurons. Ultrastructural analysis additionally revealed discrete extranuclear ERα-, ERβ- and PR-ir in neuronal and glial profiles throughout the hippocampal formation. While extranuclear profiles were detected in all animal groups examined, the amount and types of profiles varied with sex and estrous cycle phase. ERα-ir was highest in diestrus females, particularly in dendritic spines, axons and glia. Similarly, ERβ-ir was highest in estrus and diestrus females, mainly in dendritic spines and glia. Conversely, PR-ir was highest during proestrus, and mostly in axons. Except for very low levels of extranuclear ERβ-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.
electron microscopy; estrogen receptor alpha; estrogen receptor beta; extranuclear steroid receptors; axons; dendrites