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Although antagonist drugs are receiving increasing attention in the treatment of opioid withdrawal, the mechanisms of interaction of opiate agonists and antagonists remain largely to be investigated. We focused on the effects of very low quantities of opiate antagonists, following the clinical indication of their potential utility in detoxification. Upon reviewing the evidence on the administration of small doses of naloxone and naltrexone in the presence of agonist drugs, the effects of low-dose naltrexone during opiate administration and withdrawal are described. The application of a translational methodology allowed completing the clinical design with behavioral and cellular information obtained from a specifically developed animal model. The initial results indicate that low doses of naltrexone may help reducing the manifestation of opioid withdrawal, offer suggestions for further investigations and confirm the utility of a translational research approach to the clinical neurobiology of drug addiction. [ABSTRACT FROM AUTHOR]

Background: Despite some reports of cardiometabolic disorders associated with the risk of Alzheimer's disease (AD), limited studies have been conducted to examine the association between excessive sugar intake (a risk factor for cardiometabolic disorders) and AD risk., Aim: The purpose of our study was to evaluate if excessive sugar intake has a significant long-term effect on the risk of AD., Methods: A population sample of 37,689 participants, who enrolled in the United States (US) Women's Health Initiative - Dietary Modification Trial (WHI-DM) in 1993-2005 and its extended observational follow-up study through 1 March 2019, were analyzed. Dietary sugar intake was measured using food frequency questionnaires. AD was classified by reports using a standard questionnaire. A dietary pattern that explained the maxima variations in sugar intake was constructed using reduced rank regression (RRR) technique. Associations of RRR dietary pattern scores and sugar intake (g/day) by quartiles (Q1 through Q4) with AD risk were examined using Cox proportional hazards regression analysis with adjusting for key covariates., Results: During a mean follow-up of 18.7 years, 4586 participants reported having incident AD. The total incidence rate (95% confidence interval [CI]) of AD was 6.5 (6.3-6.7) per 1000 person-years (PYs). The incidence rates (95% CI) of AD by total sugar intake were 6.2 (5.8-6.6), 6.4 (6.0-6.8), 6.6 (6.3-7.0), and 6.9 (6.5-7.3) per 1000 PYs among those in quartiles (Q) 1 to Q4 (toward higher sugar consumption) of total sugar intake, respectively (test for trend of AD incident rates, p  < 0.001). Individuals in Q4 of total sugar intake had a 1.19 higher risk of incident AD than those in Q1 (hazard ratio [HR] = 1.19, 95% CI: 1.05-1.34, p  = 0.01). An estimated increase of 10 g/day in total sugar intake (about 2.4 teaspoons) was associated with an increased AD risk by 1.3-1.4%. Of six subtypes of sugar intake, lactose was significantly associated with AD risk., Conclusions: Our study indicates that excessive total sugar intake was significantly associated with AD risk in women. Of six subtypes of sugar intake, lactose had a stronger impact on AD risk.

Alzheimer's disease is a progressive neurodegenerative disorder characterized histologically in postmortem human brains by the presence of dense protein accumulations known as amyloid plaques and tau tangles. Plaques and tangles develop over decades of aberrant protein processing, post-translational modification, and misfolding throughout an individual's lifetime. We present a foundation of evidence from the literature that suggests chronic stress is associated with increased disease severity in Alzheimer's patient populations. Taken together with preclinical evidence that chronic stress signaling can precipitate cellular distress, we argue that chronic psychological stress renders select circuits more vulnerable to amyloid- and tau- related abnormalities. We discuss the ongoing investigation of systemic and cellular processes that maintain the integrity of protein homeostasis in health and in degenerative conditions such as Alzheimer's disease that have revealed multiple potential therapeutic avenues. For example, the endogenous cannabinoid system traverses the central and peripheral neural systems while simultaneously exerting anti-inflammatory influence over the immune response in the brain and throughout the body. Moreover, the cannabinoid system converges on several stress-integrative neuronal circuits and critical regions of the hypothalamic-pituitary-adrenal axis, with the capacity to dampen responses to psychological and cellular stress. Targeting the cannabinoid system by influencing endogenous processes or exogenously stimulating cannabinoid receptors with natural or synthetic cannabis compounds has been identified as a promising route for Alzheimer's Disease intervention. We build on our foundational framework focusing on the significance of chronic psychological and cellular stress on the development of Alzheimer's neuropathology by integrating literature on cannabinoid function and dysfunction within Alzheimer's Disease and conclude with remarks on optimal strategies for treatment potential., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Shade, Ross and Van Bockstaele.)

Arousal may be understood on a spectrum, with excessive sleepiness, cognitive dysfunction, and inattention on one side, a wakeful state in the middle, and hypervigilance, panic, and psychosis on the other side. However, historically, the concepts of arousal and stress have been challenging to define as measurable experimental variables. Divergent efforts to study these subjects have given rise to several disciplines, including neurobiology, neuroendocrinology, and cognitive neuroscience. We discuss technological advancements that chronologically led to our current understanding of the arousal system, focusing on the multifaceted nucleus locus coeruleus. We share our contemporary perspective and the hypotheses of others in the context of our current technological capabilities and future developments that will be required to move forward in this area of research., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Ross and Van Bockstaele.)

Emotional arousal is one of several factors that determine the strength of a memory and how efficiently it may be retrieved. The systems at play are multifaceted; on one hand, the dopaminergic mesocorticolimbic system evaluates the rewarding or reinforcing potential of a stimulus, while on the other, the noradrenergic stress response system evaluates the risk of threat, commanding attention, and engaging emotional and physical behavioral responses. Sex-specific patterns in the anatomy and function of the arousal system suggest that sexually divergent therapeutic approaches may be advantageous for neurological disorders involving arousal, learning, and memory. From the lens of the triple network model of psychopathology, we argue that post-traumatic stress disorder and opiate substance use disorder arise from maladaptive learning responses that are perpetuated by hyperarousal of the salience network. We present evidence that catecholamine-modulated learning and stress-responsive circuitry exerts substantial influence over the salience network and its dysfunction in stress-related psychiatric disorders, and between the sexes. We discuss the therapeutic potential of targeting the endogenous cannabinoid system; a ubiquitous neuromodulator that influences learning, memory, and responsivity to stress by influencing catecholamine, excitatory, and inhibitory synaptic transmission. Relevant preclinical data in male and female rodents are integrated with clinical data in men and women in an effort to understand how ideal treatment modalities between the sexes may be different., (© 2020 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Profound transcriptional heterogeneity in anatomically adjacent single cells suggests that robust tissue functionality may be achieved by cellular phenotype diversity. Single-cell experiments investigating the network dynamics of biological systems demonstrate cellular and tissue responses to various conditions at biologically meaningful resolution. Herein, we explain our methods for gathering single cells from anatomically specific locations and accurately measuring a subset of their gene expression profiles. We combine laser capture microdissection (LCM) with microfluidic reverse transcription quantitative polymerase chain reactions (RT-qPCR). We also use this microfluidic RT-qPCR platform to measure the microbial abundance of gut contents.

A culmination of evidence from the literature points to the Locus Coeruleus (LC)-Norepinephrine system as an underappreciated and understudied area of research in the context of Alzheimer's Disease (AD). Stress is a risk factor for developing AD, and is supported by multiple clinical and preclinical studies demonstrating that amplification of the stress system disrupts cellular and molecular processes at the synapse, promoting the production and accumulation of the amyloid beta (Aβ 42 ) peptide. Stress-induced activation of the LC is mediated by corticotropin releasing factor (CRF) and CRF receptors exhibit sex-biased stress signaling. Sex differences are evident in the neurochemical, morphological and molecular regulation of LC neurons by CRF, providing a compelling basis for the higher prevalence of stress-related disorders such as AD in females. In the present study, we examined the cellular substrates for interactions between Aβ and tyrosine hydroxylase a marker of noradrenergic somatodendritic processes in the LC, and Dopamine-β-Hydroxylase (DβH) in the infralimbic medial prefrontal cortex (ILmPFC) in mice conditionally overexpressing CRF in the forebrain (CRFOE) under a Doxycycline (DOX) regulated tetO promoter. CRFOE was sufficient to elicit a redistribution of Aβ peptides in the somatodendritic processes of the LC of male and female transgenic mice, without altering total Aβ 42 protein expression levels. DOX treated groups exhibited lysosomal compartments with apparent lipofuscin and abnormal morphology, indicating potential dysfunction of these Aβ 42 -clearing compartments. In female DOX treated groups, swollen microvessels with lipid-laden vacuoles were also observed, a sign of blood-brain-barrier dysfunction. Finally, sex differences were observed in the prefrontal cortex, as females responded to DOX treatment with increased frequency of co-localization of Aβ 42 and DβH in noradrenergic axon terminals compared to vehicle treated controls, while male groups showed no significant changes. We hypothesize that the observed sex differences in Aβ 42 distribution in this model of CRF hypersignaling is based on increased responsivity of female rodent CRF R1 in the LC. Aβ 42 production is enhanced during increased neuronal activation, therefore, the excitation of DOX treated female CRFOE LC neurons projecting to the mPFC may exhibit more frequent co-localization with Aβ due to increased neuronal activity of noradrenergic neurons.

BK Ca channels, originally discovered in Drosophila melanogaster as slowpoke ( slo ), are recognized for their roles in cellular and organ physiology. Pharmacological approaches implicated BK Ca channels in cellular and organ protection possibly for their ability to modulate mitochondrial function. However, the direct role of BK Ca channels in regulating mitochondrial structure and function is not deciphered. Here, we demonstrate that BK Ca channels are present in fly mitochondria, and slo mutants show structural and functional defects in mitochondria. slo mutants display an increase in reactive oxygen species and the modulation of ROS affected their survival. We also found that the absence of BK Ca channels reduced the lifespan of Drosophila , and overexpression of human BK Ca channels in flies extends life span in males. Our study establishes the presence of BK Ca channels in mitochondria of Drosophila and ascertains its novel physiological role in regulating mitochondrial structural and functional integrity, and lifespan.

Drug-seeking in opioid dependence is due in part to the severe negative emotion associated with the withdrawal syndrome. It is well-established that negative emotional states emerge from activity in the amygdala. More recently, gut microflora have been shown to contribute substantially to such emotions. We measured gene expression in single glia and neurons gathered from the amygdala using laser capture microdissection and simultaneously measured gut microflora in morphine-dependent and withdrawn rats to investigate drivers of negative emotion in opioid withdrawal. We found that neuroinflammatory genes, notably Tnf , were upregulated in the withdrawal condition and that astrocytes, in particular, were highly active. We also observe a decreased Firmicutes to Bacteroides ratio in opioid withdrawal indicating gut dysbiosis. We speculate that these inflammatory and gut microflora changes contribute to the negative emotion experienced in opioid withdrawal that motivates dependence.

The endocannabinoid (eCB) system has been implicated in a variety of physiological functions due to abundant expression of its receptors and endogenous ligands in the central nervous system. Substantial progress has been made in understanding how the eCB system influences the brain norepinephrine (NE) system, an important neurochemical target in the continued development of new therapies for stress-induced psychiatric disorders. We, and others, have characterized the neuroanatomical, biochemical and pharmacological effects of cannabinoid receptor modulation on brain noradrenergic circuitry and defined how molecular elements of the eCB system are positioned to directly impact the locus coeruleus (LC)-prefrontal cortex pathway, a neural circuit well recognized for contributing to symptoms of hyperarousal, a key pathophysiological feature of stress-related disorders. We also described molecular and electrophysiological properties of LC noradrenergic neurons and NE release in the medial prefrontal cortex under conditions of cannabinoid type 1 receptor deletion. Finally, we identified how stress influences cannabinoid modulation of the coeruleo-cortical pathway. A number of significant findings emerged from these studies that will be summarized in the present review and have important implications for clinical studies targeting the eCB system in the treatment of stress-induced psychiatric disorders.

Stress-related psychiatric diseases are nearly twice as prevalent in women compared to men. We recently showed in male rats that the resident-intruder model of social stress differentially engages stress-related circuitry that regulates norepinephrine-containing neurons of the locus coeruleus (LC) depending on coping strategy as determined by the latency to assume a defeat posture. Here, we determined whether this social stress had similar effects in female rats. LC afferents were retrogradely labeled with Fluorogold (FG) and rats had one or five daily exposures to an aggressive resident. Sections through the nucleus paragigantocellularis (PGi), a source of enkephalin (ENK) afferents to the LC, and central nucleus of the amygdala (CeA), a source of corticotropin-releasing factor (CRF) afferents to the LC, were processed for immunocytochemical detection of c-fos, a marker of neuronal activity, FG and ENK or CRF. Like male rats, female rats defeated with a relatively short latency (SL) in response to a single resident-intruder exposure and showed significant c-fos activation of LC neurons, PGi-ENK LC afferents, and CeA-CRF-LC afferents. With repeated exposure, some rats exhibited a long latency to defeat (LL). LC neurons and CeA-CRF-LC afferents were activated in SL rats compared to control and LL, whereas PGi-ENK LC afferents were not. Conversely, in LL rats, PGi-ENK LC and CeA-CRF-LC afferents were activated compared to controls but not LC neurons. CRF type 1 receptor (CRF1) and µ-opioid receptor (MOR) expression levels in LC were decreased in LL rats. Finally, electron microscopy showed a relative increase in MOR on the plasma membrane of LL rats and a relative increase in CRF1 on the plasma membrane of SL rats. Together, these results suggest that as is the case for males, social stress engages divergent circuitry to regulate the LC in female rats depending on coping strategy, with a bias towards CRF influence in more subordinate rats and opioid influence in less subordinate rats.

Background: It is now widely recognized that endogenous, picomolar concentrations of the 42 amino acid long peptide, amyloid-β (Aβ 42 ) is secreted under normal physiological conditions and exerts important functional activity throughout neuronal intracellular compartments. Transgenic animal models that overexpress Aβ 42 and its precursor, amyloid precursor protein (APP), have not provided predictive value in testing new treatments for Alzheimer's disease (AD), resulting in failed clinical trials. While these results are discouraging, they underscore the need to understand the physiological roles of Aβ 42 and APP under normal conditions as well as at early pre- symptomatic stages of AD. New method: We describe the use of acrolein-perfusion in immunoelectron microscopy in combination with novel antibodies directed against endogenous murine Aβ 42 and APP fragments to study abnormalities in the endolysosomal system at early stages of disease. The specific requirements, limitations and advantages of novel antibodies directed against human and murine Aβ 42 , APP and APP fragments are discussed as well as parameters for ultrastructural analysis of endolysosomal compartments., Results: Novel antibodies and a detailed protocol for immunoelectron microscopy using acrolein as a fixative are described. Acrolein is shown to preserve intraneuronal Aβ 42 species, as opposed to paraformaldehyde fixed tissue, which primarily preserves membrane bound species. Comparison with existing method(s): Technology sensitive enough to detect endogenous Aβ 42 under physiological conditions has not been widely available. We describe a number of novel and highly sensitive antibodies have recently been developed that may facilitate the analysis of endogenous Aβ 42 ., Conclusions: Using novel and highly specific antibodies in combination with electron microscopy may reveal important information about the timing of aberrant protein accumulation, as well as the progression of abnormalities in the endolysosomal systems that sort and clear these peptides., (Copyright © 2018. Published by Elsevier B.V.)

Brain endocannabinoids (eCB), acting primarily via the cannabinoid type 1 receptor (CB1r), are involved in the regulation of many physiological processes, including behavioral responses to stress. A significant neural target of eCB action is the stress-responsive norepinephrine (NE) system, whose dysregulation is implicated in myriad psychiatric and neurodegenerative disorders. Using Western blot analysis, the protein expression levels of a key enzyme in the biosynthesis of the eCB 2-arachidonoylglycerol (2-AG), diacylglycerol lipase-α (DGL-α), and two eCB degrading enzymes monoacylglycerol lipase (MGL) and fatty acid amide hydrolase (FAAH) were examined in a mouse model that lacks the NE-synthesizing enzyme, dopamine β-hydroxylase (DβH-knockout, KO) and in rats treated with N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine hydrochloride (DSP-4). In the prefrontal cortex (PFC), DGL-α protein expression was significantly increased in male and female DβH-KO mice (P < 0.05) compared to wild-type (WT) mice. DβH-KO male mice showed significant decreases in FAAH protein expression compared to WT male mice. Consistent with the DβH-KO results, DGL-α protein expression was significantly increased in male DSP-4-treated rats (P < 0.05) when compared to saline-treated controls. MGL and FAAH protein expression levels were significantly increased in male DSP-4 treated rats compared to male saline controls. Finally, we investigated the anatomical distribution of MGL and FAAH in the NE containing axon terminals of the PFC using immunoelectron microscopy. MGL was predominantly within presynaptic terminals while FAAH was localized to postsynaptic sites. These results suggest that the eCB system may be more responsive in males than females under conditions of NE perturbation, thus having potential implications for sex-specific treatment strategies of stress-related psychiatric disorders.

PTSD is heterogeneous disorder that can be long lasting and often has delayed onset following exposure to a traumatic event. Therefore, it is important to take a staging approach to evaluate progression of biological mechanisms of the disease. Here, we begin to evaluate the temporal trajectory of changes following exposure to traumatic stressors in the SPS rat PTSD model. The percent of animals displaying severe anxiety on EPM increased from 17.5% at one week to 57.1% two weeks after SPS stressors, indicating delayed onset or progressive worsening of the symptoms. The LC displayed prolonged activation, and dysbalance of the CRH/NPY systems, with enhanced CRHR1 gene expression, coupled with reduced mRNAs for NPY and Y2R. In the mediobasal hypothalamus, increased CRH mRNA levels were sustained, but there was a flip in alterations of HPA regulatory molecules, GR and FKBP5 and Y5 receptor at two weeks compared to one week. Two weeks after SPS, intranasal NPY at 300 µg/rat, but not 150 µg which was effective after one week, reversed SPS triggered elevated anxiety. It also reversed SPS elicited depressive/despair symptoms and hyperarousal. Overall, the results reveal time-dependent progression in development of anxiety symptoms and molecular impairments in gene expression for CRH and NPY systems in LC and mediobasal hypothalamus by SPS. With longer time afterwards only a higher dose of NPY was effective in reversing behavioral impairments triggered by SPS, indicating that therapeutic approaches should be adjusted according to the degree of biological progression of the disorder., (Copyright © 2019. Published by Elsevier B.V.)

The locus coeruleus (LC)-norepinephrine (NE) system is a key nucleus in which endogenous opioid and stress systems intersect to regulate the stress response. LC neurons of male rats become sensitized to stress following chronic morphine administration. Whether sex dictates this pattern of opioid-induced plasticity has not been demonstrated. Delineating the neurobiological adaptations produced by chronic opioids will enhance our understanding of stress vulnerability in opioid-dependent individuals, and may reveal how stress negatively impacts addiction recovery. In the present study, the effect of chronic morphine on the subcellular distribution of mu-opioid (MOR) and CRF receptors (CRFR) was investigated in the LC of male and female rats using immunoelectron microscopy. Results showed that placebo-treated females exhibited higher MOR and CRFR cytoplasmic distribution ratio when compared to placebo-treated males. Chronic morphine exposure induced a shift in the distribution of MOR immunogold-silver particles from the plasma membrane to the cytoplasm selectively in male LC neurons. Interestingly, chronic morphine exposure induced CRFR recruitment to the plasma membrane of both male and female LC neurons. These findings provide a potential mechanism by which chronic opioid administration increases stress vulnerability in males and females via an increase in surface availability of CRFR in LC neurons. However, our results also support the notion that cellular adaptations to chronic opioids differ across the sexes as redistribution of MOR following morphine exposure was only observed in male LC neurons., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The evolution of peptidergic signaling systems in the central nervous system serves a distinct and crucial role in brain processes and function. The diversity of physiological peptides and the complexity of their regulation and secretion from the dense core vesicles (DCV) throughout the brain is a topic greatly in need of investigation, though recent years have shed light on cellular and molecular mechanisms that are summarized in this review. Here, we focus on the convergence of peptidergic systems onto the Locus Coeruleus (LC), the sole provider of norepinephrine (NE) to the cortex and hippocampus, via large DCV. As the LC-NE system is one of the first regions of the brain to undergo degeneration in Alzheimer's Disease (AD), and markers of DCV have consistently been demonstrated to have biomarker potential for AD progression, here we summarize the current literature linking the LC-NE system with DCV dysregulation and Aβ peptides. We also include neuroanatomical data suggesting that the building blocks of senile plaques, Aβ monomers, may be localized to DCV of the LC and noradrenergic axon terminals of the prefrontal cortex. Finally, we explore the putative consequences of chronic stress on Aβ production and the role that DCV may play in LC degeneration. Clinical data of immunological markers of DCV in AD patients are discussed., (Copyright © 2018. Published by Elsevier B.V.)

Cannabinoids are capable of modulating mood, arousal, cognition and behavior, in part via their effects on the noradrenergic nucleus locus coeruleus (LC). Dysregulation of LC signaling and norepinephrine (NE) efflux in the medial prefrontal cortex (mPFC) can lead to the development of psychiatric disorders, and CB1r deletion results in alterations of α2- and β1-adrenoceptors in the mPFC, suggestive of increased LC activity. To determine how CB1r deletion alters LC signaling, whole-cell patch-clamp electrophysiology was conducted in LC-NE neurons of male and female wild type (WT) and CB1r-knock out (KO) mice. CB1r deletion caused a significant increase in LC-NE excitability and input resistance in male but not female mice when compared to WT. CB1r deletion also caused adaptations in several indices of noradrenergic function. CB1r/CB2r-KO male mice had a significant increase in cortical NE levels and tyrosine hydroxylase and CRF levels in the LC compared to WT males. CB1r/CB2r-KO female mice showed a significant increase in LC α2-AR levels compared to WT females. To further probe actions of the endocannabinoid system as an anti-stress neuromediator, the effect of CB1r deletion on CRF-induced responses in the LC was investigated. The increase in LC-NE excitability observed in male and female WT mice following CRF (300 nM) bath application was not observed in CB1r-KO mice. These results indicate that cellular adaptations following CB1r deletion cause a disruption in LC-NE signaling in males but not females, suggesting underlying sex differences in compensatory mechanisms in KO mice as well as basal endocannabinoid regulation of LC-NE activity., (© 2018 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

As a neurochemical mediator of stress resilience, NPY has been shown to oppose excitatory effects of the pro-stress neuropeptide corticotropin-releasing factor (CRF). Previous studies have described the anatomical organization of NPY and CRF in the central nucleus of the amygdala, which sends CRF projections to the locus coeruleus (LC), activating LC norepinephrine release. However, the cellular substrates for interactions between NPY and CRF in the LC remain unknown. In this study, we investigated these anatomical substrates in the male rat LC, using immunocytochemistry, confocal microscopy, and immunoelectron microscopy to detect NPY and CRF, as well as CRF and Y1 or Y2 receptors (Y1R or Y2R). Immunofluorescence and electron microscopy revealed both co-localization of NPY and CRF in LC axon terminals, as well as separately labeled terminals, suggesting NPY and CRF may serve as co-transmitters in a subset of terminals. Semi-quantitative analysis showed that 32.4% of CRF-labeled terminals contained NPY, while 58.2% (152/261) of NPY-labeled terminals contained CRF. With respect to Y1R and CRF, dual immunoelectron microscopy showed that 23.3% (67/288) of CRF-labeled axon terminals directly contacted Y1R-labeled dendrites, while only 6.3% (18/288) of CRF-labeled axon terminals co-localized with Y1R. Dual immunoelectron microscopy also showed Y2R co-localized with 30.4% (103/339) CRF-labeled terminals, but only with 16.2% (55/339) of dendrites post-synaptic to CRF-labeled axon terminals in the LC. Taken together, these findings indicate multiple sites of interaction between CRF and NPY in the LC and suggest that conditions or drugs that modulate the NPY:CRF balance in the LC may promote stress resilience., (Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.)

Chronic stress is a risk factor for a number of physiological disorders including cardiovascular disease, obesity and gastrointestinal disorders, as well as psychiatric and neurodegenerative disorders. There are a number of underlying molecular and cellular mechanisms altered in the course of chronic stress, which may increase the vulnerability of individuals to develop psychiatric disorders such as depression, and neurodegenerative disorders such as Alzheimer's Disease (AD). This is evident in the influence of stress on large-scale brain networks, including the resting state Default Mode Network (DMN), the effects of stress on neuronal circuitry and architecture, and the cellular and molecular adaptations to stress, which may render individuals with stress related psychiatric disorders more vulnerable to neurodegenerative disease later in life. These alterations include decreased negative feedback inhibition of the hypothalamic pituitary axis (HPA) axis, decreased dendritic arborization and spine density in the prefrontal cortex (PFC) and hippocampus, and the release of proinflammatory cytokines, which may suppress neurogenesis and promote neuronal cell death. Each of these factors are thought to play a role in stress-related psychiatric disease as well as AD, and have been observed in clinical and post-mortem studies of individuals with depression and AD. The goal of the current review is to summarize clinical and preclinical evidence supporting a role for chronic stress as a putative link between neuropsychiatric and neurodegenerative disease. Moreover, we provide a rationale for the importance of taking a medical history of stress-related psychiatric diseases into consideration during clinical trial design, as they may play an important role in the etiology of AD in stratified patient populations., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The locus coeruleus (LC)-norepinephrine (NE) system is an understudied circuit in the context of Alzheimer's disease (AD), and is thought to play an important role in neurodegenerative and neuropsychiatric diseases involving catecholamine neurotransmitters. Understanding the expression and distribution of the amyloid beta (Aβ) peptide, a primary component of AD, under basal conditions and under conditions of NE perturbation within the coeruleo-cortical pathway may be important for understanding its putative role in pathological states. Thus, the goal of this study is to define expression levels and the subcellular distribution of endogenous Aβ with respect to noradrenergic profiles in the rodent LC and medial prefrontal cortex (mPFC) and, further, to determine the functional relevance of NE in modulating endogenous Aβ 42 levels. We report that endogenous Aβ 42 is localized to tyrosine hydroxylase (TH) immunoreactive somatodendritic profiles of the LC and dopamine-β-hydroxylase (DβH) immunoreactive axon terminals of the infralimbic mPFC (ILmPFC). Male and female naïve rats have similar levels of amyloid precursor protein (APP) cleavage products demonstrated by western blot, as well as similar levels of endogenous Aβ 42 as determined by enzyme-linked immunosorbent assay. Two models of NE depletion, DSP-4 lesion and DβH knockout (KO) mice, were used to assess the functional relevance of NE on endogenous Aβ 42 levels. DSP-4 lesioned rats and DβH-KO mice show significantly lower levels of endogenous Aβ 42 . Noradrenergic depletion did not change APP-cleavage products resulting from β-secretase processing. Thus, resultant decreases in endogenous Aβ 42 may be due to decreased neuronal activity of noradrenergic neurons, or, by decreased stimulation of adrenergic receptors which are known to contribute to Aβ 42 production by enhancing γ-secretase processing under normal physiological conditions.

Elastin deficiency causes vascular stiffening, a leading risk for hypertension and chronic kidney disease (CKD). The mechanisms mediating hypertension and/or CKD pathogenesis due to elastin deficiency are poorly understood. Using the elastin heterozygous (Eln+/-) mouse model, we tested whether renal dysfunction due to elastin deficiency occurs independently of and precedes the development of hypertension. We assessed blood pressure and renal hemodynamics in 30-day and 12-week-old male and female mice. At P30, blood pressure of Eln+/- mice was similar to wild-type controls; however, renal blood flow was lower, whereas renal vascular resistance was augmented at baseline in Eln+/- mice. At 12 weeks, renal vascular resistance remained elevated while filtration fraction was higher in male Eln+/- relative to wild-type mice. Heterozygous mice showed isolated systolic hypertension that was evident only at nighttime. Acute salt loading with 6% dietary sodium increased daytime systolic blood pressure only in male Eln+/- mice, causing a rightward shift and blunted slope of the pressure-natriuresis curve. Renal interlobar artery basal tone and myogenic response to increasing intraluminal pressure at day 10 were similar, whereas they were augmented at day 30 and at 12 weeks old in Eln+/- mice, and normalized by the AT1R blocker, candesartan. Heterozygous mice also exhibited podocyte foot process damage that persisted even when blood pressure was normalized to wild-type levels with hydralazine. Thus, elastin insufficiency triggers structural defects and abnormal remodeling of renal vascular signaling involving AT1R-mediated vascular mechanotransduction and renal hyperfiltration with increased blood pressure sensitivity to dietary sodium contributing to systolic hypertension., (Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

The noradrenergic system has been shown to play a key role in the regulation of stress responses, arousal, mood, and emotional states. Corticotropin-releasing factor (CRF) is a primary mediator of stress-induced activation of noradrenergic neurons in the nucleus locus coeruleus (LC). The endocannabinoid (eCB) system also plays a key role in modulating stress responses, acting as an "anti-stress" neuro-mediator. In the present study, we investigated the cellular sites for interactions between the cannabinoid receptor type 1 (CB1r) and CRF in the LC. Immunofluorescence and high-resolution immunoelectron microscopy showed co-localization of CB1r and CRF in both the core and peri-LC areas. Semi-quantitative analysis revealed that 44% (208/468) of CRF-containing axon terminals in the core and 35% (104/294) in the peri-LC expressed CB1r, while 18% (85/468) of CRF-containing axon terminals in the core and 6.5% (19/294) in the peri-LC were presynaptic to CB1r-containing dendrites. In the LC core, CB1r + CRF axon terminals were more frequently of the symmetric (inhibitory) type; while in the peri-LC, a majority were of the asymmetric (excitatory) type. Triple label immunofluorescence results supported the ultrastructural analysis indicating that CB1r + CRF axon terminals contained either gamma amino butyric acid or glutamate. Finally, anterograde transport from the central nucleus of the amygdala revealed that CRF-amygdalar afferents projecting to the LC contain CB1r. Taken together, these results indicate that the eCB system is poised to directly modulate stress-integrative heterogeneous CRF afferents in the LC, some of which arise from limbic sources.

A neurochemical target at which cannabinoids interact to have global effects on behavior is brain noradrenergic circuitry. Acute and repeated administration of a cannabinoid receptor synthetic agonist is capable of increasing multiple indices of noradrenergic activity. This includes cannabinoid-induced 1) increases in norepinephrine (NE) release in the medial prefrontal cortex (mPFC); 2) desensitization of cortical α2-adrenoceptor-mediated effects; 3) activation of c-Fos in brainstem locus coeruleus (LC) noradrenergic neurons; and 4) increases in anxiety-like behaviors. In the present study, we sought to examine adaptations in adrenoceptor expression and function under conditions of cannabinoid receptor type 1 (CB1r) deletion using knockout (KO) mice and compare these to wild type (WT) controls. Electrophysiological analysis of α2-adrenoceptor-mediated responses in mPFC slices in WT mice showed a clonidine-induced α2-adrenoceptor-mediated increase in mPFC cell excitability coupled with an increase in input resistance. In contrast, CB1r KO mice showed an α2-adrenoceptor-mediated decrease in mPFC cell excitability. We then examined protein expression levels of α2- and β1-adrenoceptor subtypes in the mPFC as well as TH expression in the locus coeruleus (LC) of mice deficient in CB1r. Both α2- and β1-adrenoceptors exhibited a significant decrease in expression levels in CB1r KO mice when compared to WT in the mPFC, while a significant increase in TH was observed in the LC. To better define whether the same cortical neurons express α2A-adrenoceptor and CB1r in mPFC, we utilized high-resolution immunoelectron microscopy. We localized α2A-adrenoceptors in a knock-in mouse that expressed a hemoagglutinin (HA) tag downstream of the α2A-adrenoceptor promoter. Although the α2A-adrenoceptor was often identified pre-synaptically, we observed co-localization of CB1r with α2-adrenoceptors post-synaptically in the same mPFC neurons. Finally, using receptor binding, we confirmed prior results showing that α2A-adrenoceptor is unchanged in mPFC following acute or chronic exposure to the synthetic cannabinoid receptor agonist, WIN 55,212-2, but is increased, following chronic treatment followed by a period of abstinence. Taken together, these data provide convergent lines of evidence indicating cannabinoid regulation of the cortical adrenergic system., (Copyright © 2017 Elsevier Inc. All rights reserved.)

It is well established that central nervous system norepinephrine (NE) and corticotropin-releasing factor (CRF) systems are important mediators of behavioral responses to stressors. More recent studies have defined a role for delta opioid receptors (DOPR) in maintaining emotional valence including anxiety. The amygdala plays an important role in processing emotional stimuli, and has been implicated in the development of anxiety disorders. Activation of DOPR or inhibition of CRF in the amygdala reduces baseline and stress-induced anxiety-like responses. It is not known whether CRF- and DOPR-containing amygdalar neurons interact or whether they are regulated by NE afferents. Therefore, this study sought to better define interactions between the CRF, DOPR and NE systems in the basolateral (BLA) and central nucleus of the amygdala (CeA) of the male rat using anatomical and functional approaches. Irrespective of the amygdalar subregion, dual immunofluorescence microscopy showed that DOPR was present in CRF-containing neurons. Immunoelectron microscopy confirmed that DOPR was localized to both dendritic processes and axon terminals in the BLA and CeA. Semi-quantitative dual immunoelectron microscopy analysis of gold-silver labeling for DOPR and immunoperoxidase labeling for CRF revealed that 55 % of the CRF neurons analyzed contained DOPR in the BLA while 67 % of the CRF neurons analyzed contained DOPR in the CeA. Furthermore, approximately 41 % of DOPR-labeled axon terminals targeted BLA neurons that expressed CRF while 29 % of DOPR-labeled axon terminals targeted CeA neurons that expressed CRF. Triple label immunofluorescence microscopy revealed that DOPR and CRF were co-localized in common cellular profiles that were in close proximity to NE-containing fibers in both subregions. These anatomical results indicate significant interactions between DOPR and CRF in this critical limbic region and reveal that NE is poised to regulate these peptidergic systems in the amygdala. Functional studies were performed to determine if activation of DOPR could inhibit the anxiety produced by elevation of NE in the amygdala using the pharmacological stressor yohimbine. Administration of the DOPR agonist, SNC80, significantly attenuated elevated anxiogenic behaviors produced by yohimbine as measured in the rat on the elevated zero maze. Taken together, results from this study demonstrate the convergence of three important systems, NE, CRF, and DOPR, in the amygdala and provide insight into their functional role in modulating stress and anxiety responses.

While the ability to process fermented fruits and alcohols was once an adaptive trait that improved nutrition and quality of life, the availability and prevalence of high potency alcoholic drinks has contributed to alcohol abuse disorders in a vulnerable portion of the population. Although the neural reward systems take part in the initial response to alcohol, negative reinforcement and stress, which are normally adaptive responses, can intersect to promote continued alcohol use at all stages of the addiction cycle. Eventually a point is reached where these once adaptive responses become dysregulated resulting in uncontrolled intake that constitutes a clinically important condition termed alcohol use disorder (AUD). Current research is targeted at both the behavioral and molecular adaptations in AUDs in an effort to better develop novel approaches to intervention. In this review, historical context is provided demonstrating the societal burden of alcohol use and abuse disorders. The importance of gender in the mechanism of action of alcohol is discussed. Finally, the impact of alcohol on stress-related circuitry, uncovered by preclinical research, is outlined to provide insight into potential novel pharmacological approaches to the treatment of AUD., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Recent studies demonstrate a differential trajectory for cannabinoid receptor expression in cortical and sub-cortical brain areas across postnatal development. In the present study, we sought to investigate whether chronic systemic exposure to a synthetic cannabinoid receptor agonist causes morphological changes in the structure of dendrites and dendritic spines in adolescent and adult pyramidal neurons in the medial prefrontal cortex (mPFC) and medium spiny neurons (MSN) in the nucleus accumbens (Acb). Following systemic administration of WIN 55,212-2 in adolescent (PN 37-40) and adult (P55-60) male rats, the neuronal architecture of pyramidal neurons and MSN was assessed using Golgi-Cox staining. While no structural changes were observed in WIN 55,212-2-treated adolescent subjects compared to control, exposure to WIN 55,212-2 significantly increased dendritic length, spine density and the number of dendritic branches in pyramidal neurons in the mPFC of adult subjects when compared to control and adolescent subjects. In the Acb, WIN 55,212-2 exposure significantly decreased dendritic length and number of branches in adult rat subjects while no changes were observed in the adolescent groups. In contrast, spine density was significantly decreased in both the adult and adolescent groups in the Acb. To determine whether regional developmental morphological changes translated into behavioral differences, WIN 55,212-2-induced aversion was evaluated in both groups using a conditioned place preference paradigm. In adult rats, WIN 55,212-2 administration readily induced conditioned place aversion as previously described. In contrast, adolescent rats did not exhibit aversion following WIN 55,212-2 exposure in the behavioral paradigm. The present results show that synthetic cannabinoid administration differentially impacts cortical and sub-cortical neuronal morphology in adult compared to adolescent subjects. Such differences may underlie the disparate development effects of cannabinoids on behavior.

Stress increases vulnerability to psychiatric disorders, partly by affecting brain monoamine systems, such as the locus coeruleus (LC)-norepinephrine system. During stress, LC activity is coregulated by corticotropin-releasing factor (CRF) and endogenous opioids. This study identified neural circuitry that regulates LC activity of intruder rats during the resident-intruder model of social stress. LC afferents were retrogradely labeled with Fluorogold (FG) and rats were subjected to one or five daily exposures to an aggressive resident. Sections through the nucleus paragigantocellularis (PGi) and central amygdalar nucleus (CNA), major sources of enkephalin (ENK) and CRF LC afferents, respectively, were immunocytochemically processed to detect c-fos, FG, and CRF or ENK. In response to a single exposure, intruder rats assumed defeat with a relatively short latency (SL). LC neurons, PGI-ENK LC afferents, and CNA-CRF LC afferents were activated in these rats as indicated by increased c-fos expression. With repeated stress, rats exhibited either a SL or long latency (LL) to defeat and these strategies were associated with distinct patterns of neuronal activation. In SL rats, LC neurons were activated, as were CNA-CRF LC afferents but not PGI-ENK LC afferents. LL rats had an opposite pattern, maintaining activation of PGi-ENK LC afferents but not CNA-CRF LC afferents or LC neurons. Together, these results indicate that the establishment of different coping strategies to social stress is associated with changes in the circuitry that regulates activity of the brain norepinephrine system. This may underlie differential vulnerability to the consequences of social stress that characterize these different coping strategies.

Alcohol abuse and alcoholism are major medical problems affecting both men and women. Previous animal studies reported a difference in c-Fos neuronal activation after chronic alcohol exposure; however, females remain an understudied population. To model chronic alcohol exposure match-pair fed adult male and female rats were administered 14 days of a liquid ethanol containing diet. Analysis focused on the central nucleus of the amygdala (CeA), a region integral to stress sensitivity and substance abuse. Immunocytochemical approaches identified cells containing ΔFosB, a marker of sustained neuronal activation, and activity patterns within the CeA were mapped by subdivision and rostral-caudal extent. Significant interactions were present between all groups, with gender differences noted among control groups, and ethanol exposed animals having the greatest number of ΔFosB immunoreactive cells indicating baseline dysregulation. Compared with c-Fos, a marker of recent neuronal activation, male ethanol treated animals had similar activity to controls, indicating a neuronal habituation not seen in females. Next, a cohort of animals were exposed to the forced swim test (FST), and c-Fos was examined in addition to FST behavior. Neuronal activity was increased in ethanol exposed animals compared to controls, and control females compared to males, indicating a potentiated stress response. Further, a population of activated neurons were shown to contain either corticotropin releasing factor or enkephalin. The present data suggest that dysregulation in the CeA neuronal activity may underlie some of the negative sequelae of alcohol abuse, and may, in part, underlie the distinctive response seen between genders to alcohol use.

Endocannabinoids (eCBs) are involved in a myriad of physiological processes that are mediated through the activation of cannabinoid receptors, which are ubiquitously distributed within the nervous system. One neurochemical target at which cannabinoids interact to have global effects on behavior is brain noradrenergic circuitry. We, and others, have previously shown that CB type 1 receptors (CB1r) are positioned to pre-synaptically modulate norepinephrine (NE) release in the rat frontal cortex (FC). Diacylglycerol lipase (DGL) is a key enzyme in the biosynthesis of the endocannabinoid 2-arachidonoylglycerol (2-AG). While DGL-α is expressed in the FC in the rat brain, it is not known whether noradrenergic afferents target neurons expressing synthesizing enzymes for the endocannabinoid, 2-AG. In the present study, we employed high-resolution neuroanatomical approaches to better define cellular sites for interactions between noradrenergic afferents and FC neurons expressing DGL-α. Immunofluorescence microscopy showed close appositions between processes containing the norepinephrine transporter (NET) or dopamine-β-hydroxylase (DβH) and cortical neurons expressing DGL-α-immunoreactivity. Ultrastructural analysis using immunogold-silver labeling for DGL-α and immunoperoxidase labeling for NET or DβH confirmed that NET-labeled axon terminals were directly apposed to FC somata and dendritic processes that exhibited DGL-α-immunoreactivity. Finally, tissue sections were processed for immunohistochemical detection of DGL-α, CB1r and DβH. Triple label immunofluorescence revealed that CB1r and DβH were co-localized in common cellular profiles and these were in close association with DGL-α. Taken together, these data provide anatomical evidence for direct synaptic associations between noradrenergic afferents and cortical neurons exhibiting endocannabinoid synthesizing machinery., (Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.)

Understanding the neurobiological bases for sex differences in alcohol dependence is needed to help guide the development of individualized therapies for alcohol abuse disorders. In the present study, alcohol-induced adaptations in (1) anxiety-like behavior, (2) patterns of c-Fos activation and (3) subcellular distribution of corticotropin releasing factor receptor in locus coeruleus (LC) neurons was investigated in male and female Sprague-Dawley rats that were chronically exposed to ethanol using a liquid diet. Results confirm and extend reports by others showing that chronic ethanol exposure produces an anxiogenic-like response in both male and female subjects. Ethanol-induced sex differences were observed with increased c-Fos expression in LC neurons of female ethanol-treated subjects compared to controls or male subjects. Results also reveal sex differences in the subcellular distribution of the CRFr in LC-noradrenergic neurons with female subjects exposed to ethanol exhibiting a higher frequency of plasmalemmal CRFrs. These adaptations have implications for LC neuronal activity and its neural targets across the sexes. Considering the important role of the LC in ethanol-induced activation of the hypothalamo-pituitary-adrenal (HPA) axis, the present results indicate important sex differences in feed-forward regulation of the HPA axis that may render alcohol dependent females more vulnerable to subsequent stress exposure., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The present study investigated whether psychological and/or physiological measures of stress would impede induction onto methadone maintenance and predict early (<6 months) discontinuation. Compared with controls, opioid-dependent subjects displayed increased distress on the perceived stress scale (PSS) and post-traumatic stress disorder checklist (PCLC); 60% exhibited abnormal cortisol. Addiction severity index (ASI), drug-use, and stress indices explained between 17 and 37% of the variance in engagement including attendance, opioid abstinence, and methadone stabilization. Participants who discontinued treatment displayed poor engagement, abnormal cortisol, elevated withdrawal symptoms, higher distress, and increased ongoing opioid use versus compliant individuals. Discontinuation was initially related to drug-use severity; however, by 6 months, retention depended primarily upon cortisol abnormalities, which increased an individual's discontinuation risk by 7.7-fold. These findings support admission screening with the ASI/cortisol for drop out, and stress/drug-use indices for engagement that together may enable clinically-relevant early recognition and interventions for prevention of stress-induced relapse in opioid-dependent populations., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Monoaminergic brainstem systems have widespread projections that participate in many central processes and, when dysregulated, contribute to a plethora of neuropsychiatric and neurodegenerative disorders. Synapses are the foundation of these neuronal circuits, and their local dysfunction results in global aberrations leading to pathophysiological disease states. This review focuses on the locus coeruleus (LC) norepinephrine (NE) brainstem system and its underappreciated role in Alzheimer's disease (AD). Amyloid beta (Aβ), a peptide that accumulates aberrantly in AD has recently been implicated as a modulator of neuronal excitability at the synapse. Evidence is presented showing that disruption of the LC-NE system at a synaptic and circuit level during early stages of AD, due to conditions such as chronic stress, can potentially lead to amyloid accumulation and contribute to the progression of this neurodegenerative disorder. Additional factors that impact neurodegeneration include neuroinflammation, and network de-synchronization. Consequently, targeting the LC-NE system may have significant therapeutic potential for AD, as it may facilitate modulation of Aβ production, curtail neuroinflammation, and prevent sleep and behavioral disturbances that often lead to negative patient outcomes.

Amygdalar norepinephrine (NE) plays a key role in regulating neural responses to emotionally arousing stimuli and is involved in memory consolidation of emotionally charged events. Corticotropin-releasing factor (CRF) and dynorphin (DYN), two neuropeptides that mediate the physiological and behavioral responses to stress, are abundant in the central nucleus of the amygdala (CeA), and directly innervate brainstem noradrenergic locus coeruleus (LC) neurons. Whether the CRF- and DYN-containing amygdalar neurons receive direct noradrenergic innervation has not yet been elucidated. The present study sought to define cellular substrates underlying noradrenergic modulation of CRF- and DYN-containing neurons in the CeA using immunohistochemistry and electron microscopy. Ultrastructural analysis revealed that NE-labeled axon terminals form synapses with CRF- and DYN-containing neurons in the CeA. Semi-quantitative analysis showed that approximately 31 % of NET-labeled axon terminals targeted CeA neurons that co-expressed DYN and CRF. As a major source of CRF innervation to the LC, it is also not known whether CRF-containing CeA neurons are directly targeted by noradrenergic afferents. To test this, retrograde tract tracing using FluoroGold from the LC was combined with immunocytochemical detection of CRF and NET in the CeA. Our results revealed a population of LC-projecting CRF-containing CeA neurons that are directly innervated by NE afferents. Analysis showed that approximately 34 % of NET-labeled axon terminals targeted LC-projecting CeA neurons that contain CRF. Taken together, these results indicate significant interactions between NE, CRF and DYN in this critical limbic region and reveal direct synaptic interactions of NE with amygdalar CRF that influence the LC-NE arousal system.

Introduction: The endocannabinoid (eCB) system plays an important role in the control of mood, and its dysregulation has been implicated in several psychiatric disorders. Targeting the eCB system appears to represent an attractive and novel approach to the treatment of depression and other mood disorders. However, several failed clinical trials have diminished enthusiasm for the continued development of eCB-targeted therapeutics for psychiatric disorders, despite the encouraging preclinical data and promising preliminary results obtained with the synthetic cannabinoid nabilone for treating post-traumatic stress disorder., Areas Covered: This review describes the eCB system's role in modulating cell signaling within the brain. There is a specific focus on eCB's regulation of monoamine neurotransmission and the stress axis, as well as how dysfunction of this interaction can contribute to the development of psychiatric disorders. Additionally, the review provides discussion on compounds and drugs that target this system and might prove to be successful for the treatment of mood-related psychiatric disorders., Expert Opinion: The discovery of increasingly selective modulators of CB receptors should enable the identification of optimal therapeutic strategies. It should also maximize the likelihood of developing safe and effective treatments for debilitating psychiatric disorders.

Repeated, extreme, or traumatic stressors can elicit pathological effects leading to many negative physical and psychological outcomes. Stressors can precipitate the onset of psychiatric diseases, or exacerbate pre-existing disorders including various anxiety and mood disorders. As stressors can negatively impact human psychiatric health, it is essential to identify neurochemicals that may confer protection from the negative sequelae of repeated or extreme stress exposure. Elucidating the neurobiological underpinnings of stress resilience will enhance our ability to promote resilience to, or recovery from, stress-related psychiatric disease. Herein, we will review the evidence for neuropeptide Y as an endogenous mediator of resilience and its potential relevance for the treatment of stress-related psychiatric diseases.

Objective: A subarachnoid hemorrhage is neurologically devastating, with 50% of patients becoming disabled or deceased. Advent of Guglielmi detachable coils in 1995 permitted endovascular treatment of cerebral aneurysms. Coiling is efficacious and safe, but durability needs improvement, as nearly 20% of patients require further invasive intervention secondary to aneurysm recurrence. The aim of this study is to develop an in vitro model of endothelial cell (EC) proliferation and differentiation on four types of platinum-based coils, using gene expression profiling to understand EC biology as they colonize and differentiate on coils., Methods: Human umbilical vein ECs were grown in vitro on platinum coil segments. Growth patterns were assessed as a function of coil type. Gene expression profiles for coil attached versus coil unattached ECs were determined using immunohistochemistry and gene array analysis., Results: ECs showed rapid, robust attachment to all coil types. Some detachment occurred within 24-48 hours. Significant growth of remaining attached cells occurred during the next week, creating a confluence on coils and within coil grooves. Similar growth curve results were obtained with human brain ECs on platinum-based coil surfaces. Differentiation markers in attached cells (α(1), α(2), β(1) integrins) were expressed on immunostaining, whereas microarray gene expression revealed 48 up-regulated and 68 down-regulated genes after 24-hour growth on coils. Major pathways affected as a function of time of colonization on coils and coil type included those involved in regulation of cell cycle and cell signaling., Conclusions: We developed an in vitro model for evaluating endothelialization of platinum coils to optimize coil design to support robust EC colonization and differentiation., (Copyright © 2014. Published by Elsevier Inc.)

Endocannabinoids acting at the cannabinoid type 1 receptor (CB1R) are known to regulate attention, cognition and mood. Previous studies have shown that, in the rat medial prefrontal cortex (mPFC), CB1R agonists increase norepinephrine release, an effect that may be attributed, in part, to CB1Rs localised to noradrenergic axon terminals. The present study was aimed at further characterising functional interactions between CB1R and adrenergic receptor (AR) systems in the mPFC using in vitro intracellular electrophysiology and high-resolution neuroanatomical techniques. Whole-cell patch-clamp recordings of layer V/VI cortical pyramidal neurons in rats revealed that both acute and chronic treatment with the synthetic CB1R agonist WIN 55,212-2 blocked elevations in cortical pyramidal cell excitability and increases in input resistance evoked by the α2-adrenergic receptor (α2-AR) agonist clonidine, suggesting a desensitisation of α2-ARs. These CB1R-α2-AR interactions were further shown to be both action potential- and gamma-aminobutyric acid-independent. To better define sites of cannabinoid-AR interactions, we localised α2A-adrenergic receptors (α2A-ARs) in a genetically modified mouse that expressed a hemoagglutinin (HA) tag downstream of the α2A-AR promoter. Light and electron microscopy indicated that HA-α2A-AR was distributed in axon terminals and somatodendritic processes especially in layer V of the mPFC. Triple-labeling immunocytochemistry revealed that α2A-AR and CB1R were localised to processes that contained dopamine-β-hydroxylase, a marker of norepinephrine. Furthermore, HA-α2A-AR was localised to processes that were directly apposed to CB1R. These findings suggest multiple sites of interaction between cortical cannabinoid-adrenergic systems that may contribute to understanding the effect of cannabinoids on executive functions and mood., (© 2014 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Trafficking of G protein-coupled receptors (GPCRs) is a critical determinant of cellular sensitivity of neurons. To understand how endogenous or exogenous ligands impact cell surface expression of GPCRs, it is essential to employ approaches that achieve superior anatomical resolution at the synaptic level. In situations in which light and fluorescence microscopy techniques may provide only limited resolution, electron microscopy provides enhanced subcellular precision. Dual labeling immunohistochemistry employing visually distinct immunoperoxidase and immunogold markers has been an effective approach for elucidating complex receptor profiles at the synapse and to definitively establish the localization of individual receptors and neuromodulators to common cellular profiles. The immuno-electron microscopy approach offers the potential for determining membrane versus intracellular protein localization, as well as the association with various identifiable cellular organelles. Corticotropin-releasing factor (CRF) is an important regulator of endocrine, autonomic, immunological, behavioral and cognitive limbs of the stress response. Dysfunction of this neuropeptide system has been associated with several psychiatric disorders. This review summarizes findings from neuroanatomical studies, with superior spatial resolution, that indicate that the distribution of CRF receptors is a highly dynamic process that, in addition to being sexually dimorphic, involves complex regulation of receptor trafficking within extrasynaptic sites that have significant consequences for adaptations to stress, particularly within the locus coeruleus (LC), the major brain norepinephrine-containing nucleus., (Copyright © 2014 Elsevier Inc. All rights reserved.)

HIV-1 trans-acting protein Tat, an essential protein for viral replication, is a key mediator of neurotoxicity. If Tat oxidant injury and neurotoxicity have been described, consequent neuroinflammation is less understood. Rat caudate-putamens (CPs) were challenged with Tat, with or without prior rSV40-delivered superoxide dismutase or glutathione peroxidase. Tat injection caused oxidative stress. Administration of Tat in the CP induced an increase in numbers of Iba-1- and CD68-positive cells, as well as an infiltration of astrocytes. We also tested the effect of more protracted Tat exposure on neuroinflammation using an experimental model of chronic Tat exposure. SV(Tat): a recombinant SV40-derived gene transfer vector was inoculated into the rat CP, leading to chronic expression of Tat, oxidative stress, and ongoing apoptosis, mainly located in neurons. Intra-CP SV(Tat) injection induced an increase in microglia and astrocytes, suggesting that protracted Tat production increased neuroinflammation. SV(SOD1) or SV(GPx1) significantly reduced neuroinflammation following Tat administration into the CP. Thus, Tat-induced oxidative stress, CNS injury, neuron loss and inflammation may be mitigated by antioxidant gene delivery.

Opiate addiction is a devastating health problem, with approximately 2million people currently addicted to heroin or non-medical prescription opiates in the United States alone. In neurons, adaptations in cell signaling cascades develop following opioid actions at the mu opioid receptor (MOR). A novel putative target for intervention involves interacting proteins that may regulate trafficking of MOR. Morphine has been shown to induce a re-distribution of a MOR-interacting protein Wntless (WLS, a transport molecule necessary for secretion of neurotrophic Wnt proteins), from cytoplasmic to membrane compartments in rat striatal neurons. Given its opiate-sensitivity and its well-characterized molecular and cellular adaptations to morphine exposure, we investigated the anatomical distribution of WLS and MOR in the rat locus coeruleus (LC)-norepinephrine (NE) system. Dual immunofluorescence microscopy was used to test the hypothesis that WLS is localized to noradrenergic neurons of the LC and that WLS and MOR co-exist in common LC somatodendritic processes, providing an anatomical substrate for their putative interactions. We also hypothesized that morphine would influence WLS distribution in the LC. Rats received saline, morphine or the opiate agonist [d-Ala2, N-Me-Phe4, Gly-ol5]-enkephalin (DAMGO), and tissue sections through the LC were processed for immunogold-silver detection of WLS and MOR. Statistical analysis showed a significant re-distribution of WLS to the plasma membrane following morphine treatment in addition to an increase in the proximity of gold-silver labels for MOR and WLS. Following DAMGO treatment, MOR and WLS were predominantly localized within the cytoplasmic compartment when compared to morphine and control. In a separate cohort of rats, brains were obtained from saline-treated or heroin self-administering male rats for pulldown co-immunoprecipitation studies. Results showed an increased association of WLS and MOR following heroin exposure. As the LC-NE system is important for cognition as well as decisions underlying substance abuse, adaptations in WLS trafficking and expression may play a role in modulating MOR function in the LC and contribute to the negative sequelae of opiate exposure on executive function., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Withdrawal from opiates, such as heroin or oral narcotics, is characterized by a host of aversive physical and emotional symptoms. High rates of relapse and limited treatment success rates for opiate addiction have prompted a search for new approaches. For many opiate addicts, achieving abstinence may be further complicated by poly-drug use and co-morbid mental disorders. Research over the past decade has shed light on the influence of endocannabinoids (ECs) on the opioid system. Evidence from both animal and clinical studies point toward an interaction between these two systems, and suggest that targeting the EC system may provide novel interventions for managing opiate dependence and withdrawal. This review will summarize the literature surrounding the molecular effects of cannabinoids and opioids on the locus coeruleus-norepinephrine system, a key circuit implicated in the negative sequelae of opiate addiction. A consideration of the trends and effects of marijuana use in those seeking treatment to abstain from opiates in the clinical setting will also be presented. In summary, the present review details how cannabinoid-opioid interactions may inform novel interventions in the management of opiate dependence and withdrawal., (Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.)

Based on the importance of the locus coeruleus-norepinephrine (LC-NE) system and the dorsal raphe nucleus-serotonergic (DRN-5-HT) system in stress-related pathologies, additional understanding of brain regions coordinating their activity is of particular interest. One such candidate is the amygdalar complex, and specifically, the central nucleus (CeA), which has been implicated in emotional arousal and is known to send monosynaptic afferent projections to both these regions. Our present data using dual retrograde tract tracing is the first to demonstrate a population of amygdalar neurons that project in a collateralized manner to the LC and DRN, indicating that amygdalar neurons are positioned to coordinately regulate the LC and DRN, and links these brain regions by virtue of a common set of afferents. Further, we have also characterized the phenotype of a population of these collaterally projecting neurons from the amygdala as containing corticotropin releasing factor or dynorphin, two peptides heavily implicated in the stress response. Understanding the co-regulatory influences of this limbic region on 5HT and NE regions may help fill a gap in our knowledge regarding neural circuits impacting these systems and their adaptations in stress., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Stress is implicated in diverse psychiatric disorders including substance abuse. The locus coeruleus-norepinephrine (LC-NE) system is a major stress response system that is also a point of intersection between stress neuromediators and endogenous opioids and so may be a site at which stress can influence drug-taking behaviors. As social stress is a common stressor for humans, this study characterized the enduring impact of repeated social stress on LC neuronal activity. Rats were exposed to five daily consecutive sessions of social stress using the resident-intruder model or control manipulation. LC discharge rate recorded 2 days after the last manipulation was decreased in stressed rats compared with controls. By 10 days after the last manipulation, LC rates were comparable between groups. Systemic administration of the opiate antagonist, naloxone, robustly increased LC discharge rate in a manner suggestive of opiate withdrawal, selectively in stressed rats when administered 2 or 10 days after the last manipulation. This was accompanied by behavioral signs of mild opiate withdrawal. Western blot and electron microscopic studies indicated that repeated social stress decreased corticotropin-releasing factor type 1 receptor and increased μ-opioid receptor levels in the LC. Together, the results suggest that repeated social stress engages endogenous opioid modulation of LC activity and induces signs of cellular and physical opiate dependence that endure after the stress. These cellular effects may predispose individuals with a history of repeated social stress to substance abuse behaviors.