Optimization of immunolabeling and clearing techniques for indelibly labeled memory traces.

Recent genetic tools have allowed researchers to visualize and manipulate memory traces (i.e., engrams) in small brain regions. However, the ultimate goal is to visualize memory traces across the entire brain in order to better understand how memories are stored in neural networks and how multiple memories may coexist. Intact tissue clearing and imaging is a new and rapidly growing area of focus that could accomplish this task. Here, we utilized the leading protocols for whole-brain clearing and applied them to the ArcCreERT2 mice, a murine line that allows for the indelible labeling of memory traces. We found that CLARITY and PACT greatly distorted the tissue, and iDISCO quenched enhanced yellow fluorescent protein (EYFP) fluorescence and hindered immunolabeling. Alternative clearing solutions, such as tert-Butanol, circumvented these harmful effects, but still did not permit whole-brain immunolabeling. CUBIC and CUBIC with Reagent-1A produced improved antibody penetration and preserved EYFP fluorescence, but also did not allow for whole-brain memory trace visualization. Modification of CUBIC with Reagent-1A resulted in EYFP fluorescence preservation and immunolabeling of the immediate early gene (IEG) Arc in deep brain areas; however, optimized memory trace labeling still required tissue slicing into mm-thick tissue sections. In summary, our data show that CUBIC with Reagent-1A* is the ideal method for reproducible clearing and immunolabeling for the visualization of memory traces in mm-thick tissue sections from ArcCreERT2 mice.

Nucleus reuniens of the thalamus controls fear memory intensity, specificity and long-term maintenance during consolidation.

The thalamic nucleus reuniens (NR) has been shown to support bidirectional medial prefrontal cortex-hippocampus communication and synchronization relevant for cognitive processing. Using non-selective or prolonged inactivation of the NR, previous studies reported its activity positively modulates aversive memory consolidation. Here we examined the NR's role in consolidating contextual fear memories with varied strength, at both recent and more remote time points, using muscimol-induced temporary inactivation in rats. Results indicate the NR negatively modulates fear memory intensity, specificity, and long-term maintenance. The more intense, generalized, and enduring fear memory induced by NR inactivation during consolidation was less prone to behavioral suppression by extinction or reconsolidation disruption induced by clonidine, an alpha-2 adrenergic receptor agonist. Lastly, we used immunohistochemistry for Arc protein, which is involved in synaptic modifications underlying memory consolidation, to investigate whether treatment condition and/or conditioning status could change its levels not only in the NR, but also in the hippocampus (dorsal and ventral CA1 subregions) and the medial prefrontal cortex (anterior cingulate, prelimbic and infralimbic subregions). Results indicate a significant imbalance in the number of Arc-expressing neurons in the brain areas investigated in muscimol fear conditioned animals when compared with controls. Collectively, present results provide convergent evidence for the NR's role as a hub regulating quantitative and qualitative aspects of a contextual fear memory during its consolidation that seem to influence the subsequent susceptibility to experimental interventions aiming at attenuating its expression. They also indicate the selectivity and duration of a given inactivation approach may influence its outcomes.

Dopamine is Required for Activity-Dependent Amplification of Arc mRNA in Developing Postnatal Frontal Cortex.

The activity-regulated gene Arc/Arg3.1 encodes a postsynaptic protein crucially involved in glutamatergic synaptic plasticity. Genetic mutations in Arc pathway and altered Arc expression in human frontal cortex have been associated with schizophrenia. Although Arc expression has been reported to vary with age, what mechanisms regulate Arc mRNA levels in frontal cortex during postnatal development remains unclear. Using quantitative mRNA analysis of mouse frontal cortical tissues, we mapped the developmental profiles of Arc expression and found that its mRNA levels are sharply amplified near the end of the second postnatal week, when mouse pups open their eyes for the first time after birth. Surprisingly, electrical stimulation of the frontal cortex before eye-opening is not sufficient to drive the amplification of Arc mRNA. Instead, this amplification needs both electrical stimulation and dopamine D1-type receptor (D1R) activation. Furthermore, visual stimuli-driven amplification of Arc mRNA is also dependent on D1R activation and dopamine neurons located in the ventral midbrain. These results indicate that dopamine is required to drive activity-dependent amplification of Arc mRNA in the developing postnatal frontal cortex and suggest that joint electrical and dopaminergic activation is essential to establish the normal expression pattern of a schizophrenia-associated gene during frontal cortical development.

Dissociation between complete hippocampal context memory formation and context fear acquisition.

Rodents require a minimal time period to explore a context prior to footshock to display plateau-level context fear at test. To investigate whether this rapid fear plateau reflects complete memory formation within that short time-frame, we used the immediate-early gene product Arc as an indicator of hippocampal context memory formation-related activity. We found that hippocampal Arc expression continued to increase well past the minimal time required for plateau-level fear. This raises the possibility that context fear conditioning occurs more rapidly than complete memory formation. Thus, animals may be able to condition robustly to both complete and incomplete contextual representations.

From Memory Impairment to Posttraumatic Stress Disorder-Like Phenotypes: The Critical Role of an Unpredictable Second Traumatic Experience.

Arousal and stress critically regulate memory formation and retention. Increasing levels of stress produce an inverted U-shaped effect on cognitive performance, including the retention of explicit memories, and experiencing a severe stress during a traumatic event may lead to posttraumatic stress disorder (PTSD). The molecular mechanisms underlying the impairing effect of a severe stress on memory and the key contribution of traumatic experiences toward the development of PTSD are still unknown. Here, using increasing footshock intensities in an inhibitory avoidance paradigm, we reproduced the inverted U-shaped curve of memory performance in rats. We then show that the inverted U profile of memory performance correlates with an inverted U profile of corticosterone level in the circulation and of brain-derived neurotrophic factor, phosphorylated tropomyosin-receptor kinase B, and methyl CpG binding protein in the dorsal hippocampus. Furthermore, training with the highest footshock intensity (traumatic experience) led to a significant elevation of hippocampal glucocorticoid receptors. Exposure to an unpredictable, but not to a predictable, highly stressful reminder shock after a first traumatic experience resulted in PTSD-like phenotypes, including increased memory of the trauma, high anxiety, threat generalization, and resistance to extinction. Systemic corticosterone injection immediately after the traumatic experience, but not 3 d later, was sufficient to produce PTSD-like phenotypes. We suggest that, although after a first traumatic experience a suppression of the corticosterone-dependent response protects against the development of an anxiety disorder, experiencing more than one trauma (multiple hits) is a critical contributor to the etiology of PTSD.

Hunger States Control the Directions of Synaptic Plasticity via Switching Cell Type-Specific Subunits of NMDA Receptors.

It remains largely unknown whether and how hunger states control activity-dependent synaptic plasticity, such as long-term potentiation (LTP) and long-term depression (LTD). We here report that both LTP and LTD of excitatory synaptic strength within the appetite control circuits residing in hypothalamic arcuate nucleus (ARC) behave in a manner of hunger states dependence and cell type specificity. For instance, we find that tetanic stimulation induces LTP at orexigenic agouti-related protein (AgRP) neurons in ad libitum fed mice, whereas it induces LTD in food-deprived mice. In an opposite direction, the same induction protocol induces LTD at anorexigenic pro-opiomelanocortin (POMC) neurons in fed mice but weak LTP in deprived mice. Mechanistically, we also find that food deprivation increases the expressions of NR2C/NR2D/NR3-containing NMDA receptors (NMDARs) at AgRP neurons that contribute to the inductions of LTD, whereas it decreases their expressions at POMC neurons. Collectively, our data reveal that hunger states control the directions of activity-dependent synaptic plasticity by switching NMDA receptor subpopulations in a cell type-specific manner, providing insights into NMDAR-mediated interactions between energy states and associative memory. Significance statement: Based on the experiments performed in this study, we demonstrate that activity-dependent synaptic plasticity is also under the control of energy states by regulating NMDAR subpopulations in a cell type-specific manner. We thus propose a reversible memory configuration constructed from energy states-dependent cell type-specific bidirectional conversions of LTP and LTD. Together with the distinct functional roles played by NMDAR signaling in the control of food intake and energy states, these findings reveal a new reciprocal interaction between energy states and associative memory, one that might serve as a target for therapeutic treatments of the energy-related memory disorders or vice versa.

Sweet orosensation induces Arc expression in dorsal hippocampal CA1 neurons in an experience-dependent manner.

There is limited knowledge regarding how the brain controls the timing of meals. Similarly, there is a large gap in our understanding of how top-down cognitive processes, such as memory influence energy intake. We hypothesize that dorsal hippocampal (dHC) neurons, which are critical for episodic memory, form a memory of a meal and inhibit meal onset during the postprandial period. In support, we showed previously that reversible inactivation of these neurons during the period following a sucrose meal accelerates the onset of the next meal. If dHC neurons form a memory of a meal, then consumption should induce synaptic plasticity in dHC neurons. To test this, we determined (1) whether a sucrose meal increases the expression of the synaptic plasticity marker activity-regulated cytoskeleton-associated protein (Arc) in dHC CA1 neurons, (2) whether previous experience with sucrose influences sucrose-induced Arc expression, and (3) whether the orosensory stimulation produced by the noncaloric sweetener saccharin is sufficient to induce Arc expression. Male Sprague-Dawley rats were trained to consume a sweetened solution at a scheduled time daily. On the experimental day, they were given a solution for 7 min, euthanized, and then fluorescence in situ hybridization procedures were used to measure meal-induced Arc mRNA. Compared to caged control rats, Arc expression was significantly higher in rats that consumed sucrose or saccharin. Interestingly, rats given additional experience with sucrose had less Arc expression than rats with less sucrose experience, even though both groups consumed similar amounts on the experimental day. Thus, this study is the first to suggest that orosensory stimulation produced by consuming a sweetened solution and possibly the hedonic value of that sweet stimulation induces synaptic plasticity in dHC CA1 neurons in an experience-dependent manner. Collectively, these findings are consistent with our hypothesis that dHC neurons form a memory of a meal.

Calcium dysregulation via L-type voltage-dependent calcium channels and ryanodine receptors underlies memory deficits and synaptic dysfunction during chronic neuroinflammation.

BACKGROUND: Chronic neuroinflammation and calcium (Ca(+2)) dysregulation are both components of Alzheimer's disease. Prolonged neuroinflammation produces elevation of pro-inflammatory cytokines and reactive oxygen species which can alter neuronal Ca(+2) homeostasis via L-type voltage-dependent Ca(+2) channels (L-VDCCs) and ryanodine receptors (RyRs). Chronic neuroinflammation also leads to deficits in spatial memory, which may be related to Ca(+2) dysregulation. METHODS: The studies herein use an in vivo model of chronic neuroinflammation: rats were infused intraventricularly with a continuous small dose of lipopolysaccharide (LPS) or artificial cerebrospinal fluid (aCSF) for 28 days. The rats were treated with the L-VDCC antagonist nimodipine or the RyR antagonist dantrolene. RESULTS: LPS-infused rats had significant memory deficits in the Morris water maze, and this deficit was ameliorated by treatment with nimodipine. Synaptosomes from LPS-infused rats had increased Ca(+2) uptake, which was reduced by a blockade of L-VDCCs either in vivo or ex vivo. CONCLUSIONS: Taken together, these data indicate that Ca(+2) dysregulation during chronic neuroinflammation is partially dependent on increases in L-VDCC function. However, blockade of the RyRs also slightly improved spatial memory of the LPS-infused rats, demonstrating that other Ca(+2) channels are dysregulated during chronic neuroinflammation. Ca(+2)-dependent immediate early gene expression was reduced in LPS-infused rats treated with dantrolene or nimodipine, indicating normalized synaptic function that may underlie improvements in spatial memory. Pro-inflammatory markers are also reduced in LPS-infused rats treated with either drug. Overall, these data suggest that Ca(+2) dysregulation via L-VDCCs and RyRs play a crucial role in memory deficits resulting from chronic neuroinflammation.

Ethanol exposure induces neonatal neurodegeneration by enhancing CB1R Exon1 histone H4K8 acetylation and up-regulating CB1R function causing neurobehavioral abnormalities in adult mice.

BACKGROUND: Ethanol exposure to rodents during postnatal day 7 (P7), which is comparable to the third trimester of human pregnancy, induces long-term potentiation and memory deficits. However, the molecular mechanisms underlying these deficits are still poorly understood. METHODS: In the present study, we explored the potential role of epigenetic changes at cannabinoid type 1 (CB1R) exon1 and additional CB1R functions, which could promote memory deficits in animal models of fetal alcohol spectrum disorder. RESULTS: We found that ethanol treatment of P7 mice enhances acetylation of H4 on lysine 8 (H4K8ace) at CB1R exon1, CB1R binding as well as the CB1R agonist-stimulated GTPγS binding in the hippocampus and neocortex, two brain regions that are vulnerable to ethanol at P7 and are important for memory formation and storage, respectively. We also found that ethanol inhibits cyclic adenosine monophosphate response element-binding protein (CREB) phosphorylation and activity-regulated cytoskeleton-associated protein (Arc) expression in neonatal and adult mice. The blockade or genetic deletion of CB1Rs prior to ethanol treatment at P7 rescued CREB phosphorylation and Arc expression. CB1R knockout mice exhibited neither ethanol-induced neurodegeneration nor inhibition of CREB phosphorylation or Arc expression. However, both neonatal and adult mice did exhibit enhanced CREB phosphorylation and Arc protein expression. P7 ethanol-treated adult mice exhibited impaired spatial and social recognition memory, which were prevented by the pharmacological blockade or deletion of CB1Rs at P7. CONCLUSIONS: Together, these findings suggest that P7 ethanol treatment induces CB1R expression through epigenetic modification of the CB1R gene, and that the enhanced CB1R function induces pCREB, Arc, spatial, and social memory deficits in adult mice.

Enhanced prefrontal serotonin 2A receptor signaling in the subchronic phencyclidine mouse model of schizophrenia.

Prefrontal serotonin 2A receptors (5-HT2A Rs) have been linked to the pathogenesis and treatment of schizophrenia. Many antipsychotics fully occupy 5-HT2A R at clinical relevant doses, and activation of 5-HT2A receptors by lysergic acid diethylamide (LSD) and LSD-like drugs induces a schizophrenia-like psychosis in humans. Subchronic phencyclidine (PCP) administration is a well-established model for schizophrenia-like symptoms in rodents. The aim of the present study was to investigate whether subchronic PCP administration changes expression, binding, or functionality of cortical 5-HT2A Rs. As a measure of 5-HT2A R functionality, we used the 5-HT2A R agonist 2,5-dimethoxy-4-iodoamphetamine (DOI)-induced head-twitch response (HTR) and mRNA expression of the immediate-early genes (IEGs) activity-related cytoskeletal associated-protein (Arc), c-fos, and early growth response protein 2 (egr-2) in the frontal cortex. Mice were treated with PCP (10 mg/kg) or saline for 10 days, followed by a 5-day washout period. The PCP pretreatment increased the overall induction of HTR and frontal cortex IEG mRNA expression following a single challenge with DOI. These functional changes were not associated with changes in 5-HT2A R binding. Also, binding of the 5-HT1A R and the 5-HT transporter was unaffected. Finally, basal mRNA level of Arc was increased in the prefrontal cortex after subchronic PCP administration as revealed with in situ hybridization. Together these findings indicate that PCP administration produces changes in the brain that result in an increase in the absolute effect of DOI. Therefore, neurotransmission involving the 5-HT2A R could contribute to the behavioral deficits observed after PCP treatment. © 2013 Wiley Periodicals, Inc.

Emotional memory impairments in a genetic rat model of depression: involvement of 5-HT/MEK/Arc signaling in restoration.

Cognitive dysfunctions are common in major depressive disorder, but have been difficult to recapitulate in animal models. This study shows that Flinders sensitive line (FSL) rats, a genetic rat model of depression, display a pronounced impairment of emotional memory function in the passive avoidance (PA) task, accompanied by reduced transcription of Arc in prefrontal cortex and hippocampus. At the cellular level, FSL rats have selective reductions in levels of NMDA receptor subunits, serotonin 5-HT(1A) receptors and MEK activity. Treatment with chronic escitalopram, but not with an antidepressant regimen of nortriptyline, restored memory performance and increased Arc transcription in FSL rats. Multiple pharmacological manipulations demonstrated that procognitive effects could also be achieved by either disinhibition of 5-HT(1A)R/MEK/Arc or stimulation of 5-HT4R/MEK/Arc signaling cascades. Taken together, studies of FSL rats in the PA task revealed reversible deficits in emotional memory processing, providing a potential model with predictive and construct validity for assessments of procognitive actions of antidepressant drug therapies.

Enhanced function of prefrontal serotonin 5-HT(2) receptors in a rat model of psychiatric vulnerability.

Prefrontal serotonin 5-HT(2) receptors have been linked to the pathogenesis and treatment of affective disorders, yet their function in psychiatric vulnerability is not known. Here, we examine the effects of 5-HT(2) receptors in a rat model of psychiatric vulnerability using electrophysiology, gene expression, and behavior. Following the early stress of chronic maternal separation, we found that serotonin has atypical 5-HT(2) receptor-mediated excitatory effects in the adult prefrontal cortex that were blocked by the 5-HT(2A) receptor antagonist MDL 100907. In the absence of a serotonergic agonist, the intrinsic excitability of the prefrontal cortex was not enhanced relative to controls. Yet, in response to stimulation of 5-HT(2) receptors, adult animals with a history of early stress exhibit heightened prefrontal network activity in vitro, enhanced immediate early gene expression in vivo, and potentiated head shake behavior. These changes arise in the absence of any major alteration of prefrontal 5-HT(2A/C) mRNA expression or 5-HT(2) receptor binding. Our microarray results and quantitative PCR validation provide insight into the molecular changes that accompany such enhanced 5-HT(2) receptor function in adult animals following early stress. We observed persistent prefrontal transcriptome changes, with significant enrichment of genes involved in cellular developmental processes, regulation of signal transduction, and G-protein signaling. Specific genes regulated by early stress were validated in an independent cohort, and several altered genes were normalized by chronic blockade of 5-HT(2) receptors in adulthood. Together, our results demonstrate enhanced prefrontal 5-HT(2) receptor function and persistent alterations in prefrontal gene expression in a rat model of psychiatric vulnerability.

Reduced Hippocampal Neurogenesis in Mice Deficient in Apoptosis Repressor with Caspase Recruitment Domain (ARC).

In the adult hippocampal dentate gyrus (DG), the majority of newly generated cells are eliminated by apoptotic mechanisms. The apoptosis repressor with caspase recruitment domain (ARC), encoded by the Nol3 gene, is a potent and multifunctional death repressor that inhibits both death receptor and mitochondrial apoptotic signaling. The aim of the present study was to parse the role of ARC in the development of new granule cell neurons. Nol3 gene expression as revealed by in situ hybridization is present in the entire dentate granule cell layer. Moreover, a comparison of Nol3 expression between FACS-sorted Sox2-positive neural stem cells and Doublecortin (DCX)-positive immature neurons demonstrates upregulation of Nol3 during neurogenesis. Using ARC-deficient mice, we show that proliferation and survival of BrdU birth-dated cells are strongly reduced in the absence of ARC while neuronal-glial fate choice is not affected. Both the number of DCX-positive cells and the number of calretinin (CR)-positive immature postmitotic neurons are reduced in the hippocampus of ARC-/- mice. ARC knockout is not associated with increased numbers of microglia or with microglia activation. However, hippocampal brain-derived neurotrophic factor (BDNF) protein content is significantly increased in ARC-/- mice, possibly representing a compensatory response. Collectively, our results suggest that ARC plays a critical cell-autonomous role in preventing cell death during adult granule cell neogenesis.

Developmental Exposure to Psychostimulant Primes Activity-dependent Gene Induction in Frontal Cortex.

Perinatal neurodevelopment involves extensive formation of neural connections and onset of activity-dependent gene expression for synaptic function and plasticity. Exposure to psychostimulants at this stage imposes significant risks for developing cognitive and affective disorders later in life. However, how developmental exposure to psychostimulants may induce long-lasting molecular changes relevant to neural circuit function remains incompletely understood. In this study, we investigated the impacts of psychostimulant amphetamine on the activity-dependent induction of synaptic adaptor molecule Arc in the frontal cortex of neonatal mice. We found that transient exposure to amphetamine not only amplifies activity-dependent Arc mRNA expression immediately, but also potentiates subsequent induction of Arc mRNA in the absence of amphetamine. This priming effect is associated with a rapid and persistent increase in histone mono-methylation (H3K4me1), a marker for transcriptionally permissive chromatin, at the Arc locus, but not any long-lasting change in the phosphorylation of upstream transcription factor CREB. Furthermore, the increase in H3K4me1 at the Arc locus requires dopamine receptor signaling, and the priming of Arc expression correlates with the dopaminergic innervation pattern in the frontal cortex. Together, our results demonstrate that developmental exposure to psychostimulant amphetamine induces long-lasting chromatin changes and primes activity-dependent Arc gene induction. These findings reveal the molecular targets of psychostimulant during perinatal development that may contribute to long-term psychiatric risks.

Differential fear conditioning generates prefrontal neural ensembles of safety signals.

Fear discrimination is critical for survival, while fear generalization is effective for avoiding dangerous situations. Overgeneralized fear is a typical symptom of anxiety disorders, including generalized anxiety disorder and posttraumatic stress disorder (PTSD). Previous research demonstrated that fear discrimination learning is mediated by prefrontal mechanisms. While the prelimbic (PL) and infralimbic (IL) subdivisions of the medial prefrontal cortex (mPFC) are recognized for their excitatory and inhibitory effects on the fear circuit, respectively, the mechanisms driving fear discrimination are unidentified. To obtain insight into the mechanisms underlying context-specific fear discrimination, we investigated prefrontal neuronal ensembles representing distinct experiences associated with learning to disambiguate between dangerous and similar, but not identical, harmless stimuli. Here, we show distinct quantitative activation differences in response to conditioned and generalized fear experiences, as well as modulation of the neuronal ensembles associated with successful acquisition of context-safety contingencies. These findings suggest that prefrontal neuronal ensembles patterns code functional context-danger and context-safety relationships. The PL subdivision of the mPFC monitors context-danger associations to conditioned fear, whereas differential conditioning sparks additional ensembles associated with the inhibition of generalized fear in both the PL and IL subdivisions of the mPFC. Our data suggest that fear discrimination learning is associated with the modulation of prefrontal subpopulations in a subregion- and experience-specific fashion, and the learning of appropriate responses to conditioned and initially generalized fear experiences is driven by gradual updating and rebalancing of the prefrontal memory representations.

Distinct regulation pattern of Egr-1, BDNF and Arc during morphine-withdrawal conditioned place aversion paradigm: Role of glucocorticoids.

Negative affective aspects of opiate abstinence contribute to the persistence of substance abuse. Importantly, interconnected brain areas involved in aversive motivational processes, such as the ventral tegmental area (VTA) and medial prefrontal cortex (mPFC), become activated when animals are confined to withdrawal-paired environments. In the present study, place aversion was elicited in sham and adrenalectomized (ADX) animals by conditioned naloxone-precipitated drug withdrawal following exposure to chronic morphine. qPCR was employed to detect the expression of brain derived neurotrophic factor (Bdnf) and the immediate early genes (IEG) early growth response 1 (Egr-1) and activity-regulated cytoskeletal-associated protein (Arc) mRNAs in the VTA and mPFC at different time points of the conditioned place aversion (CPA) paradigm: after the conditioning phase and after the test phase. Sham + morphine rats exhibited robust CPA, which was impaired in ADX + morphine animals. Egr-1 and Arc were induced in the VTA and mPFC after morphine-withdrawal conditioning phase. Furthermore, Bdnf expression was enhanced in the VTA during the test phase. Bdnf induction seemed to be glucocorticoid-dependent, given that was correlated with HPA axis function and was not observed in morphine-dependent ADX animals. In addition, BDNF regulation and function was opposite in the VTA and mPFC during aversive-withdrawal memory retrieval. Our results suggest that IEGs and BDNF in these brain regions may play key roles in mediating the negative motivational component of opiate withdrawal.

PGE2-EP3 signaling exacerbates hippocampus-dependent cognitive impairment after laparotomy by reducing expression levels of hippocampal synaptic plasticity-related proteins in aged mice.

AIM: Multifactors contribute to the development of postoperative cognitive dysfunction (POCD), of which the most important mechanism is neuroinflammation. Prostaglandin E2 (PGE2) is a key neuroinflammatory molecule and could modulate hippocampal synaptic transmission and plasticity. This study was designed to investigate whether PGE2 and its receptors signaling pathway were involved in the pathophysiology of POCD. METHODS: Sixteen-month old male C57BL/6J mice were exposed to laparotomy. Cognitive function was evaluated by fear conditioning test. The levels of PGE2 and its 4 distinct receptors (EP1-4) were assessed by biochemical analysis. Pharmacological or genetic methods were further applied to investigate the role of the specific PGE2 receptors. RESULTS: Here, we found that the transcription and translation level of the EP3 receptor in hippocampus increased remarkably, but not EP1, EP2, or EP4. Immunofluorescence results showed EP3 positive cells in the hippocampal CA1 region were mainly neurons. Furthermore, pharmacological blocking or genetic suppression of EP3 could alleviate surgery-induced hippocampus-dependent memory deficits and rescued the expression of plasticity-related proteins, including cAMP response element-binding protein (CREB), activity-regulated cytoskeletal-associated protein (Arc), and brain-derived neurotrophic factor (BDNF) in hippocampus. CONCLUSION: This study showed that PGE2-EP3 signaling pathway was involved in the progression of POCD and identified EP3 receptor as a promising treatment target.

Different doses of sevoflurane facilitate and impair learning and memory function through activation of the ERK pathway and synthesis of ARC protein in the rat hippocampus.

BACKGROUND: Sevoflurane has been shown to stimulate or depress memory in adult rats; however, the cellular mechanism of this bidirectional effect has not been fully investigated. METHODS: We used an intra-hippocampal microinfusion of U0126 to suppress ERK activation. Male SD rats were randomly assigned to four groups: Sham, 0.11%SEV, 0.3%SEV and 0.3%+U0126. They received bilateral injections of U0126 or saline. Rats were anesthetized, and Inhibitory Avoidance (IA) training was performed immediately after anesthesia. The memory retention latency was observed 24 h later. In another experiment, the hippocampus was removed 45 min after IA training to assess ARC expression, the synapsin 1 protein levels and the phosphorylation level of ERK. RESULTS: Treatment with 0.11%SEV led to rapid phosphorylation of ERK, while 0.3%SEV inhibited phosphorylation; the latter change was reversed by the microinfusion of U0126 in the hippocampus. The memory latency result had similar tendencies. The local infusion of U0126 abolished the 0.3%SEV-induced memory impairment and ERK inhibition. Selective upregulations of ARC and synapsin 1 proteins were observed in the 0.3%SEV group compared with the 0.11%SEV group. CONCLUSIONS: The results indicate that different doses of sevoflurane trigger synaptic plasticity-related cytoskeleton proteins through the ERK signaling pathway. This novel modulation by inhalational agents may help to reduce their side-effects on memory function.

Acute stress and nicotine cues interact to unveil locomotor arousal and activity-dependent gene expression in the prefrontal cortex.

BACKGROUND: This study examines the interactive effects of acute stress and nicotine-associated contextual cues on locomotor activity and activity-dependent gene expression in subregions of the prefrontal cortex. METHODS: Locomotor activity of rats was measured in a context associated with either low-dose nicotine or saline administration with or without 5 minutes of pre-exposure to ferrets, a nonphysical stressor. After 45 minutes in the test environment, plasma corticosterone levels and mRNA levels of the immediate-early genes Arc, NGFI-B, and c-Fos in prefrontal and primary motor cortical subregions were measured. RESULTS: Stress alone increased plasma corticosterone and prefrontal cortex gene expression. Low-dose nicotine cues had no effect on corticosterone levels nor did they elicit conditioned motor activation, and they caused minor elevations in gene expression. Stress and low-dose nicotine cues, however, interacted to elicit conditioned motor activation and further increases in early response gene expression in prefrontal but not in the primary motor cortical subregions. CONCLUSIONS: Stress interacts with nicotine-associated cues to uncover locomotor arousal, a state associated with prefrontal neuronal activation and immediate early gene expression. Thus, in nicotine-experienced individuals, stress may be an important determinant of subjective reactivity and prefrontal cortex activation that occurs in response to nicotine-associated cues.

Downregulation of interferon alpha but not gamma receptor expression in vivo in the acquired immunodeficiency syndrome.

Interferons (IFN) elicit antiviral and antineoplastic activities by binding to specific receptors on the cell surface. In evaluating the role of IFN as therapeutic agents in AIDS, we investigated the expression of IFN alpha and gamma receptors on peripheral blood mononuclear cells (PBM) from patients with AIDS, ARC, and heterosexual control subjects using radioiodinated IFN alpha 2 and IFN gamma. The binding characteristics of the 125I-IFN alpha and gamma to PBM were analyzed to determine receptor numbers and dissociation constants. PBM from controls expressed 498 +/- 247 IFN alpha receptor sites/cell (n = 17). However, eight patients with ARC and seven patients with AIDS had a mean number of IFN alpha receptor/cell of 286 +/- 235 (P less than 0.05) and 92 +/- 88 (P less than 0.001), respectively. This was consistent with elevated levels of serum acid-labile IFN alpha and cellular 2-5A synthetase activity in patients. Treatment of PBM from the AIDS patients with exogenous IFN alpha in vitro resulted in minimal 2-5A synthetase induction in comparison to controls. In contrast, the expression of IFN gamma receptors in ARC (n = 5) and AIDS (n = 4) patients remained normal. Thus the decrease in IFN alpha receptor expression and consequent hyporesponsiveness to IFN alpha raises the question of the usefulness of IFN alpha therapy in end-stage AIDS. The normal expression of IFN gamma receptors in AIDS patients suggests that IFN gamma may prove useful in attempts to provide immune reconstitution.