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Background: Genomic sequencing in congenital heart disease (CHD) patients often discovers novel genetic variants, which are classified as variants of uncertain significance (VUS). Functional analysis of each VUS is required in specialised laboratories, to determine whether the VUS is disease causative or not, leading to lengthy diagnostic delays. We investigated stem cell cardiac disease modelling and transcriptomics for the purpose of genetic variant classification using a GATA4 (p.Arg283Cys) VUS in a patient with CHD., Methods: We performed high efficiency CRISPR gene editing with homology directed repair in induced pluripotent stem cells (iPSCs), followed by rapid clonal selection with amplicon sequencing. Genetic variant and healthy matched control cells were compared using cardiomyocyte disease modelling and transcriptomics., Results: Genetic variant and healthy cardiomyocytes similarly expressed Troponin T (cTNNT), and GATA4. Transcriptomics analysis of cardiomyocyte differentiation identified changes consistent with the patient's clinical human phenotype ontology terms. Further, transcriptomics revealed changes in calcium signalling, and cardiomyocyte adrenergic signalling in the variant cells. Functional testing demonstrated, altered action potentials in GATA4 genetic variant cardiomyocytes were consistent with patient cardiac abnormalities., Conclusions: This work provides in vivo functional studies supportive of a damaging effect on the gene or gene product. Furthermore, we demonstrate the utility of iPSCs, CRISPR gene editing and cardiac disease modelling for genetic variant interpretation. The method can readily be applied to other genetic variants in GATA4 or other genes in cardiac disease, providing a centralised assessment pathway for patient genetic variant interpretation., (© 2023. The Author(s).)

Long-term memory (LTM) can be induced by repeated spaced training trials. Using the weak inhibitory avoidance (wIA) task, we showed that one wIA session does not lead to a 24-h LTM, whereas two identical wIA sessions spaced by 15 min to 6 h induce a 24-h LTM. This LTM promotion depends both on hippocampal protein synthesis and the activity of several kinases. In agreement with the behavioral tagging (BT) hypothesis, our results suggest that the two training sessions induce transient learning tags and lead, via a cooperative effect, to the synthesis of plasticity-related proteins (PRPs) that become available and captured by the tag from the second session. Although ERKs1/2 are needed for PRPs synthesis and CaMKs are required for tag setting, PKA participates in both processes. We conclude that the BT mechanism accounts for the molecular constraints underlying the classic effect of spaced learning on LTM formation., Competing Interests: The authors declare no competing interests., (© 2023 The Authors.)

The globally important carbon sink of intact, old-growth tropical humid forests is declining because of climate change, deforestation and degradation from fire and logging 1-3 . Recovering tropical secondary and degraded forests now cover about 10% of the tropical forest area 4 , but how much carbon they accumulate remains uncertain. Here we quantify the aboveground carbon (AGC) sink of recovering forests across three main continuous tropical humid regions: the Amazon, Borneo and Central Africa 5,6 . On the basis of satellite data products 4,7 , our analysis encompasses the heterogeneous spatial and temporal patterns of growth in degraded and secondary forests, influenced by key environmental and anthropogenic drivers. In the first 20 years of recovery, regrowth rates in Borneo were up to 45% and 58% higher than in Central Africa and the Amazon, respectively. This is due to variables such as temperature, water deficit and disturbance regimes. We find that regrowing degraded and secondary forests accumulated 107 Tg C year -1 (90-130 Tg C year -1 ) between 1984 and 2018, counterbalancing 26% (21-34%) of carbon emissions from humid tropical forest loss during the same period. Protecting old-growth forests is therefore a priority. Furthermore, we estimate that conserving recovering degraded and secondary forests can have a feasible future carbon sink potential of 53 Tg C year -1 (44-62 Tg C year -1 ) across the main tropical regions studied., (© 2023. The Author(s), under exclusive licence to Springer Nature Limited.)

Each year, hundreds of millions of individuals are affected by respiratory disease leading to approximately 4 million deaths. Most respiratory pathologies involve substantially dysregulated immune processes that either fail to resolve the underlying process or actively exacerbate the disease. Therefore, clinicians have long considered immune-modulating corticosteroids (CSs), particularly glucocorticoids (GCs), as a critical tool for management of a wide spectrum of respiratory conditions. However, the complex interplay between effectiveness, risks and side effects can lead to different results, depending on the disease in consideration. In this comprehensive review, we present a summary of the bench and the bedside evidence regarding GC treatment in a spectrum of respiratory illnesses. We first describe here the experimental evidence of GC effects in the distal airways and/or parenchyma, both in vitro and in disease-specific animal studies, then we evaluate the recent clinical evidence regarding GC treatment in over 20 respiratory pathologies. Overall, CS remain a critical tool in the management of respiratory illness, but their benefits are dependent on the underlying pathology and should be weighed against patient-specific risks., Competing Interests: Competing interests: Octavian Ioachimescu is an Associate Editor for the Journal of Investigative Medicine., (© American Federation for Medical Research 2022. Re-use permitted under CC BY-NC. No commercial re-use. Published by BMJ.)

Complex organ development depends on single lumen formation and its expansion during tubulogenesis. This can be achieved by correct mitotic spindle orientation during cell division, combined with luminal fluid filling that generates hydrostatic pressure. Using a human 3D cell culture model, we have identified two regulators of these processes. We find that pleckstrin homology leucine-rich repeat protein phosphatase (PHLPP) 2 regulates mitotic spindle orientation, and thereby midbody positioning and maintenance of a single lumen. Silencing the sole PHLPP family phosphatase in Drosophila melanogaster, phlpp, resulted in defective spindle orientation in Drosophila neuroblasts. Importantly, cystic fibrosis transmembrane conductance regulator (CFTR) is the main channel regulating fluid transport in this system, stimulated by phosphorylation by protein kinase A and inhibited by the AMP-activated protein kinase AMPK. During lumen expansion, CFTR remains open through the action of PHLPP1, which stops activated AMPK from inhibiting ion transport through CFTR. In the absence of PHLPP1, the restraint on AMPK activity is lost and this tips the balance in the favour of channel closing, resulting in the lack of lumen expansion and accumulation of mucus., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2022. Published by The Company of Biologists Ltd.)

Background: Since the irruption of the coronavirus disease 2019 (COVID-19) the planet has submerged in a time of concern and uncertainty, with a direct impact on people's mental health. Moreover, the recurrent outbreaks that periodically harry different regions of the world constantly refocus people's concerns to the pandemic. Yet, each new wave heats the diverse countries in different situations, including the advances in their vaccination campaigns. In this research, we studied the levels of the general anxiety disorder (GAD) and depression in the Argentine population across the first and second waves of infections that occurred in our country., Methods: We conducted an on-line survey, within each peak of the pandemic. People were asked to self-report GAD and depression symptoms using the GAD-7 and PHQ-9 questioners, inform their vaccination status, the frequency they performed physical activity as well as working condition and modality. Here, we identified the more vulnerable groups and evaluated factors that could mitigate the rise of these mental disorders, focusing on vaccination., Results: Our data shows that reported GAD and depression levels were higher during the second wave than during the first one. More importantly, vaccinated people were less depressed than non-vaccinated people, while GAD levels remained equivalent in both groups. Other factors directly associated with lower GAD and depression levels were performing frequent physical activity and being employed, regardless of the employment modality. These observations were replicated in different age ranges and genders., Conclusion: This work evidences GAD and depression in different pandemic waves in Argentina, as well the factors that may contribute to reducing the magnitude of these disorders, including vaccination., 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 Benedetti, Barbuzza, Moscato, Reppucci, Goyeneche, Katche, Medina, Viola, Ballarini and Moncada.)

Background: There is an emerging understanding that coronavirus disease 2019 (COVID-19) is associated with increased incidence of pneumomediastinum. We aimed to determine its incidence among patients hospitalised with COVID-19 in the United Kingdom and describe factors associated with outcome., Methods: A structured survey of pneumomediastinum and its incidence was conducted from September 2020 to February 2021. United Kingdom-wide participation was solicited via respiratory research networks. Identified patients had SARS-CoV-2 infection and radiologically proven pneumomediastinum. The primary outcomes were to determine incidence of pneumomediastinum in COVID-19 and to investigate risk factors associated with patient mortality., Results: 377 cases of pneumomediastinum in COVID-19 were identified from 58 484 inpatients with COVID-19 at 53 hospitals during the study period, giving an incidence of 0.64%. Overall 120-day mortality in COVID-19 pneumomediastinum was 195/377 (51.7%). Pneumomediastinum in COVID-19 was associated with high rates of mechanical ventilation. 172/377 patients (45.6%) were mechanically ventilated at the point of diagnosis. Mechanical ventilation was the most important predictor of mortality in COVID-19 pneumomediastinum at the time of diagnosis and thereafter (p<0.001) along with increasing age (p<0.01) and diabetes mellitus (p=0.08). Switching patients from continuous positive airways pressure support to oxygen or high flow nasal oxygen after the diagnosis of pneumomediastinum was not associated with difference in mortality., Conclusions: Pneumomediastinum appears to be a marker of severe COVID-19 pneumonitis. The majority of patients in whom pneumomediastinum was identified had not been mechanically ventilated at the point of diagnosis., Competing Interests: Conflict of interest: James Melhorn has nothing to disclose. Conflict of interest: Andrew Achaiah has nothing to disclose. Conflict of interest: Francesca M. Conway has nothing to disclose. Conflict of interest: Elizabeth M. F. Thompson has nothing to disclose. Conflict of interest: Erik W. Skyllberg has nothing to disclose. Conflict of interest: Joseph Durrant has nothing to disclose. Conflict of interest: Neda A. Hasan has nothing to disclose. Conflict of interest: Yasser Madani has nothing to disclose. Conflict of interest: Prasheena Naran has nothing to disclose. Conflict of interest: Bavithra Vijayakumar has nothing to disclose. Conflict of interest: Matthew J. Tate has nothing to disclose. Conflict of interest: Gareth E. Trevelyan has nothing to disclose. Conflict of interest: Irfan Zaki has nothing to disclose. Conflict of interest: Catherine A. Doig has nothing to disclose. Conflict of interest: Geraldine Lynch has nothing to disclose. Conflict of interest: Gill Warwick has nothing to disclose. Conflict of interest: Avinash Aujayeb has nothing to disclose. Conflict of interest: Karl A. Jackson has nothing to disclose. Conflict of interest: Hina Iftikhar has nothing to disclose. Conflict of interest: Jonathan H. Noble has nothing to disclose. Conflict of interest: Anthony Y. K. C. Ng has nothing to disclose. Conflict of interest: Mark Nugent has nothing to disclose. Conflict of interest: Philip J. Evans has nothing to disclose. Conflict of interest: A. Hastings has nothing to disclose. Conflict of interest: Harry R. Bellenberg has nothing to disclose. Conflict of interest: Hannah Lawrence has nothing to disclose. Conflict of interest: Rachel L. Saville has nothing to disclose. Conflict of interest: Nikolas T. Johl has nothing to disclose. Conflict of interest: Adam N. Grey has nothing to disclose. Conflict of interest: Huw C. Ellis has nothing to disclose. Conflict of interest: Cheng Chen has nothing to disclose. Conflict of interest: Thomas L. Jones has nothing to disclose. Conflict of interest: Nadeem Maddekar has nothing to disclose. Conflict of interest: Shahul Leyakathali Khan has nothing to disclose. Conflict of interest: Ambreen Iqbal Muhammad has nothing to disclose. Conflict of interest: Hakim Ghani has nothing to disclose. Conflict of interest: Yadee Maung Maung Myint has nothing to disclose. Conflict of interest: Cecillia Rafique has nothing to disclose. Conflict of interest: Benjamin J. Pippard has nothing to disclose. Conflict of interest: Benjamin R. H. Irving has nothing to disclose. Conflict of interest: Fawad Ali has nothing to disclose. Conflict of interest: Viola H. Asimba has nothing to disclose. Conflict of interest: Aqeem Azam has nothing to disclose. Conflict of interest: Eleanor C. Barton has nothing to disclose. Conflict of interest: Malvika Bhatnagar has nothing to disclose. Conflict of interest: Matthew P. Blackburn has nothing to disclose. Conflict of interest: Kate J. Millington has nothing to disclose. Conflict of interest: Nicholas J. Budhram has nothing to disclose. Conflict of interest: Katherine L. Bunclark has nothing to disclose. Conflict of interest: Toshit P. Sapkal has nothing to disclose. Conflict of interest: Giles Dixon has nothing to disclose. Conflict of interest: Andrew J. E. Harries has nothing to disclose. Conflict of interest: Mohammad Ijaz has nothing to disclose. Conflict of interest: Vijayalakshmi Karunanithi has nothing to disclose. Conflict of interest: Samir Naik has nothing to disclose. Conflict of interest: Malik Aamaz Khan has nothing to disclose. Conflict of interest: Karishma Savlani has nothing to disclose. Conflict of interest: Vimal Kumar has nothing to disclose. Conflict of interest: Beatriz Lara Gallego has nothing to disclose. Conflict of interest: Noor A. Mahdi has nothing to disclose. Conflict of interest: Caitlin Morgan has nothing to disclose. Conflict of interest: Neena Patel has nothing to disclose. Conflict of interest: Elen W. Rowlands has nothing to disclose. Conflict of interest: Matthew S. Steward has nothing to disclose. Conflict of interest: Richard S. Thorley has nothing to disclose. Conflict of interest: Rebecca L. Wollerton has nothing to disclose. Conflict of interest: Sana Ullah has nothing to disclose. Conflict of interest: David M. Smith has nothing to disclose. Conflict of interest: Wojciech Lason has nothing to disclose. Conflict of interest: Anthony J Rostron has nothing to disclose. Conflict of interest: Najib M Rahman has nothing to disclose. Conflict of interest: Rob J Hallifax has nothing to disclose., (Copyright ©The authors 2022.)

The behavioral tagging (BT) hypothesis postulates that a weak learning experience, which only induces short-term memory, may benefit from another event that provides plasticity-related proteins (PRPs) to establish a long-lasting memory. According to BT, the weak experience sets a transient learning tag at specific activated sites, and its temporal and spatial convergence with the PRPs allows the long-term memory (LTM) formation. In this work, rats were subjected to a weak inhibitory avoidance (IAw) training and we observed that acute stress (elevated platform, EP) experienced 1 hr before IAw promoted IA-LTM formation. This effect was dependent on glucocorticoid-receptor activity as well as protein synthesis in the dorsal hippocampus. However, the same stress has negative effects on IA-LTM formation when training is strong, probably by competing for necessary PRPs. Furthermore, our experiments showed that EP immediately after training did not impair the setting of the learning tag and even facilitated IA-LTM formation. These findings reveal different impacts of a given acute stressful experience on the formation of an aversive memory that could be explained by BT processes., (© 2021 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

High-throughput tissue barrier models can yield critical insights on how barrier function responds to therapeutics, pathogens, and toxins. However, such models often emphasize multiplexing capability at the expense of physiologic relevance. Particularly, the distal lung's air-blood barrier is typically modeled with epithelial cell monoculture, neglecting the substantial contribution of endothelial cell feedback in the coordination of barrier function. An obstacle to establishing high-throughput coculture models relevant to the epithelium/endothelium interface is the requirement for underside cell seeding, which is difficult to miniaturize and automate. Therefore, this paper describes a scalable, low-cost seeding method that eliminates inversion by optimizing medium density to float cells so they attach under the membrane. This method generates a 96-well model of the distal lung epithelium-endothelium barrier with serum-free, glucocorticoid-free air-liquid differentiation. The polarized epithelial-endothelial coculture exhibits mature barrier function, appropriate intercellular junction staining, and epithelial-to-endothelial transmission of inflammatory stimuli such as polyinosine:polycytidylic acid (poly(I:C)). Further, exposure to influenza A virus PR8 and human beta-coronavirus OC43 initiates a dose-dependent inflammatory response that propagates from the epithelium to endothelium. While this model focuses on the air-blood barrier, the underside seeding method is generalizable to various coculture tissue models for scalable, physiologic screening., (© 2021 Wiley-VCH GmbH.)

Sarcomeric gene mutations are associated with the development of hypertrophic cardiomyopathy (HCM). Current drug therapeutics for HCM patients are effective in relieving symptoms, but do not prevent or reverse disease progression. Moreover, due to heterogeneity in the clinical manifestations of the disease, patients experience variable outcomes in response to therapeutics. Mechanistically, alterations in calcium handling, sarcomeric disorganization, energy metabolism and contractility participate in HCM disease progression. While some similarities exist, each mutation appears to lead to mutation-specific pathophysiology. Furthermore, these alterations may precede or proceed development of the pathology. This review assesses the efficacy of HCM therapeutics from studies performed in animal models of HCM and human clinical trials. Evidence suggests that a preventative rather than corrective therapeutic approach may be more efficacious in the treatment of HCM. In addition, a clear understanding of mutation-specific mechanisms may assist in informing the most effective therapeutic mode of action., (© 2020 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Tropical secondary forests sequester carbon up to 20 times faster than old-growth forests. This rate does not capture spatial regrowth patterns due to environmental and disturbance drivers. Here we quantify the influence of such drivers on the rate and spatial patterns of regrowth in the Brazilian Amazon using satellite data. Carbon sequestration rates of young secondary forests (<20 years) in the west are ~60% higher (3.0 ± 1.0 Mg C ha -1  yr -1 ) compared to those in the east (1.3 ± 0.3 Mg C ha -1  yr -1 ). Disturbances reduce regrowth rates by 8-55%. The 2017 secondary forest carbon stock, of 294 Tg C, could be 8% higher by avoiding fires and repeated deforestation. Maintaining the 2017 secondary forest area has the potential to accumulate ~19.0 Tg C yr -1 until 2030, contributing ~5.5% to Brazil's 2030 net emissions reduction target. Implementing legal mechanisms to protect and expand secondary forests whilst supporting old-growth conservation is, therefore, key to realising their potential as a nature-based climate solution.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The restoration and reforestation of 12 million hectares of forests by 2030 are amongst the leading mitigation strategies for reducing carbon emissions within the Brazilian Nationally Determined Contribution targets assumed under the Paris Agreement. Understanding the dynamics of forest cover, which steeply decreased between 1985 and 2018 throughout Brazil, is essential for estimating the global carbon balance and quantifying the provision of ecosystem services. To know the long-term increment, extent, and age of secondary forests is crucial; however, these variables are yet poorly quantified. Here we developed a 30-m spatial resolution dataset of the annual increment, extent, and age of secondary forests for Brazil over the 1986-2018 period. Land-use and land-cover maps from MapBiomas Project (Collection 4.1) were used as input data for our algorithm, implemented in the Google Earth Engine platform. This dataset provides critical spatially explicit information for supporting carbon emissions reduction, biodiversity, and restoration policies, enabling environmental science applications, territorial planning, and subsidizing environmental law enforcement.

Research on honeybee memory has led to a widely accepted model in which a single pairing of an odor stimulus with sucrose induces memories that are independent of protein synthesis but is unable to form protein-synthesis-dependent long-term memory (LTM). The latter is said to arise only after three or more pairings of odor and sucrose. Here, we show that this model underestimates the capacity of the bee brain to form LTMs after a unique appetitive experience. Using state-of-the art conditioning setups and individual-based analyses of conditioned responses, we found that protein-synthesis-dependent memories are formed already 4 h after the single conditioning trial and persist even 3 days later. These memories (4 h, 24 h, and 72 h) exhibit different dependencies on transcription and translation processes. Our results thus modify the traditional view of one-trial memories in an insect with a model status for memory research., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

One of the top challenges in education and neuroscience consists in translating laboratory results into strategies to improve learning and memory in teaching environments. In that sense, during the last two decades, researchers have discovered specific temporal windows around learning, during which the intervention with some experiences induces modulatory effects on the formation and/or persistence of memory. Based on these results, the aim of the present study was to design a specific strategy to improve the memory of students in a high-school scenario, by assessing the effect of a novel situation experienced close to learning. We found that the long-term memory about a geometrical figure was more precise in the group of students that faced a novel situation 1 h before or after learning the figure than the control group of students who did not face the novelty. This enhancement was probably triggered by processes acting on memory formation mechanisms that remained evident 45 days after learning, indicating that the improvement was sustained over time. In addition, our results showed that novelty no longer improved the memory if it was experienced 4 h before or after learning. However, far beyond this window of efficacy, when it was faced around 10 h after learning, the novel experience improved the memory persistence tested 7 days later. In summary, our findings characterized different temporal windows of the effectiveness of novelty acting on memory processing, providing a simple and inexpensive strategy that could be used to improve memory formation and persistence in high-school students., (Copyright © 2020 Ramirez Butavand, Hirsch, Tomaiuolo, Moncada, Viola and Ballarini.)

The superiority of spaced over massed learning is an established fact in the formation of long-term memories (LTM). Here we addressed the cellular processes and the temporal demands of this phenomenon using a weak spatial object recognition (wSOR) training, which induces short-term memories (STM) but not LTM. We observed SOR-LTM promotion when two identical wSOR training sessions were spaced by an inter-trial interval (ITI) ranging from 15 min to 7 h, consistently with spaced training. The promoting effect was dependent on neural activity, protein synthesis and ERKs1/2 activity in the hippocampus. Based on the "behavioral tagging" hypothesis, which postulates that learning induces a neural tag that requires proteins to induce LTM formation, we propose that retraining will mainly retag the sites initially labeled by the prior training. Thus, when weak, consecutive training sessions are experienced within an appropriate spacing, the intracellular mechanisms triggered by each session would add, thereby reaching the threshold for protein synthesis required for memory consolidation. Our results suggest in addition that ERKs1/2 kinases play a dual role in SOR-LTM formation after spaced learning, both inducing protein synthesis and setting the SOR learning-tag. Overall, our findings bring new light to the mechanisms underlying the promoting effect of spaced trials on LTM formation.

Complex in vitro models of the tissue microenvironment, termed microphysiological systems, have enormous potential to transform the process of discovering drugs and disease mechanisms. Such a paradigm shift is urgently needed in acute respiratory distress syndrome (ARDS), an acute lung condition with no successful therapies and a 40% mortality rate. Here, we consider how microphysiological systems could improve understanding of biological mechanisms driving ARDS and ultimately improve the success of therapies in clinical trials. We first discuss how microphysiological systems could explain the biological mechanisms underlying the segregation of ARDS patients into two clinically distinct phenotypes. Then, we contend that ARDS-mimetic microphysiological systems should recapitulate three critical aspects of the distal airway microenvironment, namely, mechanical force, inflammation, and fibrosis, and we review models that incorporate each of these aspects. Finally, we recognize the substantial challenges associated with combining inflammation, fibrosis, and/or mechanical force in microphysiological systems. Nevertheless, complex in vitro models are a novel paradigm for studying ARDS, and they could ultimately improve patient care., (© Author(s).)

Scar formation after wound healing is a major medical problem. A better understanding of the dynamic nuclear architecture of the genome during wound healing could provide insights into the underlying pathophysiology and enable novel therapeutic strategies. Here, we demonstrate that TGF-β-induced fibrotic stress increases formation of the dynamic secondary DNA structures called G-quadruplexes in skin fibroblasts, which is coincident with increased expression of collagen 1. This G-quadruplex formation is attenuated by a small molecule inhibitor of intracellular Ca 2+ influx and an anti-fibrotic compound. In addition, we identify G-quadruplex-forming sequences in the promoter region of COL1A1, which encodes collagen 1, and confirm their ability to form G-quadruplex structures under physiologically relevant conditions. Our findings reveal a link between G-quadruplexes and scar formation that may lead to novel therapeutic interventions., (© 2019 Federation of European Biochemical Societies.)

Honeybees are a standard model for the study of appetitive learning and memory. Yet, fewer attempts have been performed to characterize aversive learning and memory in this insect and uncover its molecular underpinnings. Here, we took advantage of the positive phototactic behavior of bees kept away from the hive in a dark environment and established a passive-avoidance task in which they had to suppress positive phototaxis. Bees placed in a two-compartment box learned to inhibit spontaneous attraction to a compartment illuminated with blue light by associating and entering into that chamber with shock delivery. Inhibitory learning resulted in an avoidance memory that could be retrieved 24 h after training and that was specific to the punished blue light. The memory was mainly operant but involved a Pavlovian component linking the blue light and the shock. Coupling conditioning with transcriptional analyses in key areas of the brain showed that inhibitory learning of phototaxis leads to an up-regulation of the dopaminergic receptor gene Amdop1 in the calyces of the mushroom bodies, consistently with the role of dopamine signaling in different forms of aversive learning in insects. Our results thus introduce new perspectives for uncovering further cellular and molecular underpinnings of aversive learning and memory in bees. Overall, they represent an important step toward comparative learning studies between the appetitive and the aversive frameworks., (© 2019 Marchal et al.; Published by Cold Spring Harbor Laboratory Press.)

Stress is known to have a critical impact on memory processes. In the present work, we focus on the effects of an acute stress event closely associated to an unrelated learning task. Here, we show that acute stress (elevated platform [EP] session) experienced 1 hr after a weak spatial object recognition (SOR) training, which only induces a short-term memory (STM), promoted the formation of SOR-long term memory (SOR-LTM) in rats. The effect induced by stress was dependent on the activation of glucocorticoid- and mineralocorticoid-receptors, brain-derived neurotrophic factor (BDNF) and protein synthesis in the dorsal hippocampus. In contrast, EP after a strong SOR impaired SOR-LTM probably by interfering with the use of necessary resources. Moreover, we show that the EP session before training induced anterograde interference, which it was not reversed by a subsequent exposure to an open field. Our findings provide novel insights into the impact of stress on LTM formation in rodents and they are discussed under the behavioral analogue of the synaptic tagging and capture hypothesis., (© 2018 Wiley Periodicals, Inc.)

Extracellular regulated kinase 1/2 (ERK1/2) has been strongly implicated in several cellular processes. In the brain ERK1/2 activity has been primarily involved in long-term memory (LTM) formation and expression. Here, we review earlier evidence and describe recent developments of ERK1/2 signaling in memory processing focusing the attention on the role of ERK1/2 in memory retrieval and reconsolidation, and in the maintenance of the memory trace including mechanisms involving the protection of labile memories. In addition, relearning requires ERK1/2 activity in selected brain regions. Its involvement in distinct memory stages points at ERK1/2 as a core element in memory processing and as one likely target to treat memory impairments associated with neurological disorders.

The biological effects of gamma radiation may exert damage beyond that of the individual through its deleterious effects on reproductive function. Impaired reproductive performance can result in reduced population size over consecutive generations. In a continued effort to investigate reproductive and heritable effects of ionizing radiation, we recently demonstrated adverse effects and genomic instability in progeny of parents exposed to gamma radiation. In the present study, genotoxicity and effects on the reproduction following subchronic exposure during a gametogenesis cycle to 60 Co gamma radiation (27 days, 8.7 and 53 mGy/h, total doses 5.2 and 31 Gy) were investigated in the adult wild-type zebrafish (Danio rerio). A significant reduction in embryo production was observed one month after exposure in the 53 mGy/h exposure group compared to control and 8.7 mGy/h. One year later, embryo production was significantly lower in the 53 mGy/h group compared only to control, with observed sterility, accompanied by a regression of reproductive organs in 100% of the fish 1.5 years after exposure. Histopathological examinations revealed no significant changes in the testis in the 8.7 mGy/h group, while in 62.5% of females exposed to this dose rate the oogenesis was found to be only at the early previtellogenic stage. The DNA damage determined in whole blood, 1.5 years after irradiation, using a high throughput Comet assay, was significantly higher in the exposed groups (1.2 and 3-fold increase in 8.7 and 53 mGy/h females respectively; 3-fold and 2-fold increase in 8.7 and 53 mGy/h males respectively) compared to controls. A significantly higher number of micronuclei (4-5%) was found in erythrocytes of both the 8.7 and 53 mGy/h fish compared to controls. This study shows that gamma radiation at a dose rate of ≥ 8.7 mGy/h during gametogenesis causes adverse reproductive effects and persistent genotoxicity (DNA damage and increased micronuclei) in adult zebrafish., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The heart is able to respond acutely to changes in oxygen tension. Since ion channels can respond rapidly to stimuli, the "ion channel oxygen sensing hypothesis" has been proposed to explain acute adaptation of cells to changes in oxygen demand. However the exact mechanism for oxygen sensing continues to be debated. Mitochondria consume the lion's share of oxygen in the heart, fuelling the production of ATP that drives excitation and contraction. Mitochondria also produce reactive oxygen species that are capable of altering the redox state of proteins. The cardiac L-type calcium channel is responsible for maintaining excitation and contraction. Recently, the reactive cysteine on the cardiac L-type calcium channel was identified. These data clarified that the channel does not respond directly to changes in oxygen tension, but rather responds to cellular redox state. This leads to acute alterations in cell signalling responsible for the development of arrhythmias and pathology., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

With the aim of analyzing if object recognition long-term memory (OR-LTM) formation is susceptible to retroactive interference (RI), we submitted rats to sequential sample sessions using the same arena but changing the identity of a pair of objects placed in it. Separate groups of animals were tested in the arena in order to evaluate the LTM for these objects. Our results suggest that OR-LTM formation was retroactively interfered within a critical time window by the exploration of a new, but not familiar, object. This RI acted on the consolidation of the object explored in the first sample session because its OR-STM measured 3h after training was not affected, whereas the OR-LTM measured at 24h was impaired. This sample session also impaired the expression of OR memory when it took place before the test. Moreover, local inactivation of the dorsal Hippocampus (Hp) or the medial Prefrontal Cortex (mPFC) previous to the exploration of the second pair of objects impaired their consolidation restoring the LTM for the objects explored in the first session. This data suggests that both brain regions are involved in the processing of OR-memory and also that if those regions are engaged in another process before finishing the first consolidation process its LTM will be impaired by RI., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Learning to avoid threats in the environment is highly adaptive. However, sometimes a dysregulation of fear memories processing may underlie fear-related disorders. Despite recent advances, a major question of how to effectively attenuate persistent fear memories in a safe manner remains unresolved. Here we show experiments employing a behavioural tool to target a specific time window after training to limit the persistence of a fear memory in rats. We observed that exposure to a novel environment 11 h after an inhibitory avoidance (IA) training that induces a long-lasting memory, attenuates the durability of IA memory but not its formation. This effect is time-restricted and not seen when the environment is familiar. In addition, novelty-induced attenuation of IA memory durability is prevented by the intrahippocampal infusion of the CaMKs inhibitor KN-93. This new behavioural approach which targets a specific time window during late memory consolidation, might represent a new tool for reducing the durability of persistent fear memories.

Severe burn injury significantly affects cardiovascular function for up to 3 years. However, whether this leads to long-term pathology is unknown. The impact of non-severe burn injury, which accounts for over 80% of admissions in developed countries, has not been investigated. Using a rodent model of non-severe burn injury with subsequent echocardiography we showed significantly increased left ventricular end systolic diameter (LVESD) and ventricular wall thickness at up to 3 months post-injury. Use of propranolol abrogated the changes in cardiac measures observed. Subsequently we investigated changes in a patient cohort with non-severe injury. Echocardiography measured at baseline and at 3 months post-injury showed increased LVESD at 3 months and significantly decreased posterior wall diameter. Finally, 32 years of Western Australian hospital records were used to investigate the incidence of cardiovascular disease admissions after burn injury. People who had experienced a burn had increased hospital admissions and length of stay for cardiovascular diseases when compared to a matched uninjured cohort. This study presents animal, patient and population data that strongly suggest non-severe burn injury has significant effects on cardiovascular function and long-term morbidity in some burn patients. Identification of patients at risk will promote better intervention and outcomes for burn patients.

Key Points: Genetic mutations in cardiac troponin I (cTnI) are associated with development of hypertrophic cardiomyopathy characterized by myocyte remodelling, disorganization of cytoskeletal proteins and altered energy metabolism. The L-type Ca(2+) channel is the main route for calcium influx and is crucial to cardiac excitation and contraction. The channel also regulates mitochondrial function in the heart by a functional communication between the channel and mitochondria via the cytoskeletal network. We find that L-type Ca(2+) channel kinetics are altered in cTnI-G203S cardiac myocytes and that activation of the channel causes a significantly greater increase in mitochondrial membrane potential and metabolic activity in cTnI-G203S cardiac myocytes. These responses occur as a result of impaired communication between the L-type Ca(2+) channel and cytoskeletal protein F-actin, involving decreased movement of actin-myosin and block of the mitochondrial voltage-dependent anion channel, resulting in a 'hypermetabolic' mitochondrial state. We propose that L-type Ca(2+) channel antagonists, such as diltiazem, might be effective in reducing the cardiomyopathy by normalizing mitochondrial metabolic activity., Abstract: Genetic mutations in cardiac troponin I (cTnI) account for 5% of families with hypertrophic cardiomyopathy. Hypertrophic cardiomyopathy is associated with disorganization of cytoskeletal proteins and altered energy metabolism. The L-type Ca(2+) channel (ICa-L ) plays an important role in regulating mitochondrial function. This involves a functional communication between the channel and mitochondria via the cytoskeletal network. We investigate the role of ICa-L in regulating mitochondrial function in 25- to 30-week-old cardiomyopathic mice expressing the human disease-causing mutation Gly203Ser in cTnI (cTnI-G203S). The inactivation rate of ICa-L is significantly faster in cTnI-G203S myocytes [cTnI-G203S: τ1  = 40.68 ± 3.22, n = 10 vs. wild-type (wt): τ1  = 59.05 ± 6.40, n = 6, P < 0.05]. Activation of ICa-L caused a greater increase in mitochondrial membrane potential (Ψm , 29.19 ± 1.85%, n = 15 vs. wt: 18.84 ± 2.01%, n = 10, P < 0.05) and metabolic activity (24.40 ± 6.46%, n = 8 vs. wt: 9.98 ± 1.57%, n = 9, P < 0.05). The responses occurred because of impaired communication between ICa-L and F-actin, involving lack of dynamic movement of actin-myosin and block of the mitochondrial voltage-dependent anion channel. Similar responses were observed in precardiomyopathic mice. ICa-L antagonists nisoldipine and diltiazem decreased Ψm to basal levels. We conclude that the Gly203Ser mutation leads to impaired functional communication between ICa-L and mitochondria, resulting in a 'hypermetabolic' state. This might contribute to development of cTnI-G203S cardiomyopathy because the response is present in young precardiomyopathic mice. ICa-L antagonists might be effective in reducing the cardiomyopathy by altering mitochondrial function., (© 2016 The Authors. The Journal of Physiology © 2016 The Physiological Society.)

Colorectal cancer, encompassing colon and rectal cancer, arises from the epithelial lining of the large bowel. It is most prevalent in Westernised societies and is increasing in frequency as the world becomes more industrialised. Unfortunately, metastatic colorectal cancer is not cured by chemotherapy and the annual number of deaths caused by colorectal cancer, currently 700,000, is expected to rise. Our understanding of the contribution that genetic mutations make to colorectal cancer, although incomplete, is reasonably well advanced. However, it has only recently become widely appreciated that in addition to the ongoing accumulation of genetic mutations, chronic inflammation also plays a critical role in the initiation and progression of this disease. While a robust and tractable genetic model of colorectal cancer in zebrafish, suitable for pre-clinical studies, is not yet available, the identification of genes required for the rapid proliferation of zebrafish intestinal epithelial cells during development has highlighted a number of essential genes that could be targeted to disable colorectal cancer cells. Moreover, appreciation of the utility of zebrafish to study intestinal inflammation is on the rise. In particular, zebrafish provide unique opportunities to investigate the impact of genetic and environmental factors on the integrity of intestinal epithelial barrier function. With currently available tools, the interplay between epigenetic regulators, intestinal injury, microbiota composition and innate immune cell mobilisation can be analysed in exquisite detail. This provides excellent opportunities to define critical events that could potentially be targeted therapeutically. Further into the future, the use of zebrafish larvae as hosts for xenografts of human colorectal cancer tissue, while still in its infancy, holds great promise that zebrafish could one day provide a practical, preclinical personalized medicine platform for the rapid assessment of the metastatic potential and drug-sensitivity of patient-derived cancers.

The medial prefrontal cortex (mPFC) is known for its role in decision making and memory processing, including the participation in the formation of extinction memories. However, little is known regarding its contribution to aversive memory consolidation. Here we demonstrate that neural activity and protein synthesis are required in the dorsal mPFC for memory formation of a conditioned taste aversion (CTA) task and that this region is involved in the retrieval of recent and remote long-term CTA memory. In addition, both NMDA receptor and CaMKII activity in dorsal mPFC are needed for CTA memory consolidation, highlighting the complexity of mPFC functions., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The synaptic tagging and capture (STC) hypothesis provides a compelling explanation for synaptic specificity and facilitation of long-term potentiation. Its implication on long-term memory (LTM) formation led to postulate the behavioral tagging mechanism. Here we show that a maintenance tagging process may operate in the hippocampus late after acquisition for the persistence of long-lasting memory storage. The proposed maintenance tagging has several characteristics: (1) the tag is transient and time-dependent; (2) it sets in a late critical time window after an aversive training which induces a short-lasting LTM; (3) exposing rats to a novel environment specifically within this tag time window enables the consolidation to a long-lasting LTM; (4) a familiar environment exploration was not effective; (5) the effect of novelty on the promotion of memory persistence requires dopamine D1/D5 receptors and Arc expression in the dorsal hippocampus. The present results can be explained by a broader version of the behavioral tagging hypothesis and highlight the idea that the durability of a memory trace depends either on late tag mechanisms induced by a training session or on events experienced close in time to this tag.

Similar molecular machinery is activated in neurons following an electrical stimulus that induces synaptic changes and after learning sessions that trigger memory formation. Then, to achieve perdurability of these processes protein synthesis is required for the reinforcement of the changes induced in the network. The synaptic tagging and capture theory provided a strong framework to explain synaptic specificity and persistence of electrophysiological induced plastic changes. Ten years later, the behavioral tagging hypothesis (BT) made use of the same argument, applying it to learning and memory models. The hypothesis postulates that the formation of lasting memories relies on at least two processes: the setting of a learning tag and the synthesis of plasticity related proteins, which once captured at tagged sites allow memory consolidation. BT explains how weak events, only capable of inducing transient forms of memories, can result in lasting memories when occurring close in time with other behaviorally relevant experiences that provide proteins. In this review, we detail the findings supporting the existence of BT process in rodents, leading to the consolidation, persistence, and interference of a memory. We focus on the molecular machinery taking place in these processes and describe the experimental data supporting the BT in humans.

Retroactive interference (RI) is a type of amnesia in which a new learning experience can impair the expression of a previous one. It has been studied in several types of memories for over a century. Here, we aimed to study in the long-term memory (LTM) formation of an object-in-context task, defined as the recognition of a familiar object in a context different to that in which it was previously encountered. We trained rats with two sample trials, each taking place in a different context in association with different objects. Test sessions were performed 24 h later, to evaluate LTM for both object-context pairs using separate groups of trained rats. Furthermore, given the involvement of hippocampus (Hp) and medial prefrontal cortex (mPFC) in several recognition memories, we also analyzed the participation of these structures in the LTM formation of this task by the local infusion of muscimol. Our results show that object-in-context LTM formation is sensitive to RI by a different either familiar or novel object-context pair trial, experienced 1 h later. This interference occurs in a restricted temporal window and works on the LTM consolidation phase, leaving intact short-term memory expression. The second sample trial did not affect the object recognition part of the memory. Besides, muscimol treatment before the second sample trial blocks its object-in-context LTM and restores the first sample trial memory. We hypothesized that LTM-RI amnesia is probably caused by resources or cellular machinery competition in these brain regions when they are engaged in memory formation of the traces. In sum, when two different object-in-context memory traces are being processed, the second trace interferes with the consolidation of the first one requiring mPFC and CA1 dorsal Hp activation., (© 2014 Wiley Periodicals, Inc.)

The synaptic tagging and capture theory (STC) was postulated by Frey and Morris in 1997 and provided a strong framework to explain how to achieve synaptic specificity and persistence of electrophysiological-induced plasticity changes. Ten years later, the same argument was applied on learning and memory models to explain the formation of long-term memories, resulting in the behavioral tagging hypothesis (BT). These hypotheses are able to explain how a weak event that induces transient changes in the brain can establish long-lasting phenomena through a tagging and capture process. In this framework, it was postulated that the weak event sets a tag that captures plasticity-related proteins/products (PRPs) synthesized by an independent strong event. The tagging and capture processes exhibit symmetry, and therefore, PRPs can be captured if they are synthesized either before or after the setting of the tag. In summary, the hypothesis provides a wide framework that gives a solid explanation of how lasting changes occur and how the interaction between different events leads to promotion, reinforcement, or impairment of such changes. In this chapter, we will summarize the postulates of STC hypothesis, the common features between synaptic plasticity and memory, as well as a detailed compilation of the findings supporting the existence of BT process. At the end, we pose some questions related to BT mechanism and LTM formation, which probably will be answered in the near future., (© 2014 Elsevier Inc. All rights reserved.)

Education is the most traditional means with formative effect on the human mind, learning and memory being its fundamental support. For this reason, it is essential to find different strategies to improve the studentś performance. Based on previous work, we hypothesized that a novel experience could exert an enhancing effect on learning and memory within the school environment. Here we show that novel experience improved the memory of literary or graphical activities when it is close to these learning sessions. We found memory improvements in groups of students who had experienced a novel science lesson 1 hour before or after the reading of a story, but not when these events were 4 hours apart. Such promoting effect on long-term memory (LTM) was also reproduced with another type of novelty (a music lesson) and also after another type of learning task (a visual memory). Interestingly, when the lesson was familiar, it failed to enhance the memory of the other task. Our results show that educationally relevant novel events experienced during normal school hours can improve LTM for tasks/activities learned during regular school lessons. This effect is restricted to a critical time window around learning and is particularly dependent on the novel nature of the associated experience. These findings provide a tool that could be easily transferred to the classroom by the incorporation of educationally novel events in the school schedule as an extrinsic adjuvant of other information acquired some time before or after it. This approach could be a helpful tool for the consolidation of certain types of topics that generally demand a great effort from the children.

Rationale: Regulator of G-protein signaling 5 (RGS5) modulates G-protein-coupled receptor signaling and is prominently expressed in arterial smooth muscle cells. Our group first reported that RGS5 is important in vascular remodeling during tumor angiogenesis. We hypothesized that RGS5 may play an important role in vessel wall remodeling and blood pressure regulation., Objective: To demonstrate that RGS5 has a unique and nonredundant role in the pathogenesis of hypertension and to identify crucial RGS5-regulated signaling pathways., Methods and Results: We observed that arterial RGS5 expression is downregulated with chronically elevated blood pressure after angiotensin II infusion. Using a knockout mouse model, radiotelemetry, and pharmacological inhibition, we subsequently showed that loss of RGS5 results in profound hypertension. RGS5 signaling is linked to the renin-angiotensin system and directly controls vascular resistance, vessel contractility, and remodeling. RGS5 deficiency aggravates pathophysiological features of hypertension, such as medial hypertrophy and fibrosis. Moreover, we demonstrate that protein kinase C, mitogen-activated protein kinase/extracellular signal-regulated kinase, and Rho kinase signaling pathways are major effectors of RGS5-mediated hypertension., Conclusions: Loss of RGS5 results in hypertension. Loss of RGS5 signaling also correlates with hyper-responsiveness to vasoconstrictors and vascular stiffening. This establishes a significant, distinct, and causal role of RGS5 in vascular homeostasis. RGS5 modulates signaling through the angiotensin II receptor 1 and major Gαq-coupled downstream pathways, including Rho kinase. So far, activation of RhoA/Rho kinase has not been associated with RGS molecules. Thus, RGS5 is a crucial regulator of blood pressure homeostasis with significant clinical implications for vascular pathologies, such as hypertension.

Recently encoded information can be lost in the presence of new information, a process called 'retrograde interference'. Retrograde interference has been extensively described for more than a century; however, little is known about its underlying mechanisms. Different approaches agree on the need of the synthesis of plasticity related proteins (PRPs) to consolidate a long-term memory (LTM). Our hypothesis is that when PRPs are limited, interference of a task over LTM formation of another may be due to the utilization of protein resources common to both tasks. Here, by combining the tasks of inhibitory avoidance (IA) and open field (OF) exploration in rats, we show that memory traces compete for their stabilization if PRPs are limited. As a result, LTM is formed for only one of the tasks with a consequent decrease in the memory for the other. Furthermore, infusing Arc antisense oligonucleotide into the dorsal hippocampus, we found that Arc is necessary for LTM formation of these two types of learning tasks and is one of the PRPs that can be shared between them when animals are trained in both OF and IA. In sum, these findings suggest that under conditions of reduced protein availability, a learning task interferes with LTM formation of another by using the available PRPs., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Long-term memory (LTM) consolidation requires the synthesis of plasticity-related proteins (PRPs). In addition, we have shown recently that LTM formation also requires the setting of a "learning tag" able to capture those PRPs. Weak training, which results only in short-term memory, can set a tag to use PRPs derived from a temporal-spatial closely related event to promote LTM formation. Here, we studied the involvement of glutamatergic, dopaminergic, and noradrenergic inputs on the setting of an inhibitory avoidance (IA) learning tag and the synthesis of PRPs. Rats explored an open field (PRP donor) followed by weak (tag inducer) or strong (tag inducer plus PRP donor) IA training. Throughout pharmacological interventions around open-field and/or IA sessions, we found that hippocampal dopamine D1/D5- and β-adrenergic receptors are specifically required to induce PRP synthesis. Moreover, activation of the glutamatergic NMDA receptors is required for setting the learning tags, and this machinery further required α-Ca(2+)/calmodulin-dependent protein kinase II and PKA but not ERK1/2 activity. Together, the present findings emphasize an essential role of the induction of PRPs and learning tags for LTM formation. The existence of only the PRP or the tag was insufficient for stabilization of the mnemonic trace.

Wnt signaling is involved in developmental processes, cell proliferation, and cell migration. Secreted frizzled-related protein 4 (sFRP4) has been demonstrated to be a Wnt antagonist; however, its effects on endothelial cell migration and angiogenesis have not yet been reported. Using various in vitro assays, we show that sFRP4 inhibits endothelial cell migration and the development of sprouts and pseudopodia as well as disrupts the stability of endothelial rings in addition to inhibiting proliferation. sFRP4 interfered with endothelial cell functions by antagonizing the canonical Wnt/beta-catenin signaling pathway and the Wnt/planar cell polarity pathway. Furthermore, sFRP4 blocked the effect of vascular endothelial growth factor on endothelial cells. sFRP4 also selectively induced apoptotic events in endothelial cells by increasing cellular levels of reactive oxygen species. In vivo assays demonstrated a reduction in vascularity after sFRP4 treatment. Most importantly, sFRP4 restricted tumor growth in mice by interfering with endothelial cell function. The data demonstrate sFRP4 to be a potent angiogenesis inhibitor that warrants further investigation as a therapeutic agent in the control of angiogenesis-associated pathology.

In daily life, memories are intertwined events. Little is known about the mechanisms involved in their interactions. Using two hippocampus-dependent (spatial object recognition and contextual fear conditioning) and one hippocampus-independent (conditioned taste aversion) learning tasks, we show that in rats subjected to weak training protocols that induce solely short term memory (STM), long term memory (LTM) is promoted and formed only if training sessions took place in contingence with a novel, but not familiar, experience occurring during a critical time window around training. This process requires newly synthesized proteins induced by novelty and reveals a general mechanism of LTM formation that begins with the setting of a "learning tag" established by a weak training. These findings represent the first comprehensive set of evidences indicating the existence of a behavioral tagging process that in analogy to the synaptic tagging and capture process, need the creation of a transient, protein synthesis-independent, and input specific tag.

The aim of this work was to evaluate if the intraperitoneal administration of the natural compound hesperidin, in a sedative dose, and neo-hesperidin, a hesperidin structural analog that exerts minor sedative effect, were able to induce changes in intracellular signaling cascades in different areas of the brain. The systemic administration of hesperidin produced a marked reduction in the phosphorylation state of extracellular signal-regulated kinases 1/2 (ERK 1/2), but not of Ca(+2)/calmodulin-dependent protein kinase II alpha subunit (alphaCaMKII), in the cerebral cortex, cerebellum and hippocampus. In contrast, neo-hesperidin did not markedly affect the activity of ERK 1/2 in both the cortex and the cerebellum. Taken together, these results demonstrated that intracellular signalling involving a selective decrease in ERK1/2 activation accompanied the depressant action of hesperidin. Even more, the low sedative action of neo-hesperidin correlates with a negligible decrease in phosphorylation state of ERK 1/2 (pERK 1/2), suggesting that low levels of pERK 1/2 in CNS could be a marker of sedative efficacy of flavonoids.

Spatial familiarization consists of a decrease in the exploratory activity over time after exposure to a place. Here, we show that a 30-min exposure to an open field led to a pronounced decrease in the exploratory behavior of rats, generating context familiarity. This behavioral output is associated with a selective decrease in hippocampal PKMzeta levels. A short 5-min exposure did not induce spatial familiarity or a decrease in PKMzeta, while inactivation of hippocampal PKMzeta by the specific inhibitor ZIP was sufficient to induce spatial familiarity, suggesting that the decrease in PKMzeta is involved in setting a given context as a familiar place.

A behavioral analog of the synaptic tagging and capture process, a key property of synaptic plasticity, has been predicted recently. Here, we demonstrate that weak inhibitory avoidance training, which induces short- but not long-term memory (LTM), can be consolidated into LTM by an exploration to a novel, but not a familiar, environment occurring close in time to the training session. This memory-promoting effect caused by novelty depends on activation of dopamine D1/D5 receptors and requires newly synthesized proteins in the dorsal hippocampus. Thus, our results indicate the existence of a behavioral tagging process in which the exploration to a novel environment provides the plasticity-related proteins to stabilize the inhibitory avoidance memory trace.

The glycan chains of glycoconjugates play important roles in cell-cell and cell-matrix interactions. In the CNS, previous studies on learning and memory suggest the importance of oligosaccharides attached to glycoconjugates in the modulation of synaptic connections. We studied the hippocampal glycan distribution of rats subject to an inhibitory avoidance task. The expression of glycans was examined by lectin-histochemistry using Vicia villosa lectin (VVL) for terminal alpha/beta N-acetylgalactosamine (alpha/beta GalNAc); Galanthus nivalus lectin (GNL) for terminal mannose alpha-1,3 (Man alpha-1,3); Peanut agglutinin (PNA) for galactose beta-1,3N-acetylgalactosamine (Gal beta-1,3 GalNAc); Erythrina cristagalli lectin (ECL) for galactose beta-1,4 N-acetylglucosamine (Gal beta-1,4 GlcNAc); Sambucus nigra lectin (SNA) for sialic acid alpha-2.6 galactose (SA alpha-2,6 Gal); Maackia amurensis lectin II (MAL II) for sialic acid alpha-2,3 (SA alpha-2,3); Wheat germ agglutinin (WGA) for terminal N-acetylglucosamine with/ without sialic acid (GlcNAc wo SA); succynilated WGA (sWGA) for terminal N-acetylglucosamine without sialic acid (terminal GlcNAc without SA); Griffonia simplicifolia lectin II (GSL II) for terminal alpha/beta N-acetylglucosamine (alpha/beta GlcNAc terminal); and Lotus tetragonolobus lectin (LTL) alpha-fucose. Two groups of 10 animals were examined: non-trained (Control) and Trained rats. ECL, sWGA and GSL II were negative for both groups in all the hippocampal subfields studied. For both groups, VVL was negative in CA4 and granular cells of the Dentate Gyrus (DG) and LTL was negative in the CA4 subfield. Expression of alpha/beta GalNAc, alpha-fucose and GlcNAc in other hippocampal subflields was positive, with no differences between groups. However, expression of Man alpha-1,3 was significantly higher in the CA1, CA2, CA3, and CA4 subfields in the Trained group. On the other hand, expression of Gal beta-1,3 GalNAc was significantly low in CA4 and DG in the Trained group. In conclusion, the results here presented indicate that the exposure of rats to an associative behavioral paradigm related to declarative memory, involves some regulatory mechanism/s for the differential patterns of glycan expression.

The activation of cAMP response element-binding protein (CREB) after a learning experience is a common feature in the formation of several associative memories. We recently demonstrated that the increase in the hippocampal phosphorylated CREB (pCREB) levels 1 h after a short exploration of an open field (OF) was associated to detection of spatial novelty and was not related to the memory formation of habituation in this non-associative learning paradigm. Moreover, after a long training of three OF sessions, hippocampal pCREB levels were below to that observed in control rats. The present results show that such decrease does not correlate with memory retrieval or improvement in long-term memory of habituation. Instead, it is associated with the familiarity to the arena. Our experiments revealed that the relevant variable to induce CREB deactivation was the prolonged exploration of the arena (30 min). A 15 min OF exploration was ineffective. Furthermore, the last 5 min period of a prolonged exploration was crucial to change CREB phosphorylation state: when exploration took place in a novel arena the level of pCREB increased; in contrast, when it was performed in the familiar OF, pCREB levels decreased. Taken as a whole, our results suggest that CREB phosphorylation state in the hippocampus switches in response to exposure to a novel or to a familiar spatial environment.

There is a growing body of evidence showing that the formation of associative memories is associated with an increase in phosphorylated cAMP response element-binding protein (pCREB) levels. We recently reported increased pCREB levels in the rat hippocampus after an exploration to a novel environment. In the present work, we studied whether this increment in CREB activation is associated with the formation of memory of habituation to a novel environment or with the detection of novelty. Rats were submitted to consecutive open field sessions at 3-h intervals. Measurement of the hippocampal pCREB level, carried out 1 h after each training session, showed that (1) it did not increase when rats explored a familiar environment; (2) it did not increase after a reexposure that improves the memory of habituation; (3) it increased after a brief novel exploration unable to form memory of habituation; and (4) it increased in amnesic rats for spatial habituation. Taken as a whole, our results suggest that the elevated pCREB level after a single open field exploration is not associated with the memory formation of habituation. It is indeed associated with the detection of a novel environment.

A pharmacist-managed clinic for treating latent tuberculosis infection (LTBI) in health care workers (HCWs) is described. The hospital-based clinic was begun in 1993 after a review showed that only 0.8% of HCWs at the institution who were prescribed isoniazid for LTBI completed it. The goals of the clinic are to improve HCWs' adherence to LTBI treatment, enhance outcomes, and minimize adverse drug reactions. The clinic is staffed by a pharmacist, a medical resident, and a nurse. If an employee is diagnosed with LTBI, he or she is referred to the clinic, where the pharmacist conducts an initial assessment and provides information on medication available for treatment. The worker is then scheduled for monthly follow-up visits with the pharmacist for the first three months of LTBI therapy, during which the pharmacist evaluates treatment adherence and potential adverse effects. Thereafter, the pharmacist interviews the patient monthly by telephone until therapy is complete. HCWs are allowed only a one-month supply of medication with each refill. From June 1993 through June 1997, of 131 employees who started treatment, 122 (93%) completed it. From June 1997 through December 2001, annual therapy completion rates ranged from 90% to 100%. No cases of active tuberculosis have been reported in treated patients, and only nine adverse drug reactions were reported for the 1997-2001 period. A pharmacist-managed clinic substantially improved rates of treatment completion among HCWs with LTBI.

Valerian is an ancient tranquillizing drug obtained from the underground organs of several Valeriana species. Its active principles were assumed to be terpenoids in the form of valepotriates and/or as components of the essential oil. However, unknown active compounds were not discarded and synergic effects were suspected. We have recently isolated 6-methylapigenin (MA) from Valeriana wallichii and proved that it is a benzodiazepine binding site (BDZ-bs) ligand [Planta Med. 68 (2002) 934]. The present paper is the first report of the presence of 2S(-)-hesperidin in valeriana and describes that it has sedative and sleep-enhancing properties. MA, in turn, was found to have anxiolytic properties and was able to potentiate the sleep-enhancing properties of hesperidin (HN).MA and HN are new members of the growing family of natural flavonoids with activity on the CNS, and their properties suggest that they are promising drug leads in the field.

Using the guidance by a competitive assay for the benzodiazepine binding site in the GABA(A) receptor, active compounds were isolated from the rhizomes and roots of Valeriana wallichii DC. The UV, NMR and mass spectral data permitted the identification of 6-methylapigenin. This flavonoid has a Ki = 495 nM for the BDZ-bs and a GABA ratio of 1.6-2.0, which suggests possible agonistic properties. The calculated percentage of 6-methylapigenin in the crude drug is in the range: 0.013% to 0.0013%.