While posttraumatic stress disorder (PTSD) is relatively common among psychiatric conditions, it only emerges in a subpopulation of persons exposed to environmental trauma . Brain nUsing a cohort of predominantly male military veterans and another with predominantly female civilians, Liberzon et al. found that individuals with a beta2-adrenergic receptor (ADRB2) gene variant and childhood trauma are at heightened risk for adult PTSD . InteresNoradrenergic system dysfunction has also been implicated in increased susceptibility to stress-induced opioid use and craving in individuals with opioid use disorders (OUD) . FurtherOne limitation of this study is insufficient information to determine whether it accounts for epigenetic and associated brain vulnerability factors from such childhood trauma that may also predispose to developing PTSD. It is difficult to determine whether this study controlled for pertinent non-specific neurobiological factors indirectly impacting response to trauma in a particular person, independent of this gene by environmental-adversity interaction. It is also unclear whether altered expression of this gene is tangential/incidental to co-occurring critical brain development changes altering PTSD vulnerability.To substantively advance knowledge, longitudinal research with sufficient statistical power is needed using advances in neuropsychiatric imaging and bioinformatics to identify brain genetic variants, imaging, pharmacogenetic and other molecular biomarkers, and neuropsychiatric vulnerability factors interacting with traumatic environmental events in childhood to predict long-term departures from normal brain and cognitive development, associated with PTSD and/or chronic pain. Such research should also identify critical brain and cognitive development windows in prodromal stages where primary prevention approaches might particularly be targeted to prevent the onset of PTSD and co-occurring OUD. For such a study to be impactful, it should include a large heterogeneous sample representative of the population of interest, control for relevant developmental, socio-demographic, psychiatric, environmental, and family-history cofactors, and include validated patient-reported and clinician-measured outcomes of PTSD and OUD. Sampling designs using probability-based sampling of subgroups at greater risk for PTSD and OUD  may be needed to achieve the above objectives. A reasonable research approach may also incorporate genetically informative designs  or subjects .To address these considerations, suitable research settings could be large healthcare system-based research networks containing existing electronic health record systems (EHRs) and virtual data warehouses, which routinely and in a consistent manner capture comprehensive longitudinal health data on a relevant population base of patients, which can be combined with genetic and environmental data from longitudinal-cohort research studies. Leveraging interoperable EHRs in such practice-based research networks can enhance recruitment time as well as allowing efficient and cost-efficient targeting of research subjects with relevant characteristics . FurtherTo conclude, to fill important precision-medicine research gaps described above, large practice-based research networks with standardized EHRs data collection, querying, extraction, and storage approaches are needed to collect and analyze relevant longitudinal data on large numbers of individuals with important phenotypic data to merge with genetic information. This could be efficiently accomplished by such systems having the facilities for, and expertise with, harmonized data collection and analysis of neuropsychiatric imaging data and genetic specimens for research, with accompanying patient-reported data routinely collected in clinical practice and EHRs/encounter data. For example, several practice-based research networks in the United States , 10 may The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Once acquired, a fearful memory can persist for a lifetime. Although learned fear can be extinguished, extinction memories are fragile. The resilience of fear memories to extinction may contribute to the maintenance of disorders of fear and anxiety, including post-traumatic stress disorder (PTSD). As such, considerable effort has been placed on understanding the neural circuitry underlying the acquisition, expression, and extinction of emotional memories in rodent models as well as in humans. A triad of brain regions, including the prefrontal cortex, hippocampus, and amygdala, form an essential brain circuit involved in fear conditioning and extinction. Within this circuit, the prefrontal cortex is thought to exert top-down control over subcortical structures to regulate appropriate behavioral responses. Importantly, a division of labor has been proposed in which the prelimbic (PL) and infralimbic (IL) subdivisions of the medial prefrontal cortex (mPFC) regulate the expression and suppression of fear in rodents, respectively. Here, we critically review the anatomical and physiological evidence that has led to this proposed dichotomy of function within mPFC. We propose that under some conditions, the PL and IL act in concert, exhibiting similar patterns of neural activity in response to aversive conditioned stimuli and during the expression or inhibition of conditioned fear. This may stem from common synaptic inputs, parallel downstream outputs, or cortico-cortical interactions. Despite this functional covariation, these mPFC subdivisions may still be coding for largely opposing behavioral outcomes, with PL biased towards fear expression and IL towards suppression. Pavlovian fear conditioning is a form of learning that serves as a robust model to explore the neurobiological underpinnings of disorders of fear and anxiety, including post-traumatic stress disorder (PTSD). In a typical rodent experiment, an innocuous conditioned stimulus  is paired with an aversive unconditioned stimulus . After one or more conditioning trials, presentation of the CS alone comes to elicit a conditioned fear response (CR) that includes freezing behavior , changes in heart rate and respiration, and potentiated acoustic startle Davis, . ImportaWhile learned fear serves an adaptive purpose aiding survival, pathological fear states are thought to underlie various stress and trauma-related disorders such as PTSD, which has a lifetime prevalence of nearly 8% in the general population  is thought to regulate fear expression, whereas the infralimbic cortex (IL) mediates fear suppression , the PL and the IL. Here, we will primarily focus on PL and IL. The rodent PFC exhibits laminar organization with deep and superficial layers  and interneurons  but to distinct populations of cells that ultimately dictate CRs. To this end, PL and IL both strongly innervate the BLA and these glutamatergic projections originate from layers II, V and VI  such as c-fos, Arc and Zif268 are activated in response to cellular stimulation, providing an indirect measure of neural activation and have been implicated in learning and memory , spontaneous firing rates are suppressed in both IL and PL, although IL suppression was more robust  PVINs causally initiated freezing behavior in unconditioned animals and also modulated fear expression in previously conditioned animals , Yau and McNally  have recThe neural circuits underlying fear conditioning and extinction in rats have also been identified in humans. For example, the dorsal anterior cingulate cortex (dACC) and the ventromedial prefrontal cortex (vmPFC) have been proposed to regulate the expression and suppression of fear in humans, respectively. While the temporal and spatial resolution of neuroimaging techniques cannot provide fine anatomical details for cross species comparison, they have provided a broad look at the human fear circuit and insight into PTSD. Using functional imaging with a standard fear conditioning paradigm, Phelps et al.  reportedAnother possibility is that those who suffer from PTSD fail to use contextual cues to appropriately guide behavioral responding, resulting in a greater degree of generalized fear , but this shifts to outcome-independent  performance after extended training. Interestingly, rats with PL lesions exhibit habitual responding that is insensitive to outcome value both early and late in training, whereas rats with IL lesions exhibit goal-directed responding even after extended training  whereas the expression of drug seeking behavior corresponds to a rapid activation of movement. However, the associative structure and psychological processes underlying these behaviors may be similar. It has been shown that “sign-trackers” (rats who approach a food predictive cue) also show increased auditory fear (compared to context fear), suggesting that these animals are “cue-directed”  of mPFC regulate fear expression and ventral regions (IL/vmPFC) fear suppression. However, findings from recent studies challenge the underlying assumptions of this model. For example, a number of recent anatomical and electrophysiological studies have shown that PL and IL project similarly to the amygdala  and a McKnight Memory and Cognitive Disorders Award to SM.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.


Research in mucosal vaccination in finfish has gained prominence in the last decade in pursuit of mucosal vaccines that would lengthen the duration of protective immunity in vaccinated fish. However, injectable vaccines have continued to dominate in the vaccination of finfish because they are perceived to be more protective than mucosal vaccines. Therefore, it has become important to identify the factors that limit developing protective mucosal vaccines in finfish as an overture to identifying key areas that require optimization in mucosal vaccine design. Some of the factors that limit the success for designing protective mucosal vaccines for finfish identified in this review include the lack optimized protective antigen doses for mucosal vaccines, absence of immunostimulants able to enhance the performance of non-replicative mucosal vaccines, reduction of systemic antibodies due to prolonged exposure to oral vaccination and the lack of predefined correlates of protective immunity for use in the optimization of newly developed mucosal vaccines. This review also points out the need to develop prime-boost vaccination regimes able to induce long-term protective immunity in vaccinated fish. By overcoming some of the obstacles identified herein, it is anticipated that future mucosal vaccines shall be designed to induce long-term protective immunity in finfish. Cyprinus carpio) in Israel  vaccine administered by immersion in carp  has been licensed in Canada  antigen delivery vehicles and (ii) immunostimulants , 20. As Immunostimulants explored for mucosal vaccine delivery in finfish include poly(lactic-co-glycolic acid) (PLGA) nanoparticles , 8, micrAs pointed out by Neutra and Kozlowski , vaccinein vivo targeting of APCs to enhance antigen uptake  prevented the establishment of pathology in target organs in Atlantic salmon vaccinated against IPN. In another study (490 (1:50 dilution) prevented against IPNV-induced pathology indicating that a threshold antibody titer can be established for vaccines against IPNV infection to serve as a cutoff limit at which protection can be defined. It is likely that similar approaches can be used to develop correlates of protective immunity for mucosal vaccines in finfish.This component of protective immunity is centered on preventing disease establishment by blocking pathogen dissemination to internal organs that include target organs that are prone to pathogen-induced pathology , 53. Alt490 (1:50 dilution) corresponded with post-challenge survival proportions (PCSP) >92.0% in Atlantic salmon vaccinated against IPNV using injectable vaccines (490 (1:50 dilution) corresponded with PCSP >94.0%, again indicating that antibody titer that can serve as a correlate of protection for IPN vaccines in Atlantic salmon. Efforts should be made to determine similar correlates of protection for mucosal vaccines.Although it has been shown that mucosal vaccines have the capacity to reduce post-challenge mortality in vaccinated fish , 56–60, vaccines . In anotvaccines , antibodSalmo salar L.) (Oncorhynchus mykiss) (C. carpio) (Carassius auratus)  , 65, rai mykiss) , common  carpio) , 68, andauratus) . In carpauratus) , 68. Mauauratus)  exposed In mammals, factors linked to immunotolerance include direct inactivation of antigen-sensitized lymphocytes via depletion or anergy, regulatory T-cells (T-regs), hepatic processing of antigens, and several other factors that include anti-idiotypes , 70, 71.Fish can be clustered into different groups based on their physiological and immunological differences. They can also be classified into omnivores, carnivores, or herbivorous based on their feeding habits and anatomical layout of the digestive system. Rombout et al.  pointed Oreochromis niloticus), reared at 28°C have a shorter production period lasting about 6 months . It is likely that cold-water species could require multiple boost vaccinations to maintain high levels of protective antibodies during the long production periods, which might not be needed for fish species having short production periods. The difference between migratory and non-migratory species is another important factor that has a significant influence on the design of protective vaccination strategies. For example, Atlantic salmon undergo smoltification and changes in water salinity after transfer from freshwater to seawater making them vulnerable to stress-related disease outbreaks, such as IPNV outbreak  lack genes that code for the MHC-II loci found in other fish species  that bind to pathogen-associated molecular patterns (PAMPs) on pathogens, surface markers of activated APCs and different regulatory cytokines and chemokines , 56, 57.Given the lack of knock-out (KO) models and limited number of surface marker monoclonal antibodies (mAbs) characterized in finfish, a large proportion of immune response studies are dependent on gene expression. To a large extent, the interpretation of gene expression data in finfish is by inference to mammalian studies , 78, 79,Prime-boost vaccination strategies can be classified into the homologous and heterologous vaccination strategies as shown below.Paralichthys olivaceus) vaccinated against VHSV using a homologous prime-boost vaccination strategy. Put together, these factors show that the efficacy of homologous prime-boost vaccination strategies is dependent on the route of vaccine delivery, type of antigen (live or inactivated), and antigen dose. In order to design protective homologous prime-boost vaccination protocols, it is important to find the most protective antigen delivery system, whether live or inactivated, antigen dose, and route of vaccination into fish for each disease and fish species.In situations where a single dose is insufficient to induce protective immunity, the same antigen can be administered repeatedly to increase and prolong the duration of protective immunity. This vaccination strategy has been widely used against different diseases in mammals , where t+ and CD8+ levels that significantly reduced the HIV viral loads in macaque monkeys. Jia et al.  optimization of antigen doses in order to pave way for identifying the correlates of protective immunity on mucosal surfaces and protection against systemic pathogen dissemination; (ii) choice of antigen delivery systems  in which mucosal vaccines can be delivered as replicative or non-replicative antigens; (iii) protecting oral vaccines against degradation in the acidic environment of the gut; (iv) identifying the most potent adjuvants having highly effective immunostimulants able to enhance antigen uptake, processing, and presentation to cells of the adaptive immune system; (v) identifying the most effective vaccine delivery route able to produce highly protective mucosal and systemic antibody responses in vaccinated fish; and (vi) designing highly protective prime-boost vaccination strategies able to produce long-term protective immunity in vaccinated fish.Host immune response factors that pose a challenge in designing protective mucosal vaccines for finfish include (i) oral tolerance – although the underlying mechanisms causing this condition in finfish have not been studied in detail, based on the limited studies carried out this far indications are that prolonged exposure to oral vaccination reduces the induction of systemic antibody responses in vaccinated fish; (ii) species variations need tailor-made vaccines to cope with individual species requirements; (iii) lack of established correlates of protective immunity required to determine the antigen dose able to prevent pathogen entry on mucosal surfaces and to prevent systemic pathogen dissemination for mucosal vaccines; and (iv) a complex host genomic response to mucosal vaccination whose network pathways can be difficult in identifying the most relevant genes required for optimization of vaccine performance in finfish.Although there might be several other factors that limit the design of protective mucosal vaccines for finfish, this review highlights some of the most significant challenges currently hindering our ability to develop highly efficacious mucosal vaccines for finfish. Therefore, future studies should seek to overcome some of the challenges put forth in this review in order to pave way for the design of protective mucosal vaccines for finfish.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.


Fshear) and compression (Fcompression) forces between lumbar vertebrae, most often between the last lumbar and the sacrum [Fshear and Fcompression can lead to painful muscle strain, joint capsule pain, disc herniation, inflammation (spondylitis), bone degeneration (spondylolysis) and vertebral displacement (spondylolisthesis) [Lower back pain (LBP) is one of the most common and costly medical problems today , 2. Paine sacrum , 5. In asthesis) .Fshear due to lordosis were offset by the benefits of positioning the upper body’s center of mass over the hips, stabilizing the trunk and decreasing the costs of upright posture. The only two known complete lumbar spines from early hominins show the same sexually dimorphic pattern present in modern humans, with males having fewer wedge-shaped, lordotic vertebrae than females [Fshear in pregnant hominin females who exaggerate lordosis to cope with increased fetal mass. Another hypothesis is that some cases of LBP are the result of a recent mismatch, in which the modern human spine is poorly adapted to recent environmental conditions. Since hominins prior to the post-industrial era were very active [Mechanically induced LBP is often thought be a consequence of trade-offs in the spine due to selection for bipedalism from a quadrupedal ancestor. According to this hypothesis, the costs of increased  females . More foy active , low levy active . In addiy active , 8. Alsoy active .If some cases of LBP are the result of a recent mismatch, LBP rates may be on the rise as sedentary behaviors increase. This may also suggest that trunk strengthening and endurance exercises can help treat and prevent some cases of LBP. However, the mismatch hypothesis has been poorly tested. More biomechanical research is needed to explore the relationship between novel types of spinal loading  and LBP. Detailed comparative studies of LBP rates around the world are necessary to test the hypothesis, comparing age-matched groups with different activity levels and subsistence patterns, such as hunter–gatherers and non-industrial populations.


Many neurodegenerative disorders are linked to irreversible protein aggregation, a process that usually comes along with toxicity and serious cellular damage. However, it is emerging that protein aggregation can also serve for physiological purposes, as impressively shown for prions. While the aggregation of this protein family was initially considered exclusively toxic in mammalians organisms, it is now almost clear that many other proteins adopt prion-like attributes to rationally polymerize into higher order complexes with organized physiologic roles. This implies that cells can tolerate at least in some measure the accumulation of inherently dangerous protein aggregates for functional profit. This review summarizes currently known strategies that living organisms adopt to preserve beneficial aggregation, and to prevent the catastrophic accumulation of toxic aggregates that frequently accompany neurodegeneration. Low structural complexity is at the basis of highly diversified molecular recognition, whereby one flexible protein region can bind to various heterogeneous ligands by conformational adaptation. Proteins can functionally benefit from binding promiscuity for key regulatory processes such as signal transduction, transcription, RNA processing and translation. Proteins situated on intersecting hubs of different pathways can undergo multifunctional interactions, functioning as molecular switches by means of conformational variability. However, the benefits of conformational freedom come along with the menace of protein misfolding and multifunctional failure. Although each living organism invests conspicuous amounts of energy for the rescue or elimination of misfolded polypeptides, a possible inability of the cell to cope with misfolded polypeptides inevitably leads to a massive functional destabilization, whereby proteins can either lose their original function (loss-of-function), or they acquire an improper, and therefore mostly lethal function (gain-of-function), eventually aggregating after the complete collapse of folding and clearance pathways.The central nervous system (CNS) is particularly susceptible to protein misfolding, but the reasons of such selective neuronal vulnerability are still elusive  a fibrous, non-branched morphology, (B) the ability to alter the spectral properties of the dyes Congo red and thioflavin T, and (C) X-ray diffraction patterns typical of cross-beta structure. The term was coined by German pathologist Rudolf Virchow during the characterization of masses in human brains described as “corpora amylacea”. Virchow perceived the relationship between amyloids and disease, when he addressed the problem of “amyloid degeneration” Virchow, . Today, Amyloid aggregation is stereotypically linked to conformational flexibility, which allows for structurally diverse polypeptides to fold from a native into an alternative structure of the same chemical composition, but with high beta-sheet content, a remarkable resistance against proteolytic and denaturing agents, and the ability to self-associate into fibers of typical cross-beta structure  as further key players of intercellular transmission .bona fide prion properties exist more frequently than it might sound reasonable for a purely toxic agent. Algorithms trained to identify possible prion signatures could pinpoint one prevailing structural pattern to amino acid stretches of low structural complexity abounding in glutamine and/or asparagine residues. The validity of these predictions could be experimentally verified case-by-case for proteins from yeast as well as from higher eukaryotes  and frontotemporal dementia  they undergo demixing phase separations typical of membrane-free organelles, (C) they are able to heterotypically incorporate or exchange additional RNA granule-associated protein components, (D) they tend to associate preferentially with extended 3’-UTR mRNAs sequences, and (E) they respond to post-translatory modifications in a way that reflects physiologic regulation of RNA granules. These findings highlight the potential of PLR-containing proteins to reproduce major hallmarks of RNA granule biogenesis and function by accessing a polymer state which is morphologically close to amyloid, but completely reversible and therefore devoid of any stable higher order aggregates characteristic of pathologic prions. As these polymers unify basic physical properties of stable amyloids  with reversibility and a minimal toxic hazard, they are potentially appealing for physiological use. First, the spontaneous polymerization of prion-like components can be initiated only upon an incisive conformational change, e.g., by molecules that act as nucleators. Second, polymerization comes along with a sharp phase transition from liquid to gel-like, which closely resembles physical processes of granule condensation in vivo  dynamics provides particular support to this assumption. SG are one type of membraneless organelles that originate transiently in the cytosol upon different forms of cellular challenge, contributing to the arrest of mRNA translation in response to generic insults such as intoxication, UV-irradiation, oxidative stress and heat shock , which represents an integrated response to different stress stimuli and is the best investigated trigger of SG assembly, abolishes translation initiation, causing polysome disruption and the passing over of untranslated mRNA to SG under persisting stress , this enzyme nucleates SG in response to a signal-induced concentration increase in vitro  key pathogen htt displays a highly variable polyglutamine (polyQ) sequence that develops amyloidogenic and pathogenic properties above a threshold length of around 40 glutamines. In a mouse model for HD, the intracellular aggregation of a 42Q htt variant caused a massive co-aggregation and functional inactivation of TIA-1  are a class of representative and ubi-quitously expressed, structurally unrelated molecular chaperones which bind to unfolded or partially folded polypeptides preventing them from aggregation. Hsps act in frequent combination with each other, thereby providing a powerful relay team, which is equipped with numerous accessory proteins accounting for fine-tuning and coordination.Two potent modifiers of amyloid fiber assembly are hsp90 and hsp70. While hsp70 family chaperones assist generic protein folding processes , hsp90 acts especially to stabilize proteins in a near-native, yet unstable conformation until their full structural maturation. The rationale behind the stabilization of partially rather than entirely unfolded polypeptides is to locally protect unstable or destabilized regions, e.g., during rearrangements that functionally require the transient exposure of hydrophobic moieties . Hsp90 thus prevents the collapse or the aggregation of metastable regions by keeping them in a stalled position (Eckl and Richter, As hsp70 and hsp90 influence amyloid fiber assembly at substoichiometric amounts, it has been proposed that a transient interaction occurs already with low-abundance amyloid precursors in order to repartition them from toxic into non-toxic early during amyloidogenesis (Wacker et al., in vitro, it is only by consumption of ATP that these chaperones actively redirect oligomeric intermediates “on- pathway” for amyloid assembly (Falsone et al., Both chaperones require ATP for functioning, and although the sole presence of nucleotide-free hsp70 and hsp90 is sufficient to suppress amyloid fiber growth Several studies highlight the relationships between hsps and amyloid diseases. The deletion of hsp70 genes exacerbates pathogenesis of HD (Wacker et al., The expression of hsp70 and hsp90 is frequently perturbed in neurodegenerative disorders, and both proteins can be recovered from different cytoplasmic inclusions along with other types of chaperones (Hauser et al., In this context, it was found that hsp70 is more inclined to precipitate depending on the state of bound nucleotide (Roodveldt et al., In close relation, pathologically stabilized amyloid species appear to compete with physiological substrates for binding to hsps. As shown for aSyn, stable oligomers can inhibit hsp70, which is therefore no longer capable of functionally folding other substrates (Hinault et al., Hsp70 and hsp90 are closely interlaced with protein degradation, and both chaperones possess the ability to redirect toxic aggregates to degradation (Hipp et al., UPS degradation of soluble monomers seems to prevail under homeostatic conditions, and consistently, amyloidogenic proteins are substrates of various E3 ubiquitin ligases (Kumar et al., While ubiquitylation usually occurs on internal lysine residues, some disease-associated fragments from tau, aSyn, TDP-43 and Abeta can also undergo an unusual N-terminal ubiquitylation (Brower et al., These findings highlight the predominant role of ubiquitin-mediated degradation in neuroprotection. Proteasome dysfunction seems central to neurodegeneration, and rather generic impairments of the UPS, whether pharmacologic, genetic, or physiologic, are sufficient for the unbiased appearance of diverse neurodegenerative phenotypes. The chemical UPS inhibition of dopaminergic cells favors the accumulation of Lewy-body like inclusions (Rideout et al., The integrity of the proteasome can be further challenged by an uncontrolled overload of amyloid aggregates. For Abeta, it was shown that aggregates can overturn the function of the UPS by physically interacting with proteasome subunits. Zhao and Yang suggest that a decreased proteasome activity is not due to inhibition, but rather to the competition of natural proteasome substrates with increasing concentrations of amyloid aggregates, as shown for Abeta peptide (Zhao and Yang,  C. elegans, aggregation and toxicity of polyQ expansions change in relation to the ageing-dependent expression levels of the orthologue MOAG-4 (van Ham et al., While the folding/degradation machineries presented above represent major aggregate modifying pathways, the recent discovery of the protein class MOAG-4/SERF suggests also the existence of more selective cellular mechanisms. MOAG-4/SERF possesses the unique ability to distinguish between amyloid and non-amyloid/amorphous aggregation, acting as a potent amyloid promoting factor (Falsone et al., At this time, a more defined functional classification of MOAG/SERF is precluded by to the lack of detailed functional data. Of note, human homologues SERF1 and SERF2 share low-level homology with the RNA-binding domain of Matrin3 (Scharf et al., The purely pathological significance of amyloid protein aggregation has been questioned by the discovery of physiological processes that exploit some aspects of amyloid polymerization apparently for functional purposes. The structural and mechanistic similarity between pathologic and functional particles, however, anticipates that benefits of amyloid aggregation go hand in hand with toxicity. Despite these imminent hazards, the cell seems to tolerate aggregation as long as strictly confined within spatially and temporally delimited boundaries, as defined by inherently low transcription and translation rates, localized changes in protein levels, and a high protein quality control and turnover. By these means, aggregation-prone polypeptides are subjected to an exceptional physical and biological supervision at almost every level of cellular life. Yet, surveillance systems can fail upon chronic exposure to abnormally resistant aggregates, and in spite of some considerable functional backup, multiple simultaneous dysfunctions can tilt the equilibrium from functional into toxic. It is therefore conceivable, that amyloid polymerization is physiological, as long as the cell is capable of buffering any inherent toxicity. The occurrence of multiple system failures, as for example during cell senescence or upon extraordinary cellular challenges, might lead to the prevalence of toxic phenotypes, reflecting the multifactorial nature of the associated clinical conditions.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.


In an era of multi-centre genomic dataset generation, CODEX provides a single database where these samples are collected, uniformly processed and vetted. The main drive of CODEX is to provide the wider scientific community with instant access to high-quality NGS data, which, irrespective of the publishing laboratory, is directly comparable. CODEX allows users to immediately visualize or download processed datasets, or compare user-generated data against the database's cumulative knowledge-base. CODEX contains four types of NGS experiments: transcription factor chromatin immunoprecipitation coupled to high-throughput sequencing (ChIP-Seq), histone modification ChIP-Seq, DNase-Seq and RNA-Seq. These are largely encompassed within two specialized repositories, HAEMCODE and ESCODE, which are focused on haematopoiesis and embryonic stem cell samples, respectively. To date, CODEX contains over 1000 samples, including 221 unique TFs and 93 unique cell types. CODEX therefore provides one of the most complete resources of publicly available NGS data for the direct interrogation of transcriptional programmes that regulate cellular identity and fate in the context of mammalian development, homeostasis and disease.CODEX ( One of the fundamental questions in biology is how a single fertilized egg cell faithfully develops into a multicellular organism containing specialized organs capable of homeostasis and regeneration, while the genomic content within each cell remains essentially unchanged. Cell-type specific transcriptional and chromatin landscapes are critical determinants of the global gene expression patterns that define cell identities and fate choices . As key Recent advances in next-generation sequencing (NGS) have allowed genome-wide analysis of TF binding and histone modifications , identification of open regions of chromatin (by DNase-Seq) and transcriptomic analysis (by RNA-Seq) . Such teHowever, the multi-centre nature of this huge data generation effort has had several unintended side effects: (i) the bioinformatic processing and analysis necessary to provide informative and biologically relevant insights from such experiments are not uniformly standardized or integrated, (ii) no public repository provides instant NGS data visualization, (iii) the large size of such NGS datasets (raw RNA-Seq datasets can be 100 GB) is prohibitive for the in-house processing necessary for visualization and/or further analysis without dedicated computer hardware or bioinformatics expertise and finally (iv) annotation of publicly available NGS data is often incomplete or non-intuitive, limiting simple data interpretation. These current failures significantly reduce the utility of such data to the wider research community.http://codex.stemcells.cam.ac.uk/), a database of NGS experiments including ChIP-Seq, RNA-Seq and DNase-Seq. CODEX provides uniformly processed data as well as online resources for NGS data visualization and bioinformatics analysis. Most importantly, CODEX uses a standardized bioinformatics-processing pipeline for all NGS datasets, and the details of each sample are manually curated to provide key information. CODEX currently includes over 1000 uniformly processed NGS datasets that can be easily viewed, interrogated and compared by the general scientific community, for both quick and informative comparisons as well as large-scale meta-analyses.In an effort to bridge this gap between the vast amounts of publicly available NGS raw data and end-user friendly information, we have developed CODEX , and a second focused on data from ES cells (ESCODE). In addition, other relevant cell types are also held in CODEX. To date, CODEX comprises over 1000 samples selected for maximal genomic coverage, with 221 unique TFs and 93 unique cell types across both human and mouse samples. CODEX therefore currently provides one of the most complete resources to directly interrogate transcriptional regulation of mammalian development, homeostasis and disease.CODEX is built on bioinformatics pipelines that uniformly process all relevant publicly available NGS experiments Figure , allowinRaw NGS data are stored on and can be downloaded from GEO and ArrayExpress in a standardized common format as Sequence Read Archive data files (SRA) . AlthougTo ensure CODEX remains up-to-date with newly published datasets we have developed a web crawler that searches the GEO database and reports any new samples of relevance to CODEX. This robot not only retrieves the GEO sample and series accessions, but also scans other databases   to colleIn addition to this semi-supervised method, individuals can also report any experiment or study from GEO and ArrayExpress. All requested analyses are vetted and if suitable for CODEX are added to the processing queue.Although valuable experimental information is provided by users at the time of GEO submission, such details are often poorly curated or incomplete. Such available information is collected by CODEX via its automated text-mining step. This information alongside details from relevant publications is then used to manually curate each sample added to CODEX. This is a time consuming but critical step in the processing pipeline because it provides end users with the information for each experiment, allowing users to distinguish between cell types and culture conditions that can otherwise be difficult to determine.General cell type: broadly defines the sample.Cell subtype: distinguishes cell lines from primary cells and includes the specific cell type name. This field also contains treatment type so users are aware that specific treatments have been utilized.Tissue Ontology classifier from the BRENDA database (BTO); hierarchically organizes cell types and cell lines under a generic category corresponding to the rules and formats of the Gene Ontology Consortium .Additional details: includes (where appropriate) any special culture conditions or drug treatments, as well as human patient cell line chromosomal abnormalities.CODEX also provides the following hierarchically structured information for each sample to allow users to easily search, choose, compare and contrast different samples. Each of these fields is individually manually curated:Central to CODEX is its processing algorithm, which removes one of the main difficulties users may find when comparing publicly available data. Raw NGS data mapped to different genome builds are not directly comparable and require a good knowledge of the plethora of bioinformatics tools to perform this task. Even when mapped to the same genome, the software used or processing details may differ, which prevents direct comparison.To facilitate the ease of use of the processed data within CODEX, we have developed our own NGS pipeline based on open-source and widely used bioinformatics tools. Although processing differs for each experiment type, the whole process shares some common steps, summarized in Figure http://www.bioinformatics.babraham.ac.uk/projects/fastqc/) whilst adapters and any overrepresented sequences are trimmed by using trimgalore . All initial steps of the pipeline are automated, but the resulting reports are manually inspected and any samples failing at any of these steps may be discarded.Public data is downloaded in SRA format and converted to fastq format using the SRA toolkit . Subsequet al.  are always aligned to the same reference genome: GRCm38 (mm10) for mouse and GRCh37 hg19) for human. Read alignment for ChIP-Seq and DNase-Seq is performed using bowtie2 , the reaet al. , whilst  for humaP-value stringencies (from 10−2 to 10−15) by comparing the experimental sample with the appropriate control (where available). At this stage, every stringency is manually inspected and an optimum P-value established. The resulting peak profiles are then transformed into a BED format with a uniform length of 400 bp (with the summit of TF binding in the centre ±200 bp).Next, using SAMtools  the SAM http://www-huber.embl.de/users/anders/HTSeq/doc/overview.html) is used, considering only exome-aligned reads using the UCSC transcriptome model, which for a given set of replicates, computes a single read count profile. This generates a text file with genes as rows and sample(s) as columns, with each element being the number of reads mapped to each gene .For RNA-Seq experiments, HTSeq  .The CODEX Correlation Analysis tool allows users to compare the peak profiles between selected ChIP-Seq experiments. This analysis converts peak profiles into binary vectors within a matrix, with columns as experiments and rows as genomic elements. Where a peak is called at a genomic region, the event is given a value of one, while no peak is annotated as zero. As peak profile matrices are represented by a large number of zero elements, the Pearson's correlation tends to provide a negative close-to-zero coefficient and fails to identify binding profile correlations, even if experiments share a significant number of peak regions. We have therefore opted to use the Dice coefficient to compute similarity, which only takes into account for the calculus the bound regions. The results from this analysis can either be displayed as heat map showing coefficient of agreement or percentage overlap.GSCA provides a complementary computational approach to the routine Gene Ontology analysis . This wede novo motif discovery analysis using Homer  are converted to Multiple Em for Motif Elicitation (MEME) format and used as input to TOMTOM. Significant similarities of enriched motifs to known motifs are reported in HTML output within CODEX.CODEX provides pre-computed ng Homer  and TOMTng Homer . For eac et al. –28). PWM. PWMde nAs experiments from different organisms are not directly comparable within a UCSC Genome Browser session, we have developed a web-tool that identifies common TF target genes for experiments from different organisms. The web-tool computes the intersection of the two TF target gene lists using the NCBI homologene database , and proCODEX provides free access to all processed and pre-computed analysis from the built-in web-tools. All intermediate and final result files are freely available from CODEX, so users can download them and perform any further custom analysis. For all NGS experiments in CODEX, we provide the alignment quality reports (FastQC) and the aligned-reads density profiles (bigWig). For ChIP-Seq experiments, CODEX additionally provides the peak files (BED) and lists of TF target gene (genes). For RNA-Seq, CODEX also supplies the binary mapped-reads files (BAM) and the lists of the number of counts per gene (counts).CODEX also provides a framework to store private data The treatment of unpublished data must follow the same guidelines as the public data using the same pipeline, because once published, experiments will become publicly available in CODEX. Users must therefore register to become a CODEX-consortium member and agree to the standardized pipeline described above, before being provided access. When registered users are logged in, their private samples are displayed along with the rest of the public experiments, and may use all the features, analyses and web-tools described above.Comparison of a users ChIP-Seq experiment for a factor which exists within CODEX to all similar experiments within the database and ask how the binding of this TF compares across cell types, treatments or disease states.To determine the similarity between an experiment for a factor not contained within CODEX, peak files from experiments of interest can be downloaded and easily compared to the peak binding files of the individuals experiment and the overlap of the experiments calculated.The standardized bioinformatic data processing, expert manual curation, user-friendly web-interface and built-in web-tools make CODEX a highly relevant resource for both experimental and computational biologists alike. The ability to instantly access the transcriptomes and epigenomes of numerous different cell types and disease states affords a unique opportunity to drive novel biological discoveries and clinical insights. We have endeavoured to produce an interface that is user-friendly and a database that is valuable to the general scientific community. By manually curating the details for every experiment so as to contain only clear and concise information, the cell-type, derivation and treatment of each sample can be quickly understood. With several simple selections, users can immediately visualize multiple experiments in the UCSC Genome Browser or take advantage of the bioinformatic web-tools available. In particular, we envision CODEX helping experimental biologists with otherwise difficult or time-consuming comparisons, for example:CODEX, which currently holds over 1000 ChIP-Seq, DNase-Seq and RNA-Seq samples, evolved from HAEMCODE , a mouseDevelopment of new web-tools, such as those to further compare DNA motif enrichment and TF binding sites, or interrogate samples from different organisms, will help further our understanding of the conservation of transcriptional regulation at the organ, organism and evolutionary level. Additionally, as more RNA-Seq datasets are published, we aim to further integrate TF ChIP-Seq and RNA-Seq datasets to help uncover combinatorial and cell-type specific TF activities and characterize core transcriptional programmes that define cellular identity. Such datasets and web-tools will make microarray databases, such as BloodExpress , all butCODEX provides ‘small science’ with the opportunity to mine the wealth of data generated by the wider scientific community and large-scale NGS projects. Interrogation of such large datasets will help drive our understanding of mammalian development, homeostasis and disease, and build towards a unified understanding of these processes.


Epithelial–mesenchymal transition (EMT) is a key process associated with the progression of epithelial cancers to metastatic disease. In melanoma, a similar process of phenotype switching has been reported and EMT-related genes have been implicated in promotion to a metastatic state. This review examines recent research on the role of signaling pathways and transcription factors regulating EMT-like processes in melanoma and their association with response to therapy in patients, especially response to BRAF inhibition, which is initially effective but limited by development of resistance and subsequent progression. We highlight studies implicating specific roles of various receptor tyrosine kinases (RTKs) in advancing melanoma progression by conferring a proliferative advantage and through promoting invasive phenotypes and metastasis. We also review the current knowledge of the mechanisms underlying resistance to BRAF inhibition and the potential role of melanoma phenotype switching in this process. In particular, we discuss how these important new insights may significantly enhance our ability to predict patterns of melanoma progression during treatment, and may facilitate rational development of combination therapies in the future. Malignant melanoma accounts for 75% of deaths from all skin cancers in the U.S . Women hEpithelial–mesenchymal transition has been suggested to play an important role in conferring metastatic properties in many solid tumors by altering the integrity of cell–cell junctions, promoting loss of polarity and epithelial markers, eventually resulting in loss of contact between neighboring cells. Through this process, tumor cells become more mesenchymal-like, exhibiting higher migratory and invasive properties that allow them to interact with the extracellular matrix and invade surrounding tissues . It is gA common signaling mechanism that induces EMT in a range of cancers is activation of the MAPK/ERK pathway, which can activate SNAI1 to repress E-cadherin expression and the epithelial phenotype . In addiin vivo  in melanoma phenotype switching and plasticity has recently been reviewed . InductiPhenotype switching in melanoma can be initiated by mechanisms other than those characterized in EMT. In epithelial cancer cell lines, increased LEF1 transcription activity by stable nuclear beta-catenin expression can induce EMT, which is reversible by removal of LEF1 . In melaV600E mutation , have been demonstrated to have inhibitory effects on migration and invasion in the BRAF-mutant cell line A375, with a reversal of EMT-like phenotypic changes orchestrated by induction of E-cadherin and suppression of N-cadherin . A seconGiven that signaling by various RTKs can mediate phenotype switching and promote migration through mechanisms distinct from those enhancing BRAF/MAPK-dependent proliferation and regulation of EMT-TFs, co-targeting of selected RTK signaling pathways and oncogenic BRAF appears to be a logical combination. For example, EGF signaling confers resistance to BRAF inhibition and induces melanoma invasion through Src pathways. Inhibition of the EGF receptor and Src re-sensitizes treatment-resistant BRAF-mutant melanoma cells to Vemurafenib and blocks their invasiveness . HGF secThe EMT process is crucial for normal development and for initiation of malignant transformation and metastasis in a wide range of epithelial cancers. It involves activation of various signaling pathways, as well as repression of E-cadherin through transcription factors. EMT-like phenotype switching is critical for melanocyte lineage differentiation and initiation of melanoma transformation and metastasis. While common EMT-TFs are implicated, their expression during the switch of melanoma to a mesenchymal-like invasive phenotype can differ from the role in classical EMT. In addition to TGFβ and WNT5A signaling, EGF, FGF, MET, and IGF signaling have established roles in conferring migratory and invasive functions in melanoma Figure . ImportaFL wrote the mini-review. PF, AD, GM, and RA provided intellectual input and contributed to editing the manuscript.GM is on uncompensated advisory boards for GSK, Roche-Genentech, Novartis, BMS, Millenium, Merck and Amgen, is a consultant for Provectus and receives research grant support from Novartis, Pfizer, Ventana, and Celgene. The other authors declare no commercial or financial relationships that could be construed as a potential conflict of interest.


Both parasitic load and resource availability can impact individual fitness, yet little is known about the interplay between these parameters in shaping body condition, a key determinant of fitness in wild mammals inhabiting seasonal environments.Sarcoptes scabiei) severity impacted body condition of 473 Iberian ibexes (Capra pyrenaica) harvested between 1995 and 2008 in the highly seasonal Alpine ecosystem of Sierra Nevada Natural Space (SNNS), southern Spain.Using partial least square regressions (PLSR), we explored how temporal variation in climatic conditions, vegetation dynamics and sarcoptic mange (Bottom-up regulation was found to only occur in healthy ibexes; the condition of infected ibexes was independent of primary productivity and snow cover. No link between ibex abundance and ibex body condition could be established when only considering infected individuals.The pernicious effects of mange on Iberian ibexes overcome the benefits of favorable environmental conditions. Even though the increase in primary production exerts a positive effect on the body condition of healthy ibexes, the scabietic individuals do not derive any advantage from increased resource availability. Further applied research coupled with continuous sanitary surveillance are needed to address remaining knowledge gaps associated with the transmission dynamics and management of sarcoptic mange in free-living populations. Body condition, i.e. energetic state and fat stores of an animal, is a major determinant of individual performance in most vertebrate species , includiSarcoptes scabiei is an excellent study model to evaluate how parasites may interfere with the bottom-up regulation of body condition in wild mammals inhabiting highly seasonal ecosystems. This mite is responsible for severe epizootic disease outbreaks in a broad range of mammals, sometimes causing increases in mortality rates. Infected animals typically suffer from dramatic structural and functional changes in the skin, becoming listless, dehydrated, emaciated and eventually dying from the infestation [Sarcoptic mange caused by the mite estation . Notwithestation  and pathestation  of sarcoCapra pyrenaica Schinz, 1838) is a medium-sized endemic mountain ungulate commonly affected by sarcoptic mange [Sierras de Cazorla, Segura y Las Villas Natural Park, southern Spain, in the late 80s. There, approximately 95 % of ibexes were killed by the parasite [Sierra Nevada Natural Space . One particularity of this extreme Alpine ecosystem is the strong seasonal climatic variation found in the area, with snow cover present for six months of the year. SNNS ibex are adapted to such contrasted conditions and do display an income breeder strategy, i.e. increasing adipose tissues during the summer in anticipation of food deprivation during winter times [Iberian ibex  experienced during infestation.Previous studies have addressed the consequences of mange infestation in SNNS Iberian ibex population: e.g. immune response to first and second exposures , effectsTaking advantage of thirteen years (1995–2008) of data on sarcoptic mange monitoring in the Iberian ibex population of SNNS, we explored whether disease severity interacted with primary productivity, snow cover and ibex abundance in determining body condition. The strength of these interactions was assessed during two periods of contrasted vegetation dynamics and snow cover: green (March – October) and dormant period (November – February). We hypothesized that the deleterious effects of mange on ibex body condition will be compensated by favorable environmental conditions, e.g. high primary productivity, little snow cover and low population abundance. To the best of our knowledge, this study provides the first explicit assessment of how a parasitic disease shapes bottom-up processes in a large mammal.Sierra Nevada National Park approved this study.This study complies with the Spanish and the Andalusian laws regarding bioethics and animal welfare. The 2 and is characterized by a heterogeneous orography, with an altitudinal range between 860 and 3.482 m.a.s.l  at a spatial resolution of 500 m and bi-monthly temporal resolution. Snow cover (percentage of surface covered by snow) was retrieved from the Observatorio Cambio Global - Sierra Nevada website (http://obsnev.es/linaria.html). Only NDVI values associated with shrubs and herbaceous layers were considered, since ibex primarily feed in areas encompassed by these landcover types in SNNS [Two environmental variables were considered while assessing the role of bottom-up processes on the body condition of ibex: the normalized difference vegetation index . The package “plspm” was used to develop and visualize the PLSR outputs [Partial least square regressions (PLSR) were used to assess the influence of disease severity, resource dynamics, intra-specific competition and snow cover on body condition. Carrascal et al.  defined  outputs .Information on the body condition of 473 Iberian ibex was considered in our analyses: 43  of these individuals were classified as healthy, 217  as mildly infested and 213  as severely infested.The monthly variations in the vegetation productivity, snow cover and ibex counts are shown in the Fig. 2 test we identified that bottom-up regulations of ibex condition only occurred in healthy ibexes  an increase in primary productivity clearly triggers an increase in body condition in healthy ibexes, ii) sarcoptic mange can disrupt the link between environmental conditions and body condition, and iii) body condition is independent from ibex abundance parameter in scabietic ibexes.Canis latrans), [Oryctolagus cuniculus), [Vulpes vulpes), [Vombatus ursinus), [The absence of a bottom-up regulation of body condition in scabietic individuals may be related to the physiopathology of this parasitic disease, which is characterized by emaciation, muscle mass losses and anemia , 29. Eveatrans), ; rabbitsiculus), ; red foxvulpes), ; wombatsrsinus), ). For inrsinus),  showed trsinus),  concludersinus), , compromRupicapra pyrenaica parva). It has been suggested that host’s abundance predisposes the population to mange infestation [When overabundance coincides with limited food availability, the ability of the hosts to cope with infestations can be compromised . Here, westation , 37, sinestation , 38. In estation  and hostestation . DespiteCaprinae species [In healthy ibexes, the negative relationship between population abundance and body condition is less pronounced in the green period than in the dormant season. This result could be explained by the habitat use of Iberian ibex. As with other closely related  species , ibexes  species . TherefoHere, we have shown how scabietic ibexes do not take advantage of increases in resource availability, which is a drawback to the implementation of management practices, at least in the short term. A very recent work  underlin


Drug-related behaviors in both humans and rodents are commonly thought to arise from aberrant learning processes. Preclinical studies demonstrate that the acquisition and expression of many drug-dependent behaviors involves the ventral tegmental area (VTA), a midbrain structure comprised of dopamine, GABA, and glutamate neurons. Drug experience alters the excitatory and inhibitory synaptic input onto VTA dopamine neurons, suggesting a critical role for VTA afferents in mediating the effects of drugs. In this review, we present evidence implicating the VTA in drug-related behaviors, highlight the diversity of neuronal populations in the VTA, and discuss the behavioral effects of selectively manipulating VTA afferents. Future experiments are needed to determine which VTA afferents and what neuronal populations in the VTA mediate specific drug-dependent behaviors. Further studies are also necessary for identifying the afferent-specific synaptic alterations onto dopamine and non-dopamine neurons in the VTA following drug administration. The identification of neural circuits and adaptations involved with drug-dependent behaviors can highlight potential neural targets for pharmacological and deep brain stimulation interventions to treat substance abuse disorders. Illicit drug use is a significant global problem, with the United Nations Office on Drugs and Crime estimating that 246 million people worldwide used illicit drugs in 2013. More problematic is the high incidence of substance use disorders (SUDs), which in 2014 was estimated to afflict roughly 21.5 million people in the US, corresponding to ~8% of the population , social impairment , risky behavior , and pharmacological effects . One of the most daunting aspects in treating SUDs is the high incidence of relapse, which occurs in ~40–60% of individuals . In drugRodent model systems are commonly employed to examine the effects of abused drugs on behavior. In this review, we will concentrate on psychostimulants and opiates, as extensive laboratory research has focused on these drug categories. The non-contingent administration of psychostimulants or opiates increases locomotor activity in rodents . RepeateDrug-paired cues exert a powerful influence over behavioral actions in individuals with a SUD . The devBehavioral sensitization and CPP paradigms are relatively easy to implement, but they require experimenter administered drug injections. Rodents can be readily trained to self-administer drugs via an intravenous catheter. A number of drug self-administration assays have been developed to model the behavioral symptoms observed in humans with a SUD. For example, rodents with limited access (1 h) to drugs in daily self-administration sessions maintain stable drug intake. However, rodents with extended access (6 h) to drugs increase their intake over multiple training sessions, similar to the escalated drug consumption that can be observed in individuals diagnosed with a SUD –18. JustJust as with humans with a SUD, drug-dependent behaviors in rodents involve a component of learning, whether it is contextual  or operant (drug self-administration). While numerous brain regions are involved with mediating learning and drug-related behaviors, we will focus on the ventral tegmental area (VTA) in this review. We will also discuss the major inputs to the VTA, how these inputs influence VTA neuron activity, and present recent findings on how these VTA afferents are involved with drug-dependent behaviors.The dopamine neurons arising from the VTA that project to the nucleus accumbens (NAc) are involved with mediating the reinforcing actions of abused substances –26. WhilThe neural circuitry mediating any behavior is complex, though extensive research over the past few decades illustrates that the VTA is critically involved with both rewarding and aversive drug-dependent behaviors. For example, the VTA is required for behavioral sensitization induced by amphetamine or mu-opioid receptor agonists, though evidence for the involvement of the VTA in cocaine behavioral sensitization is mixed . The VTAThe VTA along with the neighboring substantia nigra pars compacta are the primary dopamine producing nuclei in the brain . Early eIh current , the rate-limiting enzyme for dopamine synthesis, is found in ~60% of VTA neurons , 49. VTA current , 53. How current . Additio project –57.The second largest neuronal population in the VTA consists of GABA neurons (~25%) that are commonly identified by the presence of glutamic acid decarboxylase (GAD) , 59. WhiIn addition to dopamine and GABA neurons, a small percentage of VTA neurons contain vesicular glutamate transporter 2 (VGluT2), a marker for glutamate neurons. These neurons predominately reside in the medial aspects of the VTA and project to the ventral striatum, PFC, VP, amygdala, and LHb, as well as synapse onto local dopamine neurons , 68–72. Optogenetic modulation of VTA neurons can elicit either appetitive or aversive behavioral outcomes depending upon the neuronal population that is targeted. Activation of dopamine neurons is acutely reinforcing and sufficient for establishing a CPP, whereas silencing dopamine neurons is aversive and elicits a CPA , 79, 80.The VTA is innervated by a diverse array of inputs, many of which are interconnected. Large afferents to the VTA include the rostromedial tegmental nucleus (RMTg), VP, bed nucleus of the stria terminalis (BNST), LH, pedunculopontine tegmental nucleus (PPT), laterodorsal tegmental nucleus (LDT), dorsal raphe nucleus (DR), NAc, PFC, and amygdala , 85–87. The RMTg  is a nucleus comprised of GABA neurons that function as an inhibitory relay between the LHb and the VTA , 88–92. The RMTg is increasingly recognized as an important nucleus in mediating the effects of abused drugs. The reinforcing effect of opiates was originally thought to arise from activation of mu-opioid receptors on VTA GABA interneurons , though Psychostimulants also influence the activity of RMTg neurons . The nonThe VP is involved in processing rewarding stimuli and motivated behavior . GABA neThe BNST is involved in mediating fear and anxiety –120 and The LH is critical for the expression of motivated behaviors including feeding and drug seeking . The LH Accumulating evidence over the past decade highlights the importance of orexin-containing neurons in feeding, the sleep/wake cycle, and drug-dependent behaviors . Orexin-In vivo electrophysiological experiments illustrate that electrical stimulation of the LDT elicits burst firing in putative VTA dopamine neurons , which commonly has been utilized for the treatment of movement disorders. In preclinical studies, DBS targeting the NAc reduced cocaine behavioral sensitization , morphinThe implementation of optogenetic and chemogenetic approaches in behavioral experiments has validated and identified specific neural circuits that mediate a range of appetitive and aversive behaviors. Many of these studies manipulated brain regions implicated in SUDs , though MW and IO contributed to the writing of this review article.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.


Soda lakes contain high concentrations of sodium carbonates resulting in a stable elevated pH, which provide a unique habitat to a rich diversity of haloalkaliphilic bacteria and archaea. Both cultivation-dependent and -independent methods have aided the identification of key processes and genes in the microbially mediated carbon, nitrogen, and sulfur biogeochemical cycles in soda lakes. In order to survive in this extreme environment, haloalkaliphiles have developed various bioenergetic and structural adaptations to maintain pH homeostasis and intracellular osmotic pressure. The cultivation of a handful of strains has led to the isolation of a number of extremozymes, which allow the cell to perform enzymatic reactions at these extreme conditions. These enzymes potentially contribute to biotechnological applications. In addition, microbial species active in the sulfur cycle can be used for sulfur remediation purposes. Future research should combine both innovative culture methods and state-of-the-art ‘meta-omic’ techniques to gain a comprehensive understanding of the microbes that flourish in these extreme environments and the processes they mediate. Coupling the biogeochemical C, N, and S cycles and identifying where each process takes place on a spatial and temporal scale could unravel the interspecies relationships and thereby reveal more about the ecosystem dynamics of these enigmatic extreme environments. Soda lakes are found worldwide, predominantly in arid and semi-arid environments, such as the Rift Valley in East Africa, the rain-shadowed regions of California and Nevada, and the Kulunda Steppe in South Siberia (Russia) Fig. . Soda laNitriliruptor alkaliphilus  that are active both at a high pH and high salinity. Alkali-stable extracellular proteases, lipases, and cellulases have been used for the production of improved laundry detergents , Anabaenopsis and Cyanospira . At reduced salinity a mass development of heterocystous Anabaenopsis had been observed in East-African soda lakes  Fig. , where tCyanobacteria are traditionally considered as the only diazotrophic component of the oxygenic phototrophic community Fig. b2. HowevChromatiales  at moderate salinity and Ectothiorhodospiracea  at high salinity, also contribute to the primary production in soda lakes  or high salt concentration  , and Natroniella acetogena  represent heterotrophic fermentative haloalkaliphilic acetogens, utilizing amino acids and alcohols as substrates  heterotrophs, i.e., those that utilize monomeric organic compounds such as sugars, amino acids, organic acids, and alcohols are among the best represented groups of haloalkaliphiles isolated so far from soda lakes Fig. a4. AmongMethanolobus taylorii  and Methanosalsum zhilinae , and a highly salt-tolerant lithotroph Methanocalculus natronophilus  and the Alkalilimnicola–Alkalispirillum group of the Gammaproteobacteria   . Nitrogen fixation activity has been documented for aggregates of filamentous non-heterocystous Phormidium-like cyanobacteria and green algae belonging to the Ctenocladus in the oxic littoral zone of Mono Lake  in soda lakes  or butyrate  as e-donor/C-source with sulfate or thiosulfate as e-acceptor and forming acetate as a final product  are important biogeochemical processes within soda lakes  forming abiotically at high pH from sulfur and sulfide. The first, Desulfurispira natronophila, belongs to the phylum Chrysiogenetes  Fig. c2. In soors Fig. b1, such Thioalkalivibrio species such as Tv. thiocyanoxidans, Tv. paradoxus, and Tv. thiocyanodenitrificans, are capable of growth using thiocyanate as the sole energy, sulfur, and nitrogen source   harbor similar microbial communities to hypersaline solar saltern brines of neutral pH. The latter are characterized by a low diversity dominated by a few extremely halophilic archaea, belonging to the class Halobacteria within the phylum Euryarchaeota  and Gammaproteobacteria , Firmicutes , Bacteroidetes , the cyanobacterial genera Arthrospira and Anabaenopsis, and several purple phototrophic bacteria belonging to the families of Ectothiorhodospiraceae, Chromatiaceae and Rhodobacteraceae  harbor more diverse microbial communities than hypersaline environments and the community composition is affected by lake stratification and prevailing oxygen concentrations , Gammaproteobacteria, Bacteroidetes, Betaproteobacteria , Deltaproteobacteria , Actinobacteria , and benthic cyanobacteria , including the chemolithotrophic genus Thioalkalivibrio and the phototrophic genus Halorhodospira  and Wadi Natrun (Egypt) Fig. , KovalevNitrosomonas-like sequences. Additionally, FISH analysis revealed the presence of Crenarchaeota and the correlation of nitrification rates with crenarchaeal numbers. Although no archaeal amoA sequences were detected, it cannot be ruled out that ammonia-oxidizing archaea (AOA) contribute significantly to nitrification in Mono Lake  were most closely related to halo- and/or alkali-tolerant i et al.  challengi et al. .So far, soxB, which encodes an indispensible sulfate thiohydrolase in the Sox pathway proposed for the oxidation of thiosulfate in SOB  and denitrification (nitrite reduction to nitrous oxide) in the water column of two low saline Ethiopian soda lakes was also shown by the successful amplification of mRNA for particulate methane monooxygenase (lake Beseka), ammonia monooxygenase (amoA) and nitrite reductase  is cardiolipin (Banciu et al. Halobacteriaceae and Salinibacter, have membrane surface layers that are strongly enriched in acidic amino acids (Oren Structural adjustments within the cell membrane of haloalkaliphilic prokaryotes include an increased level of the neutral lipid squalene and the polar lipid cardiolipin in the phospholipid bilayer (Angelini et al. ids Oren . The proids Oren . EcophysNitrosopumilus maritimus (Könneke et al. Traditionally, studies on microbial communities were restricted to a few cultured isolates, whilst modern high-throughput techniques now allow the study of microbial community composition as a whole. Although a considerable number of cultured isolates has already been obtained from soda lakes, culture-independent methods have uncovered a much more diverse microbial community. Future studies should attempt to isolate members of the uncultured community (Alain and Querellou Metagenomics can be applied to obtain a high-resolution genetic inventory of the microbial community in soda lakes. Such a genetic inventory can be used to explore the overall metabolic capacity of the prokaryotic soda lake communities. Other high-throughput techniques monitor community-wide levels of gene-expression (meta-transcriptomics; Carvalhais et al. nifH and nifD genes in soda lakes, and distinguishing between the contribution of heterotrophs and primary producing phototrophs, is still lacking. Single cell techniques, like FISH-NanoSIMS (Dekas and Orphan amoA genes and their transcripts.Several metabolic processes might be present in soda lakes but have not yet been detected, such as anaerobic methane oxidation and anaerobic ammonium-oxidation. Furthermore, the importance of anaerobic polymer degradation in sediments is not well understood and the contribution of anoxygenic photosynthesis to primary production might be underestimated. Nitrogen fixation at hypersaline conditions is suspected to be limited to diazotrophic anoxygenic phototrophs and heterotrophs, but a comprehensive study targeting In conclusion, several questions regarding biogeochemical cycles in soda lakes are still open. To obtain a more comprehensive insight into the microbial diversity of soda lakes, its role in biogeochemical cycles and the molecular mechanisms by which the microorganisms adapt to the extreme environmental conditions, we have to study these habitats with a systems biology approach in which we combine novel isolation methods with state-of-the-art meta-omics techniques, and eventually with mathematical modeling to predict the response of cells and communities to environmental stimuli and to infer the interactions of co-existing populations.


Homo sapiens, Mus musculus, Rattus norvegicus, Gallus gallus, Epstein–Barr virus, Canis familiaris and Arabidopsis thaliana) from >300 references in the literature. These regulations can be divided into two parts: miR2Epi (103 entries describing how miRNA regulates epigenetic modification) and Epi2miR (1871 entries describing how epigenetic modification affects miRNA). Each entry of EpimiR not only contains basic descriptions of the validated experiment  but also clearly illuminates the regulatory pathway between epigenetics and miRNA. As a supplement to the curated information, the EpimiR extends to gather predicted epigenetic features  associated with miRNA for users to guide their future biological experiments. Finally, EpimiR offers download and submission pages. Thus, EpimiR provides a fairly comprehensive repository about the mutual regulation between epigenetic modifications and miRNAs, which will promote the research on the regulatory mechanism of epigenetics and miRNA.As two kinds of important gene expression regulators, both epigenetic modification and microRNA (miRNA) can play significant roles in a wide range of human diseases. Recently, many studies have demonstrated that epigenetics and miRNA can affect each other in various ways. In this study, we established the EpimiR database, which collects 1974 regulations between 19 kinds of epigenetic modifications  and 617 miRNAs across seven species (including Database URL:http://bioinfo.hrbmu.edu.cn/EpimiR/. Epigenetics refers to the heritable changes in gene expression without a change in the DNA sequence itself, including DNA methylation and post-translational modifications of chromatin proteins . Under tMicroRNA (miRNA) is a class of small noncoding RNA molecules that usually lead to translational repression and gene silencing through the specifically binding 3′ untranslated region of messenger RNAs based on sequence complementation at the post-transcriptional level . PrimaryRecent evidence suggested that epigenetic control is involved in the regulation of miRNA gene expression in diverse ways. DNA methylation of promoter-associated CpG dinucleotides  usually correlates with reduced transcription levels of corresponding miRNAs and subsequently induces the expression of miRNA target genes . Histoneet al. , which provided a comprehensive repertory of experimentally validated interactions between epigenetic modifications and miRNAs across multiple species and filled the vacancy in this field.Recently, many studies have revealed that epigenetics and miRNA can affect each other in many biological processes. Thus, a complete database that stores the experimentally validated interactions between epigenetic modifications and miRNAs is valuable to understand the molecular mechanisms of diseases and promote the targeted research toward epigenetics- or miRNA-related drugs. However, the relevant information is scattered in the massive body of literature, which is inconvenient for researchers to explore their mutual regulations. Accordingly, we developed a manually curated resource entitled ‘EpimiR’ .Before searching, users should choose to view a high-confidence-level or low-confidence-level record or both. Each curated entry in the EpimiR has been graded, which is more efficient for users to understand and use these records. High-confidence-level records usually provide sufficient evidence to present the regulatory process between miRNA and epigenetics. For low-confidence-level records, most of them merely demonstrate the existence of potential relationship between miRNAs and epigenetic modifications  from 309 papers. These regulations can be compartmentalized into two parts: 103 miR2Epi records and 1871 Epi2miR records. Each entry of Epi2miR contains the detailed information about epigenetics that can regulate the expression of miRNAs, such as epigenetic modifications that impact miRNA expression, regulators that affect epigenetic modification, genes that are proven to be directly targeted by miRNAs, tissues or conditions for detection, experimental method, species and PubMed identifier (ID). Similarly, for the miR2Epi entry, the EpimiR database clearly elucidates the process of miRNA regulating epigenetics through listing the regulators of epigenetics that are directly targeted by miRNAs, tissues or conditions for detection, experimental method, species, PubMed ID, etc. Additionally, we extend to gather predicted miRNA genomic features associated with epigenetic modification. [such as type of miRNA locating area, position of transcription start site , the National Natural Science Foundation of China (30900837) and the Foundation for University Key Teacher of the Education Department of Heilongjiang Province (1252G037). Funding for open access charge: The Foundation for University Key Teacher of the Education Department of Heilongjiang Province (1252G037).Funds for Creative Research Groups of the Conflict of interest. None declared.


Although treeline elevations are limited globally by growing season temperature, at regional scales treelines frequently deviate below their climatic limit. The cause of these deviations relate to a host of climatic, disturbance, and geomorphic factors that operate at multiple scales. The ability to disentangle the relative effects of these factors is currently hampered by the lack of reliable topoclimatic data, which describe how regional climatic characteristics are modified by topographic effects in mountain areas. In this study we present an analysis of the combined effects of local- and regional-scale factors on southern beech treeline elevation variability at 28 study areas across New Zealand. We apply a mesoscale atmospheric model to generate local-scale (200 m) meteorological data at these treelines and, from these data, we derive a set of topoclimatic indices that reflect possible detrimental and ameliorative influences on tree physiological functioning. Principal components analysis of meteorological data revealed geographic structure in how study areas were situated in multivariate space along gradients of topoclimate. Random forest and conditional inference tree modelling enabled us to tease apart the relative effects of 17 explanatory factors on local-scale treeline elevation variability. Overall, modelling explained about 50% of the variation in treeline elevation variability across the 28 study areas, with local landform and topoclimatic effects generally outweighing those from regional-scale factors across the 28 study areas. Further, the nature of the relationships between treeline elevation variability and the explanatory variables were complex, frequently non-linear, and consistent with the treeline literature. To our knowledge, this is the first study where model-generated meteorological data, and derived topoclimatic indices, have been developed and applied to explain treeline variation. Our results demonstrate the potential of such an approach for ecological research in mountainous environments. The tree limit, the uppermost elevation at which trees can survive, is a global bioclimatic phenomenon ultimately determined by growing season temperature . HoweverIt is relatively easy to obtain GIS-ready datasets, such as gridded climatic and topographic data, for investigating influences on treelines at a regional scale e.g., . HoweverHowever, there are limits to the extent to which local-scale topoclimatic effects, in particular, can be sufficiently represented by such DEM-derived indices. Daily variation in local wind speeds, for instance, are the result of a range of meteorological processes inducing effects such as valley and downslope winds, cold air ponding, and differential irradiation, which are highly variable in space and time . Such toOne such approach is to use predictive, numerical climate models that are capable of generating accurate estimates of meteorological parameters in complex terrain and that can be applied to different sites without the need for local parameterisation. A number of readily-available mesoscale atmospheric models are suited to this task. For example, The Air Pollution Model (TAPM) produced by CSIRO Australia  is a mesDaily and monthly variation in wind speed, temperature extremes, solar radiation, and relative humidity, and interactions among these variables, together define possible topoclimatic conditions at the local treeline. These meteorological variables rarely affect treelines in isolation, but rather work synergistically to produce conditions that affect trees’ physiological performance. For instance, although high winds can potentially cause direct physical damage to trees at high altitudes, this type of damage alone is typically not a critical factor in explaining treeline formation . More daversus other factors in describing such variation at this spatial scale. Previous studies have highlighted the considerable variability observed in the elevation of New Zealand’s southern beech (Nothofagaceae) treelines both locally and regionally, and the possible explanations for this variability  treelines, which comprise the majority of New Zealand’s treeline zones. These treelines are highly-abrupt and have had minimal above-treeline recruitment of new stems over the past 20 or more years . Thus, wiability . For exaiability . With th2) in size and were randomly located across southern beech treeline zones from approximately 46°S latitude in the south of the country to 39°S latitude in the north. Study area dimensions were determined by the requirements of the TAPM model and its application for our research aims (see below). The mean distance from each site location to the next closest site was 32.4-km. To verify that all treeline point locations within these study areas were actually located at treeline, we visually assessed sample points against georeferenced, 15-m resolution, SPOT 5 satellite imagery  V.4  was used−2), and for wind speeds at 10 m above the ground (m s−1). To obtain TAPM-generated data at each treeline sample point, we first generated geo-referenced grids of data for each topoclimatic variable within ArcGIS at each site. We then extracted these data to treeline point locations in the GIS using standard raster-to-point data extraction methods.To run TAPM at its finest resolution (200-m grid cells), square 7 × 7-km study areas centred on each site location were established, resulting in the generation of meteorological estimates for 1,225 grid cells at each study area for each of the two months. Meteorological outputs from the model comprised hourly data for screen-level (2-m) air temperatures (°C), relative humidities (%), and short- and long-wave radiation values , and topoclimatic amelioration (insolation) on tree physiological function at treeline. All indices were computed for the months of January and July in order to determine how these indices vary in summer and winter, and if this variation is important for understanding treeline variability.Thourly) were first rescaled relative to the overall observed site-level maximum temperature such that lower temperatures received a higher relative weighting in calculating the potential for photoinhibition at a given treeline location. The photoinhibition index was then calculated as the product of rescaled hourly temperatures andhourly solar radiation (SolRadhourly) summed across all daytime hours and divided by the number of monthly daytime hours (Ndaytime): There is considerable evidence from treeline research that increased sky exposure is detrimental to seedling establishment above existing treelines . This efDesiccation stress can occur at treeline in both summer and winter . During Ntotal) and with temperatures rescaled such that low temperatures (inducing possible frost drought conditions) contribute more to high index values: This index was similarly calculated as in Nothofagus treelines in New Zealand  and wind speeds (WShourly) to a reverse scale at each site. In this way, temperatures and wind speeds that are low relative to the site-level maximum for these variables contribute towards a higher frost index value, while those that approach the site-level maximum contribute towards a lower frost index. NetRadhourly is the hourly net radiation, computed by TAPM as the difference between incoming solar radiation and outgoing radiation emitted from the land surface; NetRadhourly values during night-time hours are negative as there is no incoming solar radiation. Hourly frost index values calculated in this manner are then summed across all night-time hours in each of January and July, and divided by the number of monthly night-time hours for those months (Nnight-time).Cold night-time temperatures combined with low wind speeds and high levels of outgoing, long-wave radiation can lead to frost conditions . Frost d Zealand . A frostDifferences in solar radiation loadings among treeline locations typically reflect either differences in topographic orientation relative to the sun  or differences in the amount of cloud cover over time. Insolation is a key topoclimatic variable at treeline, and can exert both positive and negative effects. In general, locations with higher solar radiation, that are not also subjected to cold night-time/early morning temperatures, are likely to have more favourable conditions for growth due to increased warmth . InsolatWe used principal components analysis (PCA) to decompose variation in, and examine correlations among, the raw, TAPM-generated topoclimatic variables across the 28 study areas. To visualise these outputs, we plotted PCA bi-plots of axis combinations that explained at least 10% of the variation in the multivariate data; the location of study areas in ordination space were also overlain onto these bi-plots and coloured by their latitudes, to determine if topoclimatic data showed broad geographic structuring.To investigate relationships between the response variable and the 17 explanatory factors, we used a two-stage approach: first we used random forest analysis  to estimTreeline elevation across the 28 study areas ranged from 763 to 1,486-m . There was a large amount of variation within and among study areas in the deviation of treeline elevations from their potential study area maxima . Across Approximately 80% of variation in the meteorological data generated by TAPM across the treeline locations was explained by the first three principal component axes. The first principal component axis was most strongly characterised by negative loadings for January and July minimum and maximum relative humidity, rain days, and night time longwave radiation, and positive loadings for January and July solar radiation . The secThe random forest analysis explained approximately 50% of variation in treeline elevation deviation based on the 17 explanatory factors see . Next, aThe conditional inference tree analysis produced twelve terminal nodes using six of the 12 explanatory variables entered into the analysis: precipitation, curvature, slope gradient, growing season temperature, July frost index, and July desiccation index . The firAlthough a range of previous treeline studies have used combinations of topography-based metrics and/or interpolated climatic datasets for treeline modelling e.g., , the preClear topoclimatic gradients were identified using TAPM-generated meteorological data, suggesting that there is a characteristic set of topographically-mediated processes acting across these treelines in New Zealand. Further, different treeline study areas were grouped by distinct topoclimatic conditions as represented by these gradients, likely reflecting the way in which geographic differences in mountain characteristics and regional climates drive these patterns. It is well recognised that the size of mountain ranges and their orientation relative to prevailing winds, valley widths and slope angles all affect valley-scale thermodynamic processes that regulate wind speeds, temperature extremes, and atmospheric moisture levels at different locations . From a Most of the treeline locations investigated in this study were positioned between 100 and 300 m below the potential study area maxima, suggesting that southern beech treelines at many locations across New Zealand are likely occurring well below their temperature-based limit. A recent GIS-based analysis of New Zealand beech treeline elevations  showed tTreelines at sites located away from the wettest areas of New Zealand with relatively high mean growing season temperatures (>11.5-°C) were more likely to be situated closer to their maximum potential elevation. Variation in these factors essentially describes differences in regional thermal regimes across the 28 study areas; the effect of thermal regime on treeline position in this study is consistent with previous research that has shown that warmer, drier regions have higher treelines . At moreAll topoclimatic stressors and ameliorators, as defined by our topoclimatic indices, were relatively highly important in explaining treeline deviations. In general, treelines deviated increasingly further downhill from their potential maximum elevations with increasing topoclimatic stress  but were closer to their maxima at locations with higher solar radiation input. The night-time movement of cold air from upper to lower elevations will increase the potential for frost and early morning photoinhibition, while strong daytime upslope wind movement will likely enhance desiccation conditions in both summer and winter. There will therefore likely be a combined effect of these stressors in certain locations at elevations lower than the maximum potential treeline. Cumulative effects can act across seasons; for instance, early summer frosts may disrupt the de-hardening of leaf tissues thereby exacerbating desiccation damage during the following winter . These tIt is well-recognised that treeline patterns are driven, in certain contexts, by non-linear, threshold-like responses to underlying environmental factors . In thisFifty percent of the variation in treeline elevation deviation could not be explained by our models; explanatory factors appeared to be most useful in predicting deviations of 300-m or less. It is highly likely that treeline elevation deviations of more than 300-m were driven by local disturbances that were not well-represented by our explanatory factors. The effects of disturbances, including those from fire , heavy wThis study has demonstrated that models such as TAPM have potential for enabling local-scale investigations of topoclimatic effects at treeline. Clearly, two of the biggest advantages are the generation of spatially-explicit data useful for characterising stress-related effects and the ability to produce these data at any location. However, it is important to recognise that meteorological data for one year may not be representative of typical topoclimatic conditions occurring at a given location, and it would therefore be useful to average model data over longer periods and for other critical parts of the year such as late spring and early autumn. Overall, we show that landform, topoclimatic, and regional factors together can explain half the variation in local treeline elevation variation and that their influences are geographically-structured and typically non-linear in nature. Ultimately, results from this study could be used to characterise sites with different topoclimatic situations where investigations of local scale microclimate and biotic interactions could be investigated.10.7717/peerj.1334/supp-1Table S1Click here for additional data file.10.7717/peerj.1334/supp-2Supplemental Information 1This file contains raw meteorological and derived topclimatic index data used for the analysis of topoclimatic effects at treeline locations. Data include: Site ID Elevation Latitude  minimum, mean, and maximum temperature, wind speed, relative humidity; (b) mean and maximum values for derived photoinhibition, frost, and desiccation indices; (c) January and July temperature ranges; (d) total and mean daily solar radiation and night time net radiation; (e) the number of frost and rain days.Click here for additional data file.