Cannibalism and competition can increase parasite abundance for parasites with complex life history strategies.

Ecological interactions among hosts are critical to consider when predicting disease dynamics. Most theory predicts that intraguild predation (IGP) and cannibalism negatively impact parasite populations, but this is based primarily on assumptions of simple or single-host life cycles. Here we investigate the effects of cannibalism in a size-structured host population on two digenean trematodes that have complex, multihost life cycles. A high incidence of cannibalism among paratenic hosts produced higher parasite infection loads and abundance, whereas cannibalism among obligate hosts reduced parasite abundances. We attributed this difference to trophic transmission aggregating parasites in larger, potentially fitter hosts and also to transmission among paratenic hosts via cannibalism. Moreover, we found evidence of indirect competitive interactions between parasites that can also increase infections at small scales. Our results show there are multiple mechanisms through which high cannibalism environments can benefit parasites that use paratenic hosts and trophic transfer to complete their life cycles.

Deficiency of lncRNA MERRICAL abrogates macrophage chemotaxis and diabetes-associated atherosclerosis.

Diabetes-associated atherosclerosis involves excessive immune cell recruitment and plaque formation. However, the mechanisms remain poorly understood. Transcriptomic analysis of the aortic intima in Ldlr-/- mice on a high-fat, high-sucrose-containing (HFSC) diet identifies a macrophage-enriched nuclear long noncoding RNA (lncRNA), MERRICAL (macrophage-enriched lncRNA regulates inflammation, chemotaxis, and atherosclerosis). MERRICAL expression increases by 249% in intimal lesions during progression. lncRNA-mRNA pair genomic mapping reveals that MERRICAL positively correlates with the chemokines Ccl3 and Ccl4. MERRICAL-deficient macrophages exhibit lower Ccl3 and Ccl4 expression, chemotaxis, and inflammatory responses. Mechanistically, MERRICAL guides the WDR5-MLL1 complex to activate CCL3 and CCL4 transcription via H3K4me3 modification. MERRICAL deficiency in HFSC diet-fed Ldlr-/- mice reduces lesion formation by 74% in the aortic sinus and 86% in the descending aorta by inhibiting leukocyte recruitment into the aortic wall and pro-inflammatory responses. These findings unveil a regulatory mechanism whereby a macrophage-enriched lncRNA potently inhibits chemotactic responses, alleviating lesion progression in diabetes.

Incubation of methamphetamine craving in punishment-resistant individuals is associated with activation of specific gene networks in the rat dorsal striatum.

Methamphetamine use disorder (MUD) is characterized by loss of control over compulsive drug use. Here, we used a self-administration (SA) model to investigate transcriptional changes associated with the development of early and late compulsivity during contingent footshocks. Punishment initially separated methamphetamine taking rats into always shock-resistant (ASR) rats that continued active lever pressing and shock-sensitive (SS) rats that reduced their lever pressing. At the end of the punishment phase, rats underwent 15 days of forced abstinence at the end of which they were re-introduced to the SA paradigm followed by SA plus contingent shocks. Interestingly, 36 percent of the initial SS rats developed delayed shock-resistance (DSR). Of translational relevance, ASR rats showed more incubation of methamphetamine craving than DSR and always sensitive (AS) rats. RNA sequencing revealed increased striatal Rab37 and Dipk2b mRNA levels that correlated with incubation of methamphetamine craving. Interestingly, Bdnf mRNA levels showed HDAC2-dependent decreased expression in the AS rats. The present SA paradigm should help to elucidate the molecular substrates of early and late addiction-like behaviors.

Warming and top-down control of stage-structured prey: Linking theory to patterns in natural systems.

Warming has broad and often nonlinear impacts on organismal physiology and traits, allowing it to impact species interactions like predation through a variety of pathways that may be difficult to predict. Predictions are commonly based on short-term experiments and models, and these studies often yield conflicting results depending on the environmental context, spatiotemporal scale, and the predator and prey species considered. Thus, the accuracy of predicted changes in interaction strength, and their importance to the broader ecosystems they take place in, remain unclear. Here, we attempted to link one such set of predictions generated using theory, modeling, and controlled experiments to patterns in the natural abundance of prey across a broad thermal gradient. To do so, we first predicted how warming would impact a stage-structured predator-prey interaction in riverine rock pools between Pantala spp. dragonfly nymph predators and Aedes atropalpus mosquito larval prey. We then described temperature variation across a set of hundreds of riverine rock pools (n = 775) and leveraged this natural gradient to look for evidence for or against our model's predictions. Our model's predictions suggested that warming should weaken predator control of mosquito larval prey by accelerating their development and shrinking the window of time during which aquatic dragonfly nymphs could consume them. This was consistent with data collected in rock pool ecosystems, where the negative effects of dragonfly nymph predators on mosquito larval abundance were weaker in warmer pools. Our findings provide additional evidence to substantiate our model-derived predictions while emphasizing the importance of assessing similar predictions using natural gradients of temperature whenever possible.

Structure-Reactivity Studies of 2-Sulfonylpyrimidines Allow Selective Protein Arylation.

Protein arylation has attracted much attention for developing new classes of bioconjugates with improved properties. Here, we have evaluated 2-sulfonylpyrimidines as covalent warheads for the mild, chemoselective, and metal free cysteine S-arylation. 2-Sulfonylpyrimidines react rapidly with cysteine, resulting in stable S-heteroarylated adducts at neutral pH. Fine tuning the heterocyclic core and exocyclic leaving group allowed predictable SNAr reactivity in vitro, covering >9 orders of magnitude. Finally, we achieved fast chemo- and regiospecific arylation of a mutant p53 protein and confirmed arylation sites by protein X-ray crystallography. Hence, we report the first example of a protein site specifically S-arylated with iodo-aromatic motifs. Overall, this study provides the most comprehensive structure-reactivity relationship to date on heteroaryl sulfones and highlights 2-sulfonylpyrimidine as a synthetically tractable and protein compatible covalent motif for targeting reactive cysteines, expanding the arsenal of tunable warheads for modern covalent ligand discovery.

Responses to saltwater exposure vary across species, populations and life stages in anuran amphibians.

To predict the impacts of environmental change on species, we must first understand the factors that limit the present-day ranges of species. Most anuran amphibians cannot survive at elevated salinities, which may drive their distribution in coastal locations. Previous research showed that coastal Hyla cinerea are locally adapted to brackish habitats in North Carolina, USA. Although Hyla squirella and Hyla chrysoscelis both inhabit coastal wetlands nearby, they have not been observed in saline habitats. We take advantage of naturally occurring microgeographic variation in coastal wetland occupancy exhibited by these congeneric tree frog species to explore how salt exposure affects oviposition site choice, hatching success, early tadpole survival, plasma osmolality and tadpole body condition across coastal and inland locations. We observed higher survival among coastal H. cinerea tadpoles than inland H. cinerea, which corroborates previous findings. But contrary to expectations, coastal H. cinerea had lower survival than H. squirella and H. chrysoscelis, indicating that all three species may be able to persist in saline wetlands. We also observed differences in tadpole plasma osmolality across species, locations and salinities, but these differences were not associated with survival rates in salt water. Instead, coastal occupancy may be affected by stage-specific processes like higher probability of total clutch loss as shown by inland H. chrysoscelis or maladaptive egg deposition patterns as shown by inland H. squirella. Although we expected salt water to be the primary filter driving species distributions along a coastal salinity gradient, it is likely that the factors dictating anuran ranges along the coast involve stage-, species- and location-specific processes that are mediated by ecological processes and life history traits.

Electrostatically assembled wound dressings deliver pro-angiogenic anti-miRs preferentially to endothelial cells.

Chronic non-healing wounds occur frequently in individuals affected by diabetes, yet standard-of-care treatment leaves many patients inadequately treated or with recurring wounds. MicroRNA (miR) expression is dysregulated in diabetic wounds and drives an anti-angiogenic phenotype, but miRs can be inhibited with short, chemically-modified RNA oligonucleotides (anti-miRs). Clinical translation of anti-miRs is hindered by delivery challenges such as rapid clearance and uptake by off-target cells, requiring repeated injections, excessively large doses, and bolus dosing mismatched to the dynamics of the wound healing process. To address these limitations, we engineered electrostatically assembled wound dressings that locally release anti-miR-92a, as miR-92a is implicated in angiogenesis and wound repair. In vitro, anti-miR-92a released from these dressings was taken up by cells and inhibited its target. An in vivo cellular biodistribution study in murine diabetic wounds revealed that endothelial cells, which play a critical role in angiogenesis, exhibit higher uptake of anti-miR eluted from coated dressings than other cell types involved in the wound healing process. In a proof-of-concept efficacy study in the same wound model, anti-miR targeting anti-angiogenic miR-92a de-repressed target genes, increased gross wound closure, and induced a sex-dependent increase in vascularization. Overall, this proof-of-concept study demonstrates a facile, translational materials approach for modulating gene expression in ulcer endothelial cells to promote angiogenesis and wound healing. Furthermore, we highlight the importance of probing cellular interactions between the drug delivery system and the target cells to drive therapeutic efficacy.

MicroRNA-375 repression of Kruppel-like factor 5 improves angiogenesis in diabetic critical limb ischemia.

Peripheral artery disease (PAD) is an occlusive disease of limb arteries. Critical limb ischemia (CLI) is an advanced form of PAD that is prognostically worse in subjects with diabetes and can result in limb loss, gangrene, and death, although the underlying signaling mechanisms that contribute to its development remain poorly understood. By comparing plasma samples from diabetic humans with PAD and mouse models of PAD, we identified miR-375 to be significantly downregulated in humans and mice during progression to CLI. Overexpression of miR-375 was pro-angiogenic in endothelial cells in vitro and induced endothelial migration, proliferation, sprouting, and vascular network formation, whereas miR-375 inhibition conferred anti-angiogenic effects. Intramuscular delivery of miR-375 improved blood flow recovery to diabetic mouse hindlimbs following femoral artery ligation (FAL) and improved neovessel growth and arteriogenesis in muscle tissues. Using RNA-sequencing and prediction algorithms, Kruppel-like factor 5 (KLF5) was identified as a direct target of miR-375 and siRNA knockdown of KLF5 phenocopied the effects of miR-375 overexpression in vitro and in vivo through regulatory changes in NF-kB signaling. Together, a miR-375-KLF5-NF-kB signaling axis figures prominently as a potential therapeutic pathway in the development CLI in diabetes.

Biochemical Neuroadaptations in the Rat Striatal Dopaminergic System after Prolonged Exposure to Methamphetamine Self-Administration.

Perturbations in striatal dopamine (DA) homeostasis might underlie the behavioral and pathobiological consequences of METH use disorder in humans. To identify potential consequences of long-term METH exposure, we modeled the adverse consequence DSM criterion of substance use disorders by giving footshocks to rats that had escalated their intake of METH during a drug self-administration procedure. Next, DA D1 receptor antagonist, SCH23390 was injected. Thereafter, rats were euthanized to measure several indices of the striatal dopaminergic system. Footshocks split the METH rats into two phenotypes: (i) shock-sensitive that decreased their METH-intake and (ii) shock-resistant that continued their METH intake. SCH23390 caused substantial dose-dependent reduction of METH taking in both groups. Stopping SCH23390 caused re-emergence of compulsive METH taking in shock-resistant rats. Compulsive METH takers also exhibited greater incubation of METH seeking than non-compulsive rats during withdrawal from METH SA. Analyses of DA metabolism revealed non-significant decreases (about 35%) in DA levels in resistant and sensitive rats. However, striatal contents of the deaminated metabolites, DOPAL and DOPAC, were significantly increased in sensitive rats. VMAT2 and DAT protein levels were decreased in both phenotypes. Moreover, protein expression levels of the D1-like DA receptor, D5R, and D2-like DA receptors, D3R and D4R, were significantly decreased in the compulsive METH takers. Our results parallel findings in post-mortem striatal tissues of human METH users who develop Parkinsonism after long-term METH intake and support the use of this model to investigate potential therapeutic interventions for METH use disorder.

A 360 Evaluation of Neonatal Care Quality at a Level II Neonatal Intensive Care Unit when Delivered Using a Hybrid Telemedicine Service.

OBJECTIVE: In 2013, the Section of Neonatal and Perinatal Medicine at the University of Oklahoma's Children's Hospital began providing advanced care to a regional level II neonatal intensive care unit (NICU), using a hybrid telemedicine program. This project compares health care providers' and parents' assessments of health care quality using this program. STUDY DESIGN: This is a prospective, anonymous, nonrandomized survey of health care providers and parents of neonates using our hybrid telemedicine services. Physicians, neonatal nurse practitioners (NNPs), nurses, and parents completed pencil-and-paper surveys based on their participatory roles. Institutional Review Board approval was obtained at OU Medical Center and Comanche County Memorial Hospital. Surveys consisted of 5-point Likert's scale questions. Descriptive statistics compared the level of agreement with each question across participant groups. A service quality (SQ) composite score was created by summing responses from six SQ questions. Between-group analysis was done on the SQ score using the Mann-Whitney U-test. RESULTS: Nine physicians, 10 NNPs, 12 nurses, and 40 parents completed the survey. Providers agreed (90%) that telemedicine can effectively deliver advanced neonatal care; the care patients receive is comparable to direct patient care (87%); telemedicine enhanced overall patient care quality (90%); providers can effectively interact with each other and families using telemedicine (90.3%), and overall telemedicine experience was good (90%). In total, 61% of providers reported telemedicine improves physician-patient interaction. Parents of newborns agreed that they were well informed about telemedicine use for their child's care (88%), were able to communicate routinely with neonatologists (85%), and were comfortable with their child's physical examinations (93%). Provider's versus family's (SQ) score was not significantly different. CONCLUSION: All survey participants, including neonatologists, NNPs, nurses, and patient families, reported high levels of satisfaction with the hybrid telemedicine model developed and implemented at this institution which may be comparable to in-person direct patient care. KEY POINTS: · Implementation of a hybrid telemedicine system provides an alternative to the transfer of newborns needing advanced care to tertiary care facilities.. · In this study, both health care providers and patient family members were satisfied with the quality of care using hybrid telemedicine.. · In this study, families of newborns could fully participate in their child's care using the hybrid telemedicine system..

Incorporating nonlinearity with generalized functional responses to simulate multiple predator effects.

Predicting the combined effects of predators on shared prey has long been a focus of community ecology, yet quantitative predictions often fail. Failure to account for nonlinearity is one reason for this. Moreover, prey depletion in multiple predator effects (MPE) studies generates biased predictions in applications of common experimental and quantitative frameworks. Here, we explore additional sources of bias stemming from nonlinearities in prey predation risk. We show that in order to avoid bias, predictions about the combined effects of independent predators must account for nonlinear size-dependent risk for prey as well as changes in prey risk driven by nonlinear predator functional responses and depletion. Historical failure to account for biases introduced by well-known nonlinear processes that affect predation risk suggest that we may need to reevaluate the general conclusions that have been drawn about the ubiquity of emergent MPEs over the past three decades.

Spatiotemporal Variation in Hatching Success and Nestling Sex Ratios Track Rapid Movement of a Songbird Hybrid Zone.

AbstractHybridization often occurs at the parapatric range interface between closely related species, but fitness outcomes vary: hybrid offspring exhibit diverse rates of viability and reproduction compared with their parental species. The mobile hybrid zone between two chickadee congeners (Poecile atricapillus × Poecile carolinensis) has been well studied behaviorally and genetically, but the viability of hybrids and the underlying mechanisms contributing to hybrid fitness have remained unclear. To better characterize the fitness costs of hybridization in this system, we analyzed 21 years of data from four sites, including more than 1,400 breeding attempts by the two species, to show that rates of hatching success changed substantially as the zone of hybridization moved across the landscape. Admixture-associated declines in hatching success correlated with reduced proportions of heterogametic (female) offspring, as predicted by Haldane's rule. Our data support an underlying mechanism implicating genetic admixture of the homogametic (male) parent as the primary determinant of offspring sex ratio, via incompatibilities on the hemizygous Z chromosome. Our long-term study is the first to directly measure changes in fitness costs as a vertebrate hybrid zone moves, and it shows that changes in these costs are a way to track the distribution of a hybrid zone across the landscape.

Biophysical Survey of Small-Molecule ß-Catenin Inhibitors: A Cautionary Tale.

The canonical Wingless-related integration site signaling pathway plays a critical role in human physiology, and its dysregulation can lead to an array of diseases. ß-Catenin is a multifunctional protein within this pathway and an attractive yet challenging therapeutic target, most notably in oncology. This has stimulated the search for potent small-molecule inhibitors binding directly to the ß-catenin surface to inhibit its protein-protein interactions and downstream signaling. Here, we provide an account of the claimed (and some putative) small-molecule ligands of ß-catenin from the literature. Through in silico analysis, we show that most of these molecules contain promiscuous chemical substructures notorious for interfering with screening assays. Finally, and in line with this analysis, we demonstrate using orthogonal biophysical techniques that none of the examined small molecules bind at the surface of ß-catenin. While shedding doubts on their reported mode of action, this study also reaffirms ß-catenin as a prominent target in drug discovery.

Timely Wound Healing Is Dependent on Endothelial but Not on Hair Follicle Stem Cell Toll-Like Receptor 2 Signaling.

As a part of innate immunity, toll-like receptor 2 (TLR2) plays an important function in most defensive responses of the organism, including but not limited to infections. Cutaneous injury, one of the most common challenges for mammals, mobilizes a number of cell types, including epithelial, immune, and vascular cells, for timely tissue repair. However, in contrast to immune cells, little is known about TLR2 function on nonimmune cells during skin regeneration. In this study, we used two tissue-specific conditional Tlr2-knockout mouse lines to address the effects of TLR2 in endothelial and hair follicle stem cells (HFSCs) on cutaneous wound healing. The loss of TLR2 on endothelial cells diminishes their ability to migrate, sprout, and proliferate in response to specific TLR2 ligands and also reduces the secretion of key proangiogenic factors. Lack of TLR2 on endothelial cells prolongs wound healing owing to diminished angiogenesis. TLR2 is expressed in key structures of hair follicles, including HFSCs, secondary hair germ, and dermal papilla. Despite the prominent role of HFSCs in skin regeneration, excision of TLR2 from HFSCs has no effects on their proliferation or wound healing potential. Our study shows that timely tissue regeneration after skin injury is dependent on endothelial TLR2 for robust angiogenesis, whereas HFSC TLR2 is dispensable.

miR-181b regulates vascular endothelial aging by modulating an MAP3K3 signaling pathway.

Endothelial cell (EC) aging plays a vital role in the pathogenesis of cardiovascular disease (CVD). MicroRNAs have emerged as crucial regulators of target gene expression by inhibiting mRNA translation and/or promoting mRNA degradation. We identify an aging-related and oxidative stress-responsive microRNA, miR-181b, that inhibits endothelial cell apoptosis and senescence. In gain- or loss-of-function studies, miR-181b regulated the expression of key apoptosis markers (Bcl2, Bax, cleaved-Caspase3) and senescence markers (p16, p21, γH2AX) and the ratio of apoptotic cells (TUNEL-positive) and senescent cells (SA-ßgal-positive) in H2 O2 -induced ECs. Mechanistically, miR-181b targets MAP3K3 and modulates a MAP3K3/MKK/MAPK signaling pathway. MAP3K3 knockdown recapitulated the phenotype of miR-181b overexpression and miR-181b was dependent on MAP3K3 for regulating EC apoptosis and senescence. In vivo, miR-181b expression showed a negative correlation with increasing age in the mouse aorta. Endothelial-specific deficiency of miR-181a2b2 increased the target MAP3K3, markers of vascular senescence (p16, p21), and DNA double-strand breaks (γH2AX) in the aorta of aged mice. Collectively, this study unveils an important role of miR-181b in regulating vascular endothelial aging via an MAP3K3-MAPK signaling pathway, providing new potential therapeutic targets for antiaging therapy in CVD.

Endothelial cell-specific deletion of a microRNA accelerates atherosclerosis.

BACKGROUND AND AIMS: Chronic vascular endothelial inflammation predisposes to atherosclerosis; however, the cell-autonomous roles for endothelial-expressing microRNAs (miRNAs) are poorly understood in this process. MiR-181b is expressed in several cellular constituents relevant to lesion formation. The aim of this study is to examine the role of genetic deficiency of the miR-181b locus in endothelial cells during atherogenesis. METHODS AND RESULTS: Using a proprotein convertase subtilisin/kexin type 9 (PCSK9)-induced atherosclerosis mouse model, we demonstrated that endothelial cell (EC)-specific deletion of miR-181a2b2 significantly promoted atherosclerotic lesion formation, cell adhesion molecule expression, and the influx of lesional macrophages in the vessel wall. Yet, endothelium deletion of miR-181a2b2 did not affect body weight, lipid metabolism, anti-inflammatory Ly6Clow or the pro-inflammatory Ly6Cinterm and Ly6Chigh fractions in circulating peripheral blood mononuclear cells (PBMCs), and pro-inflammatory or anti-inflammatory mediators in both bone marrow (BM) and PBMCs. Mechanistically, bulk RNA-seq and gene set enrichment analysis of ECs enriched from the aortic arch intima, as well as single cell RNA-seq from atherosclerotic lesions, revealed that endothelial miR-181a2b2 serves as a critical regulatory hub in controlling endothelial inflammation, cell adhesion, cell cycle, and immune response during atherosclerosis. CONCLUSIONS: Our study establishes that deficiency of a miRNA specifically in the vascular endothelium is sufficient to profoundly impact atherogenesis. Endothelial miR-181a2b2 deficiency regulates multiple key pathways related to endothelial inflammation, cell adhesion, cell cycle, and immune response involved in the development of atherosclerosis.

A miRNA cassette reprograms smooth muscle cells into endothelial cells.

Cellular reprogramming through targeting microRNAs (miRNAs) holds promise for regenerative therapy due to their profound regulatory effects in proliferation, differentiation, and function. We hypothesized that transdifferentiation of vascular smooth muscle cells (SMCs) into endothelial cells (ECs) using a miRNA cassette may provide a novel approach for use in vascular disease states associated with endothelial injury or dysfunction. miRNA profiling of SMCs and ECs and iterative combinatorial miRNA transfections of human coronary SMCs revealed a 4-miRNA cassette consisting of miR-143-3p and miR-145-5p inhibitors and miR-146a-5p and miR-181b-5p mimics that efficiently produced induced endothelial cells (iECs). Transcriptome profiling, protein expression, and functional studies demonstrated that iECs exhibit high similarity to ECs. Injected iECs restored blood flow recovery even faster than conventional ECs in a murine hindlimb ischemia model. This study demonstrates that a 4-miRNA cassette is sufficient to reprogram SMCs into ECs and shows promise as a novel regenerative strategy for endothelial repair.

Isolation and culture of murine aortic cells and RNA isolation of aortic intima and media: Rapid and optimized approaches for atherosclerosis research.

BACKGROUND AND AIMS: Isolation of cellular constituents from the mouse aorta is commonly used for expression or functional analyses in atherosclerosis research. However, current procedures to isolate primary cells are difficult, inefficient, and require separate mice. RNA extraction from aortic intima and media for transcriptomic analysis is also considered difficult with mixed RNA yields. To address these gaps, we provide: 1) a rapid, efficient protocol to isolate and culture diverse cell types concomitantly from the mouse aorta using immunomagnetic cell isolation; and 2) an optimized aortic intimal peeling technique for efficient RNA isolation from the intima and media. METHODS AND RESULTS: Aortic cells were obtained using an enzymatic solution and different cell types were isolated by magnetic beads conjugated to antibodies targeting endothelial cells (CD31+), leukocytes (CD45+), and fibroblast cells (CD90.2+), and smooth muscle cells were isolated by negative selection. Our protocol allows the isolation of relatively large numbers of cells (10,000 cells per aorta) in a predictable manner with high purity (>90%) verified by cell-marker gene expression, immunofluorescence, and flow cytometry. These cells are all functionally active when grown in cell culture. We also provide a rapid method to collect aortic intima-enriched RNA from Ldlr-/- mice utilizing an intima peeling approach and assess transcriptomic profiling associated with accelerated lesion formation. CONCLUSIONS: This protocol provides an effective means for magnetic bead-based isolation of different cell types from the mouse aortic wall, and the isolated cells can be utilized for functional and mechanistic studies for a range of vascular diseases including atherosclerosis.

Footshock-Induced Abstinence from Compulsive Methamphetamine Self-administration in Rat Model Is Accompanied by Increased Hippocampal Expression of Cannabinoid Receptors (CB1 and CB2).

Methamphetamine (METH) use disorder (MUD) is characterized by compulsive and repeated drug taking despite negative life consequences. Large intake of METH in humans and animals is accompanied by dysfunctions in learning and memory processes. The endocannabinoid system (ECS) is known to modulate synaptic plasticity and cognitive functions. In addition, the ECS has been implicated in some of the manifestations of substance use disorders (SUDs). We therefore sought to identify potential changes in the expression of various enzymes and of the receptors (CB1 and CB2) that are members of that system. Herein, we used a model of METH self-administration (SA) that includes a punishment phase (footshocks) that helps to separate rats into a compulsive METH phenotype (compulsive) that continues to take METH and a non-compulsive METH (abstinent) group that suppressed or stopped taking METH. Animals were euthanized 2 h after the last METH SA session and their hippocampi were used to measure mRNA levels of cannabinoid receptors (CB/Cnr), as well as those of synthesizing (DAGL-A, DAGL-B, NAPEPLD) and metabolizing (MGLL, FAAH, PTGS2) enzymes of the endocannabinoid cascade. Non-compulsive rats exhibited significant increased hippocampal expression of CB1/Cnr1 and CB2/Cnr2 mRNAs. mRNA levels of the synthesizing enzyme, DAGL-A, and of the metabolic enzymes, MGLL and FAAH, were also increased. Non-compulsive rats also exhibited a significant decrease in hippocampal Ptgs2 mRNA levels. Taken together, these observations implicate the hippocampal endocannabinoid system in the suppression of METH intake in the presence of adverse consequences.

Inflammation-dependent oxidative stress metabolites as a hallmark of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease, with poor prognosis and no cure. Substantial evidence implicates inflammation and associated oxidative stress as a potential mechanism for ALS, especially in patients carrying the SOD1 mutation and, therefore, lacking anti-oxidant defense. The brain is particularly vulnerable to oxidation due to the abundance of polyunsaturated fatty acids, such as docosahexaenoic acid (DHA), which can give rise to several oxidized metabolites. Accumulation of a DHA peroxidation product, CarboxyEthylPyrrole (CEP) is dependent on activated inflammatory cells and myeloperoxidase (MPO), and thus marks areas of inflammation-associated oxidative stress. At the same time, generation of an alternative inactive DHA peroxidation product, ethylpyrrole, does not require cell activation and MPO activity. While absent in normal brain tissues, CEP is accumulated in the central nervous system (CNS) of ALS patients, reaching particularly high levels in individuals carrying a SOD1 mutation. ALS brains are characterized by high levels of MPO and lowered anti-oxidant activity (due to the SOD1 mutation), thereby aiding CEP generation and accumulation. Due to DHA oxidation within the membranes, CEP marks cells with the highest oxidative damage. In all ALS cases CEP is present in nearly all astrocytes and microglia, however, only in individuals carrying a SOD1 mutation CEP marks >90% of neurons, thereby emphasizing an importance of CEP accumulation as a potential hallmark of oxidative damage in neurodegenerative diseases.