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Background and Purpose\ud “Food addiction” is the subject of intense public and research interest. However, this nosology based on neurobehavioral similarities among obese individuals and patients with eating disorders and drug addiction remains controversial. We thus sought to determine which aspects of disordered eating are causally linked to preclinical models of drug addiction. We hypothesized that extensive drug histories, known to cause addiction-like brain changes and drug motivation in rats, would also cause addiction-like food motivation.\ud \ud Experimental Approach\ud Rats underwent extensive cocaine, alcohol, caffeine or obesogenic diet histories, and were subsequently tested for punishment-resistant food self-administration or “compulsive appetite”, as a measure of addiction-like food motivation.\ud \ud Key Results\ud Extensive cocaine and alcohol (but not caffeine) histories caused compulsive appetite that persisted long after the last drug exposure. Extensive obesogenic diet histories also caused compulsive appetite, although neither cocaine nor alcohol histories caused excess calorie intake and bodyweight during abstinence. Hence, compulsive appetite and obesity appear to be dissociable, with the former sharing common mechanisms with preclinical drug addiction models.\ud \ud Conclusion and Implications\ud Compulsive appetite, as seen in subsets of obese individuals and patients with binge-eating disorder and bulimia nervosa (eating disorders that do not necessarily result in obesity), appears to epitomize “food addiction”. Because different drug and obesogenic diet histories caused compulsive appetite, overlapping dysregulations in the reward circuits, which control drug and food motivation independently of energy homeostasis, may offer common therapeutic targets for treating addictive behaviors across drug addiction, eating disorders and obesity.

Background Cannabidiol (CBD) has received attention for the treatment of substance use disorders. In preclinical models of relapse, CBD attenuates drug seeking across several drugs of abuse, including cocaine. However, in these models CBD has not been consistently effective. This inconsistency in CBD effects may be related to presently insufficient information on the full spectrum of CBD dose effects on drug-related behaviors. Methods We address this issue by establishing a full dose-response profile of CBD's actions using expression of cocaine-induced conditioned place preference as a model for drug-motivated behavior in male rats and by concurrently identifying dose-dependent effects of CBD on underlying neuronal activation and distinct neuronal phenotypes showing dose-dependent activation changes. Additionally, we established CBD levels in plasma and brain samples. Results CBD produced linear increases in CBD brain/plasma concentrations but suppressed conditioned place preference in a distinct U-shaped manner. In parallel with its behavioral effects, CBD produced U-shaped suppressant effects on neuronal activation in the prelimbic but not infralimbic cortex or nucleus accumbens core and shell. RNAscope in situ hybridization identified suppression of glutamatergic and GABAergic (gamma-aminobutyric acidergic) signaling in the prelimbic cortex as a possible cellular mechanism for the attenuation of cocaine-induced conditioned place preference by CBD. Conclusions The findings extend previous evidence on the potential of CBD in preventing drug-motivated behavior. However, CBD's dose-response profile may have important dosing implications for future clinical applications and may contribute to the understanding of discrepant CBD effects on drug seeking reported in the literature.

The concept of evolvability (the capacity of populations to evolve) has deep historical roots in evolutionary biology. Interest in the subject has been renewed recently by innovations in microbiology that permit direct tests of the causes of evolvability, and with the acknowledgement that evolvability of pathogens has important implications for human health. Here, we investigate how fluctuating selection on the virus, Bacteriophage λ, affects its evolvability. We imposed dynamic selection by altering the expression of two host outer membrane receptors. This, in turn, selected phage to alternately infect the host via a single, or multiple, receptors. Our selection regime resulted in two orthogonal evolutionary behaviors, namely enhanced or reduced evolvability. Strains with enhanced evolvability readily evolved between receptors, losing and gaining the ability to bind multiple receptors more quickly than the ancestral λ. This suggests the receptor-binding protein retained a genetic memory of past states and that evolutionary history can be used to predict future adaptation. Strains with reduced evolvability were refractory to re-specialization and remained generalists on both receptors. Consistent with this behavior, unevolvable strains had reduced rates of molecular evolution in the receptor-binding protein compared to their evolvable counterparts. We found a single mutation in the receptor-binding protein was sufficient to render these strains resistant to evolution and did so by counteracting a receptor-binding trade-off associated with generalism. In this way, cost-free generalization allowed for reduced evolution and evolvability while maximizing success in both environments. Our results suggest the response to fluctuating selection is contingent and can lead to distinct differences in evolvability. These findings contribute to a growing understanding of the causes and consequences of evolvability and have important implications for infectious disease management.

Conflicting evidence exists regarding the role of infralimbic cortex (IL) in the environmental control of appetitive behavior. Inhibition of IL, irrespective of its intrinsic neural activity, attenuates not only the ability of environmental cues predictive of reward availability to promote reward seeking, but also the ability of environmental cues predictive of reward omission to suppress this behavior. Here we report that such bidirectional behavioral modulation in rats is mediated by functionally distinct units of neurons (neural ensembles) that are concurrently localized within the same IL brain area but selectively reactive to different environmental cues. Ensemble-specific neural activity is thought to function as a memory engram representing a learned association between environment and behavior. Our findings establish the causal evidence for the concurrent existence of two distinct engrams within a single brain site, each mediating opposing environmental actions on a learned behavior. DOI: http://dx.doi.org/10.7554/eLife.21920.001

The TripleTOF 5600 System, a hybrid quadrupole time-of-flight mass spectrometer, was evaluated to explore the key figures of merit in generating peptide and protein identifications that included spectral acquisition rates, data quality, proteome coverage, and biological depth. Employing a Saccharomyces cerevisiae tryptic digest, careful consideration of several performance features demonstrated that the speed of the TripleTOF contributed most to the resultant data. The TripleTOF system was operated with 8, 20, and 50 MS/MS events in an effort to compare with other MS technologies and to demonstrate the abilities of the instrument platform.

The proteome of extremely thermophilic microorganisms affords a glimpse into the dynamics of microbial ecology of high temperature environments. The secretome, or extracellular proteome of these microorganisms, no doubt harbors technologically important enzymes and other thermostable biomolecules that, to date, have been characterized only to a limited extent. In the first of a two-part study on selected thermophiles, defining the secretome requires a sample preparation method that has no negative impact on all downstream experiments. Following efficient secretome purification, GeLC-MS(2) analysis and prediction servers suggested probable protein secretion to complement experimental data. In an effort to define the extracellular proteome of the extreme thermophilic bacterium Caldicellulosiruptor saccharolyticus, several techniques were considered regarding sample processing to achieve the most in-depth analysis of secreted proteins. Order of operation experiments, all including the C(18) bead technique, demonstrated that two levels of sample purification were necessary to effectively desalt the sample and provide sufficient protein identifications. Five sample preparation combinations yielded 71 proteins and the majority described, as enzymatic and putative uncharacterized proteins, anticipate consolidated bioprocessing applications. Nineteen proteins were predicted by Phobius, SignalP, SecretomeP, or TatP for extracellular secretion, and 11 contained transmembrane domain stretches suggested by Phobius and transmembrane hidden Markov model. The sample preparation technique demonstrating the most effective outcome for C. saccharolyticus secreted proteins in this study, involved acetone precipitation followed by the C(18) bead method in which 2.4% (63 proteins) of the predicted proteome was identified, including proteins suggested to have secretion and transmembrane moieties.

Background The epidemiological importance of varicella and zoster and the availability of an efficacious and safe vaccine have led to an important international debate regarding the suitability of mass vaccination. The objective of the study was to describe the epidemiology of varicella and zoster in Italy and to determine whether there have been changes with respect to observations provided by an analogous study conducted 8 years ago, in order to define the most appropriate vaccination strategy. Methods A number of data sources were evaluated, a cross-sectional population-based seroprevalence study was conducted on samples collected in 2004, and the results were compared with data obtained in 1996. Results The data from active and passive surveillance systems confirm that varicella is a widespread infectious disease which mainly affects children. VZV seroprevalence did not substantially differ from that found in the previous study. The sero-epidemiological profile in Italy is different from that in other European countries. In particular, the percentage of susceptible adolescents is at least nearly twice as high as in other European countries and in the age group 20–39 yrs, approximately 9% of individuals are susceptible to VZV. Conclusion The results of this study can contribute to evaluating the options for varicella vaccination. It is possible that in a few years, in all Italian Regions, there will exist the conditions necessary for implementing a mass vaccination campaign and that the large-scale availability of MMRV tetravalent vaccines will facilitate mass vaccination.

In this paper, we consider two questions. First, we address whether there was proper standing for the Article III courts to decide United States v. Windsor, 570 U.S. 133 S. Ct. 2675, 2696 (2013) and Hollingsworth v. Perry, 133 S. Ct. 2652, 2668 (2013). We conclude that the third-party appellants lacked standing in federal court. Second, we examine whether cases challenging state same-sex marriage bans were and are cases in “law and equity” or instead, barred under the domestic relations exception for the purposes of federal question jurisdiction. We conclude that the domestic relations exception to federal jurisdiction is an archaic, historical remnant that should be overruled by the U.S. Supreme Court, and thus, the Article III federal courts have jurisdiction to hear pure marital status cases despite their domestic nature. We call on the Supreme Court to eliminate the domestic relations exception as to all forms of federal jurisdiction.

The TripleTOF 5600 System, a hybrid triple quadrupole time-of-flight mass spectrometer, was evaluated to explore the key figures of merit in generating peptide and protein identifications which included spectral acquisition rates, data quality, proteome coverage, and biological depth. Employing a Saccharomyces cerevisiae tryptic digest, careful consideration of several performance features demonstrated that the speed of the TripleTOF contributed most to the resultant data. The TripleTOF system was operated with 8, 20, and 50 MS/MS events in an effort to compare to other MS technologies and to demonstrate the abilities of the instrument platform.

Fractionation prior to mass spectrometry is an indispensable step in proteomics. In this paper we report the success of performing offline reversed phase high pressure liquid chromatography (HPLC) fractionation on a C18 2.0 mm × 150 mm column at the peptide level with microliter per minute flow rates prior to online nano-flow reversed phase liquid chromatography mass spectrometry (nanoLC-MS) using the well-studied fungus Saccharomyces cerevisiae. A C18 75 µm × 150 mm column was used online and the online elution gradients for each fraction were adjusted in order to obtain well resolved separation. Comparing this method directly to only performing nanoLC-MS we observed a 61.6% increase in the number of identified proteins. At a 1% false discovery rate 1028 proteins were identified using two dimensions of RPLC versus 636 proteins identified in a single nano-flow separation. The majority of proteins identified by one dimension of nano-LC were present in the proteins identified in our two dimensional strategy. Although increasing analysis time, this non-orthogonal and facile pre-fractionation method affords a more comprehensive examination of the proteome.

Design of experiments (DOE) was used to determine improved settings for a LTQ-Orbitrap XL to maximize proteome coverage of Saccharomyces cerevisiae. A total of nine instrument parameters were evaluated with the best values affording an increase of approximately 60% in proteome coverage. Utilizing JMP software, 2 DOE screening design tables were generated and used to specify parameter values for instrument methods. DOE 1, a fractional factorial design, required 32 methods fully resolving the investigation of six instrument parameters involving only half the time necessary for a full factorial design of the same resolution. It was advantageous to complete a full factorial design for the analysis of three additional instrument parameters. Measured with a maximum of 1% false discovery rate, protein groups, unique peptides, and spectral counts gauged instrument performance. Randomized triplicate nanoLC-LTQ-Orbitrap XL MS/MS analysis of the S. cerevisiae digest demonstrated that the following five parameters significantly influenced proteome coverage of the sample: (1) maximum ion trap ionization time; (2) monoisotopic precursor selection; (3) number of MS/MS events; (4) capillary temperature; and (5) tube lens voltage. Minimal influence on the proteome coverage was observed for the remaining four parameters (dynamic exclusion duration, resolving power, minimum count threshold to trigger a MS/MS event, and normalized collision energy). The DOE approach represents a time- and cost-effective method for empirically optimizing MS-based proteomics workflows including sample preparation, LC conditions, and multiple instrument platforms.

Probing the intracellular proteome of Thermotoga maritima and Caldicellulosiruptor saccharolyticus in pure and co-culture affords a global investigation into the machinery and mechanisms enduring inside the bacterial thermophilic cell at the time of harvest. The second of a two part study, employing GeLC-MS2 a variety of proteins were confidently identified with

The circulating concentration of a biomarker for congestive heart failure, Brain (B-type) Natriuretic Peptide (BNP-32), is measured using ELISA based assays in order to rapidly diagnose and monitor disease progression. The lack of molecular specificity afforded by these assays has recently come into question as emerging studies indicate there are potentially multiple heterogeneous forms of BNP in circulation with immunoreactive capabilities. In order to better understand the molecular biology of BNP-32 as it relates to congestive heart failure, it would thus be advantageous to use a detection platform such as Fourier transform ion cyclotron resonance mass spectrometry. This high resolving power mass spectrometer can provide unparalleled molecular specificity and can facilitate identification and characterization of the various molecular forms across all disease states. Unfortunately, BNP circulates at low concentrations (as low as 3 fmol/mL). Thus, it will require a collaborative effort from a number of orthogonal front-end technologies to overcome the disconnect between the practical detection limits of this instrument platform and the physiological levels of BNP-32 and its alternative molecular forms. Herein, we begin optimization of these front-end techniques by first enhancing the conditions for online nanoLC-ESI-MS separations of BNP-32 and its proteolytic fragments. Through extensive analysis of various chromatographic parameters we determined that Michrom Magic C8 stationary phase used in conjunction with a continuous, vented column configuration provided advanced chromatographic performance for the nano-flow separations involving intact BNP-32 and its associated tryptic peptides. Furthermore, conditions for the tryptic digestion of BNP-32 were also studied. We demonstrate that the use of free cysteine as an alkylation quenching agent and a secondary digestion within the digestion scheme can provide targeted tryptic peptides with increased abundances. Combined, these data will serve to further augment the detection of BNP-32 by LC–MS.

In Italy, although vaccination has been recommended for a number of years, vaccination coverage for mumps is still sub-optimal. The objective of the present study was to evaluate the seroprevalence of mumps antibodies in the Italian population, stratified by age, gender and geographical area. The proportion of individuals positive for mumps antibodies remained stable in the age classes 0–11 months and 1 year (25.4% and 30.8%, respectively) and showed a continuous increase after the second year of life. The percentage of susceptible individuals was higher than 20% in persons 2–14 years of age and exceeded 10% in persons 15–39 years of age. No statistically significant differences were observed by gender or geographical area. Comparison between these results and the data obtained from a 1996 survey showed a statistically significant increase in seroprevalence in the age class 2–4 years. No changes were observed in the other age-groups. The results of this study confirm that the efforts made in recent years to improve vaccination coverage within the second year of life should be strengthened.

The Automated Decision-Aid System for Hazardous Incidents (ADASHI) will be a software tool for incident commanders and response teams at CB incidents to aid in the decision-making process, act as a response template for guiding the actions of responders, and used as a tool for training and planning. The purpose of the user survey was to determine what the ADASHI tool should look like and how it should work. The areas of interest for the survey include the following: ease of use, "value added," areas of improvement, and needed features/functions. The assessment process was conducted in the following stages: User Survey Development; User Feedback Compilation; Focus Group Assessment; and Summary of Results. The entire process was successful. The developers gained useful insight into the needs of the responder community. The results of this survey will enable the developers of ADASHI to make necessary improvements in the software making ADASHI a valuable tool for its users.