Divergent effects of repeated cocaine and novel environment exposure on locus coeruleus c‐fos expression and brain catecholamine concentrations in rats

IntroductionChronic administration of cocaine causes a disinhibited, hyperexploratory response to novel environments. As the norepinephrine (NE) system regulates exploration and is dysregulated following cocaine exposure, we hypothesized that this cocaine‐mediated hyperexploratory response is associated with increased locus coeruleus (LC) reactivity.MethodsTo test this hypothesis, we used dual fluorescent in situ hybridization immunofluorescence to analyze novelty‐induced c‐fos and tyrosine hydroxylase expression in the LC and high‐pressure liquid chromatography to measure dopamine (DA) and NE concentrations in key catecholamine projection regions following exposure to cocaine.ResultsRepeated cocaine exposure followed by a 14‐day drug‐free period increased exploration of novel environments, replicating previous findings. Novelty exposure increased LC c‐fos expression, increased anterior cingulate NE, and decreased ventral tegmental area DA. Cocaine exposure decreased amygdala (AMY) DA, but had no effect on LC c‐fos expression or NE in any tested brain region. No interactions between cocaine and novelty were found. Open arm exploration was positively correlated with LC c‐fos expression and NE concentrations in both the anterior cingulate and nucleus accumbens, and negatively correlated with AMY DA concentration.ConclusionsOur findings confirm that exposure to novel environments increases LC activity and NE in the anterior cingulate cortex, that long‐term exposure to cocaine dysregulates AMY DA, and that disinhibited exploration in novel environments correlates with NE and DA in regions that modulate risk‐taking and avoidance behavior. Further studies investigating the effects of cocaine on brain catecholamine systems are important in understanding the long‐lasting effects of cocaine on brain function.Chronic cocaine exposure causes a long‐lasting, disinhibited, hyperexploratory phenotype. This effect may be partially driven by changes in locus coeruleus (LC) function, as LC activation in response to novel environments is correlated with this disinhibited exploratory behavior.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/148353/1/brb31222_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/148353/2/brb31222.pd

Nonthermal Emission from a Supernova Remnant in a Molecular Cloud

In evolved supernova remnants (SNRs) interacting with molecular clouds, such as IC 443, W44, and 3C391, a highly inhomogeneous structure consisting of a forward shock of moderate Mach number, a cooling layer, a dense radiative shell and an interior region filled with hot tenuous plasma is expected. We present a kinetic model of nonthermal electron injection, acceleration and propagation in that environment and find that these SNRs are efficient electron accelerators and sources of hard X- and gamma-ray emission. The energy spectrum of the nonthermal electrons is shaped by the joint action of first and second order Fermi acceleration in a turbulent plasma with substantial Coulomb losses. Bremsstrahlung, synchrotron, and inverse Compton radiation of the nonthermal electrons produce multiwavelength photon spectra in quantitative agreement with the radio and the hard emission observed by ASCA and EGRET from IC 443. We distinguish interclump shock wave emission from molecular clump shock wave emission accounting for a complex structure of molecular cloud. Spatially resolved X- and gamma- ray spectra from the supernova remnants IC 443, W44, and 3C391 as might be observed with BeppoSAX, Chandra XRO, XMM, INTEGRAL and GLAST would distinguish the contribution of the energetic lepton component to the gamma-rays observed by EGRET.Comment: 14 pages, 4 figure, Astrophysical Journal, v.538, 2000 (in press

Bremsstrahlung Suppression due to the LPM and Dielectric Effects in a Variety of Materials

The cross section for bremsstrahlung from highly relativistic particles is suppressed due to interference caused by multiple scattering in dense media, and due to photon interactions with the electrons in all materials. We present here a detailed study of bremsstrahlung production of 200 keV to 500 MeV photons from 8 and 25 GeV electrons traversing a variety of target materials. For most targets, we observe the expected suppressions to a good accuracy. We observe that finite thickness effects are important for thin targets.Comment: 52 pages, 13 figures (incorporated in the revtex LaTeX file

Multiplex quantitative PCR for single-reaction genetically modified (GM) plant detection and identification of false-positive GM plants linked to Cauliflower mosaic virus (CaMV) infection.

BACKGROUND:Most genetically modified (GM) plants contain a promoter, P35S, from the plant virus, Cauliflower mosaic virus (CaMV), and many have a terminator, TNOS, derived from the bacterium, Agrobacterium tumefaciens. Assays designed to detect GM plants often target the P35S and/or TNOS DNA sequences. However, because the P35S promoter is derived from CaMV, these detection assays can yield false-positives from non-GM plants infected by this naturally-occurring virus. RESULTS:Here we report the development of an assay designed to distinguish CaMV-infected plants from GM plants in a single multiplexed quantitative PCR (qPCR) reaction. Following initial testing and optimization via PCR and singleplex-to-multiplex qPCR on both plasmid and plant DNA, TaqMan qPCR probes with different fluorescence wavelengths were designed to target actin (a positive-control plant gene), P35S, P3 (a CaMV-specific gene), and TNOS. We tested the specificity of our quadruplex qPCR assay using different DNA extracts from organic watercress and both organic and GM canola, all with and without CaMV infection, and by using commercial and industrial samples. The limit of detection (LOD) of each target was determined to be 1% for actin, 0.001% for P35S, and 0.01% for both P3 and TNOS. CONCLUSIONS:This assay was able to distinguish CaMV-infected plants from GM plants in a single multiplexed qPCR reaction for all samples tested in this study, suggesting that this protocol is broadly applicable and readily transferrable to any interested parties with a qPCR platform

Deploying aptameric sensing technology for rapid pandemic monitoring

The genome of virulent strains may possess the ability to mutate by means of antigenic shift and/or antigenic drift as well as being resistant to antibiotics with time. The outbreak and spread of these virulent diseases including avian influenza (H1N1), severe acute respiratory syndrome (SARS-Corona virus), cholera (Vibrio cholera), tuberculosis (Mycobacterium tuberculosis), Ebola hemorrhagic fever (Ebola Virus) and AIDS (HIV-1) necessitate urgent attention to develop diagnostic protocols and assays for rapid detection and screening. Rapid and accurate detection of first cases with certainty will contribute significantly in preventing disease transmission and escalation to pandemic levels. As a result, there is a need to develop technologies that can meet the heavy demand of an all-embedded, inexpensive, specific and fast biosensing for the detection and screening of pathogens in active or latent forms to offer quick diagnosis and early treatments in order to avoid disease aggravation and unnecessary late treatment costs. Nucleic acid aptamers are short, single-stranded RNA or DNA sequences that can selectively bind to specific cellular and biomolecular targets. Aptamers, as new-age bioaffinity probes, have the necessary biophysical characteristics for improved pathogen detection. This article seeks to review global pandemic situations in relation to advances in pathogen detection systems. It particularly discusses aptameric biosensing and establishes application opportunities for effective pandemic monitoring. Insights into the application of continuous polymeric supports as the synthetic base for aptamer coupling to provide the needed convective mass transport for rapid screening is also presented

Frequency of KCNC3 DNA Variants as Causes of Spinocerebellar Ataxia 13 (SCA13)

Gain-of function or dominant-negative mutations in the voltage-gated potassium channel KCNC3 (Kv3.3) were recently identified as a cause of autosomal dominant spinocerebellar ataxia. Our objective was to describe the frequency of mutations associated with KCNC3 in a large cohort of index patients with sporadic or familial ataxia presenting to three US ataxia clinics at academic medical centers.DNA sequence analysis of the coding region of the KCNC3 gene was performed in 327 index cases with ataxia. Analysis of channel function was performed by expression of DNA variants in Xenopus oocytes.Sequence analysis revealed two non-synonymous substitutions in exon 2 and five intronic changes, which were not predicted to alter splicing. We identified another pedigree with the p.Arg423His mutation in the highly conserved S4 domain of this channel. This family had an early-onset of disease and associated seizures in one individual. The second coding change, p.Gly263Asp, subtly altered biophysical properties of the channel, but was unlikely to be disease-associated as it occurred in an individual with an expansion of the CAG repeat in the CACNA1A calcium channel.Mutations in KCNC3 are a rare cause of spinocerebellar ataxia with a frequency of less than 1%. The p.Arg423His mutation is recurrent in different populations and associated with early onset. In contrast to previous p.Arg423His mutation carriers, we now observed seizures and mild mental retardation in one individual. This study confirms the wide phenotypic spectrum in SCA13

Opposing effects of final population density and stress on Escherichia coli mutation rate

Evolution depends on mutations. For an individual genotype, the rate at which mutations arise is known to increase with various stressors (stress-induced mutagenesis-SIM) and decrease at high final population density (density-associated mutation-rate plasticity-DAMP). We hypothesised that these two forms of mutation-rate plasticity would have opposing effects across a nutrient gradient. Here we test this hypothesis, culturing Escherichia coli in increasingly rich media. We distinguish an increase in mutation rate with added nutrients through SIM (dependent on error-prone polymerases Pol IV and Pol V) and an opposing effect of DAMP (dependent on MutT, which removes oxidised G nucleotides). The combination of DAMP and SIM results in a mutation rate minimum at intermediate nutrient levels (which can support 7 × 10  cells ml ). These findings demonstrate a strikingly close and nuanced relationship of ecological factors-stress and population density-with mutation, the fuel of all evolution

Environmental pleiotropy and demographic history direct adaptation under antibiotic selection

Evolutionary rescue following environmental change requires mutations permitting population growth in the new environment. If change is severe enough to prevent most of the population reproducing, rescue becomes reliant on mutations already present. If change is sustained, the fitness effects in both environments, and how they are associated-termed 'environmental pleiotropy'-may determine which alleles are ultimately favoured. A population's demographic history-its size over time-influences the variation present. Although demographic history is known to affect the probability of evolutionary rescue, how it interacts with environmental pleiotropy during severe and sustained environmental change remains unexplored. Here, we demonstrate how these factors interact during antibiotic resistance evolution, a key example of evolutionary rescue fuelled by pre-existing mutations with pleiotropic fitness effects. We combine published data with novel simulations to characterise environmental pleiotropy and its effects on resistance evolution under different demographic histories. Comparisons among resistance alleles typically revealed no correlation for fitness-i.e., neutral pleiotropy-above and below the sensitive strain's minimum inhibitory concentration. Resistance allele frequency following experimental evolution showed opposing correlations with their fitness effects in the presence and absence of antibiotic. Simulations demonstrated that effects of environmental pleiotropy on allele frequencies depended on demographic history. At the population level, the major influence of environmental pleiotropy was on mean fitness, rather than the probability of evolutionary rescue or diversity. Our work suggests that determining both environmental pleiotropy and demographic history is critical for predicting resistance evolution, and we discuss the practicalities of this during in vivo evolution

Removal of Misincorporated Ribonucleotides from Prokaryotic Genomes: An Unexpected Role for Nucleotide Excision Repair

Stringent steric exclusion mechanisms limit the misincorporation of ribonucleotides by high-fidelity DNA polymerases into genomic DNA. In contrast, low-fidelity Escherichia coli DNA polymerase V (pol V) has relatively poor sugar discrimination and frequently misincorporates ribonucleotides. Substitution of a steric gate tyrosine residue with alanine (umuC_Y11A) reduces sugar selectivity further and allows pol V to readily misincorporate ribonucleotides as easily as deoxynucleotides, whilst leaving its poor base-substitution fidelity essentially unchanged. However, the mutability of cells expressing the steric gate pol V mutant is very low due to efficient repair mechanisms that are triggered by the misincorporated rNMPs. Comparison of the mutation frequency between strains expressing wild-type and mutant pol V therefore allows us to identify pathways specifically directed at ribonucleotide excision repair (RER). We previously demonstrated that rNMPs incorporated by umuC_Y11A are efficiently removed from DNA in a repair pathway initiated by RNase HII. Using the same approach, we show here that mismatch repair and base excision repair play minimal back-up roles in RER in vivo. In contrast, in the absence of functional RNase HII, umuC_Y11A-dependent mutagenesis increases significantly in ΔuvrA, uvrB5 and ΔuvrC strains, suggesting that rNMPs misincorporated into DNA are actively repaired by nucleotide excision repair (NER) in vivo. Participation of NER in RER was confirmed by reconstituting ribonucleotide-dependent NER in vitro. We show that UvrABC nuclease-catalyzed incisions are readily made on DNA templates containing one, two, or five rNMPs and that the reactions are stimulated by the presence of mispaired bases. Similar to NER of DNA lesions, excision of rNMPs proceeds through dual incisions made at the 8th phosphodiester bond 5′ and 4th-5th phosphodiester bonds 3′ of the ribonucleotide. Ribonucleotides misinserted into DNA can therefore be added to the broad list of helix-distorting modifications that are substrates for NER