Sustained pain-related depression of behavior: effects of intraplantar formalin and complete freund’s adjuvant on intracranial self-stimulation (ICSS) and endogenous kappa opioid biomarkers in rats

Background: Intraplantar administration of complete Freund's adjuvant (CFA) and formalin are two noxious stimuli commonly used to produce sustained pain-related behaviors in rodents for research on neurobiology and treatment of pain. One clinically relevant manifestation of pain is depression of behavior and mood. This study compared effects of intraplantar CFA and formalin on depression of positively reinforced operant behavior in an assay of intracranial self-stimulation (ICSS) in rats. Effects of CFA and formalin on other physiological and behavioral measures, and opioid effects on formalin-induced depression of ICSS, were also examined. Results: There were four main findings. First, consistent with previous studies, both CFA and formalin produced similar paw swelling and mechanical hypersensitivity. Second, CFA produced weak and transient depression of ICSS, whereas formalin produced a more robust and sustained depression of ICSS that lasted at least 14 days. Third, formalin-induced depression of ICSS was reversed by morphine doses that did not significantly alter ICSS in saline-treated rats, suggesting that formalin effects on ICSS can be interpreted as an example of pain-related and analgesic-reversible depression of behavior. Finally, formalin-induced depression of ICSS was not associated with changes in central biomarkers for activation of endogenous kappa opioid systems, which have been implicated in depressive-like states in rodents, nor was it blocked by the kappa antagonist norbinaltorphimine. Conclusions: These results suggest differential efficacy of sustained pain stimuli to depress brain reward function in rats as assessed with ICSS. Formalin-induced depression of ICSS does not appear to engage brain kappa opioid systems. Electronic supplementary material The online version of this article (doi:10.1186/1744-8069-10-62) contains supplementary material, which is available to authorized users

Leaf-litter leachate is distinct in optical properties and bioavailability to stream heterotrophs

Dissolved organic C (DOC) leached from leaf litter contributes to the C pool of stream ecosystems and affects C cycling in streams. We studied how differences in leaf-litter chemistry affect the optical properties and decomposition of DOC. We used 2 species of cottonwoods (Populus) and their naturally occurring hybrids that differ in leaf-litter phytochemistry and decomposition rate. We measured DOC and nutrient concentration in leaf leachates and determined the effect of DOC quality on heterotrophic respiration in 24-h incubations with stream sediments. Differences in DOC composition and quality were characterized with fluorescence spectroscopy. Rapidly decomposing leaves with lower tannin and lignin concentrations leached ~40 to 50% more DOC and total dissolved N than did slowly decomposing leaves. Rates of heterotrophic respiration were 25 to 50% higher on leachate from rapidly decomposing leaf types. Rates of heterotrophic respiration were related to metrics of aromaticity. Specifically, rates of respiration were correlated negatively with the Fluorescence Index and positively with Specific Ultraviolet Absorbance (SUVA254) and T280 tryptophan-like fluorescence peak. These results reveal that leaf-litter DOC is distinctly different from ambient streamwater DOC. The relationships between optical characteristics of leaf leachate and bioavailability are opposite those found in streamwater DOC. Differences in phytochemistry among leaf types can influence stream ecosystems with respect to DOC quantity, composition, and rates of stream respiration. These patterns suggest that the relationship between the chemical structure of DOC and its biogeochemistry is more complex than previously recognized. These unique properties of leaf-litter DOC will be important when assessing the effects of terrestrial C on aquatic ecosystems, especially during leaf fall

Plug-and-play genetic access to drosophila cell types using exchangeable exon cassettes.

Genetically encoded effectors are important tools for probing cellular function in living animals, but improved methods for directing their expression to specific cell types are required. Here, we introduce a simple, versatile method for achieving cell-type-specific expression of transgenes that leverages the untapped potential of "coding introns" (i.e., introns between coding exons). Our method couples the expression of a transgene to that of a native gene expressed in the cells of interest using intronically inserted "plug-and-play" cassettes (called "Trojan exons") that carry a splice acceptor site followed by the coding sequences of T2A peptide and an effector transgene. We demonstrate the efficacy of this approach in Drosophila using lines containing suitable MiMIC (Minos-mediated integration cassette) transposons and a palette of Trojan exons capable of expressing a range of commonly used transcription factors. We also introduce an exchangeable, MiMIC-like Trojan exon construct that can be targeted to coding introns using the Crispr/Cas system.This work was supported by the Intramural Research Program of the National Institute of Mental Health (B.H.W.) and by grants from the Whitehall Foundation (C.J.P.), NIH (R01DC013070, C.J.P.), the Wellcome Trust (H.I. and M.L.), and the Sir Isaac Newton Trust, Cambridge (M.L.). J.E. was supported by FONDECYT #1141278 and the CINV, which is supported by the Millennium Scientific Initiative of the Ministerio de Economía, Fomento y Turismo. We thank the Bellen laboratory and the Drosophila Gene Disruption Project at Baylor College of Medicine, the Bloomington Stock Center (NIH P40OD018537), and Julie Simpson for fly lines. Thanks also to Aaron DiAntonio, Aaron Hsueh, and John Reinitz for antibodies and the NINDS Sequencing Core Facility for DNA sequencing. Finally, thanks to Sarah Naylor for technical help and Grace Gray, Herman Dierick, Koen Venken, and Hugo Bellen for comments on the manuscript and productive discussions.This is the final published version. It first appeared at http://www.ncbi.nlm.nih.gov/pubmed/25732830

Pharmacodynamic Assays to Facilitate Preclinical and Clinical Development of Pre-mRNA Splicing Modulatory Drug Candidates

The spliceosome has recently emerged as a new target for cancer chemotherapy and novel antitumor spliceosome targeted agents are under development. Here, we describe two types of novel pharmacodynamic assays that facilitate drug discovery and development of this intriguing class of innovative therapeutics; the first assay is useful for preclinical optimization of small-molecule agents that target the SF3B1 spliceosomal protein in animals, the second assay is an ex vivo validated, gel-based assay for the measurement of drug exposure in human leukocytes. The first assay utilizes a highly specific bioluminescent splicing reporter, based on the skipping of exons 4-11 of a Luc-MDM2 construct, which specifically yields active luciferase when treated with small-molecule spliceosome modulators. We demonstrate that this reporter can be used to monitor alternative splicing in whole cells in vitro. We describe here that cell lines carrying the reporter can be used in vivo for the efficient pharmacodynamic analysis of agents during drug optimization and development. We also demonstrate dose- and time-dependent on-target activity of sudemycin D6 (SD6), which leads to dramatic tumor regression. The second assay relies on the treatment of freshly drawn human blood with SD6 ex vivo treatment. Changes in alternative splicing are determined by RT-PCR using genes previously identified in in vitro experiments. The Luc-MDM2 alternative splicing bioluminescent reporter and the splicing changes observed in human leukocytes should allow for the more facile translation of novel splicing modulators into clinical application

Spectroscopy of Broad Line Blazars from 1LAC

We report on optical spectroscopy of 165 Flat Spectrum Radio Quasars (FSRQs) in the Fermi 1LAC sample, which have helped allow a nearly complete study of this population. Fermi FSRQ show significant evidence for non-thermal emission even in the optical; the degree depends on the gamma-ray hardness. They also have smaller virial estimates of hole mass than the optical quasar sample. This appears to be largely due to a preferred (axial) view of the gamma-ray FSRQ and non-isotropic (H/R ~ 0.4) distribution of broad-line velocities. Even after correction for this bias, the Fermi FSRQ show higher mean Eddington ratios than the optical population. A comparison of optical spectral properties with Owens Valley Radio Observatory radio flare activity shows no strong correlation.Comment: Accepted for publication in Ap

Spectroscopy of the Largest Ever γ-Ray-selected BL Lac Sample

We report on spectroscopic observations covering most of the 475 BL Lacs in the second Fermi Large Area Telescope (LAT) catalog of active galactic nuclei (AGNs). Including archival measurements (correcting several erroneous literature values) we now have spectroscopic redshifts for 44% of the BL Lacs. We establish firm lower redshift limits via intervening absorption systems and statistical lower limits via searches for host galaxies for an additional 51% of the sample leaving only 5% of the BL Lacs unconstrained. The new redshifts raise the median spectroscopic z from 0.23 to 0.33 and include redshifts as large as z = 2.471. Spectroscopic redshift minima from intervening absorbers have z = 0.70, showing a substantial fraction at large z and arguing against strong negative evolution. We find that detected BL Lac hosts are bright ellipticals with black hole masses M_• ~ 10^(8.5) – 10^9, substantially larger than the mean of optical AGNs and LAT Flat Spectrum Radio Quasar samples. A slow increase in M_• with z may be due to selection bias. We find that the power-law dominance of the optical spectrum extends to extreme values, but this does not strongly correlate with the γ-ray properties, suggesting that strong beaming is the primary cause of the range in continuum dominance

Colorectal cancer linkage on chromosomes 4q21, 8q13, 12q24, and 15q22

A substantial proportion of familial colorectal cancer (CRC) is not a consequence of known susceptibility loci, such as mismatch repair (MMR) genes, supporting the existence of additional loci. To identify novel CRC loci, we conducted a genome-wide linkage scan in 356 white families with no evidence of defective MMR (i.e., no loss of tumor expression of MMR proteins, no microsatellite instability (MSI)-high tumors, or no evidence of linkage to MMR genes). Families were ascertained via the Colon Cancer Family Registry multi-site NCI-supported consortium (Colon CFR), the City of Hope Comprehensive Cancer Center, and Memorial University of Newfoundland. A total of 1,612 individuals (average 5.0 per family including 2.2 affected) were genotyped using genome-wide single nucleotide polymorphism linkage arrays; parametric and non-parametric linkage analysis used MERLIN in a priori-defined family groups. Five lod scores greater than 3.0 were observed assuming heterogeneity. The greatest were among families with mean age of diagnosis less than 50 years at 4q21.1 (dominant HLOD = 4.51, α = 0.84, 145.40 cM, rs10518142) and among all families at 12q24.32 (dominant HLOD = 3.60, α = 0.48, 285.15 cM, rs952093). Among families with four or more affected individuals and among clinic-based families, a common peak was observed at 15q22.31 (101.40 cM, rs1477798; dominant HLOD = 3.07, α = 0.29; dominant HLOD = 3.03, α = 0.32, respectively). Analysis of families with only two affected individuals yielded a peak at 8q13.2 (recessive HLOD = 3.02, α = 0.51, 132.52 cM, rs1319036). These previously unreported linkage peaks demonstrate the continued utility of family-based data in complex traits and suggest that new CRC risk alleles remain to be elucidated. © 2012 Cicek et al