NMDA Receptor Antagonist Ketamine Impairs Feature Integration in Visual Perception

Recurrent interactions between neurons in the visual cortex are crucial for the integration of image elements into coherent objects, such as in figure-ground segregation of textured images. Blocking N-methyl-D-aspartate (NMDA) receptors in monkeys can abolish neural signals related to figure-ground segregation and feature integration. However, it is unknown whether this also affects perceptual integration itself. Therefore, we tested whether ketamine, a non-competitive NMDA receptor antagonist, reduces feature integration in humans. We administered a subanesthetic dose of ketamine to healthy subjects who performed a texture discrimination task in a placebo-controlled double blind within-subject design. We found that ketamine significantly impaired performance on the texture discrimination task compared to the placebo condition, while performance on a control fixation task was much less impaired. This effect is not merely due to task difficulty or a difference in sedation levels. We are the first to show a behavioral effect on feature integration by manipulating the NMDA receptor in humans.     

Up in the Tree – The Overlooked Richness of Bryophytes and Lichens in Tree Crowns

Assessing diversity is among the major tasks in ecology and conservation science. In ecological and conservation studies, epiphytic cryptogams are usually sampled up to accessible heights in forests. Thus, their diversity, especially of canopy specialists, likely is underestimated. If the proportion of those species differs among forest types, plot-based diversity assessments are biased and may result in misleading conservation recommendations. We sampled bryophytes and lichens in 30 forest plots of 20 m × 20 m in three German regions, considering all substrates, and including epiphytic litter fall. First, the sampling of epiphytic species was restricted to the lower 2 m of trees and shrubs. Then, on one representative tree per plot, we additionally recorded epiphytic species in the crown, using tree climbing techniques. Per tree, on average 54% of lichen and 20% of bryophyte species were overlooked if the crown was not been included. After sampling all substrates per plot, including the bark of all shrubs and trees, still 38% of the lichen and 4% of the bryophyte species were overlooked if the tree crown of the sampled tree was not included. The number of overlooked lichen species varied strongly among regions. Furthermore, the number of overlooked bryophyte and lichen species per plot was higher in European beech than in coniferous stands and increased with increasing diameter at breast height of the sampled tree. Thus, our results indicate a bias of comparative studies which might have led to misleading conservation recommendations of plot-based diversity assessments.     

Uncovering Wolbachia Diversity upon Artificial Host Transfer

The common endosymbiotic Wolbachia bacteria influence arthropod hosts in multiple ways. They are mostly recognized for their manipulations of host reproduction, yet, more recent studies demonstrate that Wolbachia also impact host behavior, metabolic pathways and immunity. Besides their biological and evolutionary roles, Wolbachia are new potential biological control agents for pest and vector management. Importantly, Wolbachia-based control strategies require controlled symbiont transfer between host species and predictable outcomes of novel Wolbachia-host associations. Theoretically, this artificial horizontal transfer could inflict genetic changes within transferred Wolbachia populations. This could be facilitated through de novo mutations in the novel recipient host or changes of haplotype frequencies of polymorphic Wolbachia populations when transferred from donor to recipient hosts. Here we show that Wolbachia resident in the European cherry fruit fly, Rhagoletis cerasi, exhibit ancestral and cryptic sequence polymorphism in three symbiont genes, which are exposed upon microinjection into the new hosts Drosophila simulans and Ceratitis capitata. Our analyses of Wolbachia in microinjected D. simulans over 150 generations after microinjection uncovered infections with multiple Wolbachia strains in trans-infected lines that had previously been typed as single infections. This confirms the persistence of low-titer Wolbachia strains in microinjection experiments that had previously escaped standard detection techniques. Our study demonstrates that infections by multiple Wolbachia strains can shift in prevalence after artificial host transfer driven by either stochastic or selective processes. Trans-infection of Wolbachia can claim fitness costs in new hosts and we speculate that these costs may have driven the shifts of Wolbachia strains that we saw in our model system.     

A Phase II,Randomized, Double-Blind,Placebo Controlled,Dose-Response Trial of the Melatonin Effect on the Pain Threshold of Healthy Subjects

Background

Previous studies have suggested that melatonin may produce antinociception through peripheral and central mechanisms. Based on the preliminary encouraging results of studies of the effects of melatonin on pain modulation, the important question has been raised of whether there is a dose relationship in humans of melatonin on pain modulation.

Objective

The objective was to evaluate the analgesic dose response of the effects of melatonin on pressure and heat pain threshold and tolerance and the sedative effects.

Methods

Sixty-one healthy subjects aged 19 to 47 y were randomized into one of four groups: placebo, 0.05 mg/kg sublingual melatonin, 0.15 mg/kg sublingual melatonin or 0.25 mg/kg sublingual melatonin. We determine the pressure pain threshold (PPT) and the pressure pain tolerance (PPTo). Quantitative sensory testing (QST) was used to measure the heat pain threshold (HPT) and the heat pain tolerance (HPTo). Sedation was assessed with a visual analogue scale and bispectral analysis.

Results

Serum plasma melatonin levels were directly proportional to the melatonin doses given to each subject. We observed a significant effect associated with dose group. Post hoc analysis indicated significant differences between the placebo vs. the intermediate (0.15 mg/kg) and the highest (0.25 mg/kg) melatonin doses for all pain threshold and sedation level tests. A linear regression model indicated a significant association between the serum melatonin concentrations and changes in pain threshold and pain tolerance (R² = 0.492 for HPT, R² = 0.538 for PPT, R² = 0.558 for HPTo and R² = 0.584 for PPTo).

Conclusions

The present data indicate that sublingual melatonin exerts well-defined dose-dependent antinociceptive activity. There is a correlation between the plasma melatonin drug concentration and acute changes in the pain threshold. These results provide additional support for the investigation of melatonin as an analgesic agent. Brazilian Clinical Trials Registry (ReBec): (U1111-1123-5109). IRB: Research Ethics Committee at the Hospital de Clínicas de Porto Alegre.     

Flexibility within the Rotor and Stators of the Vacuolar H+-ATPase

The V-ATPase is a membrane-bound protein complex which pumps protons across the membrane to generate a large proton motive force through the coupling of an ATP-driven 3-stroke rotary motor (V₁) to a multistroke proton pump (V_o). This is done with near 100% efficiency, which is achieved in part by flexibility within the central rotor axle and stator connections, allowing the system to flex to minimise the free energy loss of conformational changes during catalysis. We have used electron microscopy to reveal distinctive bending along the V-ATPase complex, leading to angular displacement of the V₁ domain relative to the V_o domain to a maximum of ~30°. This has been complemented by elastic network normal mode analysis that shows both flexing and twisting with the compliance being located in the rotor axle, stator filaments, or both. This study provides direct evidence of flexibility within the V-ATPase and by implication in related rotary ATPases, a feature predicted to be important for regulation and their high energetic efficiencies.     

Knee Adduction Moment and Medial Contact Force – Facts about Their Correlation during Gait

The external knee adduction moment is considered a surrogate measure for the medial tibiofemoral contact force and is commonly used to quantify the load reducing effect of orthopedic interventions. However, only limited and controversial data exist about the correlation between adduction moment and medial force. The objective of this study was to examine whether the adduction moment is indeed a strong predictor for the medial force by determining their correlation during gait. Instrumented knee implants with telemetric data transmission were used to measure tibiofemoral contact forces in nine subjects. Gait analyses were performed simultaneously to the joint load measurements. Skeletal kinematics, as well as the ground reaction forces and inertial parameters, were used as inputs in an inverse dynamics approach to calculate the external knee adduction moment. Linear regression analysis was used to analyze the correlation between adduction moment and medial force for the whole stance phase and separately for the early and late stance phase. Whereas only moderate correlations between adduction moment and medial force were observed throughout the whole stance phase (R² = 0.56) and during the late stance phase (R² = 0.51), a high correlation was observed at the early stance phase (R² = 0.76). Furthermore, the adduction moment was highly correlated to the medial force ratio throughout the whole stance phase (R² = 0.75). These results suggest that the adduction moment is a surrogate measure, well-suited to predicting the medial force ratio throughout the whole stance phase or medial force during the early stance phase. However, particularly during the late stance phase, moderate correlations and high inter-individual variations revealed that the predictive value of the adduction moment is limited. Further analyses are necessary to examine whether a combination of other kinematic, kinetic or neuromuscular factors may lead to a more reliable prediction of the force magnitude.     

Hippocampal Feedforward Inhibition Focuses Excitatory Synaptic Signals into Distinct Dendritic Compartments

Feedforward inhibition controls the time window for synaptic integration and ensures temporal precision in cortical circuits. There is little information whether feedforward inhibition affects neurons uniformly, or whether it contributes to computational refinement within the dendritic tree. Here we demonstrate that feedforward inhibition crucially shapes the integration of synaptic signals in pyramidal cell dendrites. Using voltage-sensitive dye imaging we studied the transmembrane voltage patterns in CA1 pyramidal neurons after Schaffer collateral stimulation in acute brain slices from mice. We observed a high degree of variability in the excitation-inhibition ratio between different branches of the dendritic tree. Many dendritic segments showed no depolarizing signal at all, especially the basal dendrites that received predominantly inhibitory signals. Application of the GABA_A receptor antagonist bicuculline resulted in the spread of depolarizing signals throughout the dendritic tree. Tetanic stimulation of Schaffer collateral inputs induced significant alterations in the patterns of excitation/inhibition, indicating that they are modified by synaptic plasticity. In summary, we show that feedforward inhibition restricts the occurrence of depolarizing signals within the dendritic tree of CA1 pyramidal neurons and thus refines signal integration spatially.     

Direct Imaging of ER Calcium with Targeted-Esterase Induced Dye Loading (TED)

Visualization of calcium dynamics is important to understand the role of calcium in cell physiology. To examine calcium dynamics, synthetic fluorescent Ca²⁺ indictors have become popular. Here we demonstrate TED (= targeted-esterase induced dye loading), a method to improve the release of Ca²⁺ indicator dyes in the ER lumen of different cell types. To date, TED was used in cell lines, glial cells, and neurons in vitro. TED bases on efficient, recombinant targeting of a high carboxylesterase activity to the ER lumen using vector-constructs that express Carboxylesterases (CES). The latest TED vectors contain a core element of CES2 fused to a red fluorescent protein, thus enabling simultaneous two-color imaging. The dynamics of free calcium in the ER are imaged in one color, while the corresponding ER structure appears in red. At the beginning of the procedure, cells are transduced with a lentivirus. Subsequently, the infected cells are seeded on coverslips to finally enable live cell imaging. Then, living cells are incubated with the acetoxymethyl ester (AM-ester) form of low-affinity Ca²⁺ indicators, for instance Fluo5N-AM, Mag-Fluo4-AM, or Mag-Fura2-AM. The esterase activity in the ER cleaves off hydrophobic side chains from the AM form of the Ca²⁺ indicator and a hydrophilic fluorescent dye/Ca²⁺ complex is formed and trapped in the ER lumen. After dye loading, the cells are analyzed at an inverted confocal laser scanning microscope. Cells are continuously perfused with Ringer-like solutions and the ER calcium dynamics are directly visualized by time-lapse imaging. Calcium release from the ER is identified by a decrease in fluorescence intensity in regions of interest, whereas the refilling of the ER calcium store produces an increase in fluorescence intensity. Finally, the change in fluorescent intensity over time is determined by calculation of ΔF/F₀.     

40S Ribosome Biogenesis Co-Factors Are Essential for Gametophyte and Embryo Development

Ribosome biogenesis is well described in Saccharomyces cerevisiae. In contrast only very little information is available on this pathway in plants. This study presents the characterization of five putative protein co-factors of ribosome biogenesis in Arabidopsis thaliana, namely Rrp5, Pwp2, Nob1, Enp1 and Noc4. The characterization of the proteins in respect to localization, enzymatic activity and association with pre-ribosomal complexes is shown. Additionally, analyses of T-DNA insertion mutants aimed to reveal an involvement of the plant co-factors in ribosome biogenesis. The investigated proteins localize mainly to the nucleolus or the nucleus, and atEnp1 and atNob1 co-migrate with 40S pre-ribosomal complexes. The analysis of T-DNA insertion lines revealed that all proteins are essential in Arabidopsis thaliana and mutant plants show alterations of rRNA intermediate abundance already in the heterozygous state. The most significant alteration was observed in the NOB1 T-DNA insertion line where the P-A3 fragment, a 23S-like rRNA precursor, accumulated. The transmission of the T-DNA through the male and female gametophyte was strongly inhibited indicating a high importance of ribosome co-factor genes in the haploid stages of plant development. Additionally impaired embryogenesis was observed in some mutant plant lines. All results support an involvement of the analyzed proteins in ribosome biogenesis but differences in rRNA processing, gametophyte and embryo development suggested an alternative regulation in plants.     

The Landscape of Candidate Driver Genes Differs between Male and Female Breast Cancer

The rapidly growing collection of diverse genome-scale data from multiple tumor types sheds light on various aspects of the underlying tumor biology. With the objective to identify genes of importance for breast tumorigenesis in men and to enable comparisons with genes important for breast cancer development in women, we applied the computational framework COpy Number and EXpression In Cancer (CONEXIC) to detect candidate driver genes among all altered passenger genes. Unique to this approach is that each driver gene is associated with several gene modules that are believed to be altered by the driver. Thirty candidate drivers were found in the male breast cancers and 67 in the female breast cancers. We identified many known drivers of breast cancer and other types of cancer, in the female dataset (e.g. GATA3, CCNE1, GRB7, CDK4). In contrast, only three known cancer genes were found among male breast cancers; MAP2K4, LHP, and ZNF217. Many of the candidate drivers identified are known to be involved in processes associated with tumorigenesis, including proliferation, invasion and differentiation. One of the modules identified in male breast cancer was regulated by THY1, a gene involved in invasion and related to epithelial-mesenchymal transition. Furthermore, men with THY1 positive breast cancers had significantly inferior survival. THY1 may thus be a promising novel prognostic marker for male breast cancer. Another module identified among male breast cancers, regulated by SPAG5, was closely associated with proliferation. Our data indicate that male and female breast cancers display highly different landscapes of candidate driver genes, as only a few genes were found in common between the two. Consequently, the pathobiology of male breast cancer may differ from that of female breast cancer and can be associated with differences in prognosis; men diagnosed with breast cancer may consequently require different management and treatment strategies than women.