Synthesis of photo-crosslinking probes and their application for the site-selective chemical modification of the 5-HT3 receptor

The 5-HT3 receptor (5-HT3R) is an important ion channel responsible for the transmission of nerve impulses in the CNS and PNS that is activated by the endogenous agonist serotonin (5-hydroxytryptamine, 5-HT). 5-HT3R is the only serotonin receptor belonging to the Cys-loop superfamily of neurotransmitter receptors. Different structural biology approaches can be applied, such as crystallization and x-ray analysis. Nonetheless, characterizing the exact ligand binding site(s) of these dynamic receptors is still challenging. The use of photo-crosslinking probes is an alternative validated approach allowing identification of regions in the protein that are important for the binding of small molecules. We designed our probes based on the core structure of the 5-HT3R antagonist granisetron, a FDA approved drug used for the treatment of chemotherapy-induced nausea and vomiting. We synthesized a small library of photo-crosslinking probes by conjugating diazirines and benzophenones via various linkers to granisetron. We were able to obtain several compounds with diverse linker lengths and different photo-crosslinking moieties that show nanomolar binding affinity for the orthosteric binding site. Furthermore we established a stable h5-HT3R expressing cell line and a purification protocol to yield the receptor in a high purity. Several experiments showed unambiguously that we are able to photo-crosslink our probes with the receptor site-specifically. The functionalised protein was analysed by Western blot and MS-analysis. This yielded the exact covalent modification site, corroborating current ligand binding models derived from mutagenesis and docking studies.

Bedload composition, transport and modification in rivers of Westland, New Zealand, with implications for the distribution of alluvial pounamu (jade)

River bedload surveyed at 50 sites in Westland is dominated by Alpine Schist or Torlesse Greywacke from the Alpine Fault hanging wall, with subordinate Pounamu Ultramafics or footwall-derived Western Province rocks. Tumbling experiments found ultramafics to have the lowest attrition rates, compared with greywacke sandstone and granite (which abrade to produce silt to medium-sand), or incompetent schist (which fragments). Arahura has greater total concentrations (103–105 t/km2) and proportions (5–40%) of ultramafic bedload compared with Hokitika and Taramakau catchments (101–104 t/km2, mostly <10%), matching relative areas of mapped Pounamu Ultramafic bedrock, but enriched relative to absolute areal proportions. Western Province rocks downthrown by the Alpine Fault are under-represented in the bedload. Enriched concentrations of ultramafic bedload decrease rapidly with distance downstream from source rock outcrops, changing near prominent ice-limit moraines. Bedload evolution with transport involves both downstream fining and dilution from tributaries, in a sediment supply regime more strongly influenced by tectonics and the imprint of past glaciation. Treasured New Zealand pounamu (jade) is associated with ultramafic rocks. Chances of discovery vary between catchments, are increased near glacial moraines, and are highest near source-rock outcrops in remote mountain headwaters.

Impact of Acquired Pellicle Modification on Adhesion of Early Colonizers

New preventive approaches against dental erosion caused by acidic drinks and beverages include fortification of beverages with natural polymers. We have shown that the mixture of casein and mucin significantly improved the erosion-inhibiting properties of the human pellicle layer. This study aimed to investigate the effect of pellicle modification by casein, mucin and a casein-mucin mixture on the adhesion of early bacterial colonizers. Test specimens of human tooth enamel were prepared, covered with saliva and coated with 0.5% aqueous (aq.) casein, 0.27% aq. mucin or with 0.5% aq. casein-0.27% aq. mucin, after which the adhesion of Streptococcus gordonii, Streptococcus oralis, and Actinomyces odontolyticus was measured after incubation for 30 min and 2 h. log10 colony-forming units were compared by nonparametric tests. All three bacterial strains adhered in higher number to pellicle-coated enamel than to native enamel. The protein modifications of pellicle all decreased the counts of adhering bacteria up to 0.34 log10/mm2, the most efficient being the casein-mucin mixture. In addition to the recently shown erosion-reducing effect by casein-mucin, modification of the pellicle may inhibit bacterial adherence compared to untreated human pellicle.

Internet-based interpretation bias modification for social anxiety: A pilot study

BACKGROUND AND OBJECTIVES: The biased interpretation of ambiguous social situations is considered a maintaining factor of Social Anxiety Disorder (SAD). Studies on the modification of interpretation bias have shown promising results in laboratory settings. The present study aims at pilot-testing an Internet-based training that targets interpretation and judgmental bias. METHOD: Thirty-nine individuals meeting diagnostic criteria for SAD participated in an 8-week, unguided program. Participants were presented with ambiguous social situations, were asked to choose between neutral, positive, and negative interpretations, and were required to evaluate costs of potential negative outcomes. Participants received elaborate automated feedback on their interpretations and judgments. RESULTS: There was a pre-to-post-reduction of the targeted cognitive processing biases (d = 0.57-0.77) and of social anxiety symptoms (d = 0.87). Furthermore, results showed changes in depression and general psychopathology (d = 0.47-0.75). Decreases in cognitive biases and symptom changes did not correlate. The results held stable accounting for drop-outs (26%) and over a 6-week follow-up period. Forty-five percent of the completer sample showed clinical significant change and almost half of the participants (48%) no longer met diagnostic criteria for SAD. LIMITATIONS: As the study lacks a control group, results lend only preliminary support to the efficacy of the intervention. Furthermore, the mechanism of change remained unclear. CONCLUSION: First results promise a beneficial effect of the program for SAD patients. The treatment proved to be feasible and acceptable. Future research should evaluate the intervention in a randomized-controlled setting.

Modification of aviary design reduces incidence of falls, collisions and keel bone damage in laying hens

Non-cage housing systems for laying hens such as aviaries provide greater freedom to perform species-specific behavior and thus are thought to improve welfare of the birds; however, aviaries are associated with a high prevalence of keel bone damage (fractures and deviations), which is a major welfare problem in commercial laying hens. Potential causes of keel bone damage are falls and collisions with internal housing structures that occur as birds move between tiers or perches in the aviary. The aim of this study was to investigate the scope for reducing keel bone damage by reducing falls and collisions through modifications of aviary design. Birds were kept in 20 pens in a laying hen house (225 hens per pen) that were assigned to four different treatments (n = 5 pens per treatment group) including (1) control pens and pens modified by the addition of (2) perches, (3) platforms and (4) ramps. Video recordings at 19, 22, 29, 36 and 43 weeks of age were used to analyze controlled movements and falls (including details on occurrence of collision, cause of fall, height of fall and behavior after fall) during the transitional dusk and subsequent dark phase. Palpation assessments (focusing on fractures and deviations) using 20 focal hens per pen were conducted at 18, 20, 23, 30, 37, 44, 52 and 60 weeks of age. In comparison to the control group, we found 44% more controlled movements in the ramp (P = 0.003) and 47% more controlled movements in the platform treatments (P = 0.014) as well as 45% fewer falls (P = 0.006) and 59% fewer collisions (P < 0.001) in the ramp treatment. There were no significant differences between the control and perch treatments. Also, at 60 weeks of age, 23% fewer fractured keel bones were found in the ramp compared with the control treatment (P = 0.0053). After slaughter at 66 weeks of age, no difference in keel bone damage was found between treatment groups and the prevalence of fractures increased to an average of 86%. As a potential mechanism to explain the differences in locomotion, we suggest that ramps facilitated movement in the vertical plane by providing a continuous path between the tiers and thus supported more natural behavior (i.e. walking and running) of the birds. As a consequence of reducing events that potentially damage keel bones, the installation of ramps may have reduced the prevalence of keel fractures for a major portion of the flock cycle. We conclude that aviary design and installation of specific internal housing structures (i.e. ramps and platforms) have considerable potential to reduce keel bone damage of laying hens in aviary systems.