Spike Firing and IPSPs in Layer V Pyramidal Neurons during Beta Oscillations in Rat Primary Motor Cortex (M1) In Vitro

Beta frequency oscillations (10–35 Hz) in motor regions of cerebral cortex play an important role in stabilising and suppressing unwanted movements, and become intensified during the pathological akinesia of Parkinson''s Disease. We have used a cortical slice preparation of rat brain, combined with concurrent intracellular and field recordings from the primary motor cortex (M1), to explore the cellular basis of the persistent beta frequency (27–30 Hz) oscillations manifest in local field potentials (LFP) in layers II and V of M1 produced by continuous perfusion of kainic acid (100 nM) and carbachol (5 µM). Spontaneous depolarizing GABA-ergic IPSPs in layer V cells, intracellularly dialyzed with KCl and IEM1460 (to block glutamatergic EPSCs), were recorded at −80 mV. IPSPs showed a highly significant (P< 0.01) beta frequency component, which was highly significantly coherent with both the Layer II and V LFP oscillation (which were in antiphase to each other). Both IPSPs and the LFP beta oscillations were abolished by the GABA_A antagonist bicuculline. Layer V cells at rest fired spontaneous action potentials at sub-beta frequencies (mean of 7.1+1.2 Hz; n = 27) which were phase-locked to the layer V LFP beta oscillation, preceding the peak of the LFP beta oscillation by some 20 ms. We propose that M1 beta oscillations, in common with other oscillations in other brain regions, can arise from synchronous hyperpolarization of pyramidal cells driven by synaptic inputs from a GABA-ergic interneuronal network (or networks) entrained by recurrent excitation derived from pyramidal cells. This mechanism plays an important role in both the physiology and pathophysiology of control of voluntary movement generation.     

Uptake of Genital Mucosal Sampling in HVTN 097, a Phase 1b HIV Vaccine Trial in South Africa

Because sexual transmission of HIV occurs across mucosal membranes, understanding the immune responses of the genital mucosa to vaccines may contribute knowledge to finding an effective candidate HIV vaccine. We describe the uptake of rectal secretion, cervical secretion and seminal mucosal secretion sampling amongst volunteers in a Phase 1b HIV vaccine trial. Age at screening, gender, study site and the designation of the person conducting the informed consent procedure were collected for volunteers who screened for the HVTN 097 study. A total of 211 volunteers (54% female) were screened at three sites in South Africa: Soweto (n = 70, 33%), Cape Town (n = 68, 32%) and Klerksdorp (n = 73, 35%). Overall uptake of optional mucosal sampling amongst trial volunteers was 71% (n = 149). Compared to Cape Town, volunteers from Soweto and Klerksdorp were less likely to consent to sampling (Soweto OR 0.08 CI: 0.03–0.25 p<0.001 and Klerksdorp OR 0.13 CI: 0.04–0.41 p = 0.001). In contrast, volunteers over 25 years of age were 2.39 times more likely to consent than younger volunteers (CI: 1.13–5.08, p = 0.02). Further studies are required to better understand the cultural, demographic and sociobehavioral factors which influence willingness to participate in mucosal sampling in HIV prevention studies.

Trial Registration

ClinicalTrials.gov: {"type":"clinical-trial","attrs":{"text":"NCT02109354","term_id":"NCT02109354"}}NCT02109354     

Endothelial Function and Serum Concentration of Toxic Metals in Frequent Consumers of Fish

Background

Endothelial dysfunction is involved in the pathogenesis of atherosclerosis. Consumption of fish is associated with reduced cardiovascular risk, but there is paucity of data concerning its effect on endothelial function. Furthermore, investigation of the effects of fish consumption on health must take into account the ingestion of contaminants, including transition metals and some metalloids, which may have unfavorable effects on health, including those on the cardiovascular system. We investigated the association between fish consumption, endothelial function (flow mediated dilation of the brachial artery), and serum concentration of some toxic metals in apparently healthy people.

Methods

Twenty-nine high fish consumers (at least 3 portions a week) were compared with 25 low fish consumers (less than 1 portion a week). All participants were free of diabetes, cardiovascular or other systemic diseases. Serum metal (antimonium, arsenic, mercury, lead, cobalt, copper, zinc, selenium, strontium) concentrations were measured in subgroups of 24 high fish consumers and 19 low fish consumers.

Results

Both groups exhibited similar habitual dietary patterns, age and anthropometric characteristics. The high fish consumers had higher flow mediated dilation (9.7±1.8 vs. 7.3±1.9%; P<0.001), but also higher serum concentrations of mercury (5.87±2.69 vs. 1.65±1.10 mcg/L; P<0.001) and arsenic (6.04±3.25 vs. 2.30±1.58 mcg/L; P<0.001). The fasting plasma glucose concentrations were significantly correlated with both mercury (r = 0.39; P = 0.01) and arsenic concentrations (r = 0.55; P<0.001).

Conclusions

Habitual consumption of high amounts of fish is associated with better endothelial function despite higher serum concentrations of mercury and arsenic.     

Glutathione-S-Transferase: A Minor Allergen in Birch Pollen due to Limited Release from Hydrated Pollen

Background

Recently, a protein homologous to glutathione-S-transferases (GST) was detected in prominent amounts in birch pollen by proteomic profiling. As members of the GST family are relevant allergens in mites, cockroach and fungi we investigated the allergenic relevance of GST from birch (bGST).

Methodology

bGST was expressed in Escherichia coli, purified and characterized by mass spectrometry. Sera from 217 birch pollen-allergic patients were tested for IgE-reactivity to bGST by ELISA. The mediator-releasing activity of bGST was analysed with IgE-loaded rat basophil leukaemia cells (RBL) expressing human FcεRI. BALB/c mice were immunized with bGST or Bet v 1. Antibody and T cell responses to either protein were assessed. IgE-cross-reactivity between bGST with GST from house dust mite, Der p 8, was studied with murine and human sera in ELISA. The release kinetics of bGST and Bet v 1 from birch pollen were assessed in water, simulated lung fluid, 0.9% NaCl and PBS. Eluted proteins were quantified by ELISA and analysed by immunoblotting.

Principle findings

Only 13% of 217 birch pollen-allergic patients showed IgE-reactivity to bGST. In RBL assays bGST induced mediator release. Immunization of mice with bGST induced specific IgE and a Th2-dominated cellular immune response comparably to immunization with Bet v 1. bGST did not cross-react with Der p 8. In contrast to Bet v 1, only low amounts of bGST were released from pollen grains upon incubation in water and the different physiological solutions.

Conclusion/Significance

Although bGST is abundant in birch pollen, immunogenic in mice and able to induce mediator release from effector cells passively loaded with specific IgE, it is a minor allergen for birch pollen-allergic patients. We refer this discrepancy to its limited release from hydrated pollen. Hence, bGST is an example demonstrating that allergenicity depends mainly on rapid elution from airborne particles.     

The endemic kelp Lessonia corrugata is being pushed above its thermal limits in an ocean warming hotspot

Kelps are in global decline due to climate change, which includes ocean warming. To identify vulnerable species, we need to identify their tolerances to increasing temperatures and determine whether tolerances are altered by co-occurring drivers such as inorganic nutrient levels. This is particularly important for those species with restricted distributions, which may already be experiencing thermal stress. To identify thermal tolerance of the range-restricted kelp Lessonia corrugata, we conducted a laboratory experiment on juvenile sporophytes to measure performance (growth, photosynthesis) across its thermal range (4–22°C). We determined the upper thermal limit for growth and photosynthesis to be ~22–23°C, with a thermal optimum of ~16°C. To determine if elevated inorganic nitrogen availability could enhance thermal tolerance, we compared the performance of juveniles under low (4.5 μmol · d⁻¹) and high (90 μmol · d⁻¹) nitrate conditions at and above the thermal optimum (16–23.5°C). Nitrate enrichment did not enhance thermal performance at temperatures above the optimum but did lead to elevated growth rates at the thermal optimum. Our results indicate L. corrugata is likely to be extremely susceptible to moderate ocean warming and marine heatwaves. Peak sea surface temperatures during summer in eastern and northeastern Tasmania can reach up to 20–21°C, and climate projections suggest that L. corrugata's thermal limit will be regularly exceeded by 2050 as southeastern Australia is a global ocean-warming hotspot. By identifying the upper thermal limit of L. corrugata, we have taken a critical step in predicting the future of the species in a warming climate.     

Determinism of nonadditive litter mixture effect on decomposition: Role of the moisture content of litters

The mechanisms behind the plant litter mixture effect on decomposition are still difficult to disentangle. To tackle this issue, we used a model that specifically addresses the role of the litter moisture content. Our model predicts that when two litters interact in terms of water flow, the difference of evaporation rate between two litters can trigger a nonadditive mixture effect on decomposition. Water flows from the wettest to the driest litter, changing the reaction rates without changing the overall litter water content. The reaction rate of the litter receiving the water increases relatively more than the decrease in the reaction rate of the litter supplying the water, leading to a synergistic effect. Such water flow can keep the microbial biomass of both litter in a water content domain suitable to maintain decomposition activity. When applied to experimental data (Sphagnum rubellum and Molinia caerulea litters), the model is able to assess whether any nonadditive effect originates from water content variation alone or whether other factors have to be taken into account.     

Crystallographic analysis reveals a novel second binding site for trimethoprim in active site double mutants of human dihydrofolate reductase

In order to produce a more potent replacement for trimethoprim (TMP) used as a therapy for Pneumocystis pneumonia and targets dihydrofolate reductase from Pneumocystis jirovecii (pjDHFR), it is necessary to understand the determinants of potency and selectivity against DHFR from the mammalian host and fungal pathogen cells. To this end, active site residues in human (h) DHFR were replaced with those from pjDHFR. Structural data are reported for two complexes of TMP with the double mutants Gln35Ser/Asn64Phe (Q35S/N64F) and Gln35Lys/Asn64Phe (Q35K/N64F) of hDHFR that unexpectedly show evidence for the binding of two molecules of TMP: one molecule that binds in the normal folate binding site and the second molecule that binds in a novel subpocket site such that the mutated residue Phe64 is involved in van der Waals contacts to the trimethoxyphenyl ring of the second TMP molecule. Kinetic data for the binding of TMP to hDHFR and pjDHFR reveal an 84-fold selectivity of TMP against pjDHFR (K_i 49 nM) compared to hDHFR (K_i 4093 nM). Two mutants that contain one substitution from pj- and one from the closely related Pneumocystis carinii DHFR (pcDHFR) (Q35K/N64F and Q35S/N64F) show K_i values of 593 and 617 nM, respectively; these K_i values are well above both the K_i for pjDHFR and are similar to pcDHFR (Q35K/N64F and Q35S/N64F) (305 nM). These results suggest that active site residues 35 and 64 play key roles in determining selectivity for pneumocystis DHFR, but that other residues contribute to the unique binding of inhibitors to these enzymes.     

Characterizing genetic risk at known prostate cancer susceptibility loci in African Americans

GWAS of prostate cancer have been remarkably successful in revealing common genetic variants and novel biological pathways that are linked with its etiology. A more complete understanding of inherited susceptibility to prostate cancer in the general population will come from continuing such discovery efforts and from testing known risk alleles in diverse racial and ethnic groups. In this large study of prostate cancer in African American men (3,425 prostate cancer cases and 3,290 controls), we tested 49 risk variants located in 28 genomic regions identified through GWAS in men of European and Asian descent, and we replicated associations (at p≤0.05) with roughly half of these markers. Through fine-mapping, we identified nearby markers in many regions that better define associations in African Americans. At 8q24, we found 9 variants (p≤6×10⁻⁴) that best capture risk of prostate cancer in African Americans, many of which are more common in men of African than European descent. The markers found to be associated with risk at each locus improved risk modeling in African Americans (per allele OR = 1.17) over the alleles reported in the original GWAS (OR = 1.08). In summary, in this detailed analysis of the prostate cancer risk loci reported from GWAS, we have validated and improved upon markers of risk in some regions that better define the association with prostate cancer in African Americans. Our findings with variants at 8q24 also reinforce the importance of this region as a major risk locus for prostate cancer in men of African ancestry.