bluuebunny/crossref_metadata_2025_split huggingface dataset:
Thorough GABAergic innervation of the entire axon initial segment revealed by an optogenetic ‘laserspritzer’
Authors: Wang Xinjun , Hooks Bryan M. , Sun Qian‐Quan 
Published: August 2014

Key points
The laserspritzer we developed activates axo‐axonic synapses (AASs) with spatial resolutions of less than 40 μm.
AASs innervate the entire length of the axon initial segment (AIS), as opposed to forming a highly concentrated cartridge.
AAS‐mediated synaptic potentials are inhibitory, with reversal potentials similar to those of basket synapses.
AAS inhibition impedes action potentials and epileptiform activity more robustly than perisomatic inhibitions.
AAS activation alone is sufficient to inhibit action potential generation and epileptiform activities in vitro.
AbstractGABAergic terminals of chandelier cells exclusively innervate the axon initial segment (AIS) of excitatory neurons. Although the anatomy of these synapses has been well‐studied in several brain areas, relatively little is known about their physiological properties. Using vesicular γ‐aminobutyric acid transporter–channelrhodopsin 2–enhanced yellow fluorescence protein (VGAT‐ChR2‐YFP)‐expressing mice and a novel fibreoptic ‘laserspritzer’ approach that we developed, we investigated the physiological properties of axo‐axonic synapses (AASs) in brain slices from the piriform cortex (PC) of mice. AASs were in close proximity to voltage‐gated Na+ (NaV) channels located at the AIS. AASs were selectively activated by a 5 μm laserspritzer placed in close proximity to the AIS. Under a minimal laser stimulation condition and using whole‐cell somatic voltage‐clamp recordings, the amplitudes and kinetics of IPSCs mediated by AASs were similar to those mediated by perisomatic inhibitions. Results were further validated with channelrhodopsin 2‐assisted circuit mapping (CRACM) of the entire inhibitory inputs map. For the first time, we revealed that the laserspritzer‐induced AAS‐IPSCs persisted in the presence of TTX and TEA but not 4‐AP. Next, using gramicidin‐based perforated patch recordings, we found that the GABA reversal potential (EGABA) was −73.6 ± 1.2 mV when induced at the AIS and −72.8 ± 1.1 mV when induced at the perisomatic site. Our anatomical and physiological results lead to the novel conclusions that: (1) AASs innervate the entire length of the AIS, as opposed to forming a highly concentrated cartridge, (2) AAS inhibition suppresses action potentials and epileptiform activity more robustly than perisomatic inhibitions, and (3) AAS activation alone can be sufficient to inhibit action potential generation and epileptiform activities in vitro.
Presynaptic actions of transcranial and local direct current stimulation in the red nucleus
Authors: Bączyk M. , Jankowska E. 
Published: August 2014

Key points
Transcranial direct current stimulation (tDCS) may affect both postsynaptic neurons and presynaptic axons providing input to these neurons.
Here, we show that presynaptic subcortical actions of tDCS outlast the duration of its application by up to 1 h and may contribute to long‐lasting facilitation of activation of neurons in the red nucleus in experiments on deeply anaesthetized animals. This is demonstrated by increased axonal excitability of the interpositorubral neurons that, in turn, activate rubrospinal neurons.
We also show that effects of electric fields induced by tDCS may be reproduced by local cathodal polarization, by applying <1 μA direct current within the red nucleus. Such polarization replicates effects of anodal tDCS in the cat as well as cathodal tDCS in the rat and evokes similar long‐lasting facilitation.
AbstractThe main aim of the present study was to examine to what extent long‐lasting subcortical actions of transcranial direct current stimulation (tDCS) may be related to its presynaptic actions. This was investigated in the red nucleus, where tDCS was recently demonstrated to facilitate transmission between interpositorubral and rubrospinal neurons. Changes in the excitability of preterminal axonal branches of interpositorubral neurons close to rubrospinal neurons were investigated during and after tDCS (0.2 mA) applied over the sensorimotor cortical area in deeply anaesthetized rats and cats. As a measure of the excitability, we used the probability of antidromic activation of individual interpositorubral neurons by electrical stimuli applied in the red nucleus. Our second aim was to compare effects of weak (≤1 μA) direct current applied within the red nucleus with effects of tDCS to allow the use of local depolarization in a further analysis of mechanisms of tDCS instead of widespread and more difficult to control depolarization evoked by distant electrodes. Local cathodal polarization was found to replicate all effects of cathodal tDCS hitherto demonstrated in the rat, including long‐lasting facilitation of trans‐synaptically evoked descending volleys and trisynaptically evoked EMG responses in neck muscles. It also replicated all effects of anodal tDCS in the cat. In both species, it increased the excitability of preterminal axonal branches of interpositorubral neurons up to 1 h post‐tDCS. Local anodal polarization evoked opposite effects. We thus show that presynaptic actions of polarizing direct current may contribute to both immediate and prolonged effects of tDCS.
Effects of hyperoxia and hypoxia on the physiological traits responsible for obstructive sleep apnoea
Authors: Edwards Bradley A. , Sands Scott A. , Owens Robert L. , White David P. , Genta Pedro R. , Butler James P. , Malhotra Atul , Wellman Andrew 
Published: August 2014

Key points
Changes in the level of inspired oxygen have dramatic effects on the pathophysiology of obstructive sleep apnoea (OSA): hyperoxia reduces the severity of OSA in some but not all patients, whereas hypoxia transforms obstructive events into central events. Given that OSA is likely to result from the interaction of key pathophysiological traits, including a compromised pharyngeal anatomy, inadequate upper airway muscle function, a large ventilatory response to a disturbance in ventilation (high loop gain) and a low arousal threshold, we examined how changes in oxygen levels alter these traits.
Our study demonstrates that the beneficial effect of hyperoxia on OSA severity is solely based on its ability to attenuate loop gain, whereas hypoxia increases loop gain and the arousal threshold in addition to improving pharyngeal collapsibility.
Such effects help to explain why oxygen therapy may not work in every patient with OSA and explain the disappearance of OSA and the emergence of central events during hypoxic conditions.
AbstractOxygen therapy is known to reduce loop gain (LG) in patients with obstructive sleep apnoea (OSA), yet its effects on the other traits responsible for OSA remain unknown. Therefore, we assessed how hyperoxia and hypoxia alter four physiological traits in OSA patients. Eleven OSA subjects underwent a night of polysomnography during which the physiological traits were measured using multiple 3‐min ‘drops’ from therapeutic continuous positive airway pressure (CPAP) levels. LG was defined as the ratio of the ventilatory overshoot to the preceding reduction in ventilation. Pharyngeal collapsibility was quantified as the ventilation at CPAP of 0 cmH2O. Upper airway responsiveness was defined as the ratio of the increase in ventilation to the increase in ventilatory drive across the drop. Arousal threshold was estimated as the level of ventilatory drive associated with arousal. On separate nights, subjects were submitted to hyperoxia (n = 9; FiO2 ∼0.5) or hypoxia (n = 10; FiO2 ∼0.15) and the four traits were reassessed. Hyperoxia lowered LG from a median of 3.4 [interquartile range (IQR): 2.6–4.1] to 2.1 (IQR: 1.3–2.5) (P < 0.01), but did not alter the remaining traits. By contrast, hypoxia increased LG [median: 3.3 (IQR: 2.3–4.0) vs. 6.4 (IQR: 4.5–9.7); P < 0.005]. Hypoxia additionally increased the arousal threshold (mean ± s.d. 10.9 ± 2.1 l min−1 vs. 13.3 ± 4.3 l min−1; P < 0.05) and improved pharyngeal collapsibility (mean ± s.d. 3.4 ± 1.4 l min−1 vs. 4.9 ± 1.3 l min−1; P < 0.05), but did not alter upper airway responsiveness (P = 0.7). This study demonstrates that the beneficial effect of hyperoxia on the severity of OSA is primarily based on its ability to reduce LG. The effects of hypoxia described above may explain the disappearance of OSA and the emergence of central sleep apnoea in conditions such as high altitude.
Hypercapnia attenuates ventilator‐induced lung injury via a disintegrin and metalloprotease‐17
Authors: Otulakowski Gail , Engelberts Doreen , Gusarova Galina A. , Bhattacharya Jahar , Post Martin , Kavanagh Brian P. 
Published: August 2014

Key points
Hypercapnia is common in mechanically ventilated patients with lung injury; while CO2 can ameliorate experimental lung injury, it can also cause harm.
Because hypercapnia can protect against ventilator‐induced lung injury (VILI), understanding its impact on key signalling pathways may provide insight into the mechanisms of VILI.
We show that hypercapnia blocks stretch‐mediated activation of p44/42 mitogen‐activated protein kinase (MAPK) signalling in alveolar epithelial cells; this occurs through inhibition of sheddase (i.e. the metalloprotease, ADAM17), which releases ligands that bind to the epidermal growth factor receptor.
In vivo pharmacological blockade of ADAM17 reduces downstream MAPK activation and attenuates VILI in a two‐hit mouse model.
Thus, hypercapnia uncovered a novel ADAM17‐dependent mechanism of VILI, and this represents a potential therapeutic target.
AbstractHypercapnic acidosis, common in mechanically ventilated patients, has been reported to exert both beneficial and harmful effects in models of lung injury. Understanding its effects at the molecular level may provide insight into mechanisms of injury and protection. The aim of this study was to establish the effects of hypercapnic acidosis on mitogen‐activated protein kinase (MAPK) activation, and determine the relevant signalling pathways. p44/42 MAPK activation in a murine model of ventilator‐induced lung injury (VILI) correlated with injury and was reduced in hypercapnia. When cultured rat alveolar epithelial cells were subjected to cyclic stretch, activation of p44/42 MAPK was dependent on epidermal growth factor receptor (EGFR) activity and on shedding of EGFR ligands; exposure to 12% CO2 without additional buffering blocked ligand shedding, as well as EGFR and p44/42 MAPK activation. The EGFR ligands are known substrates of the matrix metalloprotease ADAM17, suggesting stretch activates and hypercapnic acidosis blocks stretch‐mediated activation of ADAM17. This was corroborated in the isolated perfused mouse lung, where elevated CO2 also inhibited stretch‐activated shedding of the ADAM17 substrate TNFR1 from airway epithelial cells. Finally, in vivo confirmation was obtained in a two‐hit murine model of VILI where pharmacological inhibition of ADAM17 reduced both injury and p44/42 MAPK activation. Thus, ADAM17 is an important proximal mediator of VILI; its inhibition is one mechanism of hypercapnic protection and may be a target for clinical therapy.
Microdosimetry of proton and carbon ions
Authors: Liamsuwan Thiansin , Hultqvist Martha , Lindborg Lennart , Uehara Shuzo , Nikjoo Hooshang 
Published: August 2014

Purpose:To investigate microdosimetry properties of 160 MeV/u protons and 290 MeV/u12C ion beams in small volumes of diameters 10–100 nm.Methods:Energy distributions of primary particles and nuclear fragments in the beams were calculated from simulations with the general purpose code SHIELD‐HIT, while energy depositions by monoenergetic ions in nanometer volumes were obtained from the event‐by‐event Monte Carlo track structure ion code PITS99 coupled with the electron track structure code KURBUC.Results:The results are presented for frequencies of energy depositions in cylindrical targets of diameters 10–100 nm, dose distributionsyd(y) in lineal energy y, and dose‐mean lineal energies . For monoenergetic ions, the was found to increase with an increasing target size for high‐linear energy transfer (LET) ions, but decrease with an increasing target size for low‐LET ions. Compared to the depth dose profile of the ion beams, the maximum of the depth profile for the 160 MeV proton beam was located at ∼0.5 cm behind the Bragg peak maximum, while the peak of the 290 MeV/u 12C beam coincided well with the peak of the absorbed dose profile. Differences between the and dose‐averaged linear energy transfer (LETD) were large in the proton beam for both target volumes studied, and in the 12C beam for the 10 nm diameter cylindrical volumes. The determined for 100 nm diameter cylindrical volumes in the 12C beam was approximately equal to the LETD. The contributions from secondary particles to the of the beams are presented, including the contributions from secondary protons in the proton beam and from fragments with atomic number Z = 1–6 in the 12C beam.Conclusions:The present investigation provides an insight into differences in energy depositions in subcellular‐size volumes when irradiated by proton and carbon ion beams. The results are useful for characterizing ion beams of practical importance for biophysical modeling of radiation‐induced DNA damage response and repair in the depth profiles of protons and carbon ions used in radiotherapy.
Motion tracking in the liver: Validation of a method based on 4D ultrasound using a nonrigid registration technique
Authors: Vijayan Sinara , Klein Stefan , Hofstad Erlend Fagertun , Lindseth Frank , Ystgaard Brynjulf , Langø Thomas 
Published: August 2014

Purpose:Treatments like radiotherapy and focused ultrasound in the abdomen require accurate motion tracking, in order to optimize dosage delivery to the target and minimize damage to critical structures and healthy tissues around the target. 4D ultrasound is a promising modality for motion tracking during such treatments. In this study, the authors evaluate the accuracy of motion tracking in the liver based on deformable registration of 4D ultrasound images.Methods:The offline analysis was performed using a nonrigid registration algorithm that was specifically designed for motion estimation from dynamic imaging data. The method registers the entire 4D image data sequence in a groupwise optimization fashion, thus avoiding a bias toward a specifically chosen reference time point. Three healthy volunteers were scanned over several breathing cycles (12 s) from three different positions and angles on the abdomen; a total of nine 4D scans for the three volunteers. Well‐defined anatomic landmarks were manually annotated in all 96 time frames for assessment of the automatic algorithm. The error of the automatic motion estimation method was compared with interobserver variability. The authors also performed experiments to investigate the influence of parameters defining the deformation field flexibility and evaluated how well the method performed with a lower temporal resolution in order to establish the minimum frame rate required for accurate motion estimation.Results:The registration method estimated liver motion with an error of 1 mm (75% percentile over all datasets), which was lower than the interobserver variability of 1.4 mm. The results were only slightly dependent on the degrees of freedom of the deformation model. The registration error increased to 2.8 mm with an eight times lower temporal resolution.Conclusions:The authors conclude that the methodology was able to accurately track the motion of the liver in the 4D ultrasound data. The authors believe that the method has potential in interventions on moving abdominal organs such as MR or ultrasound guided focused ultrasound therapy and radiotherapy, pending the method is enabled to run in real‐time. The data and the annotations used for this study are made publicly available for those who would like to test other methods on 4D liver ultrasound data.
Postmortem validation of breast density using dual‐energy mammography
Authors: Molloi Sabee , Ducote Justin L. , Ding Huanjun , Feig Stephen A. 
Published: August 2014

Purpose:Mammographic density has been shown to be an indicator of breast cancer risk and also reduces the sensitivity of screening mammography. Currently, there is no accepted standard for measuring breast density. Dual energy mammography has been proposed as a technique for accurate measurement of breast density. The purpose of this study is to validate its accuracy in postmortem breasts and compare it with other existing techniques.Methods:Forty postmortem breasts were imaged using a dual energy mammography system. Glandular and adipose equivalent phantoms of uniform thickness were used to calibrate a dual energy basis decomposition algorithm. Dual energy decomposition was applied after scatter correction to calculate breast density. Breast density was also estimated using radiologist reader assessment, standard histogram thresholding and a fuzzy C‐mean algorithm. Chemical analysis was used as the reference standard to assess the accuracy of different techniques to measure breast composition.Results:Breast density measurements using radiologist reader assessment, standard histogram thresholding, fuzzy C‐mean algorithm, and dual energy were in good agreement with the measured fibroglandular volume fraction using chemical analysis. The standard error estimates using radiologist reader assessment, standard histogram thresholding, fuzzy C‐mean, and dual energy were 9.9%, 8.6%, 7.2%, and 4.7%, respectively.Conclusions:The results indicate that dual energy mammography can be used to accurately measure breast density. The variability in breast density estimation using dual energy mammography was lower than reader assessment rankings, standard histogram thresholding, and fuzzy C‐mean algorithm. Improved quantification of breast density is expected to further enhance its utility as a risk factor for breast cancer.
Clotrimazole as a Potent Agent for Treating the Oomycete Fish Pathogen Saprolegnia parasitica through Inhibition of Sterol 14α-Demethylase (CYP51)
Authors: Warrilow Andrew G. S. , Hull Claire M. , Rolley Nicola J. , Parker Josie E. , Nes W. David , Smith Stephen N. , Kelly Diane E. , Kelly Steven L. 
Published: August 2014

ABSTRACT

A candidate CYP51 gene encoding sterol 14α-demethylase from the fish oomycete pathogen
Saprolegnia parasitica
(SpCYP51) was identified based on conserved CYP51 residues among CYPs in the genome. It was heterologously expressed in
Escherichia coli
, purified, and characterized. Lanosterol, eburicol, and obtusifoliol bound to purified SpCYP51 with similar binding affinities (

K
s

, 3 to 5 μM). Eight pharmaceutical and six agricultural azole antifungal agents bound tightly to SpCYP51, with posaconazole displaying the highest apparent affinity (

K
d

, ≤3 nM) and prothioconazole-desthio the lowest (
K
d
, ∼51 nM). The efficaciousness of azole antifungals as SpCYP51 inhibitors was confirmed by 50% inhibitory concentrations (IC
50
s) of 0.17 to 2.27 μM using CYP51 reconstitution assays. However, most azole antifungal agents were less effective at inhibiting
S. parasitica
,
Saprolegnia diclina
, and
Saprolegnia ferax
growth. Epoxiconazole, fluconazole, itraconazole, and posaconazole failed to inhibit
Saprolegnia
growth (MIC
100
, >256 μg ml
−1
). The remaining azoles inhibited
Saprolegnia
growth only at elevated concentrations (MIC
100
[the lowest antifungal concentration at which growth remained completely inhibited after 72 h at 20°C], 16 to 64 μg ml
−1
) with the exception of clotrimazole, which was as potent as malachite green (MIC
100
, ∼1 μg ml
−1
). Sterol profiles of azole-treated
Saprolegnia
species confirmed that endogenous CYP51 enzymes were being inhibited with the accumulation of lanosterol in the sterol fraction. The effectiveness of clotrimazole against SpCYP51 activity (IC
50
, ∼1 μM) and the concentration inhibiting the growth of
Saprolegnia
species
in vitro
(MIC
100
, ∼1 to 2 μg ml
−1
) suggest that clotrimazole could be used against
Saprolegnia
infections, including as a preventative measure by pretreatment of fish eggs, and for freshwater-farmed fish as well as in leisure activities.
Punicalagin Inhibits Salmonella Virulence Factors and Has Anti-Quorum-Sensing Potential
Authors: Li Guanghui , Yan Chunhong , Xu Yunfeng , Feng Yuqing , Wu Qian , Lv Xiaoying , Yang Baowei , Wang Xin , Xia Xiaodong 
Published: August 2014

ABSTRACT

Punicalagin, an essential component of pomegranate rind, has been demonstrated to possess antimicrobial activity against several food-borne pathogens, but its activity on the virulence of pathogens and its anti-quorum-sensing (anti-QS) potential have been rarely reported. This study investigated the efficacy of subinhibitory concentrations of punicalagin on
Salmonella
virulence factors and QS systems. A broth microdilution method was used to determine the MICs of punicalagin for 10
Salmonella
strains. Motility assay and quantitative reverse transcription (RT)-PCR were performed to evaluate the effects of punicalagin on the virulence attributes and QS-related genes of
Salmonella
. The MICs of punicalagin for several
Salmonella
strains ranged from 250 to 1,000 μg/ml. Motility assays showed that punicalagin, at 1/16× MIC and 1/32× MIC, significantly decreased bacterial swimming and swarming motility, which corresponded to downregulation of the motility-related genes (
fliA
,
fliY
,
fljB
,
flhC
, and
fimD
) in RT-PCR assays. RT-PCR also revealed that punicalagin downregulated the expression of most of the selected genes involved in
Salmonella
virulence. Moreover, a QS inhibition assay indicated that punicalagin dose dependently inhibited the production of violacein by
Chromobacterium violaceum
and repressed the expression of QS-related genes (
sdiA
and
srgE
) in
Salmonella
. In addition, punicalagin significantly reduced
Salmonella
invasion of colonic cells (
P
< 0.01) with no impact on adhesion. These findings suggest that punicalagin has the potential to be developed as an alternative or supplemental agent for prevention of
Salmonella
infection.
Efficient PCR-Based Amplification of Diverse Alcohol Dehydrogenase Genes from Metagenomes for Improving Biocatalysis: Screening of Gene-Specific Amplicons from Metagenomes
Authors: Itoh Nobuya , Kariya Satomi , Kurokawa Junji 
Published: August 2014

ABSTRACT

Screening of gene-specific amplicons from metagenomes (S-GAM) has tremendous biotechnological potential. We used this approach to isolate alcohol dehydrogenase (
adh
) genes from metagenomes based on the
Leifsonia
species
adh
gene (
lsadh
), the enzyme product of which can produce various chiral alcohols. A primer combination was synthesized by reference to homologs of
lsadh
, and PCR was used to amplify nearly full-length
adh
genes from metagenomic DNAs. All
adh
preparations were fused with
lsadh
at the terminal region and used to construct
Escherichia coli
plasmid libraries. Of the approximately 2,000 colonies obtained, 1,200 clones were identified as
adh
positive (∼60%). Finally, 40
adh
genes,
Hladh
-001 to
Hladh
-040 (for
h
omologous

L
eifsonia

adh

), were identified from 223 clones with high efficiency, which were randomly sequenced from the 1,200 clones. The
Hladh
genes obtained via this approach encoded a wide variety of amino acid sequences (8 to 99%). After screening, the enzymes obtained (HLADH-012 and HLADH-021) were confirmed to be superior to LSADH in some respects for the production of anti-Prelog chiral alcohols.