Brain responses to violet, blue, and green monochromatic light exposures in humans: prominent role of blue light and the brainstem

Background

Relatively long duration retinal light exposure elicits nonvisual responses in humans, including modulation of alertness and cognition. These responses are thought to be mediated in part by melanopsin-expressing retinal ganglion cells which are more sensitive to blue light than violet or green light. The contribution of the melanopsin system and the brain mechanisms involved in the establishment of such responses to light remain to be established.

Methodology/Principal Findings

We exposed 15 participants to short duration (50 s) monochromatic violet (430 nm), blue (473 nm), and green (527 nm) light exposures of equal photon flux (10¹³ph/cm²/s) while they were performing a working memory task in fMRI. At light onset, blue light, as compared to green light, increased activity in the left hippocampus, left thalamus, and right amygdala. During the task, blue light, as compared to violet light, increased activity in the left middle frontal gyrus, left thalamus and a bilateral area of the brainstem consistent with activation of the locus coeruleus.

Conclusion/Significance

These results support a prominent contribution of melanopsin-expressing retinal ganglion cells to brain responses to light within the very first seconds of an exposure. The results also demonstrate the implication of the brainstem in mediating these responses in humans and speak for a broad involvement of light in the regulation of brain function.     

Efficient production of Clostridium botulinum exotoxin C3 in bacteria: a screening method to optimize production yields

Clostridium botulinum exoenzyme C3 is responsible for the inactivation of members of the Rho GTPase family that are implicated in actin-cytoskeleton reorganization. This property has been extensively used in the field to investigate the functionality of the Rho GTPases. However, systematic analysis of Rho GTPase functions requires large amounts of such inhibitors and consequently an optimization of the production yield of these proteins. Bacterial production of soluble proteins often requires a refolding step that noticeably affects the production yields and necessitates additional experiments to verify functional activity. This is particularly true for TAT-C3, the production yields of which are generally low. In this report, we describe a rapid and efficient method for the production of soluble C3 exoenzyme developed by screening a collection of bacterial strains. The recombinant C3 protein was fused to the TAT protein-transduction domain from HIV, to allow protein delivery into cells, and to a hexahistidine tag, that permitted purification by Nickel affinity chromatography. We have demonstrated the production of large amounts of soluble and functional protein using the bacterial strain AD494 (DE3)pLysS. This rapid and efficient method for the production of soluble C3 exoenzyme could also be useful for the production of other proteins with solubility problems.     

Reversible inactivation of the transcriptional function of P53 protein by farnesylation

Background  

The use of integrating viral vectors in Gene therapy clinical trials has pointed out the problem of the deleterous effect of the integration of the ectopic gene to the cellular genome and the safety of this strategy. We proposed here a way to induce the death of gene modified cells upon request by acting on a pro-apoptotic protein cellular localization and on the activation of its apoptotic function.     

Solution structure of Phrixotoxin 1, a specific peptide inhibitor of Kv4 potassium channels from the venom of the theraphosid spider Phrixotrichus auratus

Animal toxins block voltage-dependent potassium channels (Kv) either by occluding the conduction pore (pore blockers) or by modifying the channel gating properties (gating modifiers). Gating modifiers of Kv channels bind to four equivalent extracellular sites near the S3 and S4 segments, close to the voltage sensor. Phrixotoxins are gating modifiers that bind preferentially to the closed state of the channel and fold into the Inhibitory Cystine Knot structural motif. We have solved the solution structure of Phrixotoxin 1, a gating modifier of Kv4 potassium channels. Analysis of the molecular surface and the electrostatic anisotropy of Phrixotoxin 1 and of other toxins acting on voltage-dependent potassium channels allowed us to propose a toxin interacting surface that encompasses both the surface from which the dipole moment emerges and a neighboring hydrophobic surface rich in aromatic residues.     

Neural Mechanisms Underlying Breathing Complexity

Breathing is maintained and controlled by a network of automatic neurons in the brainstem that generate respiratory rhythm and receive regulatory inputs. Breathing complexity therefore arises from respiratory central pattern generators modulated by peripheral and supra-spinal inputs. Very little is known on the brainstem neural substrates underlying breathing complexity in humans. We used both experimental and theoretical approaches to decipher these mechanisms in healthy humans and patients with chronic obstructive pulmonary disease (COPD). COPD is the most frequent chronic lung disease in the general population mainly due to tobacco smoke. In patients, airflow obstruction associated with hyperinflation and respiratory muscles weakness are key factors contributing to load-capacity imbalance and hence increased respiratory drive. Unexpectedly, we found that the patients breathed with a higher level of complexity during inspiration and expiration than controls. Using functional magnetic resonance imaging (fMRI), we scanned the brain of the participants to analyze the activity of two small regions involved in respiratory rhythmogenesis, the rostral ventro-lateral (VL) medulla (pre-Bötzinger complex) and the caudal VL pons (parafacial group). fMRI revealed in controls higher activity of the VL medulla suggesting active inspiration, while in patients higher activity of the VL pons suggesting active expiration. COPD patients reactivate the parafacial to sustain ventilation. These findings may be involved in the onset of respiratory failure when the neural network becomes overwhelmed by respiratory overload We show that central neural activity correlates with airflow complexity in healthy subjects and COPD patients, at rest and during inspiratory loading. We finally used a theoretical approach of respiratory rhythmogenesis that reproduces the kernel activity of neurons involved in the automatic breathing. The model reveals how a chaotic activity in neurons can contribute to chaos in airflow and reproduces key experimental fMRI findings.     

Chicoric Acid Is an Antioxidant Molecule That Stimulates AMP Kinase Pathway in L6 Myotubes and Extends Lifespan in Caenorhabditis elegans

Chicoric acid (CA) is a caffeoyl derivative previously described as having potential anti-diabetic properties. As similarities in cellular mechanism similarities between diabetes and aging have been shown, we explored on L6 myotubes the effect of CA on the modulation of intracellular pathways involved in diabetes and aging. We also determined its influence on lifespan of Caenorhabditis elegans worm (C. elegans). In L6 myotubes, CA was a potent reactive oxygen species (ROS) scavenger, reducing ROS accumulation under basal as well as oxidative stress conditions. CA also stimulated the AMP-activated kinase (AMPK) pathway and displayed various features associated with AMPK activation: CA (a) enhanced oxidative enzymatic defences through increase in glutathion peroxidase (GPx) and superoxide dismutase (SOD) activities, (b) favoured mitochondria protection against oxidative damage through up-regulation of MnSOD protein expression, (c) increased mitochondrial biogenesis as suggested by increases in complex II and citrate synthase activities, along with up-regulation of PGC-1α mRNA expression and (d) inhibited the insulin/Akt/mTOR pathway. As AMPK stimulators (e.g. the anti-diabetic agent meformin or polyphenols such as epigallocatechingallate or quercetin) were shown to extend lifespan in C. elegans, we also determined the effect of CA on the same model. A concentration-dependant lifespan extension was observed with CA (5–100 μM). These data indicate that CA is a potent antioxidant compound activating the AMPK pathway in L6 myotubes. Similarly to other AMPK stimulators, CA is able to extend C. elegans lifespan, an effect measurable even at the micromolar range. Future studies will explore CA molecular targets and give new insights about its possible effects on metabolic and aging-related diseases.     

2′-and/or 3Y-Deoxy-β-L-pentofuranosyl Nucleoside Derivatives: Stereospecific Synthesis and Antiviral Activities

Abstract

Several L-enantiomers of nucleoside analogues were stereospecifically synthesized by a multi-step reaction from L-xylose and their antiviral properties were examined in vitro. Two of them, namely β-L-2′,3,′-dideoxycytidine (β-L-ddC) and its 5-fluoro derivative (β-L-FddC) were found to have potent anti-human immunodeficiency virus (HIV) and significant anti-hepatitis B virus (HBV) activities in cell cultures.     

Protein mutagenesis with monodispersity-based quality probing: selective inactivation of p53 degradation and DNA-binding properties of HPV E6 oncoprotein

Interpretation of protein mutagenesis experiments requires the ability to distinguish functionally relevant mutations from mutations affecting the structure. When a protein is expressed soluble in bacteria, properly folded mutants are expected to remain soluble whereas misfolded mutants should form insoluble aggregates. However, this rule may fail for proteins fused to highly soluble carrier proteins. In a previous study, we analysed the biophysical status of HPV oncoprotein E6 fused to the C-terminus of maltose-binding protein (MBP) and found that misfolded E6 moieties fused to MBP formed soluble aggregates of high molecular weight. By contrast, preparations of properly folded E6 fused to MBP were monodisperse. Here, we have used this finding to evaluate the quality of 19 MBP-fused E6 site-directed mutants by using a light scattering assay performed in a fluorimeter. This assay guided us to rule out structurally defective mutants and to obtain functionally relevant E6 mutants selectively altered for two molecular activities: degradation of tumour suppressor p53 and DNA recognition.