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Accumulating evidence suggests AKT1 and DRD2-AKT-GSK3 signaling involvement in schizophrenia. AKT1 activity is also required for lithium, a GSK3 inhibitor, to modulate mood-related behaviors. Notably, GSK3 inhibitor significantly alleviates behavioral deficits in Akt1−/− female mice, whereas typical/atypical antipsychotics have no effect. In agreement with adjunctive therapy with lithium in treating schizophrenia, our data mining indicated that the average utilization rates of lithium in the Taiwan National Health Insurance Research Database from 2002 to 2013 are 10.9% and 6.63% in inpatients and outpatients with schizophrenia, respectively. Given that lithium is commonly used in clinical practice, it is of great interest to evaluate the effect of lithium on alleviating Akt1-related deficits. Taking advantage of Akt1+/− mice to mimic genetic deficiency in patients, behavioral impairments were replicated in female Akt1+/− mice but were alleviated by subchronic lithium treatment for 13 days. Lithium also effectively alleviated the observed reduction in phosphorylated GSK3α/β expression in the brains of Akt1+/− mice. Furthermore, inhibition of Akt expression using an Akt1/2 inhibitor significantly reduced neurite length in P19 cells and primary hippocampal cell cultures, which was also ameliorated by lithium. Collectively, our findings implied the therapeutic potential of lithium and the importance of the AKT1-GSK3 signaling pathway.

The in vitro methods to recapitulate environmental cues around cells are usually optimized to test a specific property of the environment (biochemical nature or the stiffness of the extra cellular matrix (ECM), or nanotopography) for its capability to induce defined cell behaviors (lineage commitment, migration). Approaches that combine different environmental cues in 3D to assess the biological response of cells to the spatial organization of different biophysical and biochemical cues are growingly being developed. We demonstrate how the lamination of through-hole polymeric bio-functionalized membranes can be implemented to create complex bona fide micro-niches with differential 3D environmental properties using photoactive materials. Our approach enables to create micro-niches ranging in size from single cells to cell aggregates. They are bio-functionalized in 3D simultaneously with topographical featured, protein patterns and structured ECM surrogate with 1 micrometer resolution. We demonstrate how these niches extend in 3D the ability to pattern cells. We exemplify how they can be used to standardize cells shapes in 3D and to trigger the apico-basal polarization of single epithelial cells.

We demonstrate that room-temperature and low-pressure nanoimprint lithography techniques can be achieved by using Hybrane HS2550, a semi-crystalline hyperbranched resist polymer with a glass transition temperature below and a melting point above room temperature. Nanoimprint lithography at room temperature is possible with sub-100 nm resolution, as 75 nm line-and-spacing gratings were successfully fabricated with a tri-layer process and a metal lift-off. The melt viscosity of Hybrane HS2550 decreases drastically with temperature allowing nanoimprint experiments at low pressures, while maintaining imprint temperatures that are much lower than commonly required in nanoimprint technology.

International audience; Nerve growth cones (GCs) are chemical sensors that convert graded extracellular cues into oriented axonal motion. To ensure a sensitive and robust response to directional signals in complex and dynamic chemical landscapes, GCs are presumably able to amplify and filter external information. How these processing tasks are performed remains however poorly known. Here, we probe the signal-processing capabilities of single GCs during ?-Aminobutyric acid (GABA) directional sensing with a shear-free microfluidic assay that enables systematic measurements of the GC output response to variable input gradients. By measuring at the single molecule level the polarization of GABA A chemoreceptors at the GC membrane, as a function of the external GABA gradient, we find that GCs act as i), signal amplifiers over a narrow range of concentrations, and ii), low-pass temporal filters with a cutoff frequency independent of stimuli conditions. With computational modeling, we determine that these systems-level properties arise at a molecular level from the saturable occupancy response and the lateral dynamics of GABA A receptors. Cop. 2012 Biophysical Society.

5 pages; International audience; Immobilization and electrical wiring of enzymes is of particular importance for the elaboration of efficient biosensors and can be cumbersome. Here, we report a fast and easy protocol for enzyme immobilization, and as a proof of concept, we applied it to the immobilization of bilirubin oxidase, a labile enzyme. In the first step, bilirubin oxidase is mixed with a redox hydrogel "wiring" the enzyme reaction centers to electrodes. Then, this adduct is covered by an outer layer of PEGDA made by photoinitiated polymerization of poly(ethylene-glycol) diacrylate (PEGDA) and a photoclivable precursor, DAROCUR. This two-step protocol is 18 times faster than the current state-of-the-art protocol and leads to currents 25% higher. In addition, the outer layer of PEGDA acts as a protective layer increasing the lifetime of the electrode by 100% when operating continuously for 2000 s and by 60% when kept in dry state for 24 h. This new protocol is particularly appropriate for labile enzymes that quickly denaturate. In addition, by tuning the ratio PEGDA/DAROCUR, it is possible to make the enzyme electrodes even more active or more stable.

Using basic physical arguments, we present a design and method for the fabrication of microfluidic valves using multilayer soft lithography. These on-off valves have extremely low actuation pressures and can be used to fabricate active functions, such as pumps and mixers in integrated microfluidic chips. We characterized the performance of the valves by measuring both the actuation pressure and flow resistance over a wide range of design parameters, and compared them to both finite element simulations and alternative valve geometries.

Nerve growth cones (GCs) are chemical sensors that convert graded extracellular cues into oriented axonal motion. Ensuring a sensitive and robust GC response to directional signals requires the ability to amplify and filter external gradients. However, our knowledge of how these signal processing tasks are performed at the single cell level remains sparse. This is largely due to the limitations of conventional guidance assays that have precluded systematic measurements of the GC output response to variable input gradients. We developed a novel shear-free gradient-generating microfluidic device with a simple architecture that greatly facilitates the interface of cultured neurons with microcircuits. With this device, we probed the information-processing capabilities of single GCs during GABA directional sensing. By measuring at the single molecule level the polarization of GABAA chemoreceptors at the GC membrane as a function of the external GABA gradient, we found that GCs act as: (i) signal amplifiers over a narrow range of concentrations, (ii) low-pass temporal filters with a cut-off frequency independent of stimuli conditions. Furthermore, thanks to quantitative computational modeling, we related these systems-level properties to the saturable occupancy response and the lateral dynamics of GABAA receptors, and, thereby, provided an integrative view of individual GCs as sensing devices.

Tailoring hydrogels into biomimetic templates represents a crucial step to build better in-vitro models but it is to date still challenging. Indeed, these synthetic or natural polymeric networks are often so frail they can’t be processed through standard micro-fabrication. Here, we combine a ultra-violet pattern projector with gas permeable microreactors to control gas, reagents and photon distribution and in fine, the reaction kinetics in space and time. Doing so, enabled a generic chemistry that can structure, liquefy or decorate (locally functionalize) common hydrogels. Altogether these three hydrogel engineering operations form a flexible toolbox that supports the most commonly used hydrogels: i.e. Matrigel, Agar-agar, poly(ethylene-glycol) and poly(acryl-amide). We successfully applied this solution to grow cells into standardized micro-niches demonstrating that it can readily address cell culture challenges such has controlled adhesion on topographical structures, standardization of spheroids or culture on shaped Matrigel.; L’ingénierie d’hydrogels ; leur structuration et fonctionnalisation à l’échelle cellulaire, est une étape clé pour aboutir à de modèles in-vitro plus physiologiques. À ce jour, elle reste difficile car ces matériaux polymères, mous et riches en eau, sont souvent trop fragiles pour la micro-fabrication traditionnelle. Pour pallier à ce fait, nous avons combinée illumination ultraviolette structurée et chambres de réaction perméables au gaz nous offrant la maitrise sur la distribution de photons, les réactifs et les gaz présents à chaque instant et en tout point d’un champ d’illumination. Nous pouvons ainsi contrôler une photochimie adaptée aux hydrogels les plus répandus et structurer, décorer ou liquéfier ces matériaux. Ensemble ces trois opérations forment une boite à outil complète adaptée aux substrats les plus communs que sont Matrigel, Agar, Poly(acrylamide) et Poly(éthylène-glycol). Nous avons par la suite fabriqué des micro-niches en hydrogel permettant la culture standardisée de lignées cellulaire et de neurones primaires soit par adhésion sur des topographies ou par auto-organisation en sphéroïdes. Ceci démontre que la plateforme est à même de répondre à des enjeux importants de culture cellulaire tridimensionnelle.