Search

Realizing the full potential of organoids and assembloids to model neural development and disease will require improved methods for long-term, minimally invasive recording of electrical activity. Current technologies, such as patch clamp, penetrating microelectrodes, planar electrode arrays and substrate-attached flexible electrodes, do not allow chronic recording of organoids in suspension, which is necessary to preserve architecture. Inspired by kirigami art, we developed flexible electronics that transition from a two-dimensional to a three-dimensional basket-like configuration with either spiral or honeycomb patterns to accommodate the long-term culture of organoids in suspension. Here we show that this platform, named kirigami electronics (KiriE), integrates with and enables chronic recording of cortical organoids for up to 120 days while preserving their morphology, cytoarchitecture and cell composition. We demonstrate integration of KiriE with optogenetic and pharmacological manipulation and modeling phenotypes related to a genetic disease. Moreover, KiriE can capture corticostriatal connectivity in assembloids following optogenetic stimulation. Thus, KiriE will enable investigation of disease and activity patterns underlying nervous system assembly. Electrical activity in neural organoids is captured with a device inspired by a paper-cutting art. [ABSTRACT FROM AUTHOR]

Morphogenesis requires building stable macromolecular structures from highly dynamic proteins. Muscles are anchored by long-lasting integrin adhesions to resist contractile force. However, the mechanisms governing integrin diffusion, immobilization, and activation within developing tissues remain elusive. Here, we show that actin polymerization-driven membrane protrusions form nanotopographies that enable strong adhesion at Drosophila muscle attachment sites (MASs). Super-resolution microscopy reveals that integrins assemble adhesive belts around Arp2/3-dependent actin protrusions, forming invadosome-like structures with membrane nanotopographies. Single protein tracking shows that, during MAS development, integrins become immobile and confined within diffusion traps formed by the membrane nanotopographies. Actin filaments also display restricted motion and confinement, indicating strong mechanical connection with integrins. Using isolated muscle cells, we show that substrate nanotopography, rather than rigidity, drives adhesion maturation by regulating actin protrusion, integrin diffusion and immobilization. These results thus demonstrate that actin-polymerization-driven membrane protrusions are essential for the formation of strong integrin adhesions sites in the developing embryo, and highlight the important contribution of geometry to morphogenesis. The mechanisms controlling the formation of cellular adhesions are not fully understood. Here, the authors use single molecule tracking and super-resolution imaging to show that the actin cytoskeleton regulates integrin diffusion in the developing Drosophila embryo. [ABSTRACT FROM AUTHOR]

Subcellular protein localization regulates protein function and can be corrupted in cancers1 and neurodegenerative diseases2,3. The rewiring of localization to address disease-driving phenotypes would be an attractive targeted therapeutic approach. Molecules that harness the trafficking of a shuttle protein to control the subcellular localization of a target protein could enforce targeted protein relocalization and rewire the interactome. Here we identify a collection of shuttle proteins with potent ligands amenable to incorporation into targeted relocalization-activating molecules (TRAMs), and use these to relocalize endogenous proteins. Using a custom imaging analysis pipeline, we show that protein steady-state localization can be modulated through molecular coupling to shuttle proteins containing sufficiently strong localization sequences and expressed in the necessary abundance. We analyse the TRAM-induced relocalization of different proteins and then use nuclear hormone receptors as shuttles to redistribute disease-driving mutant proteins such as SMARCB1Q318X, TDP43ΔNLS and FUSR495X. TRAM-mediated relocalization of FUSR495X to the nucleus from the cytoplasm correlated with a reduction in the number of stress granules in a model of cellular stress. With methionyl aminopeptidase 2 and poly(ADP-ribose) polymerase 1 as endogenous cytoplasmic and nuclear shuttles, respectively, we demonstrate relocalization of endogenous PRMT9, SOS1 and FKBP12. Small-molecule-mediated redistribution of nicotinamide nucleotide adenylyltransferase 1 from nuclei to axons in primary neurons was able to slow axonal degeneration and pharmacologically mimic the genetic WldS gain-of-function phenotype in mice resistant to certain types of neurodegeneration4. The concept of targeted protein relocalization could therefore inspire approaches for treating disease through interactome rewiring.Targeted protein relocalization using shuttle proteins with potent ligands amenable to incorporation into targeted relocalization activating molecules could be used to regulate cellular physiology and correct disease states in neurodegenerative diseases, cancer and genetic disorders. [ABSTRACT FROM AUTHOR]

Self-organizing neural organoids represent a promising in vitro platform with which to model human development and disease1–5. However, organoids lack the connectivity that exists in vivo, which limits maturation and makes integration with other circuits that control behaviour impossible. Here we show that human stem cell-derived cortical organoids transplanted into the somatosensory cortex of newborn athymic rats develop mature cell types that integrate into sensory and motivation-related circuits. MRI reveals post-transplantation organoid growth across multiple stem cell lines and animals, whereas single-nucleus profiling shows progression of corticogenesis and the emergence of activity-dependent transcriptional programs. Indeed, transplanted cortical neurons display more complex morphological, synaptic and intrinsic membrane properties than their in vitro counterparts, which enables the discovery of defects in neurons derived from individuals with Timothy syndrome. Anatomical and functional tracings show that transplanted organoids receive thalamocortical and corticocortical inputs, and in vivo recordings of neural activity demonstrate that these inputs can produce sensory responses in human cells. Finally, cortical organoids extend axons throughout the rat brain and their optogenetic activation can drive reward-seeking behaviour. Thus, transplanted human cortical neurons mature and engage host circuits that control behaviour. We anticipate that this approach will be useful for detecting circuit-level phenotypes in patient-derived cells that cannot otherwise be uncovered.Human stem cell-derived cortical organoids transplanted into rats mature and integrate into sensory and motivation circuits to influence behaviour. [ABSTRACT FROM AUTHOR]

Neurotransmitters play essential roles in regulating neural circuit dynamics both in the central nervous system as well as at the peripheral, including the gastrointestinal tract1–3. Their real-time monitoring will offer critical information for understanding neural function and diagnosing disease1–3. However, bioelectronic tools to monitor the dynamics of neurotransmitters in vivo, especially in the enteric nervous systems, are underdeveloped. This is mainly owing to the limited availability of biosensing tools that are capable of examining soft, complex and actively moving organs. Here we introduce a tissue-mimicking, stretchable, neurochemical biological interface termed NeuroString, which is prepared by laser patterning of a metal-complexed polyimide into an interconnected graphene/nanoparticle network embedded in an elastomer. NeuroString sensors allow chronic in vivo real-time, multichannel and multiplexed monoamine sensing in the brain of behaving mouse, as well as measuring serotonin dynamics in the gut without undesired stimulations and perturbing peristaltic movements. The described elastic and conformable biosensing interface has broad potential for studying the impact of neurotransmitters on gut microbes, brain–gut communication and may ultimately be extended to biomolecular sensing in other soft organs across the body.NeuroString, a tissue-like biological interface created by laser patterning of polyimide into a graphene/nanoparticle network embedded in an elastomer, is introduced, allowing in vivo real-time detection of neurotransmitters in the brain and gut. [ABSTRACT FROM AUTHOR]

The glycocalyx is a shell of heavily glycosylated proteins and lipids distributed on the cell surface of nearly all cell types. Recently, it has been found that bulky transmembrane glycoproteins such as MUC1 can modulate membrane shape by inducing membrane protrusions. In this work, we examine the reciprocal relationship of how membrane shape affects MUC1's spatial distribution on the cell membrane and its biological significance. By employing nanopatterned surfaces and membrane-sculpting proteins to manipulate membrane curvature, we show that MUC1 avoids positively-curved membranes (membrane invaginations) and accumulates on negatively-curved membranes (membrane protrusions). MUC1's curvature sensitivity is dependent on the length and the extent of glycosylation of its ectodomain, with large and highly glycosylated forms preferentially staying out of positive curvature. Interestingly, MUC1's avoidance of positive membrane curvature enables it to escape from endocytosis and being removed from the cell membrane. These findings also suggest that the truncation of MUC1's ectodomain, often observed in breast and ovarian cancers, may enhance its endocytosis and potentiate its intracellular accumulation and signaling. MUC1 is a heavily glycosylated protein on the cell surface. Here the authors show that MUC1 prefers negative over positive membrane curvature due to its bulky size, enabling MUC1 to avoid endocytosis and surface removal based on curvature preference. [ABSTRACT FROM AUTHOR]

Drug-induced cardiotoxicity arises primarily when a compound alters the electrophysiological properties of cardiomyocytes. Features of intracellular action potentials (iAPs) are powerful biomarkers that predict proarrhythmic risks. In the last decade, a number of vertical nanoelectrodes have been demonstrated to achieve parallel and minimally-invasive iAP recordings. However, the large variability in success rate and signal strength have hindered nanoelectrodes from being broadly adopted for proarrhythmia drug assessment. In this work, we develop vertically-aligned nanocrown electrodes that are mechanically robust and achieve > 99% success rates in obtaining intracellular access through electroporation. We validate the accuracy of nanocrown electrode recordings by simultaneous patch clamp recording from the same cell. Finally, we demonstrate that nanocrown electrodes enable prolonged iAP recording for continual monitoring of the same cells upon the sequential addition of four incremental drug doses. Our technology development provides an advancement towards establishing an iAP screening assay for preclinical evaluation of drug-induced arrhythmogenicity. Nanoelectrodes for measuring intracellular action potentials suffer from issues with success rate, signal strength and fabrication. Here, the authors report on a scalable technique which creates robust nanocrown electrodes with high success rates by electroporation and demonstrate the advance towards preclinical drug evaluation. [ABSTRACT FROM AUTHOR]

Bioelectronics aims to design electronic devices which can be fully integrated within tissues to monitor or stimulate specific cell functions. The main challenge is the engineering of the cell–chip interface and diverse materials, and devices have been developed to recapitulate biological architectures and functionalities. In this Prospective article, the authors give an overview on how the bioelectronics community has exploited biomimetic approaches to emulate cell morphologies, interactions, and functions to design optimal electrical platforms to be coupled to living cells. [ABSTRACT FROM AUTHOR]

Hierarchically ordered scaffold has a great impact on cell patterning and tissue engineering. The introduction of controllable coils into a scaffold offers an additional unique structural feature compared to conventional linear patterned scaffolds and can greatly increase interior complexity and versatility. In this work, 3D coil compacted scaffolds with hierarchically ordered patterns and tunable coil densities created using speed-programmed melt electrospinning writing (sMEW) successfully led to in vitro cell growth in patterns with tunable cell density. Subcutaneous implantation in mice showed great in vivo biocompatibility, as evidenced by no significant increase in tumor necrosis factor α (TNF-α) and interleukin 6 (IL-6) levels in mouse serum. In addition, a lumbar vertebra was successfully printed for mesenchymal stem cells to grow in the desired pattern. A long-range patterned matrix composed of programmable short-range compacted coils enabled the design of complex structures, e.g., for tailored implants, by readily depositing short-range coil-compacted secondary architectures along with customized primary design. Tissue engineering: melt electrospinning writing helps cells grow a backbone Improvements to a recently developed technique for printing biomaterials could make it easier to fabricate complex scaffolds for tissue regrowth. By shaping biocompatible polymers into thin microfibers and then collecting the strands on a moving platform, researchers can build tissue scaffolds layer-by-layer into 3D structures. Yingchun Su from the Harbin Institute of Technology in Harbin, China, and Aarhus University, Denmark, and colleagues report that materials printed by this 'speed-programmed melt electrospinning writing' technique can be composed of coil-shaped microfibers. The team found that the coil-like strands, produced by manipulating platform collection speeds, were effective at introducing pores into 3D scaffolds to better mimic natural tissue. Experiments showed this approach enabled better control over the cell density and growth behavior of stem cells implanted in a scaffold shaped like an artificial lumbar vertebra. Speed-programmed melt electrospinning writing (sMEW) is used to create a hierarchically ordered biomimetic scaffold with long-range patterned and short-range porous architectures for cell growth in patterns with tunable cell density. [ABSTRACT FROM AUTHOR]

Tungsten diselenide (WSe2) has many excellent properties and provides superb potential in applications of valley-based electronics, spin-electronics, and optoelectronics. To facilitate the digital and analog application of WSe2 in CMOS, it is essential to understand the underlying ambipolar hole and electron transport behavior. Herein, the electric field screening of WSe2 with a thickness range of 1-40 layers is systemically studied by electrostatic force microscopy in combination with non-linear Thomas-Fermi theory to interpret the experimental results. The ambipolar transport behavior of 1-40 layers of WSe2 transistors is systematically investigated with varied temperature from 300 to 5 K. The thickness-dependent transport properties (carrier mobility and Schottky barrier) are discussed. Furthermore, the surface potential of WSe2 as a function of gate voltage is performed under Kelvin probe force microscopy to directly investigate its ambipolar behavior. The results show that the Fermi level will upshift by 100 meV when WSe2 transmits from an insulator to an n-type semiconductor and downshift by 340 meV when WSe2 transmits from an insulator to a p-type semiconductor. Finally, the ambipolar WSe2 transistor-based analog circuit exhibits phase-control by gate voltage in an analog inverter, which demonstrates practical application in 2D communication electronics. Transistors made from two-dimensional tungsten selenide (WSe2) crystals may simplify fabrication of electronic communication devices. Modern transistors amplify and manipulate current by applying electric fields to semiconductor films known as channels that are designed to transport either positive or negative charges. Mingdong Dong from Aarhus University in Denmark and colleagues have developed a new transistor that can move both types of charge. The team found that when individual flakes of WSe2 were stacked into a multi-layered channel structure, the dominant type of charge transported could be switched using an external electrode. A prototype amplifier constructed from two WSe2-based transistors showed this approach enabled fundamental control over analog signals, and could potentially be used in complex circuits that require fewer materials and smaller chip footprints than usual. Compared with unipolar transistors, ambipolar transistors, which can easily switch between n-type and p-type behavior by applying an electric field, are most promising candidates since they can effectively simplify circuit design and save the layout area in CMOS. In this study, we take a deep insight into the thickness dependent physical properties of WSe2, where the optical properties, electric field screening effect, and the ambipolar transport behavior are systematically studied. Furthermore, the investigation of ambipolar WSe2 transistors in analogue circuits exhibiting gate-controlled phase change directly explores its practical application in 2D communication electronics. [ABSTRACT FROM AUTHOR]

The mechanical stability and deformability of the cell nucleus are crucial to many biological processes, including migration, proliferation and polarization. In vivo, the cell nucleus is frequently subjected to deformation on a variety of length and time scales, but current techniques for studying nuclear mechanics do not provide access to subnuclear deformation in live functioning cells. Here we introduce arrays of vertical nanopillars as a new method for the in situ study of nuclear deformability and the mechanical coupling between the cell membrane and the nucleus in live cells. Our measurements show that nanopillar-induced nuclear deformation is determined by nuclear stiffness, as well as opposing effects from actin and intermediate filaments. Furthermore, the depth, width and curvature of nuclear deformation can be controlled by varying the geometry of the nanopillar array. Overall, vertical nanopillar arrays constitute a novel approach for non-invasive, subcellular perturbation of nuclear mechanics and mechanotransduction in live cells. [ABSTRACT FROM AUTHOR]

Existing methods for human induced pluripotent stem cell (hiPSC) cardiac differentiation are efficient but require complex, undefined medium constituents that hinder further elucidation of the molecular mechanisms of cardiomyogenesis. Using hiPSCs derived under chemically defined conditions on synthetic matrices, we systematically developed an optimized cardiac differentiation strategy, using a chemically defined medium consisting of just three components: the basal medium RPMI 1640, L-ascorbic acid 2-phosphate and rice-derived recombinant human albumin. Along with small molecule-based induction of differentiation, this protocol produced contractile sheets of up to 95% TNNT2+ cardiomyocytes at a yield of up to 100 cardiomyocytes for every input pluripotent cell and was effective in 11 hiPSC lines tested. This chemically defined platform for cardiac specification of hiPSCs will allow the elucidation of cardiomyocyte macromolecular and metabolic requirements and will provide a minimal system for the study of maturation and subtype specification. [ABSTRACT FROM AUTHOR]

Action potentials have a central role in the nervous system and in many cellular processes, notably those involving ion channels. The accurate measurement of action potentials requires efficient coupling between the cell membrane and the measuring electrodes. Intracellular recording methods such as patch clamping involve measuring the voltage or current across the cell membrane by accessing the cell interior with an electrode, allowing both the amplitude and shape of the action potentials to be recorded faithfully with high signal-to-noise ratios. However, the invasive nature of intracellular methods usually limits the recording time to a few hours, and their complexity makes it difficult to simultaneously record more than a few cells. Extracellular recording methods, such as multielectrode arrays and multitransistor arrays, are non-invasive and allow long-term and multiplexed measurements. However, extracellular recording sacrifices the one-to-one correspondence between the cells and electrodes, and also suffers from significantly reduced signal strength and quality. Extracellular techniques are not, therefore, able to record action potentials with the accuracy needed to explore the properties of ion channels. As a result, the pharmacological screening of ion-channel drugs is usually performed by low-throughput intracellular recording methods. The use of nanowire transistors, nanotube-coupled transistors and micro gold-spine and related electrodes can significantly improve the signal strength of recorded action potentials. Here, we show that vertical nanopillar electrodes can record both the extracellular and intracellular action potentials of cultured cardiomyocytes over a long period of time with excellent signal strength and quality. Moreover, it is possible to repeatedly switch between extracellular and intracellular recording by nanoscale electroporation and resealing processes. Furthermore, vertical nanopillar electrodes can detect subtle changes in action potentials induced by drugs that target ion channels. [ABSTRACT FROM AUTHOR]

The article focuses the use of kinked nanowire transistors to record the intracellular signals of the targeted cells in human body. Topics discussed include using the transistor can make intracellular recording of cardiac muscle cells and used to identify change in action potential on treating ion channel blockers. It also provides information on the use of nanowire transistors in the measurement of excitatory and inhibitory postsynaptic potentials.