High-Aspect-Ratio Nanoelectrodes Enable Long-Term Recordings of Neuronal Signals with Subthreshold Resolution.

The further development of neurochips requires high-density and high-resolution recordings that also allow neuronal signals to be observed over a long period of time. Expanding fields of network neuroscience and neuromorphic engineering demand the multiparallel and direct estimations of synaptic weights, and the key objective is to construct a device that also records subthreshold events. Recently, 3D nanostructures with a high aspect ratio have become a particularly suitable interface between neurons and electronic devices, since the excellent mechanical coupling to the neuronal cell membrane allows very high signal-to-noise ratio recordings. In the light of an increasing demand for a stable, noninvasive and long-term recording at subthreshold resolution, a combination of vertical nanostraws with nanocavities is presented. These structures provide a spontaneous tight coupling with rat cortical neurons, resulting in high amplitude sensitivity and postsynaptic resolution capability, as directly confirmed by combined patch-clamp and microelectrode array measurements.

High Aspect Ratio and Light-Sensitive Micropillars Based on a Semiconducting Polymer Optically Regulate Neuronal Growth.

Many nano- and microstructured devices capable of promoting neuronal growth and network formation have been previously investigated. In certain cases, topographical cues have been successfully complemented with external bias, by employing electrically conducting scaffolds. However, the use of optical stimulation with topographical cues was rarely addressed in this context, and the development of light-addressable platforms for modulating and guiding cellular growth and proliferation remains almost completely unexplored. Here, we develop high aspect ratio micropillars based on a prototype semiconducting polymer, regioregular poly(3-hexylthiophene-2,5-diyl) (P3HT), as an optically active, three-dimensional platform for embryonic cortical neurons. P3HT micropillars provide a mechanically compliant environment and allow a close contact with neuronal cells. The combined action of nano/microtopography and visible light excitation leads to effective optical modulation of neuronal growth and orientation. Embryonic neurons cultured on polymer pillars show a clear polarization effect and, upon exposure to optical excitation, a significant increase in both neurite and axon length. The biocompatible, microstructured, and light-sensitive platform developed here opens up the opportunity to optically regulate neuronal growth in a wireless, repeatable, and spatio-temporally controlled manner without genetic modification. This approach may be extended to other cell models, thus uncovering interesting applications of photonic devices in regenerative medicine.

Mechanical and Electronic Cell-Chip Interaction of APTES-Functionalized Neuroelectronic Interfaces.

In this work, we analyze the impact of a chip coating with a self-assembled monolayer (SAM) of (3-aminopropyl)triethoxysilane (APTES) on the electronic and mechanical properties of neuroelectronic interfaces. We show that the large signal transfer, which has been observed for these interfaces, is most likely a consequence of the strong mechanical coupling between cells and substrate. On the one hand, we demonstrate that the impedance of the interface between Pt electrodes and an electrolyte is slightly reduced by the APTES SAM. However, this reduction of approximately 13% is definitely not sufficient to explain the large signal transfer of APTES coated electrodes demonstrated previously. On the other hand, the APTES coating leads to a stronger mechanical clamping of the cells, which is visible in microscopic images of the cell development of APTES-coated substrates. This stronger mechanical interaction is most likely caused by the positively charged amino functional group of the APTES SAM. It seems to lead to a smaller cleft between substrate and cells and, thus, to reduced losses of the cell's action potential signal at the electrode. The disadvantage of this tight binding of the cells to the rigid, planar substrate seems to be the short lifetime of the cells. In our case the density of living cells starts to decrease together with the visual deformation of the cells typically at DIV 9. Solutions for this problem might be the use of soft substrates and/or the replacement of the short APTES molecules with larger molecules or molecular multilayers.

Engineering Biocompatible Interfaces via Combinations of Oxide Films and Organic Self-Assembled Monolayers.

In this paper, we demonstrate that cell adhesion and neuron maturation can be guided by patterned oxide surfaces functionalized with organic molecular layers. It is shown that the difference in the surface potential of various oxides (SiO2, Ta2O5, TiO2, and Al2O3) can be increased by functionalization with a silane, (3-aminopropyl)-triethoxysilane (APTES), which is deposited from the gas phase on the oxide. Furthermore, it seems that only physisorbed layers (no chemical binding) can be achieved for some oxides (Ta2O5 and TiO2), whereas self-assembled monolayers (SAM) form on other oxides (SiO2 and Al2O3). This does not only alter the surface potential but also affects the neuronal cell growth. The already high cell density on SiO2 is increased further by the chemically bound APTES SAM, whereas the already low cell density on Ta2O5 is even further reduced by the physisorbed APTES layer. As a result, the cell density is ∼8 times greater on SiO2 compared to Ta2O5, both coated with APTES. Furthermore, neurons form the typical networks on SiO2, whereas they tend to cluster to form neurospheres on Ta2O5. Using lithographically patterned Ta2O5 layers on SiO2 substrates functionalized with APTES, the guided growth can be transferred to complex patterns. Cell cultures and molecular layers can easily be removed, and the cell experiment can be repeated after functionalization of the patterned oxide surface with APTES. Thus, the combination of APTES-functionalized patterned oxides might offer a promising way of achieving guided neuronal growth on robust and reusable substrates.

Surface Functionalization of Platinum Electrodes with APTES for Bioelectronic Applications.

The interface between electronic components and biological objects plays a crucial role in the success of bioelectronic devices. Since the electronics typically include different elements such as an insulating substrate in combination with conducting electrodes, an important issue of bioelectronics involves tailoring and optimizing the interface for any envisioned applications. In this paper, we present a method for functionalizing insulating substrates (SiO2) and metallic electrodes (Pt) simultaneously with a stable monolayer of organic molecules ((3-aminopropyl)triethoxysilane (APTES)). This monolayer is characterized by high molecule density, long-term stability, and positive surface net charge and most likely represents a self-assembled monolayer (SAM). It facilitates the conversion of biounfriendly Pt surfaces into biocompatible surfaces, which allows cell growth (neurons) on both functionalized components, SiO2 and Pt, which is comparable to that of reference samples coated with poly-L-lysine (PLL). Moreover, the functionalization greatly improves the electronic cell-chip coupling, thereby enabling the recording of action potential signals of several millivolts at APTES-functionalized Pt electrodes.

Composite Lipid Bilayers from Cell Membrane Extracts and Artificial Mixes as a Cell Culture Platform.

An artificial lipid bilayer is the closest possible model for the cell membrane. Despite that, current methods of lipid bilayer assembly and functionalization do not provide a satisfactory mimic of the cell-cell contact due to the inability to recreate an asymmetrical multicomponent system. In the current work, a method to produce an integrated solid-supported lipid bilayer combining natural extracts from cell membranes and artificially made lipid vesicles is proposed. This simple method allows delivery of transmembrane proteins and components of the extracellular matrix into the substrate. Biocompatibility of the composite natural/artificial lipid bilayers is evaluated by their interactions with the cardiomyocyte-like HL-1 cell line. Compared with fully artificial mixes, composite lipid bilayers allow cells to adhere and develop a morphologically more normal cytoskeleton.