Yoonkey Nam

Gold-coated microelectrode array with thiol linked self-assembled monolayers for engineering neuronal cultures

We report the use of a gold coating on microelectrode arrays (MEAs) to enable the use of the relatively reliable surface modification chemistry afforded by alkanethiol self-assembled monolayers (SAMs). The concept is simple and begins with planar MEAs, which are commercially available for neuronal cell culture and for brain slice studies. A gold film, with an intermediate adhesive layer of titanium, is deposited over the insulation of an existing MEA in a manner so as to be thin enough for transmission light microscopy as well as to avoid electrical contact to the electrodes. The alkanethiol-based linking chemistry is then applied for the desired experimental purpose. Here we show that polylysine linked to alkanethiol SAM can control the geometry of an in vitro hippocampal neuronal network grown on the MEA. Furthermore, recordings of neuronal action potentials from random and patterned networks suggest that the gold coating does not significantly alter the electrode properties. This design scheme may be useful for increasing the number of neurons located in close proximity to the electrodes. Realization of in vitro neuronal circuits on MEAs may significantly benefit basic neuroscience studies, as well as provide the insight relevant to applications such as neural prostheses or cell-based biosensors. The gold coating technique makes it possible to use the rich set of thiol-based surface modification techniques in combination with MEA recording.

Stimulus-Artifact Elimination in a Multi-Electrode System

To fully exploit the recording capabilities provided by current and future generations of multi-electrode arrays, some means to eliminate the residual charge and subsequent artifacts generated by stimulation protocols is required. Custom electronics can be used to achieve such goals, and by making them scalable, a large number of electrodes can be accessed in an experiment. In this work, we present a system built around a custom 16-channel IC that can stimulate and record, within 3 ms of the stimulus, on the stimulating channel, and within 500 mus on adjacent channels. This effectiveness is achieved by directly discharging the electrode through a novel feedback scheme, and by shaping such feedback to optimize electrode behavior. We characterize the different features of the system that makes such performance possible and present biological data that show the system in operation. To enable this characterization, we present a framework for measuring, classifying, and understanding the multiple sources of stimulus artifacts. This framework facilitates comparisons between artifact elimination methodologies and enables future artifact studies.

Surface-modified microelectrode array with flake nanostructure for neural recording and stimulation.

A novel microelectrode modification method is reported for neural electrode engineering with a flake nanostructure (nanoflake). The nanoflake-modified electrodes are fabricated by combining conventional lithography and electrochemical deposition to implement a microelectrode array (MEA) on a glass substrate. The unique geometrical properties of nanoflake sharp tips and valleys are studied by optical, electrochemical and electrical methods in order to verify the advantages of using nanoflakes for neural recording devices. The in vitro recording and stimulation of cultured hippocampal neurons are demonstrated on the nanoflake-modified MEA and the clear action potentials are observed due to the nanoflake impedance reduction effect.

In Vitro Developmental Acceleration of Hippocampal Neurons on Nanostructures of Self-Assembled Silica Beads in Filopodium-Size Ranges†

Abstract Topographical cues play an important role in in vitro neuronal development. In their Communication (DOI: 10.1002/anie.201106271), Y. Nam, I. S. Choi, J. S. Lee, and co-workers show that neuritogenetic acceleration occurs on silica-bead monolayers made up of beads with a diameter of more than 200 nm, but not on monolayers of beads with smaller diameters. The biochemical study indicates neurons sense topographical differences in nanostructures and alter their behavior accordingly.

Directional neurite growth using carbon nanotube patterned substrates as a biomimetic cue.

Researchers have made extensive efforts to mimic or reverse-engineer in vivo neural circuits using micropatterning technology. Various surface chemical cues or topographical structures have been proposed to design neuronal networks in vitro. In this paper, we propose a carbon nanotube (CNT)-based network engineering method which naturally mimics the structure of extracellular matrix (ECM). On CNT patterned substrates, poly-L-lysine (PLL) was coated, and E18 rat hippocampal neurons were cultured. In the early developmental stage, soma adhesion and neurite extension occurred in disregard of the surface CNT patterns. However, later the majority of neurites selectively grew along CNT patterns and extended further than other neurites that originally did not follow the patterns. Long-term cultured neuronal networks had a strong resemblance to the in vivo neural circuit structures. The selective guidance is possibly attributed to higher PLL adsorption on CNT patterns and the nanomesh structure of the CNT patterns. The results showed that CNT patterned substrates can be used as novel neuronal patterning substrates for in vitro neural engineering.

Gated Luminescence Imaging of Silicon Nanoparticles

The luminescence lifetime of nanocrystalline silicon is typically on the order of microseconds, significantly longer than the nanosecond lifetimes exhibited by fluorescent molecules naturally present in cells and tissues. Time-gated imaging, where the image is acquired at a time after termination of an excitation pulse, allows discrimination of a silicon nanoparticle probe from these endogenous signals. Because of the microsecond time scale for silicon emission, time-gated imaging is relatively simple to implement for this biocompatible and nontoxic probe. Here a time-gated system with ∼10 ns resolution is described, using an intensified CCD camera and pulsed LED or laser excitation sources. The method is demonstrated by tracking the fate of mesoporous silicon nanoparticles containing the tumor-targeting peptide iRGD, administered by retro-orbital injection into live mice. Imaging of such systemically administered nanoparticles in vivo is particularly challenging because of the low concentration of probe in the targeted tissues and relatively high background signals from tissue autofluorescence. Contrast improvements of >100-fold (relative to steady-state imaging) is demonstrated in the targeted tissues.

Electrochemically Driven, Electrode-Addressable Formation of Functionalized Polydopamine Films for Neural Interfaces†

The electrode-specific formation of polydopamine films is achieved by applying positive voltage to the target electrodes at pH 6.0. The functionalization of the films is simultaneously carried out by co-depositing dopamine with molecules of interest onto the electrode.

Beyond the SERS: Raman Enhancement of Small Molecules Using Nanofluidic Channels with Localized Surface Plasmon Resonance

This work was supported by KRIBB (Korea Research Institute of Bioscience & Biotechnology) and the National Research Foundation of Korea (NRF) grant funded by the Korea government (MEST) (No. 2010–0017693).

Pitch‐Dependent Acceleration of Neurite Outgrowth on Nanostructured Anodized Aluminum Oxide Substrates

Nervous systems are composed of microstructured scaffolds with three-dimensional nanofeatured textures. These textures enable the systems to give nanometer-scaled physical cues to the overlying cells, along with biochemical cues. However, the topographical effects on the neurons are still an unexplored territory, although there have been many reports on the biochemical cues for neuronal behavior. It is practically very difficult to investigate the topographical environments in vivo in the biological systems and/or to mimic them precisely in vitro. There is much recent evidence that the cellular response is affected by the physical properties of artificial materials. Studies with such materials could therefore provide us with new insight into the developmental processes of the brain and enable elucidation of the unexplored nanotopographical effects on neuronal behavior. The responses of nerve cells to surface roughness have been studied on various substrates, such as porous silicon, thin titanium nitride films, carbon nanotubes, topographically molded poly(dimethylsiloxane), silicon pillar arrays, gallium phosphide nanowires, aligned nanofiber arrays, and silicon nanowires. Previous studies showed that nerve cells exhibit enhanced attachment and viability as well as the axonal guidance effect on rough surfaces, as opposed to topographically flat surfaces. However, there have been few reports on how nanometer-scaled features regulate neuronal behavior in terms of neurite development. To generate nanotopographical stimuli to neurons in a controllable and systematic manner, it is necessary to make reproducible, rigid structures with variable topographical features. Among the methods for creating topographies on surfaces at the nanometer scale, the fabrication of anodized aluminum oxide (AAO) is highly effective, straightforward,

Neural recording and stimulation of dissociated hippocampal cultures using microfabricated three-dimensional tip electrode array.

There is increasing interest in interfacing dissociated neuronal cultures with planar multielectrode arrays (MEAs) for the study of the dynamics of neuronal networks. Here we report on the successful use of three-dimensional tip electrode arrays (3D MEAs), originally developed for use with brain slices, for recording and stimulation of cultured neurons. We observed that many neurons grew directly on protruding electrode surface, appearing to make excellent contact. A larger than usual portion of extracellular spikes had large positive peaks, while most of the spikes from conventional two-dimensional electrode arrays had large negative spikes. This may be due to the direct capacitive coupling situation provided by relatively large electrode surface area.