Immunohistological and Ultrastructural Study of the Inflammatory Response to Perforated Polyimide Cortical Implants: Mechanisms Underlying Deterioration of Electrophysiological Recording Quality.

The deterioration of field potential (FP) recording quality and yield by in vivo multielectrode arrays (MEA) within days to weeks of implantation severely limits progress in basic and applied brain research. The prevailing hypothesis is that implantation of MEA platforms initiate and perpetuate inflammatory processes which culminate in the formation of scar tissue (the foreign body response, FBR) around the implant. The FBR leads to progressive degradation of the recording qualities by displacing neurons away from the electrode surfaces, increasing the resistance between neurons (current source) and the sensing pads and by reducing the neurons' excitable membrane properties and functional synaptic connectivity through the release of pro-inflammatory cytokines. Meticulous attempts to causally relate the cellular composition, cell density, and electrical properties of the FBR have failed to unequivocally correlate the deterioration of recording quality with the histological severity of the FBR. Based on confocal and electron microscope analysis of thin sections of polyimide based MEA implants along with the surrounding brain tissue at different points in time after implantation, we propose that abrupt FP amplitude attenuation occurs at the implant/brain-parenchyma junction as a result of high seal resistance insulation formed by adhering microglia to the implant surfaces. In contrast to the prevailing hypothesis, that FP decrease occurs across the encapsulating scar of the implanted MEA, this mechanism potentially explains why no correlations have been found between the dimensions and density of the FBR and the recording quality. Recognizing that the seal resistance formed by adhering-microglia to the implant constitutes a downstream element undermining extracellular FP recordings, suggests that approaches to mitigate the formation of the insulating glial could lead to improved recording quality and yield.

Multisite Intracellular Recordings by MEA.

The enormous advances made over the last 50 years in materials science, microelectronics, and nanoelectronics, together with the acknowledgment that substrate-integrated planar multielectrode arrays (MEA) are limited to recording of extracellular field potentials (FPs) rather than the entire electrophysiological signaling repertoire of the brain, have prompted a number of laboratories to merge the advantages of planar MEA technologies (non-damaging and durable) with those of the classical sharp and patch electrodes for intracellular recordings. Unlike extracellular planar electrode-based MEAs, the new generation of three-dimensional (3D) vertical nanoelectrodes are designed to functionally penetrate the plasma membrane of cultured cells and operate in a similar manner to classical intracellular microelectrodes. Although only approximately 10 years has elapsed since the development of the first vertical 3D nanostructure-based MEAs, this technology has progressed to enable recordings of attenuated intracellular action potentials (APs) and synaptic potentials from individual neurons, cardiomyocytes, and striated myotubes. Furthermore, recently the scaling advantages of nanochip/microchip fabrication technologies enabled simultaneously intracellular recordings of APs from hundreds of cultured cardiomyocytes, thus heralding a new milestone in MEA technology.In this chapter we present the earliest and today's cutting-edge achievements of this "young vertical nano-sensors MEA technology" at the single-cell and network levels, explain the biophysical principles and the various configurations used to form functional nanoelectrode/cell hybrids, and describe the quality and characteristic features of the recorded intracellular APs and subthreshold synaptic potentials by the vertical nanoelectrode-based MEA. Basic cell-biological mechanisms that curtail the length of time intracellular access by the nanoelectrodes are discussed, and approaches to overcome this problem are offered.Recent development of biotechnologies that use induced human pluripotent stem cells taken from healthy subjects and patients, and in vitro drug screening for the development of personalized medicine as well as basic brain research will benefit tremendously from the use of MEAs that record the entire brain electrophysiological signaling repertoire from individual cells within an operational network rather than only extracellular FPs.

Multisite Attenuated Intracellular Recordings by Extracellular Multielectrode Arrays, a Perspective.

Multielectrode arrays (MEA) are used extensively for basic and applied electrophysiological research of neuronal- and cardiomyocyte-networks. Whereas immense progress has been made in realizing sophisticated MEA platforms of thousands of addressable, high-density, small diameter, low impedance sensors, the quality of the interfaces formed between excitable cells and classical planar sensor has not improved. As a consequence in vitro and in vivo MEA are "blind" to the rich and important "landscape" of sub-threshold synaptic potentials generated by individual neurons. Disregarding this essential fraction of network signaling repertoire has become the standard and almost the "scientific ideology" of MEA users. To overcome the inherent limitations of substrate integrated planar MEA platforms that only record extracellular field potentials, a number of laboratories have developed in vitro MEA for intracellular recordings. Most of these novel devices use vertical nano-rods or nano-wires that penetrate the plasma membrane of cultured cells and record the electrophysiological signaling in a manner similar to sharp intracellular microelectrodes. In parallel, our laboratory began to develop a bioinspired approach in-which cell biological energy resources are harnessed to self-force a cell into intimate contact with extracellular gold mushroom-shaped microelectrodes to record attenuated synaptic- and action-potentials with characteristic features of intracellular recordings. Here we describe some of the experiments that helped evolve the approach and elaborate on the biophysical principles that make it possible to record intracellular potentials by an array of extracellular electrode. We illustrate the qualities and limitations of the method and discuss prospects for further improvement of this technology.

Impedimetric monitoring of IGF-1 protection of in vitro cortical neurons under ischemic conditions.

Microelectrode arrays (MEAs) incorporated with the electric cell substrate impedance sensing (ECIS) technique provide a method for acquiring cellular electrophysiological information, which is useful for the time-course monitoring of cellular outgrowth and damage. This research utilizes the ECIS technique for monitoring the time-course impedimetric changes in normal and insulin-like growth factor 1 (IGF-1)-protected cortical neurons under the ischemic insult of oxygen glucose deprivation (OGD) created in a microperfusion environment. The neuronal apoptosis is reflected by the relatively low cell viability (28 ± 11.5 %) after 30-min OGD followed by 24 h of re-oxygenation. Also the hyperpolarization phase of mitochondrial membrane potential (MMP) occurs during 2 h of the re-oxygenation period. In contrast, cortical neurons treated with 50 and 100 ng/mL IGF-1 show higher survival rates of 45 ± 5.2 % and 49 ± 9.2 %, respectively, and no occurrence of the hyperpolarization of MMP during the re-oxygenation period. The ECIS results demonstrate that the measured impedance of cortical neurons decreased from 826 ± 86 kΩ to 224 ± 32 kΩ due to cell detachment under the insult of OGD. The measured impedance of IGF-1-protected cortical neurons slowly decreased to about 50 % of the original value (560 ± 45 kΩ for 50 ng/mL and 593 ± 44 kΩ for 100 ng/mL) compared to saline control of 232 ± 37 kΩ, which indicates improved cell adhesion under OGD conditions. The time-course impedimetric results show that the proposed ECIS-based MEAs platform incorporated with a microperfusion environment can be used for the real-time monitoring of cortical neurons under in vitro OGD and the IGF-1 protective effect against OGD-induced ischemic neuronal death.

Electrophysiologic studies of neronal activities under ischemia condition.

Substrate with integrated microelectrode arrays (MEAs) provides an alternative electrophysiological method. With MEAS, one can measure the impedance and elicit electrical stimulation from multiple sites of MEAs to determine the electrophysiological conditions of cells. The aims of this research were to construct an impedance and action potential measurement system for neurons cultured on MEAs for observing the electrophysiological signal transmission in neuronal network during glucose and oxygen deprivation (OGD). An extracellular stimulator producing the biphasic micro-current pulse for neuron stimulation was built in this study. From the time-course recording of impedance, OGD condition effectively induced damage in neurons in vitro. It is known that the results of cell stimulation are affected by electrode impedance, so does the result of neuron cells covered on the electrode can measure the sealing resistance. For extracellular stimulation study, cortical neuronal activity was recorded and the suitable stimulation window was determined. However, the stimulation results were affected by electrode impedance as well as sealing impedance resulting from neuron cells covering the electrode. Further development of surface modification for cultured neuron network should provide a better way for in vitro impedance and electrophysiological measurements.