Post-explant profiling of subcellular-scale carbon fiber intracortical electrodes and surrounding neurons enables modeling of recorded electrophysiology.

Objective.Characterizing the relationship between neuron spiking and the signals that electrodes record is vital to defining the neural circuits driving brain function and informing clinical brain-machine interface design. However, high electrode biocompatibility and precisely localizing neurons around the electrodes are critical to defining this relationship.Approach.Here, we demonstrate consistent localization of the recording site tips of subcellular-scale (6.8µm diameter) carbon fiber electrodes and the positions of surrounding neurons. We implanted male rats with carbon fiber electrode arrays for 6 or 12+ weeks targeting layer V motor cortex. After explanting the arrays, we immunostained the implant site and localized putative recording site tips with subcellular-cellular resolution. We then 3D segmented neuron somata within a 50µm radius from implanted tips to measure neuron positions and health and compare to healthy cortex with symmetric stereotaxic coordinates.Main results.Immunostaining of astrocyte, microglia, and neuron markers confirmed that overall tissue health was indicative of high biocompatibility near the tips. While neurons near implanted carbon fibers were stretched, their number and distribution were similar to hypothetical fibers placed in healthy contralateral brain. Such similar neuron distributions suggest that these minimally invasive electrodes demonstrate the potential to sample naturalistic neural populations. This motivated the prediction of spikes produced by nearby neurons using a simple point source model fit using recorded electrophysiology and the mean positions of the nearest neurons observed in histology. Comparing spike amplitudes suggests that the radius at which single units can be distinguished from others is near the fourth closest neuron (30.7 ± 4.6µm,X-± S) in layer V motor cortex.Significance.Collectively, these data and simulations provide the first direct evidence that neuron placement in the immediate vicinity of the recording site influences how many spike clusters can be reliably identified by spike sorting.

Carbon fiber electrodes for intracellular recording and stimulation.

Objective.To understand neural circuit dynamics, it is critical to manipulate and record many individual neurons. Traditional recording methods, such as glass microelectrodes, can only control a small number of neurons. More recently, devices with high electrode density have been developed, but few of them can be used for intracellular recording or stimulation in intact nervous systems. Carbon fiber electrodes (CFEs) are 8µm-diameter electrodes that can be assembled into dense arrays (pitches ⩾ 80µm). They have good signal-to-noise ratios (SNRs) and provide stable extracellular recordings both acutely and chronically in neural tissuein vivo(e.g. rat motor cortex). The small fiber size suggests that arrays could be used for intracellular stimulation.Approach.We tested CFEs for intracellular stimulation using the large identified and electrically compact neurons of the marine molluskAplysia californica. Neuron cell bodies inAplysiarange from 30µm to over 250µm. We compared the efficacy of CFEs to glass microelectrodes by impaling the same neuron's cell body with both electrodes and connecting them to a DC coupled amplifier.Main results.We observed that intracellular waveforms were essentially identical, but the amplitude and SNR in the CFE were lower than in the glass microelectrode. CFE arrays could record from 3 to 8 neurons simultaneously for many hours, and many of these recordings were intracellular, as shown by simultaneous glass microelectrode recordings. CFEs coated with platinum-iridium could stimulate and had stable impedances over many hours. CFEs not within neurons could record local extracellular activity. Despite the lower SNR, the CFEs could record synaptic potentials. CFEs were less sensitive to mechanical perturbations than glass microelectrodes.Significance.The ability to do stable multi-channel recording while stimulating and recording intracellularly make CFEs a powerful new technology for studying neural circuit dynamics.

Compositional and morphological properties of platinum-iridium electrodeposited on carbon fiber microelectrodes.

Objective. Neural interfaces based on carbon fiber (CF) electrodes have demonstrated key positive attributes such as minimal foreign body response and mechanical strength to self-insert in brain tissue. However, carbon does not form a low impedance electrode interface with neural tissue. Electrodeposited platinum iridium (PtIr) has been used to improve electrode interface properties for metallic bioelectrodes.Approach. In this study, a PtIr electrodeposition process has been performed on CF microelectrode arrays to improve the interfacial properties of these arrays. We study the film morphology and composition as well as electrode durability and impedance.Results. A PtIr coating with a composition of 70% Pt, 30% Ir and a thickness of ∼400 nm was observed. Pt and Ir were evenly distributed within the film. Impedance was decreased by 89% @ 1 kHz. Accelerated soak testing in a heated (T= 50∘C) saline solution showed impedance increase (@ 1 kHz) of ∼12% after 36 days (89 equivalent) of soaking.

Ultra-small carbon fiber electrode recording site optimization and improved in vivo chronic recording yield.

OBJECTIVE: Carbon fiber electrodes may enable better long-term brain implants, minimizing the tissue response commonly seen with silicon-based electrodes. The small diameter fiber may enable high-channel count brain-machine interfaces capable of reproducing dexterous movements. Past carbon fiber electrodes exhibited both high fidelity single unit recordings and a healthy neuronal population immediately adjacent to the recording site. However, the recording yield of our carbon fiber arrays chronically implanted in the brain typically hovered around 30%, for previously unknown reasons. In this paper we investigated fabrication process modifications aimed at increasing recording yield and longevity. APPROACH: We tested a new cutting method using a 532nm laser against traditional scissor methods for the creation of the electrode recording site. We verified the efficacy of improved recording sites with impedance measurements and in vivo array recording yield. Additionally, we tested potentially longer-lasting coating alternatives to PEDOT:pTS, including PtIr and oxygen plasma etching. New coatings were evaluated with accelerated soak testing and acute recording. MAIN RESULTS: We found that the laser created a consistent, sustainable 257 ± 13.8 µm2 electrode with low 1 kHz impedance (19 ± 4 kΩ with PEDOT:pTS) and low fiber-to-fiber variability. The PEDOT:pTS coated laser cut fibers were found to have high recording yield in acute (97% > 100 µV pp , N = 34 fibers) and chronic (84% > 100 µV pp , day 7; 71% > 100 µV pp , day 63, N = 45 fibers) settings. The laser cut recording sites were good platforms for the PtIr coating and oxygen plasma etching, slowing the increase in 1 kHz impedance compared to PEDOT:pTS in an accelerated soak test. SIGNIFICANCE: We have found that laser cut carbon fibers have a high recording yield that can be maintained for over two months in vivo and that alternative coatings perform better than PEDOT:pTS in accelerated aging tests. This work provides evidence to support carbon fiber arrays as a viable approach to high-density, clinically-feasible brain-machine interfaces.

Nanosecond pulsed electric signals can affect electrostatic environment of proteins below the threshold of conformational effects: The case study of SOD1 with a molecular simulation study.

Electric fields can be a powerful tool to interact with enzymes or proteins, with an intriguing perspective to allow protein manipulation. Recently, researchers have focused the interest on intracellular enzyme modifications triggered by the application of nanosecond pulsed electric fields. These findings were also supported by theoretical predictions from molecular dynamics simulations focussing on significant variations in protein secondary structures. In this work, a theoretical study utilizing molecular dynamics simulations is proposed to explore effects of electric fields of high intensity and very short nanosecond duration applied to the superoxide dismutase (Cu/Zn-SOD or SOD-1), an important enzyme involved in the cellular antioxidant defence mechanism. The effects of 100-nanosecond pulsed electric fields, with intensities ranging from 108 to 7x108 V/m, on a single SOD1 enzyme are presented. We demonstrated that the lowest intensity of 108 V/m, although not inducing structural changes, can produce electrostatic modifications on the reaction centre of the enzyme, as apparent from the dipolar response and the electric field distribution of the protein active site. Electric pulses above 5x108 V/m produced a fast transition between the folded and a partially denatured state, as inferred by the secondary structures analysis. Finally, for the highest field intensity used (7x108 V/m), a not reversible transition toward an unfolded state was observed.

Feasibility of Drug Delivery Mediated by Ultra-Short and Intense Pulsed Electric Fields.

The increasing interest towards biocompatible nanotechnologies in medicine, combined with electric fields stimulation, is leading to the development of electro-sensitive smart systems for drug delivery applications. Common examples of electro-sensitive materials include phospholipids that can be used to design nano-sized vesicles suitable for external electric actuation. To this regard, recently the use of pulsed electric fields to trigger release across phospholipid membranes has been numerically studied, for a deeper understanding of the phenomena at the molecular scale. Aim of this work is to give an experimental validation of the feasibility of controlling drug release from liposomes mediated by nanosecond pulsed electric fields.

Can Pulsed Electromagnetic Fields Trigger On-Demand Drug Release from High-Tm Magnetoliposomes?

Recently, magnetic nanoparticles (MNPs) have been used to trigger drug release from magnetoliposomes through a magneto-nanomechanical approach, where the mechanical actuation of the MNPs is used to enhance the membrane permeability. This result can be effectively achieved with low intensity non-thermal alternating magnetic field (AMF), which, however, found rare clinic application. Therefore, a different modality of generating non-thermal magnetic fields has now been investigated. Specifically, the ability of the intermittent signals generated by non-thermal pulsed electromagnetic fields (PEMFS) were used to verify if, once applied to high-transition temperature magnetoliposomes (high-Tm MLs), they could be able to efficiently trigger the release of a hydrophilic model drug. To this end, hydrophilic MNPs were combined with hydrogenated soybean phosphatidylcholine and cholesterol to design high-Tm MLs. The release of a dye was evaluated under the effect of PEMFs for different times. The MNPs motions produced by PEMF could effectively increase the bilayer permeability, without affecting the liposomes integrity and resulted in nearly 20% of release after 3 h exposure. Therefore, the current contribution provides an exciting proof-of-concept for the ability of PEMFS to trigger drug release, considering that PEMFS find already application in therapy due to their anti-inflammatory effects.

Technological and Theoretical Aspects for Testing Electroporation on Liposomes.

Recently, the use of nanometer liposomes as nanocarriers in drug delivery systems mediated by nanoelectroporation has been proposed. This technique takes advantage of the possibility of simultaneously electroporating liposomes and cell membrane with 10-nanosecond pulsed electric fields (nsPEF) facilitating the release of the drug from the liposomes and at the same time its uptake by the cells. In this paper the design and characterization of a 10 nsPEF exposure system is presented, for liposomes electroporation purposes. The design and the characterization of the applicator have been carried out choosing an electroporation cuvette with 1 mm gap between the electrodes. The structure efficiency has been evaluated at different experimental conditions by changing the solution conductivity from 0.25 to 1.6 S/m. With the aim to analyze the influence of device performances on the liposomes electroporation, microdosimetric simulations have been performed considering liposomes of 200 and 400 nm of dimension with different inner and outer conductivity (from 0.05 to 1.6 S/m) in order to identify the voltage needed for their poration.

Exploring the Applicability of Nano-Poration for Remote Control in Smart Drug Delivery Systems.

Smart drug delivery systems represent an interesting tool to significantly improve the efficiency and the precision in the treatment of a broad category of diseases. In this context, a drug delivery mediated by nanosecond pulsed electric fields seems a promising technique, allowing for a controlled release and uptake of drugs by the synergy between the electropulsation and nanocarriers with encapsulated drugs. The main concern about the use of electroporation for drug delivery applications is the difference in dimension between the liposome (nanometer range) and the cell (micrometer range). The choice of liposome dimension is not trivial. Liposomes larger than 500 nm of diameter could be recognized as pathogen agents by the immune system, while liposomes of smaller size would require external electric field of high amplitudes for the membrane electroporation that could compromise the cell viability. The aim of this work is to theoretically study the possibility of a simultaneous cell and liposomes electroporation. The numerical simulations reported the possibility to electroporate the cell and a significant percentage of liposomes with comparable values of external electric field, when a 12 nsPEF is used.

Polymeric nanoparticles for hemoglobin-based oxygen carriers.

This article reports on the current status of the research on blood substitutes with particular attention on hemoglobin-based oxygen carriers (HBOCs). Insights on the physiological role of hemoglobin are reported in the view of the development of both acellular and cellular hemoglobin-based oxygen carriers. Attention is then focused on biocompatible polymeric materials that find application as matrices for cellular based HBOCs and on the strategies employed to avoid methemoglobin formation. Results are reported regarding the use of bioerodible polymeric matrices based on hemiesters of alternating copolymer (maleic anhydride-co-butyl vinyl ether) for the preparation of hemoglobin loaded nanoparticles.