Nanocarbon-Iridium Oxide Nanostructured Hybrids as Large Charge Capacity Electrostimulation Electrodes for Neural Repair

Abstract

Nanostructuring nanocarbons with IrO_x yields to material coatings with large charge capacities for neural electrostimulation, and large reproducibility in time, that carbons do not exhibit. This work shows the contributions of carbon and the different nanostructures present, as well as the impact of functionalizing graphene with oxygen and nitrogen, and the effects of including conducting polymers within the hybrid materials. Different mammalian neural growth models differentiate the roles of the substrate material in absence and in presence of applied electric fields and address optimal electrodes for the future clinical applications.

Keywords: charge capacity; electrostimulation; iridium oxide; nanocarbons; repair.

Figure 1

(A–D) Time evolution of TEM images at about 10 min intervals, showing TEM images of dry drops of Iridium oxo solutions obtained from hydrolysis of IrCl₃, evolve under the electron microscope (120 KV Jeol). Last two images, (E,F) Diffraction rings obtained at this low resolution match those of quasiamorphous K_xIrO₂ and later metallic Ir. Thermal evolution has also been observed before in Ar atmosphere (in O₂ yielding IrO₂ rutile [21]) Images show two different time intervals. Global time in the order of minutes. (Original results).

Figure 2

Top: (A) Simultaneous cyclic voltammetry and ECQM mass changes during dynamic Electrodeposition of IrO_x involving mass deposition and K⁺ intercalation/deintercalation, (B) SEM lateral images of resulting coatings on Pt (12 nm) glass slides, (C) Macroscopic images of IrO_x deposited on Pt (12 nm)-Ti (5 nm)-glass substrates. Bottom (D) Typical electrodeposition Cyclic voltammetry of IrO_x-Nanocarbon hybrids (this case IrO_x-NGO) (E) macroscopic images showing that the first layers are mostly IrO_x and (F) SEM lateral images of the coatings. From ref. [24,29] Published with permission of Elsevier.

Figure 3

(A) Exfoliated graphite yielding pristine graphene and (B,C) adhesion of Ir oxoparticles adhered to electrochemically exfoliated graphene nanoparticles, eG ((A,B) HRTEM, (C) SEM). (Note that Iridium species evolve under the microscope).

Figure 4

SEM images of IrO_x-CNT (top A–C) and IrO_x-CNT-PEDOT hybrids (bottom D–F) [18,27]. With permission from Elsevier.

Figure 5

(A–D) SEM images for IrO_x-GO hybrid at various scales showing typical cracks and the millfeuille nanostructure [28]. Yellow big square shows a magnified vision of the zone at small square. With permission from Elsevier.

Figure 6

(A) SEM images of IrO_x-eG hybrid showing the millfeuille nanostructure. (inset: macro scale showing the large homogeneity of the coating). (B) Crack developed under the SEM electron beam with millfeuille ordering. (Inset showing secondary electrons image, where carbon is not seen vs iridium) [24]. With permission from Elsevier.

Figure 7

SEM images of IrO_x-NGO hybrid coatings for one of the IrO_x—NGO hybrid (NGO treated at 100 °C (A,D), 220 °C (B,E) and 300 °C (C,F) [29]. With permission from Elsevier.

Figure 8

(A) Cyclic voltammetries in biologically emulating sodium phosphate buffer of representative nanostructured IrO_x–graphene hybrids during electric field application in electrostimulation processes and (B) associated charge storage capacity changes during 1000 cycles. (C) Graphite hybrid, not shown sees the CSC decrease after a few cycles, to the IrO_x values. (D) Impedance comparison for several materials [34]. With permission from Elsevier.

Figure 9

CV showing the significant decrease in current when PEDOT is incorporated to the IrO_x-CNT hybrid. (IrO_x-CNT compared with IrO_x-CNT-PEDOT and with individual components). The lowest currents always correspond to the composites or hybrids containing PEDOT [18]. With permission from Elsevier.

Figure 10

Neuronal survival and functionality on IrO_x-eG. Representative fluorescent microphotographs of neurons growing on: (A) Borosilicate glass, (B) IrO_x, (C) IrOx-eG and (D) Platinum, stained against Tau (red) and NeuN (green) showing dendrites and living cell nucleii. Scale bar = 100 µm [24]. With permission from Elsevier.

Figure 11

Astrocyte influence on neuronal survival on IrO_x.nanocarbon nanostructured materials containing conducting polymer PEDOT with respect to IrO_x. (A) Dissociated neural cells were (seeded at 115,000 cells × cm⁻² on different materials coated with poly-L-lysine (white bars) or with a monolayer of confluent astrocytes (cross bars) and grown for 5 DIV. (B) Representative fluorescent photomicrographs of primary cultures of enriched neurons and of neuron-astrocyte co-culture growing on IrO_x and IrO_x-CNTPEDOT. Cells were processed for tau (green) and GFAP (red) immunocytochemistry to label neurons and astrocytes, respectively. Scale bar = 100 μm [18]. *** p < 0.001 vs. neurons grown on poly-L-lysine coated materials. With permission from Elsevier.

Figure 12

(A) Quantification of neural cell growth in absence and presence of EF with Q delivered below (80% of the total CSC) and above CSC (based on Tau imunostaining of scratched cortical neuron cultures showing spontaneous regeneration of surface covered by new neurites after scratching the neuronal monolayer at 5 DIV in each case) [34]. (B) Representative images of cell growth with no field and using (80% and 200% of the CSC value for the charge delivery). (B) Neurite and nuclei images for comparison (Cells were processed for Tau inmunocitochemistry and Bis-benzimide staining of the nuclei). With permission from Elsevier.

Figure 13

Spontaneous regeneration of an in vitro “wound like” scratch in neural cell cultures over IrO_x-eG hybrid. (A) Tau imunostaining of scratched cortical neuron cultures showing spontaneous regeneration of surface covered by new neurites after scratching the neuronal monolayer at 5 DIV. (B1) Neurite and (B2) nuclei images for comparison (Cells were processed for Tau inmunocitochemistry and Bis-benzimide staining of the nuclei). (C) Quantitative integration of the area covered by new neurites at different days after scratch. Results are mean ± sem (n = 5). * p < 0.05; ** p < 0.01, *** p < 0.001 vs. t = 0 after significant one-way ANOVA. Electric field application would be carried out in a zone where a more drastic change is seen for growth (d2, marked in the red arrow) [34].

Figure 14

Scratch model repair under the application of an electric field using IrO_x-graphene as substrate, (A) cin cathode and (B) anode areas. The firs image in each set corresponds to the reference spontaneous scratch growth using polypyrrole surface on PEDOT with lysine counterions. (Note that cells are near polypyrrole and not PEDOT in the polymer bilayer). Scale bars are equal for all images. [22] B-lys is a PEDOT-Polypyrrole bilayer with lysine counterion, IrO_x-eG is the hybrid described in this work. With permission from Elsevier.

Figure 15

(A) Quantification of scratch repair (as area reoccupied by neurites) for various pairs of electrode materials, using symmetrical cells (equal electrodes), or reducing first the electrode that will act as anode, as compared with spontaneous repair [34]. Results are mean ± sem. * p < 0.05 and ** p < 0.01 after significant one-way ANOVA (p < 0.05; F3,15 = 3.486) and post-comparison tests. (B) Quantification of scratch repair (as area occupied), as compared with the use of Pt electrodes. With permission from Elsevier.