In vivo and in vitro electrochemical impedance spectroscopy analysis of acute and chronic intracranial electrodes

Electrochemical impedance spectroscopy- This technique involves measurement of an electrochemical interface by applying a sinusoidal voltage perturbation across a range of frequencies. For potentiostatic modulation, a single perturbation amplitude is maintained across the frequency spectrum, typically lower than amplitudes used in biological stimulation. The frequency range chosen for these measurements was 10 Hz-100 kHz, with an amplitude of 0.01 V. Data were sampled at 39 frequency intervals across this spectrum, or 9.5/dec. Using the complex impedance values obtained by our instrument, a fitted model was created to approximate the components of the biotic-abiotic interface. Data collection- Data were primarily collected using a Palmsens4 Potentiostat instrument in Electrochemical Impedance Spectroscopy mode, both in vitro and in vivo in non-human primates (NHP). NHP subjects included two adult male rhesus macaque monkeys chronically implanted with both penetrating deep brain stimulation (DBS) electrodes and surface-contact electrocorticography (ECoG) electrodes placed epidurally. Measurements in vivo were performed only in 2-electrode mode, using a titanium screw or rod mounted to each animal’s skullcap as a return electrode. For in vitro measurements, a 2-electrode system was employed for direct comparison to typical in vivo data collection methods. A 3-electrode system including an Ag/AgCl reference electrode was also used in vitro to verify the reliability of the 2-electrode system. The Palmsens device was operated using battery power to isolate it from electrical line noise. Sample fluids used in vitro included phosphate-buffered saline (PBS) at 1x concentration and artificial cerebrospinal fluid (aCSF). Sample fluid temperature was regulated to approximate biological conditions, specifically NHP internal body temperature of approximately 37° C. Temperature regulation was performed using a Gamry electrochemical flow-cell connected to a thermally regulated bath, with measured temperature verified to within plus or minus 0.1°C. Working electrodes included a selection of commercially available intracranial electrodes, designed for both human and NHP applications. Electrodes used for in vivo measurements were chronically implanted, and connection to the Palmsens device was achieved via an external connector. The counter electrode used in vivo consisted of titanium screw for Animal 1 and a titanium rod for Animal 2, with the same approximate surface area. Counter electrodes used in vitro included a titanium screw similar to that used in vivo as well as platinum wire (7.5 cm length/0.5 mm diameter) and platinum foil (625 mm2). When a 3-electrode measurement system was applied, the “reference” consisted of a silver-silver chloride (Ag/AgCl) electrode, as is typically employed in non-biological EIS applications. For comparison to standard methods of electrode impedance verification, measurements were also performed using a Bak Electrode Impedance Tester calibrated with a 1 kOhm resistor. In this setup, impedance was computed as a function of two contacts: a working electrode and a counter electrode. Setup and solution preparation- Gray and white matter have measured conductivities of 0.33 and 0.142 S/m. The required salt concentration to mimic gray and white matter solutions was found by measuring the conductivity of dissolved salt at intervals of 0.5 g/L at 37°C +/- 0.1°C until the measured conductivity was greater than 0.33 S/m. Laboratory-grade salt (Thermo Fischer) was added between intervals, mixed using a vortex at 300 rpm until clear. A line fit to these conductivity values was used to find salt concentrations that approximated the conductivity of gray and white matter. The aCSF was made using 12.6 mM NaCl, 3mM KCl, 26 mM NaHCO3, 1.4 mM NaH2PO4, 10mM Glucose, 2 mM CaCl2, and 2 mM MgSO4, osmolarity of 306 mOsm/L, and pH of 7.36. For in vitro recordings, electrodes were suspended in solution with all contacts fully immersed in electrolyte and not touching glass side walls. EIS was measured using the Bak Electrode Impedance Tester and the PalmSens device. The PalmSens device delivered 10 Hz-10kHz, 0.1 nA-100 mAmps, 3 or 10 cycles each. The electrodes included in this solution comparison were AdTech Depth, AdTech Strip, AdTech Grid, Dixi Depth, Neuropace Depth, Neuropace Strip, and AdTech Behnke-Fried. To facilitate experimental determination of the stochastic error structure, trials were repeated 3x per electrode. For in vivo recordings, the exposed distal end of the working electrode was accessed, and a titanium screw mounted in the NHP skullcap served as a combined counter/reference electrode. A Palmsens MUX8/R2 Multiplexer facilitated switching between channels of the working electrode, with three scans performed per channel to facilitate error regression.