Bioelectronic medicine, using electricity to target neuronal circuits and modulate neural communication, offers a promising therapeutic approach for various neuronal disorders and dysfunctions. While cortical interfaces, sensory and motor prostheses, and vagus nerve interfaces have been extensively researched, fewer approaches exist for addressing autonomic nerve fibers of the enteric nervous system, particularly those regulating digestive and metabolic processes. These small pelvic nerves, with their complex branched structures, are involved in peristalsis and other vital functions. Dysfunction in these circuits leads to chronic diseases like constipation and urinary dysfunction, which are often treated with medication or surgery without addressing the root cause. Effective treatment requires identifying and mapping pathological neuronal signaling chains and modulating them based on healthy signals in a closed-loop manner. The challenge lies in selectively recording signals from these small, submillimeter nerves surrounded by supportive layers (epineurium, perineurium, and endoneurium), which are mainly composed of unmyelinated C-fibers generating low-amplitude signals. Innovative, low-invasive neural interfaces with micrometer-sized electrodes are crucial for this purpose. Reducing electrode impedance is important to improve the signal-to-noise ratio (SNR) and allow for the measurement of lower signal amplitudes. This can be achieved by increasing the electrode size or increasing the active electrochemical surface area by coating it with materials like titanium nitride (TiN). Microscopic electrodes with refined surfaces increase spatial resolution and electrode density. The electrode-to-signal source distance also impacts SNR, with intraneural electrodes (inside the nerve) providing better selectivity and SNR compared to extraneural electrodes (outside the nerve). Existing intrafascicular electrodes like LIFE and TIME offer increased selectivity but are limited in their ability to address multiple fascicles. The 3D Utah slanted electrode array (USEA) addresses this with varying electrode heights, but its rigidity and electrode size limit its application to small-diameter autonomic nerves. Flexible polymer substrates are therefore needed to match the mechanical properties of the surrounding tissue. This research proposes a novel 3D MEA with protruding electrodes on a flexible substrate to improve signal recording from autonomic nerves.

The paper reviews existing neural interfaces for peripheral nerve stimulation and recording. It highlights the limitations of extraneural electrodes, such as cuff electrodes, which lack the selectivity to record single-unit activity. Intraneural electrodes like the Longitudinal Intrafascicular Electrode (LIFE) and Transverse Intrafascicular Multichannel Electrode (TIME) offer improved selectivity but are limited in the number of fascicles they can address. The authors discuss the Utah Slanted Electrode Array (USEA), a 3D approach with varying electrode heights, but note its rigidity and large electrode size as drawbacks for small autonomic nerves. The literature review establishes the need for a flexible, 3D MEA with small, low-impedance electrodes for improved recording from delicate autonomic nerves.

The study developed a flexible, 3D microelectrode array (MEA) with 32 cylindrical microelectrodes on a polyimide substrate. The fabrication process involved creating a 2D MEA with gold conductive tracks, then depositing cylindrical gold pillars via electroplating. A titanium/gold seed layer was sputtered and patterned using photoresist, forming a template for gold pillar electrodeposition. The process included steps for hydrophilization, cleaning via cyclic voltammetry, ultrasound treatment to remove air bubbles, and chronoamperometry for controlled gold deposition. Sidewall insulation was achieved with parylene C coating. Pillar head modification involved photoresist application, flood exposure and development to expose pillar heads, parylene C removal via reactive ion etching (RIE), gold wet etching to roughen the surface, and finally Ti/TiN sputter deposition and lift-off. Two electrode types were created: Type A (wet etched only) and Type B (wet etched and TiN coated). The impedance was characterized, and ex vivo testing was performed using mouse retinae to validate functionality. The MEA design incorporated a finger structure connector for encapsulation and connection to an external adapter, a flexible ribbon cable for mechanical decoupling, and oval holes to prevent air entrapment during implantation. The authors also describe the microfabrication process in detail. They utilized microelectromechanical systems (MEMS) processes and materials like polyimide, parylene C, and titanium nitride (TiN) to develop a flexible and biocompatible device. The fabrication process involves steps like thin-film deposition, photolithography, electroplating, reactive ion etching (RIE), and wet etching. The MEA design is optimized for the targeted application with a specific layout and dimensions to effectively interface with pelvic neural structures.

The fabricated 3D MEAs exhibited low impedances (around 7 kΩ at 1 kHz for a 50 µm diameter electrode with ~5000 µm² exposed surface area) and low intrinsic noise. Ex vivo testing with mouse retinae successfully recorded spontaneous neuronal spikes with amplitudes up to 66 µV, demonstrating the functionality of the device. The combined techniques of wet etching and TiN coating on the electrode tips proved effective in lowering impedance. The flexible design and small size of the electrodes make it suitable for recording from delicate structures. The study successfully demonstrates the feasibility of using a flexible, 3D MEA to record low-amplitude neural signals from tissue, indicating its suitability for applications in bioelectronic medicine and the study of neural circuits.The different fabrication processes, including electroplating, parylene C deposition, and TiN coating, were optimized to obtain the desired properties. The researchers characterized the impedance of the electrodes and found that the modified electrodes (type B) had significantly lower impedance values compared to the unmodified electrodes (type A). Furthermore, the results of the ex vivo experiments demonstrated that the flexible 3D MEA was capable of recording spontaneous neuronal spikes from mouse retinae.

The results demonstrate the successful fabrication and validation of a flexible, 3D protruding MEA capable of recording low-amplitude signals from delicate neural structures. The low impedance and high SNR achieved through the combination of electrode design and surface modifications are key advantages over existing technologies. The flexibility of the device is crucial for minimizing tissue damage and improving long-term stability in vivo. The successful ex vivo recordings in mouse retinae, a tissue with similar delicate structure to autonomic nerves, provide strong evidence for the applicability of the MEA to the intended target (pelvic nerves). This technology could enable significant advances in bioelectronic medicine, facilitating the study of neural circuits involved in digestive function and potentially leading to novel treatment strategies for related disorders. The device’s potential for chronic implantation allows for long-term monitoring and manipulation of nerve signals, which holds great promise for advanced diagnostics and therapy.

This study presents a novel fabrication method for a flexible 3D MEA with low-impedance microelectrodes suitable for recording low-amplitude signals from delicate autonomic nerves. The successful ex vivo recordings demonstrate the potential of this technology for applications in bioelectronic medicine, particularly for the study and treatment of digestive disorders. Future work should focus on in vivo studies to evaluate the long-term performance and biocompatibility of the MEA in animal models and eventually in human clinical trials. Further refinement of the MEA design and fabrication process is also warranted to optimize electrode density and spatial resolution.

The study currently only presents ex vivo results. In vivo testing in relevant animal models is needed to confirm the long-term performance and biocompatibility of the MEA. The long-term stability of the device within the tissue and the potential for tissue response remain to be fully investigated. The small sample size of the ex vivo pilot study limits the generalizability of the findings. Further studies with larger sample sizes and different nerve preparations are warranted.