Autonomic nerves convey essential neural signals that regulate vital body functions. Recording clearly distinctive physiological neural signals from autonomic nerves will help develop new treatments for restoring regulatory functions. However, this is very challenging due to the small nature of autonomic nerves and the low-amplitude signals from their small axons. We developed a multi-channel, high-density, intraneural carbon fiber microelectrode array (CFMA) with ultra-small electrodes (8–9 µm in diameter, 150–250 µm in length) for recording physiological action potentials from small autonomic nerves. In this study, we inserted CFMA with up to 16 recording carbon fibers in the cervical vagus nerve of 22 isoflurane-anesthetized rats. We recorded action potentials with peak-to-peak amplitudes of 15.1–91.7 µV and signal-to-noise ratios of 2.0–8.3 on multiple carbon fibers per experiment, determined conduction velocities of some vagal signals in the afferent (0.7–4.4 m/s) and efferent (0.7–8.8 m/s) directions, and monitored firing rate changes in breathing and blood glucose modulated conditions. Overall, these experiments demonstrated that CFMA is a novel interface for in-vivo intraneural action potential recordings. This work is considerable progress towards the comprehensive understanding of physiological neural signaling in vital regulatory functions controlled by autonomic nerves.

Peripheral nerve mapping tools with higher spatial resolution are needed to advance systems neuroscience, and potentially provide a closed‐loop biomarker in neuromodulation applications. Two critical challenges of microscale neural interfaces are 1) how to apply them to small peripheral nerves, and 2) how to minimize chronic reactivity. A flexible microneedle nerve array (MINA) is developed, which is the first high‐density penetrating electrode array made with axon‐sized silicon microneedles embedded in low‐modulus thin silicone. The design, fabrication, acute recording, and chronic reactivity to an implanted MINA, are presented. Distinctive units are identified in the rat peroneal nerve. The authors also demonstrate a long‐term, cuff‐free, and suture‐free fixation manner using rose bengal as a light‐activated adhesive for two time‐points. The tissue response is investigated at 1‐week and 6‐week time‐points, including two sham groups and two MINA‐implanted groups. These conditions are quantified in the left vagus nerve of rats using histomorphometry. Micro computed tomography (micro‐CT) is added to visualize and quantify tissue encapsulation around the implant. MINA demonstrates a reduction in encapsulation thickness over previously quantified interfascicular methods. Future challenges include techniques for precise insertion of the microneedle electrodes and demonstrating long‐term recording.

This protocol is for obtaining physiological action potential recordings in rat vagus nerves using carbon fiber microelectrode arrays (CFMAs) in spontaneous and blood glucose and breathing modulated conditions. The rats were anesthetized with isoflurane, which maintained consistent and stable depth of anesthesia for recording vagal nerve activity with ultra-small carbon fibers. Blood glucose levels were modulated by intraperitoneal (IP) injection of glucose, insulin, or 2-deoxy-D-glucose (2-DG). Breathing was modulated by increasing anesthesia depth. Carbon fiber microelectrode arrays are available through the Multimodal Integrated Neural Technologies (MINT) technology hub (https://mint.engin.umich.edu/), which is supported by the National Science Foundation (Award 1707316). This research was also supported by the National Institute of Health SPARC Program (Award OT2OD024907).

25Autonomic nerves convey essential neural signals that regulate vital body functions. Recording 26 clearly distinctive physiological neural signals from autonomic nerves will help develop new 27 treatments for restoring regulatory functions. However, this is very challenging due to the small 28 nature of autonomic nerves and the low-amplitude signals from their small axons. We developed 29 a multi-channel, high-density, intraneural carbon fiber microelectrode array (CFMA) with ultra-30 small electrodes (8-9 µm in diameter, 150-250 µm in length) for recording physiological action 31 potentials from small autonomic nerves. In this study, we inserted CFMA with up to 16 32 recording carbon fibers in the cervical vagus nerve of 22 isoflurane-anesthetized rats. We 33 recorded action potentials with peak-to-peak amplitudes of 15.1-91.7 µV and signal-to-noise 34 ratios of 2.0-8.3 on multiple carbon fibers per experiment, determined conduction velocities of 35 some vagal signals in the afferent (0.7-1.0 m/sec) and efferent (0.7-8.8 m/sec) directions, and 36 monitored firing rate changes in breathing and blood glucose modulated conditions. Overall, 37 these experiments demonstrated that CFMAs are a novel interface for in-vivo intraneural action 38 potential recordings from autonomic nerves. This work is a milestone towards the 39 comprehensive understanding of physiological neural signaling and the development of 40 innovative treatment modalities for restoring vital functions controlled by autonomic nerves. 41 42