Objective Our goal is to develop an interface that integrates chronic monitoring of lower urinary tract (LUT) activity with stimulation of peripheral pathways. Approach Penetrating microelectrodes were implanted in sacral dorsal root ganglia (DRG) of adult male felines. Peripheral electrodes were placed on or in the pudendal nerve, bladder neck and near the external urethral sphincter. Supra-pubic bladder catheters were implanted for saline infusion and pressure monitoring. Electrode and catheter leads were enclosed in an external housing on the back. Neural signals from microelectrodes and bladder pressure of sedated or awake-behaving felines were recorded under various test conditions in weekly sessions. Electrodes were also stimulated to drive activity. Main results LUT single- and multi-unit activity was recorded for 4 to 11 weeks in four felines. As many as 18 unique bladder pressure single-units were identified in each experiment. Some channels consistently recorded bladder afferent activity for up to 41 days, and we tracked individual single-units for up to 23 days continuously. Distension-evoked and stimulation-driven (DRG and pudendal) bladder emptying was observed, during which LUT sensory activity was recorded. Significance This chronic implant animal model allows for behavioral studies of LUT neurophysiology and will allow for continued development of a closed-loop neuroprosthesis for bladder control.

Bioelectric medicine treatments target disorders of the nervous system unresponsive to pharmacological methods. While current stimulation paradigms effectively treat many disorders, the underlying mechanisms are relatively unknown, and current neuroscience recording electrodes are often limited in their specificity to gross averages across many neurons or axons. Here, we develop a novel, durable carbon fiber electrode array adaptable to many neural structures for precise neural recording. Carbon fibers (6.8 µm diameter) were sharpened using a reproducible blowtorch method that uses the reflection of fibers against the surface of a water bath. The arrays were developed by partially embedding carbon fibers in medical-grade silicone to improve durability. We recorded acute spontaneous electrophysiology from the rat cervical vagus nerve (CVN), feline dorsal root ganglia (DRG), and rat brain. Blowtorching resulted in fibers of 72.3 ± 33.5-degree tip angle with 146.8 ± 17.7 µm exposed carbon. Observable neural clusters were recorded using sharpened carbon fiber electrodes from rat CVN (41.8 µVpp), feline DRG (101.1 µVpp), and rat brain (80.7 µVpp). Recordings from the feline DRG included physiologically relevant signals from increased bladder pressure and cutaneous brushing. These results suggest that this carbon fiber array is a uniquely durable and adaptable neural recording device. In the future, this device may be useful as a bioelectric medicine tool for diagnosis and closedloop neural control of therapeutic treatments and monitoring systems.

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).

ObjectiveBioelectric medicine offers therapeutic diagnoses and treatments for disorders of the nervous system unresponsive to pharmacological treatments. While current neural interfaces effectively treat many disorders with stimulation, recording specificity is often limited to gross averages across many neurons or axons. Here, we develop and describe a novel, robust carbon fiber electrode array adaptable to many neural structures for precise neural recording.ApproachCarbon fibers were sharpened using a blowtorch method made reproducible by using the reflection of fibers against the surface of a water bath. Arrays of carbon fibers were developed by partially embedding carbon fibers in medical-grade silicone to improve robustness to fracture. Acute spontaneous electrophysiology was recorded from the rat cervical vagus nerve, feline dorsal root ganglia, and rat brain. Acute brushing and bladder pressure electrophysiology was recorded from feline dorsal root ganglia as well.Main resultsBlowtorching resulted in fibers of 72.3 ± 33.5 degree tip angle with 146.8 ± 17.7 μm exposed carbon. Silicone-embedded carbon fiber arrays were robust to bending (87.5% of fibers remained unbroken, 50,000 passes). Observable neural clusters were recorded using sharpened carbon fiber electrodes from rat cervical vagus nerve (41.8 μVpp, N=3 electrodes), feline dorsal root ganglia (101.1 μVpp, N=32 electrodes), and rat brain (80.7 μVpp, N=7 electrodes). Recordings from the feline dorsal root ganglia included physiologically-relevant signals from increased bladder pressure and cutaneous brushing.SignificanceThese results suggest that this carbon fiber array is a uniquely robust and adaptable neural recording device, useful for specific electrophysiology measurements. In the future, this device may be useful as a bioelectric medicine tool for diagnosis and closed-loop neural control of therapeutic treatments and monitoring systems.

Background: The role of the kidney in glucose homeostasis has gained global interest. Kidneys are innervated by renal nerves, and renal denervation animal models have shown improved glucose regulation. We hypothesized that stimulation of renal nerves at kilohertz frequencies, which can block propagation of action potentials, would increase urine glucose excretion. Conversely, we hypothesized that low frequency stimulation, which has been shown to increase renal nerve activity, would decrease urine glucose excretion. Methods: We performed non-survival experiments on male rats under thiobutabarbital anesthesia. A cuff electrode was placed around the left renal artery, encircling the renal nerves. Ureters were cannulated bilaterally to obtain urine samples from each kidney independently for comparison. Renal nerves were stimulated at kilohertz frequencies (1-50 kHz) or low frequencies (2-5 Hz), with intravenous administration of a glucose bolus shortly into the 25-40-min stimulation period. Urine samples were collected at 5-10-min intervals, and colorimetric assays were used to quantify glucose excretion and concentration between stimulated and non-stimulated kidneys. A KruskalWallis test was performed across all stimulation frequencies (α = 0.05), followed by a post-hoc Wilcoxon rank sum test with Bonferroni correction (α = 0.005). Results: For kilohertz frequency trials, the stimulated kidney yielded a higher average total urine glucose excretion at 33 kHz (+ 24.5%; n = 9) than 1 kHz (− 5.9%; n = 6) and 50 kHz (+ 2.3%; n = 14). In low frequency stimulation trials, 5 Hz stimulation led to a lower average total urine glucose excretion (− 40.4%; n = 6) than 2 Hz (− 27.2%; n = 5). The average total urine glucose excretion between 33 kHz and 5 Hz was statistically significant (p < 0.005). Similar outcomes were observed for urine flow rate, which may suggest an associated response. No trends or statistical significance were observed for urine glucose concentrations. Conclusion: To our knowledge, this is the first study to investigate electrical stimulation of renal nerves to modulate urine glucose excretion. Our experimental results show that stimulation of renal nerves may modulate urine glucose excretion, however, this response may be associated with urine flow rate. Future work is needed to examine the underlying mechanisms and identify approaches for enhancing regulation of glucose excretion.

State‐of‐the‐art intraneural peripheral nerve electrodes are large, silicon‐based structures that can cause substantial tissue response and are ill‐suited for recording from small autonomic nerves. The focus of this work is to adapt our minimally‐scarring carbon fiber brain electrodes to a chronic intraneural nerve array. The objective of this study is to increase the durability of the carbon fiber electrodes to withstand the surgical handling necessary for nerve implantation, while maintaining a cellular scale electrode that can insert through the outer epineurium layer of peripheral nerves. Toward that end, we embedded carbon fibers in silicone to increase robustness, sharpened the fibers to penetrate epineurium, and tested fibers coated with poly(3,4‐ethylene‐dioxythiophene):sodium p‐toluenesulfonate (PEDOT:pTS) in vivo. We tested the durability of carbon fibers by embedding uninsulated fibers in a body of silicone and inducing a 90‐degree bend to simulate the expected shear forces seen during surgery. We found that fibers of 175μm length were robust to thousands of bend cycles. After 3000 bends, 15 of 16 fibers remained intact. We blunt cut carbon fibers using a 532nm green laser and inserted fibers into silicone of stiffness similar to epineurium to determine a length for in vivo testing. We found that blunt fibers reliably insert into silicone without breaking when less than 200μm in length (N=8 fibers). We hypothesized that sharpened fibers would insert into tissue more easily, and therefore sharpened fibers with a 5mm butane flame when protruding from the surface of water. The dimensions of sharpened fibers were 224μm in length with 147μm of exposed carbon and a tip angle of 72 degrees (N=24 fibers). We inserted functionalized sharpened carbon fibers coated with PEDOT:pTS into peroneal and cervical vagus nerves of anesthetized rats (N=5 arrays, 71 fibers, 7 animals). In all experiments, we observed that the sharpened carbon fiber electrodes penetrated the epineurium for successful insertion. The average 1kHz impedance of coated fibers prior to and upon insertion was 38kΩ and 56kΩ, respectively. In both cases, the Z1kHz of roughly 90% of fibers measured below 100kΩ. We recorded spontaneous and evoked neural activity in each experiment and are analyzing the data for discernable single units. We also inserted sharpened fibers into anesthetized feline sacral dorsal root ganglia (N=3) and confirmed insertion with evoked neural recordings and 1kHz impedances (Z=59kΩ). This work shows that the durable nature of carbon fibers embedded in silicone allows them to withstand surgical handling and insert into various types of nerves. Our next step will be the fabrication of an array for chronic implantation. Support or Funding Information NIH OT2OD024907, NINDS U01NS094375, NINDS UF1NS107659, NSF 1707316