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Existing approaches for constructing chronically implantable peripheral nerve cuff electrodes for use in small rodents often require specialized equipment and/or highly trained personnel. In this protocol we demonstrate a simple, low-cost approach for fabricating chronically implantable cuff electrodes, and demonstrate their effectiveness for vagus nerve stimulation (VNS) in rats.
Peripheral nerve cuff electrodes have long been used in the neurosciences and related fields for stimulation of, for example, vagus or sciatic nerves. Several recent studies have demonstrated the effectiveness of chronic VNS in enhancing central nervous system plasticity to improve motor rehabilitation, extinction learning, and sensory discrimination. Construction of chronically implantable devices for use in such studies is challenging due to rats’ small size, and typical protocols require extensive training of personnel and time-consuming microfabrication methods. Alternatively, commercially available implantable cuff electrodes can be purchased at a significantly higher cost. In this protocol, we present a simple, low-cost method for construction of small, chronically implantable peripheral nerve cuff electrodes for use in rats. We validate the short and long-term reliability of our cuff electrodes by demonstrating that VNS in ketamine/xylazine anesthetized rats produces decreases in breathing rate consistent with activation of the Hering-Breuer reflex, both at the time of implantation and up to 10 weeks after device implantation. We further demonstrate the suitability of the cuff electrodes for use in chronic stimulation studies by pairing VNS with skilled lever press performance to induce motor cortical map plasticity.
Recently, the demand for chronically implantable cuff electrodes for stimulation of peripheral nerves has grown, as studies increasingly demonstrate the preclinical usefulness of this technique for the treatment of numerous inflammatory diseases1,2,3 and neurological disorders4,5,6,7,8,9,10,11,12,13,14,15. Chronic VNS, for example, has been shown to enhance neocortical plasticity in a variety of learning contexts, improving motor rehabilitation4,5,6,7,8, extinction learning10,11,12,13,14, and sensory discrimination15. Commercially available peripheral nerve cuff electrodes are often associated with extended times for order fulfillment and relatively high costs, which can limit their accessibility. Alternatively, protocols for “in-house” fabrication of chronically implantable cuff electrodes remain limited, and rodent anatomy presents particular challenges due to their small size. Current protocols for constructing cuff electrodes for chronic rodent experiments often require the use of complex equipment and techniques, as well as extensively trained personnel. In this protocol, we demonstrate a simplified approach to cuff electrode fabrication based on previously published and widely used methods16,17. We validate the functionality of our chronically implanted electrodes in rats by demonstrating that, at the time of cuff implantation around the left cervical vagus nerve, stimulation applied to the cuff electrodes successfully produced a cessation of breathing and drop in SpO2. Stimulation of afferent pulmonary receptor vagal fibers is known to engage the Hering-Breuer reflex, in which the inhibition of several respiratory nuclei in the brainstem results in the suppression inspiration18. Thus, cessation of breathing consistent with the Hering-Breuer reflex, and the resulting drop in SpO2, provide a straightforward test for proper electrode implantation and cuff function in anesthetized rats. To validate the long-term functionality of chronically implanted cuff electrodes, reflex responses were measured at the time of implantation and compared to the responses obtained in the same animals six weeks after implantation. A second group of rats was implanted with VNS cuff electrodes after behavioral training on a lever pressing task. In these rats, VNS paired with correct task performance produced reorganization of the cortical motor map, consistent with previously published studies19,20,21,22. At the time of motor cortical mapping under anesthesia, which occurred 5–10 weeks after device implantation, we further validated cuff function in VNS-treated animals by confirming that VNS successfully induced a cessation of breathing and a greater than 5% drop in SpO2.
The recently published protocols from Childs et al.17 and Rios et al.16 provide a well-validated starting point for a simplified cuff electrode fabrication approach, as this popular method has been utilized by multiple labs conducting chronic VNS studies in rodents1,2,3,4,5,6,7,8,9,10,11. The original method involves several high-precision steps for manipulating the fine microwires such that cuff electrode fabrication takes over an hour to complete, and extensive training to perform reliably. The simplified approach described here requires significantly fewer materials and tools and can be completed in under one hour by minimally trained personnel.
All procedures described in this protocol are carried out in accordance with the NIH Guide for the Care and Use of Laboratory Animals and were approved by the Institutional Animal Care and Use Committee of The University of Texas at Dallas.
1. Stimulating cuff electrode fabrication
2. Head-cap construction
NOTE: Headcap assembly procedures are similar to those published previously (Childs et al.17), and are summarized here for convenience.
3. Device usage
Vagus nerve cuff electrodes and headcaps were chronically implanted in rats according to previously published surgical procedures17,19,20,21,22. Prior to implantation, impedance at 1 kHz was measured across the cuff leads with the cuff tubing submerged in saline (impedance = 1.2 ± 0.17 kΩ [mean ± std]; N = 9). Only cuffs with impedances less tha...
Here we describe a simple, low-cost approach for assembly of chronically implantable stimulating cuff electrodes for use in rodents, facilitating preclinical investigations of this emerging therapy. This simplified method requires no specialized training or equipment, and uses a small number of tools and supplies that are easily accessible to most research labs, reducing both the monetary and labor costs of device manufacture compared to other approaches16,26
The authors have nothing to disclose.
This work was funded by the University of Texas at Dallas and the UT Board of Regents. We thank Solomon Golding, Bilaal Hassan, Marghi Jani, and Ching-Tzu Tseng for technical assistance.
Name | Company | Catalog Number | Comments |
Biocompatible polyurethane-based polymer tubing, 0.080" OD x 0.040" ID | Braintree Scientific | MRE080 36 FT | |
Dissecting microscope | AM Scopes | #SM-6T-FRL | |
Fine Serrated Scissors, straight, 22mm cutting edge | Fine Science Tools | #14058-09 | for cutting Pt/Ir wire and suture thread |
Forceps, #5 Dumont forceps, straight, 11 cm, 0.1 x 0.06 mm tip | Fine Science Tools | #11626-11 | |
Forceps, ceramic tipped forceps, 0.3 mm x 30 mm tips | Electron Microscopy Sciences | #78127-71 | |
Gold Pins, PCB Press Fit Socket | Mill-Max | #1001-0-15-15-30-27-04-0 | or similar small pins for connecting cuff leads to headcap |
Isobutane lighter | BIC | #LCP21-AST | for de-insulating Pt/Ir wire |
Micro strip connector with latch, 4-pin | Omnetics | A24002-004 / PS1-04-SS-LT | |
Pipette tip, 10 uL | VWR | 89079-464 | |
Platinum-Iridium (90/10%) Wire, 0.001" (diameter) x 9 strands, PTFE insulated | Sigmund Cohn | 10IR9/49T | |
Razor Blade, Single Edge, Surgical Carbon Steel No.9 | VWR | #55411-050 | for cutting MicroRenathane tubing |
Sewing needle, ca. 4.0 cm length x 0.7 mm diameter (size 6-7) | Singer | 00276 | Smaller needle for threading Pt/Ir wire |
Sewing needle, ca. 4.5 cm length x 0.8 mm diameter (size 2-3) | Singer | 00276 | Larger needle for pinning cuff during assembly and for threading suture |
Small foam board | Juvo+/Amazon | B07C9637SJ | for fabrication platform; our dimensions are ca. 2.5" x 3.5" x 1" (L x W x H) |
Solder, multicore lead-free, 0.38mm diameter | Loctite/Multicore | #796037 | |
Soldering station | Weller | WES51 | or similar soldering iron compatible with long conical tips (this part has been discontinued) |
Soldering tip, long conical, 0.01" / 0.4 mm | Weller | 1UNF8 | |
Suture, nonabsorbable braided silk ,size 6/0 | Fine Science tools | #18020-60 | |
UV (405 nm) spot light | Henkel/Loctite | #2182207 | |
UV Light Cure Adhesive 25 ml | Henkel/Loctite | AA 3106 | or similar biocompatible UV cure adhesive |
Wire wrapping wire, 30 AWG | Digikey | K396-ND |
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