Development of the rat abdominal VNS implant was funded by the Defense Advanced Research Projects Agency (DARPA) BTO, under the auspices of Dr. Doug Weber and Dr. Eric Van Gieson through the Space and Naval Warfare Systems Center (Contract No. N66001-15-2-4060). Research reported in this publication was supported by the Bionics Institute Incubation Fund. The Bionics Institute acknowledges the support they receive from the Victorian Government through its Operational Infrastructural Support Program. We would like to thank Mr. Owen Burns for mechanical design, Prof. John B Furness for anatomical expertise, Prof. Robert K Shepherd for peripheral interface, neuromodulation and recording expertise, Ms. Philippa Kammerer and Ms. Amy Morley for animal husbandry and testing, Ms. Fenella Muntz and Dr. Peta Grigsby for their advice on post-operative animal care, and Ms. Jenny Zhou and the electrode fabrication team from NeoBionica for production of the VNS arrays.

All procedures involving animals were approved by the Animal Ethics Committee of St. Vincent&#39;s Hospital (Melbourne) and complied with the Australian Code for the Care and Use of Animals for Scientific Purposes (National Health and Medical Research Council of Australia) and the Prevention of Cruelty to Animals (1986) Act. In total, 24 female Dark Agouti rats (8-9 weeks old) were used for this study. The experimental groups consisted of: a normal cohort (n = 8) that received no collagen injection or VNS implant; an uns...

Abdominal VNS for Treating Inflammatory Conditions in Rat Models

<ol>
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T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuroimmunomodulation+of+vagus+nerve+stimulation+and+the+therapeutic+implications.">Neuroimmunomodulation of vagus nerve stimulation and the therapeutic implications.</a> Front Aging Neurosci. 15, 1173987 (2023).</li><li>Fudim, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Device+therapy+in+chronic+heart+failure:+JACC+state-of-the-art+review.">Device therapy in chronic heart failure: JACC state-of-the-art review.</a> J Am Coll Cardiol. 78 (9), 931-956 (2021).</li><li>Sinniger, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+12-month+pilot+study+outcomes+of+vagus+nerve+stimulation+in+Crohn's+disease.">A 12-month pilot study outcomes of vagus nerve stimulation in Crohn's disease.</a> Neurogastroenterol Motil. 32 (10), 13911 (2020).</li><li>Koopman, F. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vagus+nerve+stimulation+in+patients+with+rheumatoid+arthritis:+24+month+safety+and+efficacy.">Vagus nerve stimulation in patients with rheumatoid arthritis: 24 month safety and efficacy.</a> Arthritis Rheumatol. 70, (2018).</li><li>Genovese, M. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Safety+and+efficacy+of+neurostimulation+with+a+miniaturised+vagus+nerve+stimulation+device+in+patients+with+multidrug-refractory+rheumatoid+arthritis:+a+two-stage+multicentre,+randomised+pilot+study.">Safety and efficacy of neurostimulation with a miniaturised vagus nerve stimulation device in patients with multidrug-refractory rheumatoid arthritis: a two-stage multicentre, randomised pilot study.</a> Lancet Rheumatol. 2 (9), e527-e538 (2020).</li><li>Lu, J. Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+randomized+trial+on+the+effect+of+transcutaneous+electrical+nerve+stimulator+on+glycemic+control+in+patients+with+type+2+diabetes.">A randomized trial on the effect of transcutaneous electrical nerve stimulator on glycemic control in patients with type 2 diabetes.</a> Sci Rep. 13 (1), 2662 (2023).</li><li>Huang, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+transcutaneous+auricular+vagus+nerve+stimulation+on+impaired+glucose+tolerance:+a+pilot+randomized+study.">Effect of transcutaneous auricular vagus nerve stimulation on impaired glucose tolerance: a pilot randomized study.</a> BMC Complement Altern Med. 14, 203 (2014).</li><li>Chang, R. B., Strochlic, D. E., Williams, E. K., Umans, B. D., Liberles, S. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vagal+sensory+neuron+subtypes+that+differentially+control+breathing.">Vagal sensory neuron subtypes that differentially control breathing.</a> Cell. 161 (3), 622-633 (2015).</li><li>McAllen, R. M., Shafton, A. D., Bratton, B. O., Trevaks, D., Furness, J. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Calibration+of+thresholds+for+functional+engagement+of+vagal+A,+B+and+C+fiber+groups+in+vivo.">Calibration of thresholds for functional engagement of vagal A, B and C fiber groups in vivo.</a> Bioelectron Med (Lond). 1 (1), 21-27 (2018).</li><li>Payne, S. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anti-inflammatory+effects+of+abdominal+vagus+nerve+stimulation+on+experimental+intestinal+inflammation).">Anti-inflammatory effects of abdominal vagus nerve stimulation on experimental intestinal inflammation).</a> Front Neurosci. 13, 418 (2019).</li><li>Ben-Menachem, E., Revesz, D., Simon, B. J., Silberstein, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Surgically+implanted+and+non-invasive+vagus+nerve+stimulation:+a+review+of+efficacy,+safety+and+tolerability.">Surgically implanted and non-invasive vagus nerve stimulation: a review of efficacy, safety and tolerability.</a> Eur J Neurol. 22 (9), 1260-1268 (2015).</li><li>Parhizgar, F., Nugent, K., Raj, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Obstructive+sleep+apnea+and+respiratory+complications+associated+with+vagus+nerve+stimulators.">Obstructive sleep apnea and respiratory complications associated with vagus nerve stimulators.</a> J Clin Sleep Med. 7 (4), 401-407 (2011).</li><li>Mao, H., Chen, Y., Ge, Q., Ye, L., Cheng, H. S. h. o. r. t. -. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=and+long-term+response+of+vagus+nerve+stimulation+therapy+in+drug-resistant+epilepsy:+A+systematic+review+and+meta-analysis.">and long-term response of vagus nerve stimulation therapy in drug-resistant epilepsy: A systematic review and meta-analysis.</a> Neuromodulation. 25 (3), 327-342 (2022).</li><li>Payne, S. C., Furness, J. B., Stebbing, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioelectric+neuromodulation+for+gastrointestinal+disorders:+effectiveness+and+mechanisms.">Bioelectric neuromodulation for gastrointestinal disorders: effectiveness and mechanisms.</a> Nat Rev Gastroenterol Hepatol. 16 (2), 89-105 (2019).</li><li>Payne, S. C., Romas, E., Hyakumura, T., Muntz, F., Fallon, J. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Abdominal+vagus+nerve+stimulation+alleviates+collagen-induced+arthritis+in+rats.">Abdominal vagus nerve stimulation alleviates collagen-induced arthritis in rats.</a> Front Neurosci. 16, 1012133 (2022).</li><li>Payne, S. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Blood+glucose+modulation+and+safety+of+efferent+vagus+nerve+stimulation+in+a+type+2+diabetic+rat+model.">Blood glucose modulation and safety of efferent vagus nerve stimulation in a type 2 diabetic rat model.</a> Physiol Rep. 10 (8), 15257 (2022).</li><li>Shepherd, R. K., Fallon, J. B., Payne, S. C., Burns, O., Furness, J. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Peripheral+nerve+electrode+array.">Peripheral nerve electrode array.</a> US patent. , (2019).</li><li>Castoro, M. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Excitation+properties+of+the+right+cervical+vagus+nerve+in+adult+dogs.">Excitation properties of the right cervical vagus nerve in adult dogs.</a> Exp Neurol. 227 (1), 62-68 (2011).</li><li>Payne, S. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Differential+effects+of+vagus+nerve+stimulation+strategies+on+glycemia+and+pancreatic+secretions.">Differential effects of vagus nerve stimulation strategies on glycemia and pancreatic secretions.</a> Physiol Rep. 8 (11), 14479 (2020).</li><li>Prechtl, J. C., Powley, T. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+fiber+composition+of+the+abdominal+vagus+of+the+rat.">The fiber composition of the abdominal vagus of the rat.</a> Anat Embryol (Berl). 181 (2), 101-115 (1990).</li><li>Gasser, H. S., Erlanger, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+played+by+the+sizes+of+the+constituent+fibers+of+a+nerve+trunk+in+determining+the+form+of+its+action+potential+wave.">The role played by the sizes of the constituent fibers of a nerve trunk in determining the form of its action potential wave.</a> Am J Physiol-Legacy Content. 80 (3), 522-547 (1927).</li><li>Parker, J. L., Shariati, N. H., Karantonis, D. 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H., Schnitzlein, H. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+numbers+of+nerve+fibers+in+the+vagus+nerve+of+man.">The numbers of nerve fibers in the vagus nerve of man.</a> Anat Rec. 139, 429-435 (1961).</li><li>Bassi, G. 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This method of abdominal VNS implant surgery and chronic stimulation of the vagus nerve and recording of ECAPs have been successfully used and well-tolerated for 5 weeks in rats following implantation10,15,16. Retraction of the stomach, liver, and gut to gain a good view of the esophagus and the vagus nerve is one of the key steps in the surgery. Once these organs are retracted, the vagus nerve becomes accessible. Retraction of ...

Vagus nerve stimulation (VNS) delivered at the cervical site in the neck is The United States Food and Drug Administration (FDA)-approved treatment for refractory epilepsy, refractory depression, and post-ischemic stroke rehabilitation1, and European Commission-approved for heart failure in Europe2. Non-invasive cervical VNS is FDA-approved for migraine and headache1. Its application is expected to expand, with recent clinical trials showing efficacy of VNS in other indications such as Crohn&#39;s disease3, rheumatoid arthritis4,5 and impaired glucose tolerance and type 2 diabetes6,7. Although promising, cervical VNS can cause bradycardia and apnea due to off-target activation of the nerve fibers that innervate the lungs and the heart8,9,10. Side effects such as cough, pain, voice alteration, headache and increase in apnea-hypopnea index are commonly reported in patients receiving cervical VNS11,12. Reduction in stimulation strength is a common strategy for reducing these side effects, however reduced charge may limit efficacy of VNS therapy by failing to activate therapeutic fibers11. In support of this hypothesis, the responder rate of patients receiving high intensity stimulation for the treatment of epilepsy was higher than that of patients receiving low intensity stimulation13.
Abdominal VNS is applied on the subdiaphragmatic vagus nerve, above the hepatic and celiac branches14 (Figure 1). Our previous study demonstrated that in rats abdominal VNS does not cause cardiac or respiratory side effects associated with cervical VNS10. Earlier studies also demonstrate anti-inflammatory effects of abdominal VNS in a rat model of inflammatory bowel disease and rheumatoid arthritis10,15 as well as reduction in glycemia in a rat model of type 2 diabetes16. Recently, the abdominal VNS technology has been translated for a first-in-human clinical trial for the treatment of inflammatory bowel disease (NCT05469607).
The peripheral nerve electrode array used to deliver stimulation to the abdominal vagus nerve (WO201909502017) has been custom developed for use in rats, and comprises of two to three platinum electrode pairs placed 4.7 mm apart, supported by a medical-grade silicone elastomer cuff, a suturing tab to anchor the array to the esophagus, a lead wire and a percutaneous connector to be mounted on the lumbar region (Figure 2). The lead wire is tunneled under the skin on the left side of the animal. Multiple electrode pair design allows for electrical stimulation of the nerve as well as recording electrically evoked compound action potentials (ECAPs), which confirms correct placement of the implant onto the nerve and suprathreshold stimulation intensities. Abdominal VNS is well tolerated in freely moving rats for months10,15,16. This allows for assessment of its efficacy on disease models.
This manuscript describes the methods for the electrode array sterilization, abdominal vagus nerve implantation surgery, and chronic stimulation and recording of ECAPs in awake rats for studying the efficacy of abdominal VNS in a variety of disease models. These methods were originally developed for studying the efficacy of abdominal VNS in the rat model of inflammatory bowel disease10 and have also been successfully used for a rat model of rheumatoid arthritis15 and diabetes16.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>0.9% saline</td>
			<td>Briemarpak</td>
			<td>SC3050</td>
			<td></td>
		</tr>
		<tr>
			<td>Baytril</td>
			<td>Bayer</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Betadine</td>
			<td>Sanofi-Aventis Healthcare</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Buprelieve (Buprenorphine)</td>
			<td>Jurox</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Data acquisition device</td>
			<td>National Instruments</td>
			<td>USB-6210</td>
			<td></td>
		</tr>
		<tr>
			<td>DietGel Boost (dietary gel supplement)</td>
			<td>ClearH2O</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Dumont tweezer, style 5</td>
			<td>ProSciTech</td>
			<td>T05-822</td>
			<td></td>
		</tr>
		<tr>
			<td>Dumont tweezer, style N7, self-closing</td>
			<td>ProSciTech</td>
			<td>EMS72864-D</td>
			<td></td>
		</tr>
		<tr>
			<td>Elmasonic P sonicator</td>
			<td>Elma</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Hartmann&#39;s solution</td>
			<td>Baxter</td>
			<td>AHB2323</td>
			<td></td>
		</tr>
		<tr>
			<td>Hemostat</td>
			<td>ProSciTech</td>
			<td>TS1322-140</td>
			<td></td>
		</tr>
		<tr>
			<td>HPMC/PAA Moisturising Eye Gel</td>
			<td>Alcon</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Igor Pro-8 software</td>
			<td>Wavemetrics, Inc</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Isoflo (Isoflurane)</td>
			<td>Zoetis</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Isolated differential amplifier</td>
			<td>World Precision Instruments</td>
			<td>ISO-80</td>
			<td></td>
		</tr>
		<tr>
			<td>Liquid pyroneg</td>
			<td>Diversey</td>
			<td>HH12291</td>
			<td>cleaning solution</td>
		</tr>
		<tr>
			<td>Marcaine (Bupivacaine)</td>
			<td>Aspen</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Plastic drape</td>
			<td>Multigate</td>
			<td>22-203</td>
			<td></td>
		</tr>
		<tr>
			<td>Rat vagus nerve implant</td>
			<td>Neo-Bionica</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Rimadyl (Carprofen)</td>
			<td>Zoetis</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Silk suture 3-0</td>
			<td>Ethicon</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Silk suture 7-0</td>
			<td>Ethicon</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>SteriClave autoclave</td>
			<td>Cominox</td>
			<td>24S</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile disposable surgical gown</td>
			<td>Zebravet</td>
			<td>DSG-S</td>
			<td></td>
		</tr>
		<tr>
			<td>Suicide Nickel hooks</td>
			<td>Jarvis Walker</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Ultrapure water</td>
			<td>Merck Millipre</td>
			<td>Milli-Q Direct</td>
			<td></td>
		</tr>
		<tr>
			<td>Underpads</td>
			<td>Zebravet</td>
			<td>UP10SM</td>
			<td></td>
		</tr>
		<tr>
			<td>Vannas scissors</td>
			<td>ProSciTech</td>
			<td>EMS72933-01</td>
			<td></td>
		</tr>
		<tr>
			<td>Vicryl suture 4-0</td>
			<td>Ethicon</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

implantation surgery for abdominal vagus nerve stimulation and recording studies in awake rats

Abdominal vagus nerve stimulation (VNS) can be applied to the subdiaphragmatic branch of the vagus nerve of rats. Due to its anatomical location, it does not have any respiratory and cardiac off-target effects commonly associated with cervical VNS. The lack of respiratory and cardiac off-target effects means that the intensity of stimulation does not need to be lowered to reduce side effects commonly experienced during cervical VNS. Few recent studies demonstrate the anti-inflammatory effects of abdominal VNS in rat models of inflammatory bowel disease, rheumatoid arthritis, and glycemia reduction in a rat model of type 2 diabetes. Rat is a great model to explore the potential of this technology because of the well-established anatomy of the vagus nerve, the large size of the nerve that allows easy handling, and the availability of many disease models. Here, we describe the methods for cleaning and sterilizing the abdominal VNS electrode array and surgical protocol in rats. We also describe the technology required for confirmation of suprathreshold stimulation by recording evoked compound action potentials. Abdominal VNS has the potential to offer selective, effective treatment for a variety of conditions, including inflammatory diseases, and the application is expected to expand similarly to cervical VNS.

Author Spotlight: Exploring Abdominal VNS for Inflammatory Conditions

Implantation Surgery for Abdominal Vagus Nerve Stimulation and Recording Studies in Awake Rats

Abdominal VNS has the advantage of not causing cardiac and respiratory side effects associated with cervical VNS. The aim of the study was to investigate the anti-inflammatory effect of abdominal VNS in a rat model of rheumatoid arthritis. Previously, we have shown that abdominal VNS reduces symptoms of a rat model of inflammatory bowel disease.A first-in-human clinical trial is currently underway to investigate the safety and efficacy of abdominal VNS as a treatment of inflammatory bowel disease. We are expanding our neuromodulation research to investigate whether we can record spontaneous inflammation-induced vagus nerve activity as the next step for developing an adaptive abdominal VNS system for the treatment of inflammatory conditions. We have demonstrated that abdominal VNS causes no cardiac or respiratory off-target effects that are associated with cervical VNS, and it has anti-inflammatory effects in rat models of inflammatory bowel disease and rheumatoid arthritis.The effects of abdominal VNS on the body are not well understood compared to cervical VNS. This protocol may help to establish experimental protocols for testing the effect of abdominal VNS on different physiological parameters in healthy animals or inflammatory conditions such as systemic lupus and chronic kidney disease.

Recording evoked compound action potentials (ECAPs, Figure 3A,B) immediately after surgery is a technique that can be used to help confirm correct placement of the nerve within the array channel, and that stimulation is efficacious in activating the vagus nerve.
In Figure 3, female dark agouti rats (8-9 weeks of age) were implanted with the VNS electrode array. In rats randomly selected to receive therapeutic stimulat...

Watch this Scientific Journal Video about Author Spotlight: Exploring Abdominal VNS for Inflammatory Conditions at JoVE.com

The present protocol describes the surgical technique for implanting an electrode array onto the abdominal vagus nerve in rats, along with methods for chronic electrophysiology testing and stimulation using the implanted device.

Abdominal vagus nerve stimulation (VNS) can be applied to the subdiaphragmatic branch of the vagus nerve of rats. Due to its anatomical location, it does ...

implantation-surgery-for-abdominal-vagus-nerve-stimulation-recording

Research

JoVE Journal

Neuroscience

741 vistas.  Bionics Institute. La ENV abdominal tiene la ventaja de no causar efectos secundarios cardíacos y respiratorios asociados con la ENV cervical. El objetivo del estudio fue investigar el efecto antiinflamatorio de la ENV abdominal en un modelo de rata con artritis reumatoide. Anteriormente, hemos demostrado que la ENV abdominal reduce los síntomas de un modelo de enfermedad inflamatoria intestinal en ratas.Actualmente se está llevando a cabo un primer ensayo clínico en humanos para investigar la seguri...