This study was funded by grants from the National Natural Science Foundation of China (Nos. 81970890) and the Chongqing Scientific Research Institution Performance incentive project (Nos. 19540). We thank Anandhan Dhanasingh and Zhi Shu from the MED-EL company for their support.

All procedures and animal surgeries were conducted according to the guidelines of the Ethics Committee of the PLA General Hospital and were approved.
1. Anesthesia and surgical preparation
<ol>
	<li>Inject the pig (male, 2 months old, 5 kg) muscularly with tiletamine and zolazepam with a dosage of 10-15 mg/kg and intubate it with a 5.5-French endotracheal tube. Maintain anesthesia by ventilator-assisted respiration with isofluorane inhalation. Monitor the oxygen saturation (&#62;90%), breathing (15-20/min), and heart rate (60-120 beats/min) using the pulse oximetry clamp of an ECG monitor, which is connected to the pig&#39;s tongue.</li>
	<li>Put the mini-pig in a left lateral position (when the right side is to be implanted) on&#160;a thermostatically regulated heating pad to prevent&#160;hypothermia. Confirm the pig is adequately anesthetized using various stimuli. Ensure the absence of all responses (e.g., toe pinch&#160;reflex). Apply artificial tear ointment to the eyes of the minipig to keep the cornea from drying. Keep the eyes closed using a medical patch.</li>
	<li>Shave the surgical area around the earlobe, keeping it 10 cm in diameter&#160;(Figure 1) and disinfect&#160;it with three alternating swabs of iodine&#160;and alcohol in a circular motion from the center toward the outside. Cover the surgical area with sterile surgical drapes.</li>
	<li>Cover the microscope with a sterile plastic sleeve and remove the parts covering the eyepieces and objective.</li>
</ol>
2. Surgical procedure
<ol>
	<li>Locate the surface projection site of the cochlea 1 cm behind the posterior auricular sulcus at the level of the earlobe. Make a postauricular incision about 5 cm long with the projection site as the center using a #15 scalpel. Divide the subcutaneous tissue, parotid gland, and sternocleidomastoid muscle with micro-scissors to expose the surface of the mastoid bone (Figure 2A). Use bipolar cautery when necessary to minimize bleeding.</li>
	<li>Cortical mastoidectomy
	<ol>
		<li>Drill the mastoid at the surface projection of the cochlea on the mastoid bone (Figure 2B) to the external auditory canal (EAC), which is dense and pale blue (Figure 2C). Be careful not to damage the pale or reddish vertical segment of the facial nerve dorsal to the EAC to avoid bleeding (Figure 2C). 
		NOTE: If the facial nerve is damaged, bipolar cautery is a good choice to stop bleeding.</li>
	</ol>
	</li>
	<li>Expose the tympanum by drilling the bone surrounding the posterior bony EAC (Figure 2D). Separate the skin of the EAC and the facial nerve with a hypodermic needle to avoid damaging the facial nerve. Carefully push the skin of the EAC away to expose the tympanum (including the ossicular chain) and round window niche (Figure 3A).</li>
	<li>Expose the round window membrane. Remove the round window niche with a small diamond burr, and keep continuous suction-irrigation to expose the basal turn of the cochlea and round window membrane (Figure 3B).</li>
	<li>Fix the receiver package. Separate the cranial parietal muscle to form a pocket just large enough for the receiver. Place the internal receiver package in the muscular pocket and fix it with a fixation suture.</li>
	<li>Insert the electrode array, which is connected to a receiver fixed in a muscular pocket, by opening the round window membrane with a sharp microsurgical knife and inserting the array using micro forceps slowly, steadily, and continuously in relationship to the modiolus of the cochlea (Figure 3C). Suture the surgical incision with an absorbable&#160;suture 2-0.</li>
	<li>ECAP measurements 
	NOTE: The setup is composed of a PC with MAESTRO Software connected to the patient&#39;s cochlear implant (CI) through the stimulation device (MAX Programming Interface) and the CI coil.
	<ol>
		<li>Magnetically connect the CI coil to the CI receiver through the skin. Confirm the integrity of the CI and check the electrode impedance for all channels before ECAP measurements using the telemetry function of the CI, which is conducted by MAESTRO Software automatically (Figure 4A,B).</li>
		<li>Conduct the ECAP measurements as described previously19. Select the ECAP module, choose all 12 electrodes for stimulation, and wait for the ECAP tests of the electrodes to be completed. See the Table of Materials for the software and stimulation device used to measure ECAP responses. Stimulate all 12 electrodes for ECAP measurements using biphasic stimuli of 30 &#181;s phase duration, with an alternating-polarity paradigm, averaging over 25 iterations, and a stimulation rate of 45.1 pulses/s.</li>
	</ol>
	</li>
</ol>
3. Postoperative care
<ol>
	<li>Keep monitoring the mini-pig to avoid harm due to unconscious movement until it has regained sufficient consciousness to maintain sternal recumbency. Put&#160;the minipig on the thermostatically regulated heating pad to prevent&#160;hypothermia.</li>
	<li>Place the mini-pig back into its home cage alone.</li>
	<li>Inject antibiotics to prevent postoperative infection for 7 days.</li>
	<li>Check the mini-pig for symptoms of vestibular injury such as nystagmus, circling, or rolling over.</li>
</ol>
4. Postoperative CT scan
<ol>
	<li>&#160;Administer intramuscular injection of 3% sodium pentobarbital 1 ml/kg, and 0.1 ml/kg of Sulforaphane to minipig to induce anesthesia. Use&#160;a 37&#8451; warming plate to keep it warm. After&#160;3 or 5 minutes,&#160;a CT scan can be&#160;conducted.&#160;</li>
	<li>To confirm the correct position of the electrode array, narcotize the mini-pig with tiletamine and zolazepam 1 week after the operation. Perform the CT scan and 3D reconstruction20 using the 3D slicer image computing platform (see the Table of Materials). Import DICOM data of the CT and conduct the Volume rendering module to achieve 3D images of the CI.</li>
</ol>

<ol>
	<li>World report on hearing. World Health Organization Available from: <a target="_blank" href="https://www.who.int/publications/i/item/world-report-on-hearing">https://www.who.int/publications/i/item/world-report-on-hearing</a> (2021)</li><li>Lee, S. 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=Natural+course+of+residual+hearing+preservation+with+a+slim,+modiolar+cochlear+implant+electrode+array.">Natural course of residual hearing preservation with a slim, modiolar cochlear implant electrode array.</a> American Journal of Otolaryngology. 43 (2), 103382 (2022).</li><li>Lorens, 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=Binaural+advantages+in+using+a+cochlear+implant+for+adults+with+profound+unilateral+hearing+loss.">Binaural advantages in using a cochlear implant for adults with profound unilateral hearing loss.</a> Acta Oto-Laryngologica. 139 (2), 153-161 (2019).</li><li>Lally, J. W., Adams, J. K., Wilkerson, B. 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+use+of+cochlear+implantation+in+the+elderly.">The use of cochlear implantation in the elderly.</a> Current Opinion in Otolaryngology &amp; Head and Neck Surgery. 27 (5), 387-391 (2019).</li><li>Rhodes, R. M., Tsai Do, B. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Future+of+implantable+auditory+devices.">Future of implantable auditory devices.</a> Otolaryngologic Clinics of North America. 52 (2), 363-378 (2019).</li><li>Colesa, D. J., 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=Development+of+a+chronically-implanted+mouse+model+for+studies+of+cochlear+health+and+implant+function.">Development of a chronically-implanted mouse model for studies of cochlear health and implant function.</a> Hearing Research. 404, 108216 (2021).</li><li>Toulemonde, P., 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=Evaluation+of+the+efficacy+of+dexamethasone-eluting+electrode+array+on+the+post-implant+cochlear+fibrotic+reaction+by+three-dimensional+immunofluorescence+analysis+in+Mongolian+gerbil+cochlea.">Evaluation of the efficacy of dexamethasone-eluting electrode array on the post-implant cochlear fibrotic reaction by three-dimensional immunofluorescence analysis in Mongolian gerbil cochlea.</a> Journal of Clinic Medicine. 10 (15), 3315 (2021).</li><li>King, J., Shehu, I., Roland, J. T., Svirsky, M. A., Froemke, R. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+physiological+and+behavioral+system+for+hearing+restoration+with+cochlear+implants.">A physiological and behavioral system for hearing restoration with cochlear implants.</a> Journal of Neurophysiology. 116 (2), 844-858 (2016).</li><li>Chen, M., Min, S., Zhang, C., Hu, X., Li, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Using+extracochlear+multichannel+electrical+stimulation+to+relieve+tinnitus+and+reverse+tinnitus-related+auditory-somatosensory+plasticity+in+the+cochlear+nucleus.">Using extracochlear multichannel electrical stimulation to relieve tinnitus and reverse tinnitus-related auditory-somatosensory plasticity in the cochlear nucleus.</a> Neuromodulation. , (2021).</li><li>Yi, H., 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=Miniature+pigs:+A+large+animal+model+of+cochlear+implantation.">Miniature pigs: A large animal model of cochlear implantation.</a> American Journal of Translational Research. 8 (12), 5494-5502 (2016).</li><li>Vollmer, M., Beitel, R. E., Schreiner, C. E., Leake, P. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Passive+stimulation+and+behavioral+training+differentially+transform+temporal+processing+in+the+inferior+colliculus+and+primary+auditory+cortex.">Passive stimulation and behavioral training differentially transform temporal processing in the inferior colliculus and primary auditory cortex.</a> Journal of Neurophysiology. 117 (1), 47-64 (2017).</li><li>Sunwoo, W., Delgutte, B., Chung, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+bilateral+cochlear+implant+stimulation+partially+restores+neural+binaural+sensitivity+in+neonatally-deaf+rabbits.">Chronic bilateral cochlear implant stimulation partially restores neural binaural sensitivity in neonatally-deaf rabbits.</a> The Journal of Neuroscience. 41 (16), 3651-3664 (2021).</li><li>Mantokoudis, G., 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=Lamb+temporal+bone+as+a+surgical+training+model+of+round+window+cochlear+implant+electrode+insertion.">Lamb temporal bone as a surgical training model of round window cochlear implant electrode insertion.</a> Otology &amp; Neurotology. 37 (1), 52-56 (2016).</li><li>de Abajo, J., 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=Effects+of+implantation+and+reimplantation+of+cochlear+implant+electrodes+in+an+in+vivo+animal+experimental+model+(Macaca+fascicularis).">Effects of implantation and reimplantation of cochlear implant electrodes in an in vivo animal experimental model (Macaca fascicularis).</a> Ear and Hearing. 38 (1), 57-68 (2017).</li><li>Johnson, L. A., Della Santina, C. C., Wang, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Temporal+bone+characterization+and+cochlear+implant+feasibility+in+the+common+marmoset+(Callithrix+jacchus).">Temporal bone characterization and cochlear implant feasibility in the common marmoset (Callithrix jacchus).</a> Hearing Research. 290 (1-2), 37-44 (2012).</li><li>Yin, P., Li, S., Li, X. J., Yang, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+pathogenic+insights+from+large+animal+models+of+neurodegenerative+diseases.">New pathogenic insights from large animal models of neurodegenerative diseases.</a> Protein &amp; Cell. , (2022).</li><li>Yu, S. M., Wang, C. W., Zhao, D. M., Zhang, Q. C., Pei, D. Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Raising+and+pathogen+purification+of+Chinese+experimental+mini-pig.">Raising and pathogen purification of Chinese experimental mini-pig.</a> Laboratory Animal Science and Administration. 20, 44-46 (2003).</li><li>Guo, W., 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=The+morphology+and+electrophysiology+of+the+cochlea+of+the+miniature+pig.">The morphology and electrophysiology of the cochlea of the miniature pig.</a> The Anatomical Record. 298 (3), 494-500 (2015).</li><li>Christov, 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=Electric+compound+action+potentials+(ECAPs)+and+impedances+in+an+open+and+closed+operative+site+during+cochlear+implantation.">Electric compound action potentials (ECAPs) and impedances in an open and closed operative site during cochlear implantation.</a> Cochlear Implants International. 20 (1), 23-30 (2019).</li><li>Zhong, L. L., 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=Inner+ear+structure+of+miniature+pigs+measured+by+multi-planar+reconstruction+techniques.">Inner ear structure of miniature pigs measured by multi-planar reconstruction techniques.</a> American Journal of Translational Research. 10 (3), 709-717 (2018).</li><li>The Lancet. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hearing+loss:+An+important+global+health+concern.">Hearing loss: An important global health concern.</a> The Lancet. 387 (10036), 2351 (2016).</li><li>Guo, R., 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=Cochlear+implant-based+electric-acoustic+stimulation+modulates+neural+stem+cell-derived+neural+regeneration.">Cochlear implant-based electric-acoustic stimulation modulates neural stem cell-derived neural regeneration.</a> Journal of Materials Chemistry B. 9 (37), 7793-7804 (2021).</li><li>Gabrielpillai, J., Geissler, C., Stock, B., St&#246;ver, T., Diensthuber, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Growth+hormone+promotes+neurite+growth+of+spiral+ganglion+neurons.">Growth hormone promotes neurite growth of spiral ganglion neurons.</a> Neuroreport. 29 (8), 637-642 (2018).</li><li>Li, H., 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=Guided+growth+of+auditory+neurons:+Bioactive+particles+towards+gapless+neural+-+electrode+interface.">Guided growth of auditory neurons: Bioactive particles towards gapless neural - electrode interface.</a> Biomaterials. 122, 1-9 (2017).</li><li>Wille, I., 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=Development+of+neuronal+guidance+fibers+for+stimulating+electrodes:+Basic+construction+and+delivery+of+a+growth+factor.">Development of neuronal guidance fibers for stimulating electrodes: Basic construction and delivery of a growth factor.</a> Frontiers in Bioengineering and Biotechnology. 10, 776890 (2022).</li></ol>

The authors declare that they have no conflicts of interest.

Around 15% of the world's population have some degree of hearing loss, and over 5% have disabling hearing loss21. CI provision is the most efficient treatment for both adult and pediatric patients with severe and profound sensorineural hearing loss. As the first successful implantable cranial nerve stimulator, over the past 2 decades, CIs have offered thousands of people with hearing loss the opportunity to return to the world of sound and (re)integrate into mainstream society. Even though CIs are...

According to the World Health Organization (WHO), over 1 billion people are at risk of hearing loss globally, and it is estimated that, by 2050, one out of four people will suffer hearing loss1. Over the last 2 decades, CIs have been the most effective intervention for people with permanent severe and profound sensorineural hearing loss (SNHL). A CI converts physical signals of sound into bioelectrical signals that stimulate the spiral ganglion neurons (SGNs), bypassing hair cells. Over time, the indications for a CI have been broadened so that they now include people with residual hearing, unilateral hearing loss, and very old or young people2,3,4. Meanwhile, totally implantable CIs and advanced arrays have been developed5. There is, however, no economically feasible large animal model for investigating the electrophysiology and histopathology of the inner ear with a CI. This lack of a large animal model limits research seeking to improve CIs and gain insights into the electrophysiological impact of CIs on the inner ear.
Several rodent animal models have been applied in CI research, such as mouse6, gerbil7, rat8, and guinea pig9; however, the characteristics of morphology and electrophysiological responses are different from that in humans. Cochlear structures of animal models traditionally used for CI studies, such as cats, guinea pigs, and other animals, differ greatly from those of human cochlear structures10. Although array insertion has been conducted on cats11 and rabbits12, because of their smaller cochleae, this was done with arrays that were not designed for use in humans. Several large animal models have also been explored for CI. Lambs are well suited as a training model for atraumatic cochlear implantation, but the smaller size of the cochlea makes full array insertion impossible13. Primates might be the most suitable animals for CI research because of their anatomical similarity to humans14,15; however, the sexual maturity of monkeys is delayed (4-5 years), the gestation period is up to about 165 days, and each female usually produces only one offspring per year16. These reasons, and the expensive cost, hinder the extensive application of primates in CI research.
In contrast, pigs reach sexual maturity at 5-8 months and have a gestation period of ~114 days, making pigs more accessible for CI research as a large animal model16. Bama mini pigs (mini-pigs) originated from a small-sized pig species in China in 1985, whose genetic background is well understood. They are characterized by an inherent small size, early sexual maturity, rapid breeding, and ease of management17. The mini-pig is an ideal model for otology and audiology because of its similarity to humans in morphology and electrophysiology18. The scala tympani length of a Bama mini-pig is 38.58 mm, which is close to the 36 mm length in humans10. The mini-pig cochlea has 3.5 turns, which is similar to the 2.5-3 turns seen in humans10. In addition to morphology, the electrophysiology of Bama mini-pigs is also highly similar to that of humans18. Therefore, in the present study, we inserted arrays designed for human use into the mini-pig cochlea via the round window membrane and followed a similar surgical approach to that used in human CI recipients. Intraoperative ECAP measurements were applied to evaluate the procedure. The process we describe herein could be used both for preclinical translational research associated with CIs and as a platform for resident training.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>0.5 mm diamond burr</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>1 mm diamond burr</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>5 mm diamond burr</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>2-0 suture silk</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>3D Slicer image computing platform</td>
			<td></td>
			<td></td>
			<td>3D reconstruction of CT image</td>
		</tr>
		<tr>
			<td>Alcohol</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Bipolar cautery</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Bipolar electrocoagulation</td>
			<td></td>
			<td></td>
			<td>Stop bleeding</td>
		</tr>
		<tr>
			<td>CI designed for human use (CONCERTO FLEX28)</td>
			<td>MED-EL</td>
			<td>&#160;Concerto F28</td>
			<td></td>
		</tr>
		<tr>
			<td>Dressing forceps</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>ECG monitor</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Iodine tincture</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td></td>
			<td></td>
			<td>3.6 mL/h</td>
		</tr>
		<tr>
			<td>Laryngoscope</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>MAESTRO Software</td>
			<td>MED-EL</td>
			<td></td>
			<td>Measure ECAP responses</td>
		</tr>
		<tr>
			<td>Micro forceps</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Micro spatula</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Mosquito forceps</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Needle holder</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Needle probe</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Negative pressure suction device</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Otological surgical instruments&#160;</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Respiratory Anesthesia Machine</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Scalpel with blade No. 15</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Scissors</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Shaver</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Stimulation device (MAX Programming Interface)</td>
			<td>MED-EL</td>
			<td></td>
			<td>Measure ECAP responses</td>
		</tr>
		<tr>
			<td>Surgery microscope</td>
			<td>Leica</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical drill</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical Power Device</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Tiletamine and zolazepan</td>
			<td></td>
			<td></td>
			<td>10-15 mg/kg</td>
		</tr>
		<tr>
			<td>Tissue forceps</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Trachea cannula</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

the miniature pig: a large animal model for cochlear implant research

Cochlear implants (CI) are the most effective method to treat people with severe-to-profound sensorineural hearing loss. Although CIs are used worldwide, no standard model exists for investigating the electrophysiology and histopathology in patients or animal models with a CI or for evaluating new models of electrode arrays. A large animal model with cochlea characteristics similar to those of humans may provide a research and evaluation platform for advanced and modified arrays before their use in humans. 
To this end, we established standard CI methods with Bama mini-pigs, whose inner ear anatomy is highly similar to that of humans. Arrays designed for human use were implanted into the mini pig cochlea through a round window membrane, and a surgical approach followed that was similar to that used for human CI recipients. Array insertion was followed by evoked compound action potential (ECAP) measurements to evaluate the function of the auditory nerve. This study describes the preparation of the animal, surgical steps, array insertion, and intraoperative electrophysiological measurements. 
The results indicated that the same CI used for humans could be easily implanted in mini-pigs via a standardized surgical approach and yielded similar electrophysiological outcomes as measured in human CI recipients. Mini-pigs could be a valuable animal model to provide initial evidence of the safety and potential performance of novel electrode arrays and surgical approaches before applying them to human beings.

The integrity (Figure 4A)&#160;and impedances (Figure 4B) of the CI were confirmed by MAESTRO Software. ECAP results showed that all 12 electrodes demonstrated good neural responses (Figure 4C), meaning the electrode array was well attached to the cochlear axis and stimulated the auditory nerve. Figure 5 demonstrates postoperative 3D reconstructed electrode coils in the right cochlea. The array did not ...

Watch this Scientific Journal Video about The Miniature Pig: A Large Animal Model for Cochlear Implant Research at JoVE.com

The Miniature Pig: A Large Animal Model for Cochlear Implant Research

Miniature pigs (mini-pigs) are an ideal large animal model for research into cochlear implants. Cochlear implantation surgery in mini-pigs can be utilized to provide initial evidence of the safety and potential performance of novel electrode arrays and surgical approaches in a living system similar to human beings.

Cochlear implants (CI) are the most effective method to treat people with severe-to-profound sensorineural hearing loss. Although CIs are used worldwide, ...

the-miniature-pig-a-large-animal-model-for-cochlear-implant-research

Research

JoVE Journal

Medicine

2.5K Views.  Chinese PLA General Hospital. Miniature pigs (mini-pigs) are an ideal large animal model for research into cochlear implants. Cochlear implantation surgery in mini-pigs can be utilized to provide initial evidence of the safety and potential performance of novel electrode arrays and surgical approaches in a living system similar to human beings.

Video: The Miniature Pig: A Large Animal Model for Cochlear Implant Research

An Isolated Working Heart System for Large Animal Models

Since its introduction in the late 19th century, the Langendorff isolated heart perfusion apparatus, and the subsequent development of the working heart model, have been invaluable tools for studying cardiovascular function and disease1-15. Although the Langendorff heart preparation can be used for any mammalian heart, most studies involving this apparatus use small animal models (e.g., mouse, rat, and rabbit) due to the increased complexity of systems for larger mammals1,3,11. One major difficulty is ensuring a constant coronary perfusion pressure over a range of different heart sizes &#8211; a key component of any experiment utilizing this device1,11. By replacing the classic hydrostatic afterload column with a centrifugal pump, the Langendorff working heart apparatus described below allows for easy adjustment and tight regulation of perfusion pressures, meaning the same set-up can be used for various species or heart sizes. Furthermore, this configuration can also seamlessly switch between constant pressure or constant flow during reperfusion, depending on the user&#8217;s preferences. The open nature of this setup, despite making temperature regulation more difficult than other designs, allows for easy collection of effluent and ventricular pressure-volume data.

Most studies involving the Langendorff apparatus use small animal models due to the increased complexity of systems for larger mammals. We describe a Langendorff system for large animal models that allows for use across a range of species, including humans, and relatively easy data acquisition.

Since its introduction in the late 19th century, the Langendorff isolated heart perfusion apparatus, and the subsequent development of the working heart ...

Techniques for Processing Eyes Implanted with a Retinal Prosthesis for Localized Histopathological Analysis: Part 2 Epiretinal Implants with Retinal Tacks

Retinal prostheses for the treatment of certain forms of blindness are gaining traction in clinical trials around the world with commercial devices currently entering the market. In order to evaluate the safety of these devices, in preclinical studies, reliable techniques are needed. However, the hard metal components utilised in some retinal implants are not compatible with traditional histological processes, particularly in consideration for the delicate nature of the surrounding tissue. Here we describe techniques for assessing the health of the eye directly adjacent to a retinal implant secured epiretinally with a metal tack.
Retinal prostheses feature electrode arrays in contact with eye tissue. The most commonly used location for implantation is the epiretinal location (posterior chamber of the eye), where the implant is secured to the retina with a metal tack that penetrates all the layers of the eye. Previous methods have not been able to assess the proximal ocular tissue with the tack in situ, due to the inability of traditional histological techniques to cut metal objects. Consequently, it has been difficult to assess localized damage, if present, caused by tack insertion.
Therefore, we developed a technique for visualizing the tissue around a retinal tack and implant. We have modified an established technique, used for processing and visualizing hard bony tissue around a cochlear implant, for the soft delicate tissues of the eye. We orientated and embedded the fixed eye tissue, including the implant and retinal tack, in epoxy resin, to stabilise and protect the structure of the sample. Embedded samples were then ground, polished, stained, and imaged under various magnifications at incremental depths through the sample. This technique allowed the reliable assessment of eye tissue integrity and cytoarchitecture adjacent to the metal tack.

Here we describe histological techniques for visualising ocular tissue directly adjacent to a metal epiretinal tack and retinal prosthesis.

Retinal prostheses for the treatment of certain forms of blindness are gaining traction in clinical trials around the world with commercial devices ...

Surgical Angiogenesis in Porcine Tibial Allotransplantation: A New Large Animal Bone Vascularized Composite Allotransplantation Model

Segmental bone loss resulting from trauma, infection malignancy and congenital anomaly remains a major reconstructive challenge. Current therapeutic options have significant risk of failure and substantial morbidity.
Use of bone vascularized composite allotransplantation (VCA) would offer both a close match of resected bone size and shape and the healing and remodeling potential of living bone. At present, life-long drug immunosuppression (IS) is required. Organ toxicity, opportunistic infection and neoplasm risks are of concern to treat such non-lethal indications.
We have previously demonstrated that bone and joint VCA viability may be maintained in rats and rabbits without the need of long-term-immunosuppression by implantation of recipient derived vessels within the VCA. It generates an autogenous, neoangiogenic circulation with measurable flow and active bone remodeling, requiring only 2 weeks of IS. As small animals differ from man substantially in anatomy, bone physiology and immunology, we have developed a porcine bone VCA model to evaluate this technique before clinical application is undertaken. Miniature swine are currently widely used for allotransplantation research, given their immunologic, anatomic, physiologic and size similarities to man. Here, we describe a new porcine orthotopic tibial bone VCA model to test the role of autogenous surgical angiogenesis to maintain VCA viability.
The model reconstructs segmental tibial bone defects using size- and shape-matched allogeneic tibial bone segments, transplanted across a major swine leukocyte antigen (SLA) mismatch in Yucatan miniature swine. Nutrient vessel repair and implantation of recipient derived autogenous vessels into the medullary canal of allogeneic tibial bone segments is performed in combination with simultaneous short-term IS. This permits a neoangiogenic autogenous circulation to develop from the implanted tissue, maintaining flow through the allogeneic nutrient vessels for a short time. Once established, the new autogenous circulation maintains bone viability following cessation of drug therapy and subsequent nutrient vessel thrombosis.

Currently any kind of vascularized composite allotransplantation depends on long-term-immunosuppression, difficult to support for non-life-critical indications. We present a new porcine tibial VCA model that can be used to study bone VCA and demonstrate the use of surgical angiogenesis to maintain bone viability without the need of long-term immune-modulation.

Segmental bone loss resulting from trauma, infection malignancy and congenital anomaly remains a major reconstructive challenge. Current therapeutic ...

Cochlear Implantation in the Guinea Pig

Cochlear implants are highly efficient devices that can restore hearing in subjects with profound hearing loss. Due to improved speech perception outcomes, candidacy criteria have been expanded over the last few decades. This includes patients with substantial residual hearing that benefit from electrical and acoustical stimulation of the same ear, which makes hearing preservation during cochlear implantation an important issue. Electrode impedances and the related issue of energy consumption is another major research field, as progress in this area could pave the way for fully implantable auditory prostheses. To address these issues in a systematic way, adequate animal models are essential. Therefore, the goal of this protocol is to provide an animal model of cochlear implantation, which can be used to address various research questions. Due to its large tympanic bulla, which allows easy surgical access to the inner ear, as well as its hearing range which is relatively similar to the hearing range of humans, the guinea pig is a commonly used species in auditory research. Cochlear implantation in the guinea pig is performed via a retroauricular approach. Through the bullostomy a cochleostomy is drilled and the cochlear implant electrode is inserted into the scala tympani. This electrode can then be used for electrical stimulation, determination of electrode impedances and the measurement of compound action potentials of the auditory nerve. In addition to these applications, cochlear implant electrodes can also be used as drug delivery devices, if a topical delivery of pharmaceutical agents to the cells or fluids of the inner ear is intended.

The goal of this protocol is to provide an animal model of cochlear implantation, which can be used to address a multitude of research questions. Potential applications include the evaluation of pharmaceutical interventions or electrical stimulation for beneficial effects on hearing thresholds or electrode impedances.

Cochlear implants are highly efficient devices that can restore hearing in subjects with profound hearing loss. Due to improved speech perception ...

Assessment of Plasma Coagulation on Liver Tissue in a Large Animal Model In Vivo

Plasma coagulation as a form of electrocautery is used in liver surgery for decades to seal the large liver cut surface after major hepatectomy to prevent hemorrhages at a later stage. The exact effects of plasma coagulation on liver tissue are only poorly examined. In our porcine model, the coagulation effects can be examined close to the clinical application. A combined laser Doppler flowmeter and spectrophotometer documents microcirculation changes during coagulation at 8 mm tissue depth noninvasively, providing quantifiable information about hemostasis beyond the subjective clinical impression. The temperature at coagulation site is assessed with an infrared thermometer prior and post coagulation and with a thermographic camera during coagulation, a measurement of the gas beam temperature is not possible due to the upper threshold of the devices. The depth of coagulation is measured microscopically on hematoxylin/eosin stained sections after calibration with an object micrometer and gives an exact information about the power setting-coagulation depth-relation. The sealing effect is examined on the bile ducts as it is not possible for a plasma coagulator to seal larger blood vessels. Burst pressure experiments are carried out on explanted organs to rule out blood pressure related effects.

Assessment of Plasma Coagulation on Liver Tissue in a Large Animal Model In Vivo

Here we present a protocol to experimentally assess plasma coagulation in liver tissue in vivo. In a porcine model, microcirculation is examined by laser Doppler, coagulation depth is measured histologically, temperature at coagulation site by infrared thermometer and thermographic camera, and duct sealing effect is documented by burst pressure experiments.

Plasma coagulation as a form of electrocautery is used in liver surgery for decades to seal the large liver cut surface after major hepatectomy to prevent ...

Ferric Chloride-induced Canine Carotid Artery Thrombosis: A Large Animal Model of Vascular Injury

Occlusive arterial thrombosis leading to cerebral ischemic stroke and myocardial infarction contributes to ~13 million deaths every year globally. Here, we have translated a vascular injury model from a small animal into a large animal (canine), with slight modifications that can be used for pre-clinical screening of prophylactic and thrombolytic agents. In addition to the surgical methods, the modified protocol describes the step-by-step methods to assess carotid artery canalization by angiography, detailed instructions to process both the brain and carotid artery for histological analysis to verify carotid canalization and cerebral hemorrhage, and specific parameters to complete an assessment of downstream thromboembolic events by utilizing magnetic resonance imaging (MRI). In addition, specific procedural changes from the previously well-established small animal model necessary to translate into a large animal (canine) vascular injury are discussed.

Here, we present the modifications necessary to a well characterized and commonly used small animal ferric chloride-induced (FeCl3) carotid artery injury model for use in a large animal vascular injury model. The resulting model can be utilized for pre-clinical trial assessment of both prophylactic and thrombolytic pharmacological and mechanical interventions.

Occlusive arterial thrombosis leading to cerebral ischemic stroke and myocardial infarction contributes to ~13 million deaths every year globally. Here, ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

Flow Cytometry Analysis of Immune Cell Subsets within the Murine Spleen, Bone Marrow, Lymph Nodes and Synovial Tissue in an Osteoarthritis Model

Osteoarthritis (OA) is one of the most prevalent musculoskeletal diseases, affecting patients suffering from pain and physical limitations. Recent evidence indicates a potential inflammatory component of the disease, with both T-cells and monocytes/macrophages potentially associated with the pathogenesis of OA. Further studies postulated an important role for subsets of both inflammatory cell lineages, such as Th1, Th2, Th17, and T-regulatory lymphocytes, and M1, M2, and synovium-tissue-resident macrophages. However, the interaction between the local synovial and systemic inflammatory cellular response and the structural changes in the joint is unknown. To fully understand how T-cells and monocytes/macrophages contribute towards OA, it is important to be able to quantitively identify these cells and their subsets simultaneously in synovial tissue, secondary lymphatic organs and systemically (the spleen and bone marrow). Nowadays, the different inflammatory cell subsets can be identified by a combination of cell-surface markers making multi-color flow cytometry a powerful technique in investigating these cellular processes. In this protocol, we describe detailed steps regarding the harvest of synovial tissue and secondary lymphatic organs as well as generation of single cell suspensions. Furthermore, we present both an extracellular staining assay to identify monocytes/macrophages and their subsets as well as an extra- and intra-cellular staining assay to identify T-cells and their subsets within the murine spleen, bone marrow, lymph nodes and synovial tissue. Each step of this protocol was optimized and tested, resulting in a highly reproducible assay that can be utilized for other surgical and non-surgical OA mouse models.

Here, we describe a detailed and reproducible flow cytometry protocol to identify monocyte/macrophage and T-cell subsets using both extra- and intracellular staining assays within the murine spleen, bone marrow, lymph nodes and synovial tissue, utilizing an established surgical model of murine osteoarthritis.

Osteoarthritis (OA) is one of the most prevalent musculoskeletal diseases, affecting patients suffering from pain and physical limitations. Recent ...

A Large Animal Model for Acute Kidney Injury by Temporary Bilateral Renal Artery Occlusion

Acute kidney injury (AKI) is associated with higher risk for morbidity and mortality post-operatively. Ischemia-reperfusion injury (IRI) is the most common cause of AKI. To mimic this clinical scenario, this study presents a highly reproducible large animal model of renal IRI in swine using temporary percutaneous bilateral balloon-catheter occlusion of the renal arteries. The renal arteries are occluded for 60 min by introducing the balloon-catheters through the femoral and carotid artery and advancing them into the proximal portion of the arteries. Iodinated contrast is injected in the aorta to assess any opacification of the kidney vessels and confirm the success of the artery occlusion. This is furtherly confirmed by the flattening of the pulse waveform at the tip of the balloon catheters. The balloons are deflated and removed after 60 min of bilateral renal artery occlusion, and the animals are allowed to recover for 24 h. At the end of the study, plasma creatinine and blood urea nitrogen significantly increase, while eGFR and urine output significantly decrease. The need for iodinated contrast is minimal and does not affect renal function. Bilateral renal artery occlusion better mimics the clinical scenario of perioperative renal hypoperfusion, and the percutaneous approach minimizes the impact of the inflammatory response and the risk of infection seen with an open approach, such as a laparotomy. The ability to create and reproduce this clinically relevant swine model eases the clinical translation to humans.

This study presents a highly reproducible large animal model of renal ischemia-reperfusion injury in swine using temporary percutaneous bilateral balloon-catheter occlusion of the renal arteries for 60 min and reperfusion for 24 h.

Acute kidney injury (AKI) is associated with higher risk for morbidity and mortality post-operatively. Ischemia-reperfusion injury (IRI) is the most ...

Large Animal Model for Evaluating the Efficacy of the Gene Therapy in Ischemic Heart

Coronary artery disease is one of the significant causes of mortality and morbidity worldwide. Despite the progression of current therapeutics, a considerable proportion of coronary artery disease patients remain symptomatic. Gene therapy-mediated therapeutic angiogenesis offers a novel therapeutic method for improving myocardial perfusion and relieving symptoms. Gene therapy with different angiogenic factors has been studied in few clinical trials. Due to the novelty of the method, the progress of myocardial gene therapy is a continuous path from bench to bedside. Therefore, large animal models are needed for evaluating the safety and efficacy. The more the large animal model identifies the original disease and the endpoints used in clinics, the more predictable outcomes are from clinical trials. Here, we introduce a large animal model for evaluating the efficacy of the gene therapy in the ischemic porcine heart. We use clinically relevant imaging methods such as ultrasound imaging and 15H2O-PET. For targeting the gene transfers into the desired area, electroanatomical mapping is used. The aim of this method is: (1) to mimic chronic coronary artery disease, (2) to induce therapeutic angiogenesis at hypoxic areas of the heart, and (3) to evaluate the safety and efficacy of the gene therapy by using relevant endpoints.

Myocardial gene therapy for ischemic heart disease holds great promise for future therapeutics. Here, we introduce a large animal model for evaluating the efficacy of gene therapy in the ischemic heart.