This study was supported by NIH-NINDS (R21NS130241), IND DEPT HLTH (55051, 74247, 74244), and US ARMY (HT94252310700).

The animal study protocol was approved by the Institutional Animal Care and Use Committee of Indiana University School of Medicine. Approval Code: 21098MD/R/MSS/HZ Approval Date: 29 September 2021.
1. Preparation of catheter
<ol>
	<li>Cut a 30 cm polyethylene PE-30 tube (.017 inch x .030 inch). Use a lighter to flare one end of the tube, ensuring it does not touch the flame and withdrawing the lighter once the tube has formed an appropriately round, bell-shaped tip. Any residue from the flaring procedure is then carefully cleaned or removed before proceeding with the surgery.</li>
	<li>Carefully insert about 3/4 of the 25G needle into the other end of the tube. Prepare a 1 mL syringe and fill it with sterile 0.9% NaCl. Connect the syringe to the 25 G needle.</li>
	<li>Gently infuse saline to check for a proper round tip and no leakage from the ends of the needle. Ensure that no pressure is felt and saline flows smoothly through the catheter.</li>
</ol>
2. Preparation of electrodes
<ol>
	<li>Prepare 2 steel wires of 20 cm in length. Take the steel wires and apply sand polish to both ends of the coating zone to strip 5 mm of the wire.</li>
	<li>Take a 25G needle and insert it on one side of the wire. Make sure to insert the needle carefully to avoid damaging the wire. Bend the stripped part of the wire like a hook. The hook will help to connect the wire to the EUS muscle.</li>
	<li>Use solder to attach the pin to the other end of the striped wire. Soldering will help secure the pin to the wire and ensure a strong connection. Make sure to heat the tin-lead soldering until it melts and covers the wire and pin.</li>
</ol>
3. Preparation of animal
<ol>
	<li>House female C57BL/6 mouse (8 weeks old, 18-20 g body weight) in the animal facility according to Institutional Animal Care with a 12 h light-dark cycle and unlimited access to water and standard food pellets. 
	NOTE: Prior to the experimental procedures, mice were acclimated to the restraint apparatus over a period of three days. During this time, each mouse was placed in the restraint for 10-15 minutes daily, allowing them to become familiar with the setup. Mice were monitored for signs of distress, and adjustments were made to promote a calm experience. This acclimation aimed to reduce anxiety and enhance animal welfare during the experiments</li>
</ol>
4. Anesthesia during surgery
<ol>
	<li>Place the animals in a 2% isoflurane and pure oxygen (1 L/min) chamber. Confirm the animal&#39;s complete anesthesia using negative toe-pinch examination before transferring it to the mask. Once confirmed, change the gas condition to a mask.</li>
	<li>Ensure that the anesthesia mask is fixed in the 
	appropriate position on the sterile surgical field. Place the animal supine on the heating pad covered with the sterile drape with its nose in an inhalation small mask (0.8-1 L/min with 2% isoflurane) to continue administering anesthesia.</li>
</ol>
5. Surgical preparation
<ol>
	<li>Fix the limbs of the animal with tape. Use an electric razor to shave the fur of the lower abdomen and around the urethral meatus (genital region).&#160;Administer 3 mg/kg Meloxicam (5 mg/ml) subcutaneously as a pre-emptive analgesic to the mouse before surgery to ensure effective pain management.</li>
	<li>Apply an eye ointment to prevent any potential dryness in the eyes. Prepare the shaved area using the povidone-iodine solution and wipe away the solution with 70% ethanol. Put a sterile drape on the surgery region.</li>
	<li>Autoclave all surgical tools and electrode wires&#160;at 121&#176;C for 15&#8211;20 minutes to ensure sterility prior to the procedure. Soak the polyethylene tube in a&#160;glutaraldehyde or ethylene oxide&#160;for&#160;20&#8211;30 minutes,&#160;then rinse thoroughly with sterile saline to remove any residues</li>
</ol>
6. Surgical procedure
<ol>
	<li>Bladder catheter implantation
	<ol>
		<li>Under the surgical microscope, utilizing Metzenbaum straight, blunt scissors, create a 1-2 cm incision in the midline of the abdominopelvic skin. Proceed to incise the fascia and muscles in the midline to expose the bladder through the incisional wound.</li>
		<li>Once the bladder is visible through the incisional wound, proceed to retract any surrounding organs or tissues as needed to obtain a clear view of the surgical field. Take care to avoid any unnecessary manipulation or tension on the bladder, as this can lead to complications such as urinary leakage or injury to surrounding structures.</li>
		<li>Grasp the bladder dome and place a purse string by utilizing a 5-0 nonabsorbable monofilament suture with a taper-tip needle.</li>
		<li>Using micro scissors, create a small cystostomy within the purse string and make a hole until urine flows out.</li>
		<li>Grasp the round tip end of the catheter and insert it through the hole. Once the tip of the tube has passed through the hole, suture the purse string around the tube. Then, gently pull the tube outward until the tip is felt under the suture.</li>
		<li>Slowly infuse 1 mL of saline from the other end of the tube to distend the bladder. Check for any leakage around the catheter. If leakage is present, place an additional suture.</li>
		<li>Once the saline comes out of the urethra, withdraw the saline to decompress the bladder.</li>
	</ol>
	</li>
	<li>EUS electrodes implantation (Figure 1)
	<ol>
		<li>Use surgical scissors to extend the abdominal incision up to the pelvic floor.</li>
		<li>In line with the bladder, move the muscles and membranes to the pudendal canals and locate the urethra and external sphincter muscle. Be careful not to hurt iliac and middle caudal vessels and pudendal nerves.</li>
		<li>Puncture the skin bilaterally, 1 cm away from the urethral meatus, using the needle containing the electrode.</li>
		<li>Carefully grab the hook tip with forceps and gently pull the needle away from the skin.</li>
		<li>Using the electrode&#39;s tip, carefully hook the EUS muscle bilaterally. Avoid punching too deep, as this may harm the muscle, which could lead to possible urine leakage.</li>
		<li>Use the 3-0 nonabsorbable monofilament to suture the pelvic and abdomen muscles and skin.</li>
	</ol>
	</li>
	<li>Waterproofing the skin
	<ol>
		<li>Apply a thin layer of cyanoacrylate glue to the skin where the electrodes exit to fix the electrodes in place.</li>
		<li>Apply the cyanoacrylate glue 1 cm away surrounding the urethral meatus and 3 cm further extending to the abdomen and sutured region. To avoid contact with the glue, carefully hold the meatus up with forceps.</li>
		<li>Use a 0.5-10 &#181;L micropipette to withdraw the accelerator liquid to dry the glue. 
		CAUTION: Accelerator liquid is a combustible liquid.</li>
		<li>Add the accelerator liquid to ensure proper adhesive reaction. This will help to dry the glue more quickly and ensure that it sets securely.</li>
	</ol>
	</li>
	<li>Urodynamic preparation
	<ol>
		<li>Prepare an inverted polystyrene weighing boat 4.5 cm in length, width, and depth. Cut it into a triangle shape with a base of 4 cm to put the mouse&#39;s urethra meatus in this space. Put the disposable base mold, 37 mm x 24 mm x 5 mm, under the space for collecting the urine.</li>
		<li>Reposition the mouse in a prone position and carefully move it onto a custom-made plate equipped with a gas mask.</li>
		<li>Ensure that the urethral meatus is properly positioned within the groove. After gently restraining the head and limbs of the mouse with tape, ensuring that the chest and nose are unobstructed for breathing, remove the inhalant anesthesia. Use pulse oximetry or monitor chest wall movement to check respiration. Place the plate on a heating pad for heat support and allow the mouse to recover until it regains full consciousness (Figure 2).</li>
		<li>Perform cystometry only when the mouse is fully awake, which is at least 40 min after recovery from anesthesia.</li>
	</ol>
	</li>
</ol>
7. Cystometry and EUS-EMG recording preparation
<ol>
	<li>Set up and calibrate the infusion pump according to the manufacturer&#39;s instructions.</li>
	<li>Take a 20 mL syringe with a diameter of 19.05 mm and fill it with sterile 0.9% NaCl at room temperature. Secure the syringe to the infusion pump. Set the infusion speed to 0.01 mL/min.</li>
	<li>Connect the syringe by the PE-30 tube to one side of the three-way connector. Connect the bladder catheter to the other side to a pressure transducer. Before connecting the bladder catheter, make sure to remove all air bubbles.</li>
	<li>Fix the pressure transducer at the same level as the mouse bladder. The pressure transducer is connected via an amplifier to the data acquisition system.</li>
	<li>Attach one ground line hook to the skin and the other to the electrode connector sites. Record the pressure in the software.</li>
	<li>After starting the software, check the intravesical pressure (IVP) and EUS-EMG signals. Save the sample name and set the time.</li>
	<li>Start the pump infusion. Record the signals for 40 minutes. At the end of the recording, euthanize the animal, as this is an end-stage procedure.</li>
</ol>

Precise Urodynamic Measurement in Awake Mice Using Cyanoacrylate Skin Barriers

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O., 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=Best+practices+for+cystometric+evaluation+of+lower+urinary+tract+function+in+muriform+rodents.">Best practices for cystometric evaluation of lower urinary tract function in muriform rodents.</a> Neurourol Urodyn. 39 (6), 1868-1884 (2020).</li><li>Hashimoto, 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=Sex+differences+in+lower+urinary+tract+function+in+mice+with+or+without+spinal+cord+injury.">Sex differences in lower urinary tract function in mice with or without spinal cord injury.</a> Neurourol Urodyn. 43 (1), 267-275 (2024).</li><li>Kadekawa, K., 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=Characterization+of+bladder+and+external+urethral+activity+in+mice+with+or+without+spinal+cord+injury-a+comparison+study+with+rats.">Characterization of bladder and external urethral activity in mice with or without spinal cord injury-a comparison study with rats.</a> Am J Physiol Regul Integr Comp Physiol. 310 (8), R752-R758 (2016).</li><li>Lee, 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=The+effects+of+periurethral+muscle-derived+stem+cell+injection+on+leak+point+pressure+in+a+rat+model+of+stress+urinary+incontinence.">The effects of periurethral muscle-derived stem cell injection on leak point pressure in a rat model of stress urinary incontinence.</a> Int Urogynecol J. 14, 31-37 (2003).</li><li>Mann-Gow, T. K., 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=Evaluating+the+procedure+for+performing+awake+cystometry+in+a+mouse+model.">Evaluating the procedure for performing awake cystometry in a mouse model.</a> J Vis Exp. (123), e55588 (2017).</li><li>Saito, 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=Time-dependent+progression+of+neurogenic+lower+urinary+tract+dysfunction+after+pinal+cord+injury+in+the+mouse+model.">Time-dependent progression of neurogenic lower urinary tract dysfunction after pinal cord injury in the mouse model.</a> Am J Physiol Renal Physioly. 321 (1), F26-F32 (2021).</li><li>Schneider, M. 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=A+novel+urodynamic+model+for+lower+urinary+tract+assessment+in+awake+rats.">A novel urodynamic model for lower urinary tract assessment in awake rats.</a> BJU Int. 115, 8-15 (2015).</li><li>Habib, A., Mehanna, A., Medra, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cyanoacrylate:+A+handy+tissue+glue+in+maxillofacial+surgery:+Our+experience+in+alexandria,+egypt.">Cyanoacrylate: A handy tissue glue in maxillofacial surgery: Our experience in alexandria, egypt.</a> J Maxillofac Oral Surg. 12, 243-247 (2013).</li><li>Sunjic Roguljic, V., Roguljic, L., Jukic, I., Kovacic, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+influence+of+wound+closure+techniques+after+surgical+decompression+in+patients+with+carpal+tunnel+syndrome+on+sleep+disturbance+and+life+quality:+A+prospective+comparison+of+surgical+techniques.">The influence of wound closure techniques after surgical decompression in patients with carpal tunnel syndrome on sleep disturbance and life quality: A prospective comparison of surgical techniques.</a> Clin Pract. 14 (2), 546-555 (2024).</li><li>Sohn, J. J., Gruber, T. M., Zahorsky-Reeves, J. L., Lawson, G. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+2-ethyl-cyanoacrylate+and+2-butyl-cyanoacrylate+for+use+on+the+calvaria+of+cd1+mice.">Comparison of 2-ethyl-cyanoacrylate and 2-butyl-cyanoacrylate for use on the calvaria of cd1 mice.</a> J Am Assoc Lab Anim Sci. 55 (2), 199-203 (2016).</li><li>Ren, 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=Injectable,+self-healing+hydrogel+adhesives+with+firm+tissue+adhesion+and+on-demand+biodegradation+for+sutureless+wound+closure.">Injectable, self-healing hydrogel adhesives with firm tissue adhesion and on-demand biodegradation for sutureless wound closure.</a> Sci Adv. 9 (33), 4327 (2023).</li><li>Ito, H., Pickering, A. E., Kanai, A., Fry, C. H., Drake, 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=Muro-neuro-urodynamics;+a+review+of+the+functional+assessment+of+mouse+lower+urinary+tract+function.">Muro-neuro-urodynamics; a review of the functional assessment of mouse lower urinary tract function.</a> Front Physiol. 8, 240395 (2017).</li><li>Abdelkhalek, A. S., Youssef, H. A., Saleh, A. S., Bollen, P., Zvara, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anesthetic+protocols+for+urodynamic+studies+of+the+lower+urinary+tract+in+small+rodents-a+systematic+review.">Anesthetic protocols for urodynamic studies of the lower urinary tract in small rodents-a systematic review.</a> PloS One. 16 (6), e0253192 (2021).</li><li>Saab, B. 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=Short-term+memory+impairment+after+isoflurane+in+mice+is+prevented+by+the+&#945;5+&#947;-aminobutyric+acid+type+a+receptor+inverse+agonist+l-655,708.">Short-term memory impairment after isoflurane in mice is prevented by the &#945;5 &#947;-aminobutyric acid type a receptor inverse agonist l-655,708.</a> J Am Soc Anesthesiol. 113 (5), 1061-1071 (2010).</li><li>Cannon, T. W., Damaser, M. S. <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+anesthesia+on+cystometry+and+leak+point+pressure+of+the+female+rat.">Effects of anesthesia on cystometry and leak point pressure of the female rat.</a> Life Sci. 69 (10), 1193-1202 (2001).</li><li>Weiss, D. 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=Morphology+of+the+external+genitalia+of+the+adult+male+and+female+mice+as+an+endpoint+of+sex+differentiation.">Morphology of the external genitalia of the adult male and female mice as an endpoint of sex differentiation.</a> Mol Cell Endocrinol. 354 (1-2), 94-102 (2012).</li><li>Leggat, P. A., Kedjarune, U., Smith, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Toxicity+of+cyanoacrylate+adhesives+and+their+occupational+impacts+for+dental+staff.">Toxicity of cyanoacrylate adhesives and their occupational impacts for dental staff.</a> Ind Health. 42 (2), 207-211 (2004).</li></ol>

The authors have nothing to disclose.

This urodynamic technique describes an improved procedure for measuring urine volume and EUS- EMG signal in awake and restrained mice. The presence of fur around the urethral meatus and abdominal area can interfere with the accuracy of the VV measurement by absorbing urine. Although the fur surrounding the urethral meatus and abdomen had been carefully shaved before surgery, the remaining small furs within these areas and the skin still absorbed urine, usually leaving a wet area in the abdomen after recording. This issue...

The storage and release of urine are dependent on the coordinated activity of the urinary bladder and external urethral sphincter (EUS). In some pathologies such as neurogenic bladder, both the bladder detrusor muscles and the sphincter can become dysfunctional, leading to significant bladder problems, especially after traumatic spinal cord injury(SCI)1.
Small rodents are commonly used as an experimental model to study the preclinical function of the lower urinary tract (LUT)2. Filling Cystometry (FC) and EUS electromyography (EUS-EMG) recording techniques can provide precise objective information depending on the choice of methods, accurate measurement, and interpretation of results3. Urodynamic tests are commonly used to evaluate the voiding volume (VV), voiding efficiency (VE), and bladder capacity4. VE measures how effectively the bladder can empty itself. It is calculated by dividing the voided volume by the sum of voided and residual volumes (VV+RV). On the other hand, bladder capacity is calculated by adding the VV (the amount of urine expelled during urination) to the RV (the amount of urine left in the bladder after urination)5. Therefore, the measurement of VV and RV are the keys to deducing other parameters.
Precisely measuring VV in mice during urodynamic tests presents various challenges. The urine of rodents, when physically restrained in a prone position, tends to be drawn downwards through the ventral abdominal wall due to the influence of gravity6. This phenomenon can lead to the absorption of urine by the abdominal fur and skin, which, in turn, underestimates the volume of urine excreted. Considering the small amount of urine produced by mouse, the impact of this absorbance on the accuracy of results is even more pronounced7. Furthermore, in models of SCI, VV is often lower than in normal mice due to the impact of the detrusor sphincter dyssynergia (DSD), which increases the risk of leak point pressures and urine absorption by the fur8. These factors have a significant impact on the results. Therefore, accurate measurement of VV and RV during terminal urodynamic studies in mice is crucial9. Currently, there is a lack of details in the methodologies provided in published literature about how to measure urine volume accurately in mouse models.
Cyanoacrylate adhesive is a type of glue that is commonly used in surgical procedures in human and animal models due to its quick and effective bonding properties10,11,12. This adhesive is particularly useful for closing wounds and lacerations, as it forms a strong and flexible bond when applied to the skin13. Moreover, it can be a great barrier against urine and wetness that may come into contact with fur and wounds11.
In this article, we have developed a novel and cost-effective technique that utilizes cyanoacrylate adhesive to achieve precise results in cystometry and EUS-EMG recordings in awake mice. This method will be beneficial in understanding the underlying causes of bladder dysfunction and devising more effective treatments for LUT disorders.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Accelerator</td>
			<td>BOB SMITH INDUSTRIES</td>
			<td>BSI-152</td>
			<td></td>
		</tr>
		<tr>
			<td>Cyanoacrylate&#160;</td>
			<td>TED PELLA, Inc</td>
			<td>14478</td>
			<td></td>
		</tr>
		<tr>
			<td>Disposable base mold</td>
			<td>TED PELLA, Inc</td>
			<td>27147-4</td>
			<td></td>
		</tr>
		<tr>
			<td>Infusion pump</td>
			<td>Harvard Apparatus PHD ULTRA</td>
			<td>70-3006</td>
			<td></td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Henry Schein Inc</td>
			<td>1182097</td>
			<td></td>
		</tr>
		<tr>
			<td>PIN</td>
			<td>World Precision Instruments</td>
			<td>5482</td>
			<td></td>
		</tr>
		<tr>
			<td>Polyethylene Tubing 30</td>
			<td>Braintree Scientific Inc</td>
			<td>PE30</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile Weighing Boat</td>
			<td>HEATHROW SCIENTIFIC</td>
			<td>797CK2</td>
			<td></td>
		</tr>
		<tr>
			<td>Windaq/Lite&#160;</td>
			<td>DATAQ INSTRUMENTS</td>
			<td>249022</td>
			<td></td>
		</tr>
	</tbody>
</table>

optimizing mouse urodynamic techniques for improved accuracy 

Accurate measurement of urinary parameters in awake mice is crucial for understanding lower urinary tract (LUT) dysfunction, particularly in conditions like neurogenic bladder post-traumatic spinal cord injury (SCI). However, conducting cystometry recordings in mice presents notable challenges. When mice are in a prone and restricted position during recording sessions, urine tends to be absorbed by the fur and skin, leading to an underestimation of voided volume (VV). The goal of this study was to enhance the accuracy of cystometry and external urethral sphincter electromyography (EUS-EMG) recordings in awake mice. We developed a unique method utilizing cyanoacrylate adhesive to create a waterproof skin barrier around the urethral meatus and abdomen, preventing urine absorption and ensuring precise measurements. Results show that after applying the cyanoacrylate, the sum of VV and RV remained consistent with the infused saline volume, and there were no wet areas observed post-experiment, indicating successful prevention of urine absorption. Additionally, the method simultaneously stabilized the electrodes connected with the external urethral sphincter (EUS), ensured stable electromyography (EMG) signals, and minimized artifacts caused by the movement of the awakened mouse and manipulation of the experimenter. Methodological details, results, and implications are discussed, highlighting the importance of improving urodynamic techniques in preclinical research.

Author Spotlight: Enhanced Urodynamic Method for Precise Urine Measurement in Awake Mice with Neurogenic Bladder

Optimizing Mouse Urodynamic Techniques for Improved Accuracy 

The goal of this study is to improve the urodynamic technology to increase the accuracy of the urine measurement. Accurate measurement of the urinary parameter awake mice is crucial understanding lower urinary tract dysfunction, particularly in condition like a neurogenic bladder, post-traumatic spinal injury. We developed a unique method utilizing cyanoacrylate adhesive to create a waterproof skin barrier around the and abdomen, preventing urine absorption and ensuring precise measurements of voided volume.We would use this more accurate assay to evaluate urinary function in the spinal cord injury animal models, and another neurodegenerative disease.

Cystometry and EUS-EMG activity tracings were used to analyze the data. The continuous cystometry method involves the infusion of saline into the bladder and simultaneously measuring the pressure and volume changes in the bladder. To measure VV, 0.4 mL of saline was infused at a speed of 0.01 mL/min, and urine was collected over 40 min in a cap. The post-void residual (PVR) can be obtained by aspirating the saline through the catheter. In normal mice without glue, the sum of VV and RV was often less than 0.4 mL. After th...

This protocol provides a guide for waterproofing the skin with cyanoacrylate to prevent urine absorption by fur and skin. It includes instructions for applying the glue to the skin, implanting a bladder catheter, and electrodes for cystometry and external urethral sphincter electromyography recordings in awake mice.

Accurate measurement of urinary parameters in awake mice is crucial for understanding lower urinary tract (LUT) dysfunction, particularly in conditions ...

optimizing-mouse-urodynamic-techniques-for-improved-accuracy

Research

JoVE Journal

Neuroscience

319 Views.  Indiana University School of Medicine. The goal of this study is to improve the urodynamic technology to increase the accuracy of the urine measurement. Accurate measurement of the urinary parameter awake mice is crucial understanding lower urinary tract dysfunction, particularly in condition like a neurogenic bladder, post-traumatic spinal injury. We developed a unique method utilizing cyanoacrylate adhesive to create a waterproof skin barrier around the and abdomen, preventing urine absorption and ensuring precise measurements o...

Video: Author Spotlight: Enhanced Urodynamic Method for Precise Urine Measurement in Awake Mice with Neurogenic Bladder

Preparation of Catheter and Electrodes for Cystometry and External Urethral Sphincter Electromyography Recordings

To begin, cut a 30-centimeter polyethylene PE-30 tube. Carefully insert about three-fourths of the 25-gauge needle into one end of the tube. After preparing a 1-milliliter syringe, fill it with sterile 0.9%sodium chloride solution, and connect it to the 25-gauge needle.Then, gently infuse saline to check for a proper round tip and no leakage from the ends of the needle. Prepare two steel wires, each 20 centimeters in length. Then, insert a 25-gauge needle on one side of the wire.Use solder to attach the pin to the other end of the striped wire.

Surgical Procedure for Bladder Catheter and EUS Electrode Implantation for External Urethral Sphincter Electromyography Recordings

To begin, fix the limbs of the anesthetized animal with tape. Use an electric razor to shave the fur of the lower abdomen and around the urethral meatus. Under the surgical microscope, use Metzenbaum straight blunt scissors to create a one to two centimeter incision in the midline of the abdominal pelvic skin.Incise the fascia and muscles in the midline to expose the bladder through the incisional wound. Once the bladder is visible through the wound, retract any surrounding organs or tissues, as needed, to obtain a clear view of the surgical field. Then, grasp the bladder dome and use a 5-0 non-absorbable monofilament suture with a taper tip needle to place a purse string.Using micro scissors, create a small cystostomy within the purse string and make a hole until urine flows out. Now, grasp the round tip end of the catheter and insert it through the hole. Once the tip of the tube has passed through the hole, suture the purse string around the tube.After that, gently pull the tube outward until the tip is felt under the suture. Then, slowly infuse one milliliter of saline from the other end of the tube to distend the bladder, and check for any leakage around the catheter. Once the saline comes out of the urethra, withdraw the saline to decompress the bladder.Use surgical scissors to extend the abdominal incision up to the pelvic floor. In line with the bladder, move the muscles and membranes to the pudendal canals and locate the urethra and external sphincter muscle. Using the needle containing the electrode, puncture the skin bilaterally one centimeter away from the urethral meatus.Then, carefully grab the hook tip with forceps and gently pull the needle away from the skin. Using the electrode's tip, carefully hook the external urethral sphincter muscle bilaterally. Use the 3-0 non-absorbable monofilament to suture the pelvic and abdomen muscles and skin.To fix the electrodes in place, apply a thin layer of cyanoacrylate glue to the skin where the electrodes exit. Next, use a 0.5 to 10 microliter micro pipette to withdraw the accelerator liquid to dry the glue. Then, add the accelerator liquid to ensure a proper adhesive reaction.Prepare an inverted polystyrene weighing boat, 4.5 centimeters in length, width and depth. Cut it into a triangle shape with a base of four centimeters to place the mouse's urethral meatus in this space. Put the disposable base mold under the space for collecting the urine.Then, reposition the mouse in a prone position and carefully move it onto a custom-made plate equipped with a gas mask. Ensure that the urethral meatus is properly positioned within the groove. Gently restrain the head and limbs of the mouse with tape and place the plate on a heating pad until the mouse regains full consciousness.

Cystometry and EUS-EMG Recording Preparation in Awake Mice

To begin, implant a bladder catheter and EUS electrode in the anesthetized mouse. After waterproofing the mouse's skin, prepare it for the urodynamic procedure. Ensure that the mouse is fully recovered from anesthesia, monitoring respiratory function throughout.Then take a 20-milliliter syringe and fill it with sterile 0.9%sodium chloride solution at room temperature. Secure the syringe to the infusion pump and set the infusion speed to 0.01 milliliters per minute. Now connect the syringe by the PE-30 tube to one side of the three-way connector.Connect the bladder catheter to the other side and to a pressure transducer. Fix the pressure transducer at the same level as the mouse bladder. Then attach one ground line hook to the skin and the other to the electrode connector sites.Record the pressure in the software. After starting the software, check the intravesical pressure and external urethral sphincter electromyography signals. Save the sample name and set the time.Finally, start the pump infusion and record the signals.