Name: Ex Vivo Method for Assessing the Mouse Reproductive Tract Spontaneous Motility and a MATLAB-based Uterus Motion Tracking Algorithm for Data Analysis (Video) | JoVE Journal
Uploaded: 2019-09-01T13:00:00.000+00:00
Duration: 6 min 22 s
Description: Learn an ex vivo method for assessing uterine motility using mouse reproductive tract tissue. This technique enables quantitative testing of smooth muscle relaxants for dysmenorrhea research with intact cellular interactions.

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5 minutes

This work was supported by internal IU funds. AGO conceived the study. XC and AGO were involved in the design of the described experiments.&#160; FL and AGO analyzed and interpreted the data. KLL, JOB, FL performed all of the ex vivo experiments. FL wrote the MATLAB script. KLL, JOB, and AGO wrote the manuscript.&#160; All authors read and approved the final version of the manuscript.&#160;&#160;

All procedures with animals have been approved by the Institutional Animal Care and Use Committee at the Indiana University School of Medicine (Indianapolis, IN). 2-5 month-old F2-129S-C57BL/6 sexually-mature female mice were used in the study.
CAUTION: Ensure safety by wearing a lab coat, mask, and gloves when working with animals and biohazardous materials.
1. Solution Preparation
<ol>
	<li>Prepare the Krebs Buffer, which contains:&#160; 130 mM NaCl, 5 mM KCl, 2 mM CaCl2, 1.2 mM NaH2PO4, 0.56 mM MgCl2, 25 mM NaHCO3, and 5 mM glucose, pH 7.4. Continuously oxygenate the Krebs buffer with a mixture of compressed gases containing 5% CO2 and 95% O2 while maintaining the buffer temperature at 37 &#176;C using a circulating water bath.</li>
	<li>Prepare Dulbecco&#39;s Phosphate-Buffered Saline (DPBS), which contains: 2.68 mM KCl, 1.47 mM KH2PO4, 136.89 mM NaCl, and 8.1 mM Na2HPO4, pH 7.4.</li>
</ol>
2. Animal Preparation
<ol>
	<li>Anesthetize mice using inhalation of isoflurane (3%) with waste gas scavenging. Ensure adequate anesthesia by assessing withdrawal reflexes. Pinch the rear toe to affirm no movements are elicited, indicating loss of reflex responses. After the deep anesthesia is achieved, euthanize the animal by decapitation. &#160; 
	NOTE: Isoflurane may cause smooth muscle dilation. Therefore, the reproductive tract preparation should be extensively washed and incubated in the Krebs buffer for at least 15-30 min before beginning the ex vivo experiments. Isoflurane may cause irritation and discomfort when it is in contact with the skin, so proceed with caution.</li>
	<li>Place the body on a large weigh boat lined with a paper towel.&#160; 
	NOTE: Pregnant female laboratory personnel should not be involved in experiments with isoflurane because it can decrease fetal weight, decrease fetal skeletal ossification, and increase the risk of spontaneous abortion8,9. CO2 inhalation can be used as a substitute to euthanize mice.&#160;</li>
</ol>
3. Determination of Estrous Cycle Stage
<ol>
	<li>With small forceps, lift the clitoris to gain access to the vaginal ostium and slowly insert a micropipette tip containing 10 &#181;L of DPBS into the vagina.
	<ol>
		<li>Ensure the micropipette tip is inserted through the ostium at an angle of 10 - 30&#176; to avoid puncturing the vaginal wall. The liquid should still be visible in the tip after insertion. If the liquid is not visible, the tip was inserted too far into the vagina, and the paracervical area of the vagina might have been perforated.</li>
		<li>Lightly pull down on the vaginal ostium muscles with the tip of the micropipette to allow air to exit from the vagina.</li>
	</ol>
	</li>
	<li>Slowly flush the vaginal cavity by pipetting up and down 2-3 times with 10 &#181;L of DPBS and place the drawn cell suspension on a glass slide.</li>
	<li>Use an Inverted Phase Contrast Microscope to determine the estrous cycle stage through cytologic analysis. The procedure is done as described elsewhere10,11. Ensure the cell suspension does not dry out before cytologic analysis can be performed. The suspension may be diluted with fresh DPBS, if necessary.</li>
</ol>
4. Mouse Reproductive Tract Ddissection
<ol>
	<li>Arrange the mouse in a supine position and spread its extremities to expose the abdominopelvic region.</li>
	<li>Spray with 70% ethanol to moisten and disinfect the abdominopelvic area.&#160;</li>
	<li>Using forceps, carefully lift the skin located superior to the clitoris. Make small transverse incisions on the lateral aspects of the lower abdominal area, up to the upper extremities to expose the peritoneum (Figure 1A). During this process, an external flap will form. As small incisions are continually made, the flap will increase in size.</li>
	<li>Carefully cut through the peritoneum to expose the gastrointestinal tract (Figure 1B). It is important to note that the uterine horns can oftentimes be located directly under the peritoneum, so make incisions cautiously and do not touch&#160;the horns as this can affect uterine motility.</li>
	<li>Using forceps, remove the fascia and adipose tissue covering the gastrointestinal tract. Remove the following gastrointestinal tract segments from the abdominal cavity: the duodenum, jejunum, ileum, cecum, ascending and transverse colon (Figure 1C).</li>
	<li>To locate the reproductive organs, first identify the urinary bladder (Figure 1C, &#34;4&#34;), which may have a deflated appearance due to voiding after euthanasia. The vagina will be right under the urinary bladder.</li>
	<li>Locate the pubic symphysis at the confluence of the pubic bones (caudally to the bladder).</li>
	<li>Using scissors, remove the pubic symphysis by carefully making incisions on its lateral sides through the interpubic fibrocartilaginous tissue to gain access to and provide a route for vagina extraction (Figure 1D).</li>
	<li>Cut through the perineum located between the anus and lower part of the vulva.</li>
	<li>Using forceps, lift the vagina and slowly excise the rectum.</li>
	<li>Identify two uterine horns that bifurcate into a fork, rostrally to the vagina. Locate a convoluted oviduct and ovary at the end of each horn, which may be hidden under the remaining gastrointestinal tract segments. Use small dissecting scissors to remove any ligaments that connect to and support the horns, oviducts, and ovaries within the abdominal cavity.</li>
	<li>Remove the reproductive tract, which includes the vagina, uterus, oviducts, and ovaries, from the abdominal cavity.</li>
	<li>Transfer the isolated reproductive tract (Figure 1E) into a 100 mm Petri dish filled with 10 mL of DPBS. Do not compress the uterine horns to avoid damaging the myometrium.</li>
	<li>Use forceps and surgical scissors to remove any connective and adipose tissue surrounding the uterine horns and vagina as well as any fur in the pubic region that could hinder the image quality. Remove the broad ligament to allow motility of the uterine horns.</li>
	<li>Wash the isolated reproductive tract with fresh DPBS two times and transfer it into a 35 mm Petri dish filled with 3 mL of oxygenated Krebs solution.</li>
</ol>
5. Tissue Imaging
<ol>
	<li>Place the Petri dish containing the reproductive tract in oxygenated Krebs buffer on a black surface. Maintain the dish at room temperature. 
	NOTE: It is possible to use an infrared warming pad to maintain the tissue at 37 &#176;C.</li>
	<li>Allow 15-30 min for spontaneous contractions to begin. Record spontaneous uterine motility for 10 min from an axial plane using any type of digital video equipment.</li>
	<li>Transfer the preparation into a Petri dish containing the oxygenated Krebs buffer supplemented with a test compound. Record spontaneous uterine motility for about 10 min from an axial plane using any type of digital video equipment.</li>
	<li>Wash the entire reproductive tract in 100 mm Petri dish with 10 mL of DPBS to assess the reversibility of treatment.</li>
	<li>Transfer the preparation into a Petri dish with the freshly oxygenated Krebs buffer. Record spontaneous uterine motility for about 10 min from an axial plane using any type of digital video equipment.</li>
	<li>Transfer the preparation to another 35 mm Petri dish filled with oxygenated Krebs buffer supplemented with the vehicle for the test compound to ensure that there are no mechanically induced changes in spontaneous uterine motility. This is an important control.</li>
	<li>Transfer the video footage to a computer hard drive.</li>
</ol>
6. Data Analysis
<ol>
	<li>Make clips using any video editing software from the original video footage containing the control, treatment, and wash episodes.</li>
	<li>Use the MATLAB software and the provided script (see online Supplemental material) to quantify spontaneous uterine motility. 
	NOTE: Computer Vision Toolbox add-on for MATLAB must be installed for the script to be fully functional.
	<ol>
		<li>Open the MATLAB script, go to the Editor tab, and click Run.</li>
		<li>Select the first video file and click Open.</li>
		<li>Enter a label for the video file in the pop-up dialog box and click OK.</li>
		<li>Enter the time interval (s) needed for calculating the rate of horn movement (&#916; Euclidean distance / &#916;s).</li>
		<li>Use the mouse cursor to select two points on the first frame of the video. A pop-up window will ask to confirm that the selected points should be used for tracking. Click Start to initiate the real time tracking process that will be displayed in the pop-up window. Alternatively, click Reselect Points to reselect the two points.</li>
		<li>Monitor the accuracy of the tracking process on the pop-up window.</li>
		<li>Observe a rate vs. time scatter plot and distance vs. time scatter plot in a new pop-up window. Save the two figures by selecting File | Save As in the same window to document the data.</li>
		<li>Find a folder named PointTrackerData, automatically created by MATLAB, in the same directory where the MATLAB script is located. Identify an Excel file named label_Data containing data points collected from the video in two separate worksheet tabs. 
		NOTE: Any alternative motion tracking software can be used to quantify spontaneous uterus motility.</li>
	</ol>
	</li>
	<li>Use an appropriate software (e.g., Excel or SigmaPlot 13) to perform the statistical analysis.</li>
</ol>

<ol><li>Kuijsters, N. P. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Uterine+peristalsis+and+fertility%3A+current+knowledge+and+future+perspectives%3A+a+review+and+meta-analysis%5BTitle%5D)+AND+%22Reproductive+BioMedicine+Online%22%5BJournal%5D)">Uterine peristalsis and fertility: current knowledge and future perspectives: a review and meta-analysis.</a> Reproductive BioMedicine Online. 35 (1), 50-71 (2017).</li>
<li>Kural, M., Noor, N. N., Pandit, D., Joshi, T., Patil, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Menstrual+characteristics+and+prevalence+of+dysmenorrhea+in+college+going+girls%5BTitle%5D)+AND+%22Journal+of+Family+Medicine+and+Primary+Care%22%5BJournal%5D)">Menstrual characteristics and prevalence of dysmenorrhea in college going girls.</a> Journal of Family Medicine and Primary Care. 4 (3), 426-431 (2015).</li>
<li>Dehnavi, Z. M., Jafarnejad, F., Kamali, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+Effect+of+aerobic+exercise+on+primary+dysmenorrhea%3A+A+clinical+trial+study%5BTitle%5D)+AND+%22Journal+of+education+and+health+promotion%22%5BJournal%5D)">The Effect of aerobic exercise on primary dysmenorrhea: A clinical trial study.</a> Journal of education and health promotion. 7, 3(2018).</li>
<li>Lindner, H. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Significance+of+prostaglandins+in+the+regulation+of+cyclic+events+in+the+ovary+and+uterus%5BTitle%5D)+AND+%22Advances+in+prostaglandin+and+thromboxane+research%22%5BJournal%5D)">Significance of prostaglandins in the regulation of cyclic events in the ovary and uterus.</a> Advances in prostaglandin and thromboxane research. 8, 1371-1390 (1980).</li>
<li>Bernardi, M., Lazzeri, L., Perelli, F., Reis, F. M., Petraglia, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Dysmenorrhea+and+related+disorders%5BTitle%5D)+AND+%22F1000+research%22%5BJournal%5D)">Dysmenorrhea and related disorders.</a> F1000 research. 6, 1645(2017).</li>
<li>Marjoribanks, J., Ayeleke, R. O., Farquhar, C., Proctor, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Nonsteroidal+anti-inflammatory+drugs+for+dysmenorrhoea%5BTitle%5D)+AND+%22The+Cochrane+database+of+systematic+reviews%22%5BJournal%5D)">Nonsteroidal anti-inflammatory drugs for dysmenorrhoea.</a> The Cochrane database of systematic reviews. 7, 001751(2015).</li>
<li>Oladosu, F. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Abdominal+skeletal+muscle+activity+precedes+spontaneous+menstrual+cramping+pain+in+primary+dysmenorrhea%5BTitle%5D)+AND+%22American+journal+of+obstetrics+and+gynecology%22%5BJournal%5D)">Abdominal skeletal muscle activity precedes spontaneous menstrual cramping pain in primary dysmenorrhea.</a> American journal of obstetrics and gynecology. 219 (1), 91(2018).</li>
<li>Lawson, C. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Occupational+exposures+among+nurses+and+risk+of+spontaneous+abortion%5BTitle%5D)+AND+%22American+journal+of+obstetrics+and+gynecology%22%5BJournal%5D)">Occupational exposures among nurses and risk of spontaneous abortion.</a> American journal of obstetrics and gynecology. 206 (4), 327-328 (2012).</li>
<li>Mazze, R. I., Wilson, A. I., Rice, S. A., Baden, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Fetal+development+in+mice+exposed+to+isoflurane%5BTitle%5D)+AND+%22Teratology%22%5BJournal%5D)">Fetal development in mice exposed to isoflurane.</a> Teratology. 32 (3), 339-345 (1985).</li>
<li>Byers, S. L., Wiles, M. V., Dunn, S. L., Taft, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Mouse+estrous+cycle+identification+tool+and+images%5BTitle%5D)+AND+%22PLoS+One%22%5BJournal%5D)">Mouse estrous cycle identification tool and images.</a> PLoS One. 7 (4), 35538(2012).</li>
<li>Caligioni, C. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Assessing+reproductive+status%2Fstages+in+mice%5BTitle%5D)+AND+%22Current+protocols+in+neuroscience%22%5BJournal%5D)">Assessing reproductive status/stages in mice.</a> Current protocols in neuroscience. , Appendix 4, Appendix (2009).</li>
<li>Segal, S., Csavoy, A. N., Datta, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+tocolytic+effect+of+catecholamines+in+the+gravid+rat+uterus%5BTitle%5D)+AND+%22Anesthesia+and+analgesia%22%5BJournal%5D)">The tocolytic effect of catecholamines in the gravid rat uterus.</a> Anesthesia and analgesia. 87 (4), 864-869 (1998).</li>
<li>Chen, X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Phenylephrine%2C+a+common+cold+remedy+active+ingredient%2C+suppresses+uterine+contractions+through+cAMP+signalling%5BTitle%5D)+AND+%22Scientific+reports%22%5BJournal%5D)">Phenylephrine, a common cold remedy active ingredient, suppresses uterine contractions through cAMP signalling.</a> Scientific reports. 8 (1), 11666(2018).</li>
<li>Domino, M., Pawlinski, B., Gajewski, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Biomathematical+pattern+of+EMG+signal+propagation+in+smooth+muscle+of+the+non-pregnant+porcine+uterus%5BTitle%5D)+AND+%22PLoS+One%22%5BJournal%5D)">Biomathematical pattern of EMG signal propagation in smooth muscle of the non-pregnant porcine uterus.</a> PLoS One. 12 (3), 0173452(2017).</li>
<li>Crane, L. H., Martin, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Pace-maker+activity+in+the+myometrium+of+the+oestrous+rat%3A+in+vivo+studies+using+video-laparoscopy%5BTitle%5D)+AND+%22Reproduction%2C+fertility%2C+and+development%22%5BJournal%5D)">Pace-maker activity in the myometrium of the oestrous rat: in vivo studies using video-laparoscopy.</a> Reproduction, fertility, and development. 3 (5), 519-527 (1991).</li>
</ol>

The authors have nothing to disclose.

Here, we described a method for assessing spontaneous contractility of the entire rodent reproductive tract, which includes the ovaries, oviducts, uterine horns, and the vagina. We used a similar method to demonstrate the relaxant effect of phenylephrine on spontaneous uterine motility13, however, in the past we were unable to provide quantitative analysis of the data. In this work, we developed an algorithm for quantitative motility data analysis using the MATLAB motion tracking module. This is a...

As a major female organ, the uterus is crucial for reproduction and essential for the nourishment of the fetus1. The uterus consists of three layers: the perimetrium, myometrium and endometrium. The myometrium is the major contractile layer of the uterus and plays a key role in fetus delivery. The endometrium is the innermost layer lining the uterine cavity and is essential for embryo implantation. In non-pregnant females of reproductive age, the endometrial layer is shed monthly at the beginning of the menstrual cycle. The myometrium aids in this shedding process by maintaining the spontaneous myometrial contractions needed for clearing the necrotic endometrial tissue from the uterus1.
Unfortunately, increased myometrial contractility can result in negative side effects such as dysmenorrhea, or painful menstrual cramps. This is especially seen in young females and nulliparous women2. However, dysmenorrhea is different for every woman and depends on the strength of their myometrial contractions; stronger contractions are often associated with the sensation of severe cramping3. Myometrial contractility can be visualized using uterine ultrasound and is often recognized as endometrial waves. Enhanced release of prostaglandins during menstruation4 in a uterus undergoing endometrial sloughing is believed to contribute to increased myometrial hypercontractility, resulting in ischemia and hypoxia of the uterine muscle and thus increased pain3.
Severe dysmenorrhea can hinder the day-to-day activity of some women and 3 to 33% of women have very severe pain, which could cause a woman to be bedridden for 1 to 3 days each menstrual cycle5. Dysmenorrhea is the leading cause of gynecological morbidity in women of reproductive age regardless of age, nationality, and economic status5. The estimated prevalence of dysmenorrhea is both high and variable, ranging from 45% to 93% in women of reproductive age5.&#160; Dysmenorrhea-associated pain has an effect on the daily life of women and may result in poor academic performance in adolescents, lower quality of sleep, restriction of daily activities, and mood changes5.
Many women who experience severe dysmenorrhea resort to over-the-counter medications to relieve their pain. Such over-the-counter medications contain cyclooxygenase (COX) inhibitors which prevent the formation of prostaglandins6. However, COX inhibitors are associated with adverse cardiovascular events, and about 18% of women with dysmenorrhea are unresponsive to these inhibitors7. Therefore, there is a need for new medications to reduce menstrual cramps. Since over contractility of the uterus contributes to the pathogenesis of dysmenorrhea, one possible strategy may be the usage of uterine relaxants.
It is beneficial to quantify the effects of potential relaxant drugs in a model of naturally occurring spontaneous myometrial wave-like contractions. However, thus far, no efficient ex vivo method for testing muscle-relaxing drugs in the intact uterus has been described. Currently, isometric tension measurements are used to evaluate relaxant drug effects. During such measurements, a uterine muscle strip is maintained at a constant length under preload in a tissue bath while the force of uterine muscle contractions is recorded before and after oxytocin stimulation in the presence or absence of a relaxant drug. Although this approach is very useful, it requires expensive equipment. Furthermore, isometric contractions do not resemble the spontaneous myometrial wave-like contractions that naturally occur in the intact uterus. Uniquely, the uterine myometrial waves in rodents can be visualized as uterine horn motility when the entire reproductive tract (ovaries, oviducts, uterus, and vagina) is maintained in a buffer solution. Here, we present an ex vivo method for monitoring the spontaneous motility of the intact mouse uterus placed in a Petri dish containing oxygenated Krebs buffer. We also describe a motility quantification algorithm utilizing the MATLAB motion tracker. This novel approach provides an easy and less expensive alternative to test the relaxant potential of naturally occurring remedies and synthetic compounds.

<table><tbody><tr><td>Epinephrine hydrochloride</td><td>Sigma-Aldrich</td><td>&amp;nbsp;E4642&amp;nbsp;</td><td></td></tr><tr><td>Dulbecco&#039;s PBS</td><td>Fisher Sceintific&amp;nbsp;</td><td>17-512Q</td><td></td></tr><tr><td>Ethanol 200 PROOF</td><td>Decon Laboratories</td><td>2701</td><td></td></tr><tr><td>NaCl</td><td>Sigma-Aldrich</td><td>S7653</td><td></td></tr><tr><td>Glucose</td><td>Sigma-Aldrich</td><td>G7528</td><td></td></tr><tr><td>KCl</td><td>Sigma-Aldrich</td><td>P9333</td><td></td></tr><tr><td>CaCl&lt;sub&gt;2&lt;/sub&gt; &amp;middot; 2H&lt;sub&gt;2&lt;/sub&gt;O</td><td>Sigma-Aldrich</td><td>C5080</td><td></td></tr><tr><td>NaH&lt;sub&gt;2&lt;/sub&gt;PO&lt;sub&gt;4&lt;/sub&gt;</td><td>Sigma-Aldrich</td><td>S0751</td><td></td></tr><tr><td>MgCl&lt;sub&gt;2&lt;/sub&gt;&amp;nbsp; &amp;middot; 6H&lt;sub&gt;2&lt;/sub&gt;O</td><td>Sigma-Aldrich</td><td>M9272</td><td></td></tr><tr><td>NaHCO&lt;sub&gt;3&lt;/sub&gt;</td><td>Sigma-Aldrich</td><td>S6297</td><td></td></tr><tr><td>Isoflurane, USP</td><td>Patterson Veterinary</td><td>07-893-2374</td><td></td></tr><tr><td>Dissecting Extra-Fine-Pointed Precision Splinter Forceps</td><td>Fisher Sceintific</td><td>13-812-42</td><td></td></tr><tr><td>Curved Hardened Fine Iris Scissors</td><td>Fine Science Tools</td><td>14091-09</td><td></td></tr><tr><td>Dissection High-performance Modular Stereomicroscope</td><td>Leica</td><td>MZ6&amp;nbsp;</td><td></td></tr><tr><td>Digital 5 Megapixel Color Microscope Camera with active cooling system</td><td>Leica</td><td>&amp;nbsp;DFC425 C</td><td></td></tr><tr><td>Stereomaster Microscope Fiber-Optic Light Sources</td><td>Fisher Sceintific&amp;nbsp;</td><td>12-562-21</td><td></td></tr><tr><td>Weigh Boat</td><td>Fisher Sceintific&amp;nbsp;</td><td>WB30304</td><td></td></tr><tr><td>Convertors Astound Standard Surgical Gown&amp;nbsp;</td><td>Cardinal Health&amp;nbsp;</td><td>9515</td><td>Small, Medium or Large</td></tr><tr><td>Gloves</td><td>McKesson Corporation</td><td>20-1080</td><td>Small, Medium, or Large; powder-free sterile latex or nitrile surgical gloves</td></tr><tr><td>&amp;nbsp;Petri Dish</td><td>Corning Falcon</td><td>351029</td><td>100 mm</td></tr><tr><td>&amp;nbsp;Petri Dish</td><td>Corning Falcon</td><td>353001</td><td>35 mm</td></tr><tr><td>95% O&lt;sub&gt;2&lt;/sub&gt;- 5% CO&lt;sub&gt;2&lt;/sub&gt; gas mixture</td><td>Praxair</td><td>&amp;nbsp;MM OXCD5-K</td><td></td></tr><tr><td>Ear-loop Masks</td><td>Valumax International</td><td>5430E-PP</td><td></td></tr><tr><td>DSLR 24.2 MP Camera</td><td>Canon</td><td>EOS Rebel T6i</td><td></td></tr><tr><td>MATLAB</td><td>MathWorks</td><td>N/A</td><td>version 2019 or later</td></tr></tbody></table>

ex vivo method for assessing the mouse reproductive tract spontaneous motility and a matlab-based uterus motion tracking algorithm for data analysis

Ex Vivo Method for Assessing the Mouse Reproductive Tract Spontaneous Motility and a MATLAB-based Uterus Motion Tracking Algorithm for Data Analysis

Dysmenorrhea, or painful cramping, is the most common symptom associated with menses in females and its severity can hinder women&#39;s everyday lives. Here, we present an easy and inexpensive method that would be instrumental for testing new drugs decreasing uterine contractility. This method utilizes the unique ability of the entire mouse reproductive tract to exhibit spontaneous motility when maintained ex vivo in a Petri dish containing oxygenated Krebs buffer. This spontaneous motility resembles the wave-like myometrial activity of the human uterus, referred to as endometrial waves. To demonstrate the effectiveness of the method, we employed a well-known uterine relaxant drug, epinephrine. We demonstrate that the spontaneous motility of the entire mouse reproductive tract can be quickly and reversibly inhibited by 1 &#181;M epinephrine in this Petri dish model. Documenting the changes of uterine motility can be easily done using an ordinary smart phone or a sophisticated digital camera. We developed a MATLAB-based algorithm allowing motion tracking to quantify spontaneous uterine motility changes by measuring the rate of uterine horn movements. A major advantage of this ex vivo approach is that the reproductive tract remains intact throughout the entire experiment, preserving all intrinsic intrauterine cellular interactions. The major limitation of this approach is that up to 10-20% of uteri may exhibit no spontaneous motility. Thus far, this is the first quantitative ex vivo method for assessing spontaneous uterine motility in a Petri dish model.

Figure 1 shows representative images taken during the entire reproductive tract isolation procedure that is described in this protocol. To avoid contaminating the buffer with fur, which would decrease video quality, we moistened the mouse body with 70% ethanol. The major benchmark for the dissection section of the protocol is to find the urinary bladder. The uterus and vagina will be located inferior to the urinary bladder.
To test the protocol, we treated the ent...

Uterine contractions are important for the well-being of females. However, pathologically increased contractility may result in dysmenorrhea, especially in younger females. Here, we describe a simple ex vivo preparation allowing quick assessment of the efficacy of smooth muscle relaxants that may be used for treating dysmenorrhea.

Dysmenorrhea, or painful cramping, is the most common symptom associated with menses in females and its severity can hinder women&#39;s everyday lives. ...

ex-vivo-method-for-assessing-mouse-reproductive-tract-spontaneous

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Measuring Sperm Guidance and Motility within the Caenorhabditis elegans Hermaphrodite Reproductive Tract

This protocol describes a method for monitoring the progression of morphological changes over time in the uterus in an inducible mouse model of endometrial cancer using ultrasound imaging with correlation to gross and histological changes.

Non-Invasive Ultrasound Assessment of Endometrial Cancer Progression in Pax8-Directed Deletion of the Tumor Suppressors Arid1a and Pten in Mice

Pregnancy establishment is a dynamic process involving complex embryo and uterine crosstalk. The precise contributions of the maternal uterine environment to these processes remain an active area of investigation. Here, detailed protocols are provided to aid in designing in vivo animal models to address these research questions.

Methods for Studying Uterine Contributions to Pregnancy Establishment in an Ovariectomized Mouse Model

This study introduces a novel mouse model specifically designed to simulate endometrioma, a clinically significant subtype of endometriosis. By utilizing C57BL/6J mice, the model aims to elucidate the pathophysiological mechanisms underlying endometrioma-related infertility, offering a refined tool to bridge the current knowledge gap in reproductive medicine.

Establishment of an Experimental Mouse Model of Endometrioma to Study its Related Infertility

Learn an ex vivo method for assessing uterine motility using mouse reproductive tract tissue. This technique enables quantitative testing of smooth muscle relaxants for dysmenorrhea research with intact cellular interactions.

Ex Vivo Method for Assessing the Mouse Reproductive Tract Spontaneous Motility and a MATLAB-based Uterus Motion Tracking Algorithm for Data Analysis (Video) | JoVE Journal

Ex Vivo Method for Assessing the Mouse Reproductive Tract Spontaneous Motility and a MATLAB-based Uterus Motion Tracking Algorithm for Data Analysis

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