Name: ultrasonography in experimental reproductive investigations on rats
Uploaded: 2017-12-02T13:00:00
Description: Watch this Scientific Journal Video about Ultrasonography in Experimental Reproductive Investigations on Rats at JoVE.com

This research was supported by Animal Experimentation Section of Murcia University and the Obstetrics and Gynecology Department of Murcia University. We thank all the technicians working at the CEIB (Centro Experimental en Investigaciones Biom&#233;dicas), the section of animal experimentation of the University of Murcia, who have collaborated on this project.

This study was carried out in strict accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health and in accordance with the ARRIVE (Animal Research: Reporting In Vivo Experiments) guidelines. The protocol received a license for animal experimentation complying with Directive 2010/63/EU with the authorization number A13170404 (Anexo 1). All experiments were performed in a European Union-certified laboratory following national guidelines for the ethical c...

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	<li>Hunter, R. 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=Adipose-Derived+Stromal+Vascular+Fraction+Cell+Effects+on+a+Rodent+Model+of+Thin+Endometrium.">Adipose-Derived Stromal Vascular Fraction Cell Effects on a Rodent Model of Thin Endometrium.</a> PLoS ONE. 10 (12), e0144823 (2015).</li><li>Wang, H., Dey, S. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Roadmap+to+embryo+implantation:+clues+from+mouse+models.">Roadmap to embryo implantation: clues from mouse models.</a> Nat Rev Genet. 7 (3), 185-199 (2006).</li><li>Pistner, A., Belmonte, S., Coulthard, T., Blaxall, B. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Murine+Echocardiography+and+Ultrasound+Imaging.">Murine Echocardiography and Ultrasound Imaging.</a> J Vis Exp. (42), (2010).</li><li>Lohmiller, J. J., Swing, S. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reproduction+and+Breeding.">Reproduction and Breeding.</a> The Laboratory Rat. , 147-164 (2006).</li><li>Jaiswal, R. S., Singh, J., Adams, G. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-resolution+ultrasound+biomicroscopy+for+monitoring+ovarian+structures+in+mice.">High-resolution ultrasound biomicroscopy for monitoring ovarian structures in mice.</a> Reprod Biol Endocrinol. 7 (69), (2009).</li><li>Kim, G. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Murine+Fetal+Echocardiography.">Murine Fetal Echocardiography.</a> J Vis Exp. (72), (2013).</li><li>Jing, Z., Qiong, Z., Yonggang, W., Yanping, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rat+bone+marrow+mesenchymal+stem+cells+improve+regeneration+of+thin+endometrium+in+rat.">Rat bone marrow mesenchymal stem cells improve regeneration of thin endometrium in rat.</a> Fertil Steril. 101 (2), 587-594 (2014).</li><li>Mu, J., Adamson, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Developmental+changes+in+hemodynamics+of+uterine+artery,+utero-+and+umbilicoplacental,+and+vitelline+circulations+in+mouse+throughout+gestation.">Developmental changes in hemodynamics of uterine artery, utero- and umbilicoplacental, and vitelline circulations in mouse throughout gestation.</a> Am J Physiol Heart Circ Physiol. 291 (3), H1421-H1428 (2006).</li><li>Anderson, C. M., Lopez, F., Zhang, H. Y., Pavlish, K., Benoit, J. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reduced+uteroplacental+perfusion+alters+uterine+arcuate+artery+function+in+the+pregnant+Sprague-Dawley+rat.">Reduced uteroplacental perfusion alters uterine arcuate artery function in the pregnant Sprague-Dawley rat.</a> Biol Reprod. 72 (3), 762-766 (2005).</li><li>Hongmei, 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=Ultrasound+Molecular+Imaging+of+Vascular+Endothelial+Growth+Factor+Receptor+2+Expression+for+Endometrial+Receptivity+Evaluation.">Ultrasound Molecular Imaging of Vascular Endothelial Growth Factor Receptor 2 Expression for Endometrial Receptivity Evaluation.</a> Theranostics. 5 (2), 206-217 (2015).</li></ol>

Due to the procedural modifications and troubleshooting that was required in this study, despite our aim of identifying all the phases of the estrus cycle in rats using ultrasound, we were unable to find any significant differences. We hypothesize that these difficulties may be because the estrus cycle only lasts a few days in rats, unlike the cycle in women. We are sure that all measurements were done at exactly the right time to determine any differences. Therefore, we regrouped the estrus cycle phases into only two ph...

Rat animal models have been widely used in reproductive medicine, including in embryo and ovarian transplantation1,2. However, in the past, the study of reproductive system development in rodents has been based on one-time histological examination of excised tissues, and the longitudinal study of day-to-day reproductive organ changes has not been possible in rats3. Ultrasound has been widely used in assisted reproductive technology in humans for over 30 years, but this valuable technology has only been applied to rats recently.
Our objective was to establish an ultrasonographic approach for evaluating the reproductive organs of Sprague-Dawley rats to design experimental models for reproduction and gynecology investigation and to demonstrate the procedure because to our knowledge, there are no current visualized publications regarding this procedure. We describe the procedure of ultrasonographic examination of the female rat&#39;s reproductive system and present ultrasonographic findings on anatomy and uterine artery blood flow using high-definition ultrasound. We monitored the characteristics of the endometrium, ovaries and uterine artery blood flow in non-pregnant animals at different stages of estrus cycle to evaluate the significant differences in endometrial thickness, ovarian morphology and uterine blood flow in different stages of the estrus cycle, similar to women. We used high-quality ultrasound equipment with a 70 MHz frequency and a resolution level of 30 &#181;m. Our other objective was to evaluate changes in the resistance of uterine blood flow in pregnant rats. This technique allows for the study of daily changes in the reproductive organs without sacrificing animals.
There are several technical difficulties in utilizing ultrasound on rats. These difficulties include: the rat endometrium is much thinner than a human female4. Difficulty in imaging the ovaries of rats has been attributed to thicker skin and abdominal wall musculature in rats, which resulted in near-complete attenuation of the ultrasound5, and the uterine artery is much more difficult to find in non-pregnant rats. We have solved many technical problems with the procedure, and for those problems that remain, we show how to minimize them.
Successful monitoring of sonographic changes in the reproductive organs of rats without the need to sacrifice animals will open the possibility of building future animal models of reproductive medicine and other surgical procedures.

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			<td>Vevo3100 high-resolution in vivo micro-imaging system*</td>
			<td>Visual Sonics, inc.</td>
			<td></td>
			<td>www.visualsonics.com/products</td>
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			<td>Vevo integrated rail system including physiological monitoring unit.</td>
			<td>Visual Sonics, inc.</td>
			<td></td>
			<td>www.visualsonics.com/products</td>
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			<td>MX400 Transducter</td>
			<td>Visual Sonics, inc.</td>
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			<td>www.visualsonics.com/products</td>
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			<td>Vevo Lab Software</td>
			<td>Visual Sonics, inc.</td>
			<td></td>
			<td>www.visualsonics.com/products</td>
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		<tr>
			<td>HSD: Sprague Dawley SD</td>
			<td>Envigo, inc.</td>
			<td></td>
			<td>Rat strain</td>
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			<td>Lubricating Gel</td>
			<td>General Supply</td>
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			<td>CIBERTEC CA-EAC20 Anesthesia Trolley System</td>
			<td>Cibertec S.A&#160;</td>
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			<td>Anesthesia Machine</td>
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			<td>Ecogel 100 ultrasound gel</td>
			<td>Eco-Med Pharmaceuticals Inc.</td>
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			<td>Hair removal lotion (Nair)&#160;</td>
			<td>General Supply</td>
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			<td>Isoflurane</td>
			<td>Esteve Veterinaria</td>
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			<td>Inhalatory anesthesia</td>
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			<td>* Required software is Vevo software including B-Mode application, pulse wave Doppler application, and vascular strain analysis tools package.</td>
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ultrasonography in experimental reproductive investigations on rats

With the development of assisted reproductive technology and the ethical limitations of research on humans, rat animal models have been widely used in reproductive medicine. In the past, the study of reproductive system development in rodents has been based on one-time histological examination of excised tissues. Recently, with the development of high-resolution transabdominal ultrasound, high-quality sonography can now be performed to evaluate the reproductive organs of rats, allowing a new method for studying the reproductive system. Images were obtained using a high-resolution ultrasonographic system. Gynecological ultrasonography was performed on 28 eight-week-old non-pregnant rats and 5 pregnant Sprague-Dawley rats. We describe how to recognize organs of the reproductive system and associated structures in typical views during different phases of the estrus cycle. Color flow Doppler was used to measure uterine artery blood flow and evaluate uterine blood flow pattern changes during different stages of pregnancy. We have demonstrated that ultrasound exploration is a useful method for evaluating changes in internal reproductive organs. Its use raises the possibility of conducting additional experiments, including medical or surgical procedures, and provides the ability to monitor sonographic changes to internal organs without sacrificing animals.

There were no significant differences in antero-posterior uterine horn diameters or in the thickness of the endometrium between the two sides of the uterine horn (Table 1). Compared with the group 2, the mean endometrial thickness in the group 1 was thicker, but no significant differences (P&#62;0.05) were found between the two groups (Figure 3). Nevertheless, we found fluid inside the uterus (in 8 out of 28 rats) near the estrus cycle associated with changes in endometrium ...

Watch this Scientific Journal Video about Ultrasonography in Experimental Reproductive Investigations on Rats at JoVE.com

Ultrasonography in Experimental Reproductive Investigations on Rats

This manuscript describes the utility of ultrasound performed on female rats to design experimental models for reproductive and gynecological investigation. A step-by-step explanation of how to perform ultrasonographic evaluation is shown.

With the development of assisted reproductive technology and the ethical limitations of research on humans, rat animal models have been widely used in ...

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