This work is supported by a Hamilton-Fairley NHMRC Fellowship (RH, AL); the Victorian Government's Operational Infrastructure Support Program (RH, EW) and the Marie Curie Industria-Academia Partnership and Pathways grant sponsored by the European Commission (ME, PD). The authors would like to thank Dr. Andre Miyague, Dr. Francesca Russo, Ms. Rosita Kinnart and Mr. Ivan Laermans for their technical expertise in producing this video.

The following experimental protocol is approved by the Animal Ethics Committee, Katholieke Universiteit Leuven, Leuven, Belgium. We followed previously described surgical procedure13 including some methodological changes, especially in anesthesia procedure.
1. Inducing Intrauterine Growth Restriction (IUGR)
<ol>
	<li>Time dated pregnant rabbits (hybrid Dendermonde and New Zealand White) are housed in individual cages on a 12/12 hr light schedule with access to water and standard r...

We have used a previously validated approach of surgically reducing uteroplacental blood flow in a rabbit to produce IUGR13-16 and later examining fetal cardiac function14 to describe microultrasound technology and cardiac function analysis available on the VisualSonics VEVO 2100. The ability to reproduce fetoplacental Doppler changes of human IUGR fetuses in a small animal model and to then allow examination of cardiac function with recently described echocardiography is likely to progress fetal ca...

The burden of cardiovascular disease that results from fetal intrauterine growth restriction (IUGR) cannot be overstated. It is the leading cause of stillbirth after congenital abnormalities.1 IUGR refers to a fetus that fails to reach its growth potential and is commonly a result of placental insufficiency.2 In survivors, cardiovascular ill health is evident across the life span with myocardial dysfunction apparent in infancy and childhood3,4 and subsequent hypertension5, diabetes6, and obesity developing in adult life - all cumulative cardiac risk factors from birth towards premature death from ischemic heart disease.7
Developing animal models to characterize the maternal-fetal communication that establishes IUGR and the subsequent fetal response to reduced substrate availability is necessary if we are to both better understand the pathophysiology of IUGR-related cardiac impairment and to develop cardio-protective strategies to improve fetal and postnatal health. In this regard, many different models across different species have been described.8 IUGR is commonly induced by maternal under nutrition with a low protein diet in rodents,9 surgical ablation or ligation of uterine blood flow in rodents and guinea pigs10 or occlusion of the umbilical artery in sheep.11 However, it is apparent that no model fully recapitulates the human IUGR.12
In this current methodological study, we used a well validated approach of selective uteroplacental vascular interruption in a rabbit13-16 that not only produces fetal cardiovascular responses observed with ultrasound clinically14, but also allows interrogation of fetal cardiac function with novel echocardiography using microultrasound technology of the VisualSonics VEVO 2100. While Doppler ultrasound of fetoplacental vessels remains the cornerstone of current antenatal surveillance of IUGR fetuses17, functional echocardiography is increasingly being utilized to provide new insights into disease pathophysiology and to assess fetal wellbeing.18 Accordingly, here we take these advances from clinical research and describe an animal model that harbors not only this imaging sophistication but also provides the experimental platform to investigate mechanistic pathways and novel therapeutics.

<table border="1">
	<tbody>
 <tr>
 <td>Name of Reagent/Material</td>
 <td>Company</td>
 <td>Catalog Number</td>
 <td>Comments</td>
 </tr>
 <tr>
 <td>Ketamine</td>
 <td>Ceva Sante Animale</td>
 <td></td>
 <td><a href="http://www.ceva.com/en" target="_blank">http://www.ceva.com/en</a></td>
 </tr>
 <tr>
 <td>Xylazine</td>
 <td>Ceva Sante Animale</td>
 <td></td>
 <td><a href="http://www.ceva.com/en" target="_blank">http://www.ceva.com/en</a></td>
 </tr>
 <tr>
 <td>Depot Provera</td>
 <td>Pharmacia Upjohn</td>
 <td></td>
 <td></td>
 </tr>
 <tr>
 <td>Penicillin G</td>
 <td>Kela Pharma</td>
 <td></td>
 <td><a href="http://www.kela.be" target="_blank">http://www.kela.be</a></td>
 </tr>
 <tr>
 <td >Lidocaine</td>
 <td>B Braun Medical</td>
 <td></td>
 <td><a href="http://www.bbraun.com/" target="_blank">http://www.bbraun.com/</a></td>
 </tr>
 <tr>
 <td>Temgesic</td>
 <td>Schering-Plough</td>
 <td></td>
 <td><a href="http://www.merck-animal-health-usa.com/" target="_blank">http://www.merck-animal-health-usa.com/</a></td>
 </tr>
 <tr>
 <td>Isolurane</td>
 <td>Isoba Vet; Abbott Laboratories Ltd</td>
 <td></td>
 <td><a href="http://www.abbottanimalhealth.com/index.html" target="_blank">http://www.abbottanimalhealth.com/index.html</a></td>
 </tr>
 <tr>
 <td>Ethicon</td>
 <td>Johnson and Johnson</td>
 <td></td>
 <td><a href="http://www.ethiconproducts.co.uk/products/sutures" target="_blank">http://www.ethiconproducts.co.uk/products/sutures</a></td>
 </tr>
 <tr>
 <td>Ethicon</td>
 <td>Johnson and Johnson</td>
 <td></td>
 <td><a href="http://www.ethiconproducts.co.uk/products/sutures" target="_blank">http://www.ethiconproducts.co.uk/products/sutures</a></td>
 </tr>
 <tr>
 <td>Ethicon</td>
 <td>Johnson and Johnson</td>
 <td></td>
 <td><a href="http://www.ethiconproducts.co.uk/products/sutures" target="_blank">http://www.ethiconproducts.co.uk/products/sutures</a></td>
 </tr>
 <tr>
 <td>VEVO 2100</td>
 <td>VisualSonics</td>
 <td>SN100-0032</td>
 <td><a href="http://www.visualsonics.com/" target="_blank">http://www.visualsonics.com/</a></td>
 </tr>
 <tr>
 <td>Aquasonic Gel</td>
 <td>Parker Laboratories</td>
 <td>01 02</td>
 <td><a href="http://www.parkerlabs.com/ultrasound_products.html" target="_blank">http://www.parkerlabs.com/ultrasound_products.html</a></td>
 </tr>
 <tr>
 <td>Nellcor N-20PA Pulse oximeter</td>
 <td>Covidien</td>
 <td></td>
 <td><a href="http://www.nellcor.com/prod/PRODUCT.ASPX?S1=POX&amp;S2=MON&amp;id=282&amp;V" target="_blank">http://www.nellcor.com/prod/PRODUCT.ASPX?S1=POX&amp;S2=MON&amp;id=282&amp;V</a></td>
 </tr>
	 </tbody>
 </table>

fetal echocardiography and pulsed-wave doppler ultrasound in a rabbit model of intrauterine growth restriction

Fetal intrauterine growth restriction (IUGR) results in abnormal cardiac function that is apparent antenatally due to advances in fetoplacental Doppler ultrasound and fetal echocardiography. Increasingly, these imaging modalities are being employed clinically to examine cardiac function and assess wellbeing in utero, thereby guiding timing of birth decisions. Here, we used a rabbit model of IUGR that allows analysis of cardiac function in a clinically relevant way. Using isoflurane induced anesthesia, IUGR is surgically created at gestational age day 25 by performing a laparotomy, exposing the bicornuate uterus and then ligating 40-50% of uteroplacental vessels supplying each gestational sac in a single uterine horn. The other horn in the rabbit bicornuate uterus serves as internal control fetuses. Then, after recovery at gestational age day 30 (full term), the same rabbit undergoes examination of fetal cardiac function. Anesthesia is induced with ketamine and xylazine intramuscularly, then maintained by a continuous intravenous infusion of ketamine and xylazine to minimize iatrogenic effects on fetal cardiac function. A repeat laparotomy is performed to expose each gestational sac and a microultrasound examination (VisualSonics VEVO 2100) of fetal cardiac function is performed. Placental insufficiency is evident by a raised pulsatility index or an absent or reversed end diastolic flow of the umbilical artery Doppler waveform. The ductus venosus and middle cerebral artery Doppler is then examined. Fetal echocardiography is performed by recording B mode, M mode and flow velocity waveforms in lateral and apical views. Offline calculations determine standard M-mode cardiac variables, tricuspid and mitral annular plane systolic excursion, speckle tracking and strain analysis, modified myocardial performance index and vascular flow velocity waveforms of interest. This small animal model of IUGR therefore affords examination of in utero cardiac function that is consistent with current clinical practice and is therefore useful in a translational research setting.

An asymmetrical growth restricted fetus and placenta from uteroplacental vascular ligation is compared to a normal control fetus and placenta in Figure 1F. Asymmetrical growth is confirmed by reduced neonatal birth weight and increased head circumference:abdominal circumference ratio to controls. Representative results from fetoplacental Doppler studies are shown in Figure 2. A normal low resistance pattern of positive end-diastolic flow in a control fetus is shown. With progressive incr...

Watch this Scientific Journal Video about Fetal Echocardiography and Pulsed-wave Doppler Ultrasound in a Rabbit Model of Intrauterine Growth Restriction at JoVE.com

Fetal Echocardiography and Pulsed-wave Doppler Ultrasound in a Rabbit Model of Intrauterine Growth Restriction

We describe examination of fetal cardiac function with contemporary functional fetal echocardiography and fetoplacental Doppler ultrasound using the VisualSonics VEVO 2100 microultrasound in a surgically induced model of intrauterine fetal growth restriction in a rabbit.

Fetal intrauterine growth restriction (IUGR) results in abnormal cardiac function that is apparent antenatally due to advances in fetoplacental Doppler ...

fetal-echocardiography-pulsed-wave-doppler-ultrasound-rabbit-model

Research

JoVE Journal

Medicine

28.1K ビュー.  University Hospitals Leuven. 私たちは、現代の機能胎児心エコー検査とウサギ子宮内胎児発育制限の外科的に誘発されるモデルでVisualSonics VEVO 2100 microultrasoundを使って胎児胎盤ドップラー超音波で胎児の心臓機能の検査について説明します。

ビデオ: Fetal Echocardiography and Pulsed-wave Doppler Ultrasound in a Rabbit Model of Intrauterine Growth Restriction

We describe examination of fetal cardiac function with contemporary functional fetal echocardiography and fetoplacental Doppler ultrasound using the VisualSonics VEVO 2100 microultrasound in a surgically induced model of intrauterine fetal growth restriction in a...

murine fetal echocardiography

Murine Fetal Echocardiography

Fetal and perinatal death is a common feature when studying genetic alterations affecting cardiac development. High-frequency ultrasound imaging has improved 2-D resolution and can provide excellent information on early cardiac development and is an ideal method to detect the impact on cardiac structure and function prior to...

murine echocardiography and ultrasound imaging

Murine Echocardiography and Ultrasound Imaging

This video demonstrates use of a rail-mounted high-frequency ultrasound probe to perform echocardiography on an anesthetized mouse.  The methods describe both conventional two-dimensional and M-mode measurements of cardiac function in addition to newer, more powerful tools such as color Doppler, strain analysis, as well as general and targeted contrast...

application of ultrasound and shear wave elastography imaging in a rat model of nafld/nash

Application of Ultrasound and Shear Wave Elastography Imaging in a Rat Model of NAFLD/NASH

This protocol describes the use of an enhanced ultrasound technique to non-invasively observe and quantify liver tissue changes in rodent models of nonalcoholic fatty liver disease.

high frequency ultrasound for the analysis of fetal and placental development in vivo

High Frequency Ultrasound for the Analysis of Fetal and Placental Development In Vivo

Here we describe the technique of high frequency ultrasound for in vivo analysis of fetuses in mice. This method allows the follow-up of fetuses and the analysis of placental parameters as well as maternal and fetal blood flow throughout...

High Frequency Ultrasound for the Analysis of Fetal and Placental Development In Vivo

fetal mouse cardiovascular imaging using a high-frequency ultrasound (30/45mhz) system

Fetal Mouse Cardiovascular Imaging Using a High-frequency Ultrasound 30/45MHZ System

High-frequency ultrasound imaging of the fetal mouse has improved imaging resolution and can provide precise non-invasive characterization of cardiac development and structural defects. The protocol outlined herein is designed to perform real-time fetal mice echocardiography in...

Fetal Mouse Cardiovascular Imaging Using a High-frequency Ultrasound (30/45MHZ) System

high-frequency ultrasound echocardiography to assess zebrafish cardiac function

High-Frequency Ultrasound Echocardiography to Assess Zebrafish Cardiac Function

We describe a protocol to assess heart morphology and function in adult zebrafish using high-frequency echocardiography. The method allows visualization of the heart and subsequent quantification of functional parameters, such as heart rate (HR), cardiac output (CO), fractional area change (FAC), ejection fraction (EF), and blood inflow and outflow...

continuous venous-arterial doppler ultrasound during a preload challenge

Continuous Venous-Arterial Doppler Ultrasound During a Preload Challenge

The Frank-Starling-Sarnoff curve is clinically important and describes the relationship between cardiac preload and output. This report illustrates a novel method of simultaneous jugular venous and carotid arterial Doppler velocimetry as transient surrogates of cardiac preload and output, respectively; this approach is enabled by wireless, wearable Doppler...

transthoracic echocardiographic examination in the rabbit model

Transthoracic Echocardiographic Examination in the Rabbit Model

Here we describe, step by step, a detailed protocol for performing echocardiography in the rabbit model. We show how to correctly obtain the different echocardiographic views and imaging planes, as well as the different imaging modes available in a clinical echocardiography system routinely used in human and veterinary...

a rat model of mild intrauterine hypoperfusion with microcoil stenosis

A Rat Model of Mild Intrauterine Hypoperfusion with Microcoil Stenosis

Mild intrauterine hypoperfusion was produced by artery stenosis with metal microcoils wrapped around the uterine and ovarian arteries in rats at embryonic day 17. This procedure produced prenatal hypoperfusion and intrauterine growth...

calvarial model of bone augmentation in rabbit for assessment of bone growth and neovascularization in bone substitution materials

Calvarial Model of Bone Augmentation in Rabbit for Assessment of Bone Growth and Neovascularization in Bone Substitution Materials

Here we present a surgical protocol in rabbits with the aim to assess bone substitution materials in terms of bone regeneration capacities. By using PEEK cylinders fixed onto rabbit skulls, osteoconduction, osteoinduction, osteogenesis and vasculogenesis induced by the materials may be evaluated either on live or euthanized...

percutaneous contrast echocardiography-guided intramyocardial injection and cell delivery in a large preclinical model

Percutaneous Contrast Echocardiography-guided Intramyocardial Injection and Cell Delivery in a Large Preclinical Model

Novel therapeutic strategies in cardiac regenerative medicine require extensive and detailed studies in large preclinical animal models before they can be considered for use in humans. Here, we demonstrate a percutaneous contrast echocardiography-guided intramyocardial injection technique in rabbits, which is valuable for hypothesis testing the efficacy of such novel...

application of acupotomy in a knee osteoarthritis model in rabbit

Author Spotlight: Investigating the Mechanism of Action of Acupotomy in Treating Knee Osteoarthritis

In this protocol, a knee osteoarthritis model was prepared using the modified Videman method, and the operation procedures and precautions of acupotomy are detailed. The effectiveness of acupotomy has been demonstrated by testing the mechanical properties of quadriceps femoris and tendon and the mechanical and morphological properties of...

Application of Acupotomy in a Knee Osteoarthritis Model in Rabbit

ultrasound-based pulse wave velocity evaluation in mice

Ultrasound-based Pulse Wave Velocity Evaluation in Mice

Arterial stiffness represents a key factor in cardiovascular disease and pulse wave velocity (PWV) can be considered as a surrogate index for arterial stiffness. This protocol describes an image processing algorithm for calculating PWV in mice based on ultrasound image processing that is applicable at different arterial...

assessment of cerebral lateralization in children using functional transcranial doppler ultrasound (ftcd)

Assessment of Cerebral Lateralization in Children using Functional Transcranial Doppler Ultrasound fTCD

Functional transcranial Doppler sonography (fTCD) is a simple and non-invasive ultrasound technique which can be used to assess the lateralization of cognitive functions, especially language, and is suitable for use with children.

Assessment of Cerebral Lateralization in Children using Functional Transcranial Doppler Ultrasound (fTCD)

doppler ultrasound scan and blood flow quantitation in leg-extension exercises

Doppler Ultrasound Scan and Blood Flow Quantitation in Leg-Extension Exercises  

doppler ultrasound-based leg blood flow assessment during single-leg knee-extensor exercise in an uncontrolled setting

Author Spotlight: Assessing the Reliability of Doppler Ultrasound in Measuring Leg Blood Flow 

This test-retest study evaluated leg blood flow measured by the Doppler ultrasound technique during single-leg knee-extensor exercise. The within-day, between-day, and inter-rater reliability of the method was investigated. The approach demonstrated high within-day and acceptable between-day reliability. However, the inter-rater reliability was unacceptably low during rest and at low...

Assessing Leg Blood Flow in Training Workloads

https://cloudfront.jove.com/CDNSource/teasers/65746_b.jpg

assessing teratogenic changes in a zebrafish model of fetal alcohol exposure

Assessing Teratogenic Changes in a Zebrafish Model of Fetal Alcohol Exposure

In order to understand the molecular mechanisms of the ethanol-induced developmental damage, we have developed a zebrafish model of ethanol exposure and are exploring the physical, cellular, and genetic alterations that occur after ethanol exposure1.  We then seek to find potential interventions and rapidly test them in this animal...

imaging and quantification of the hepatic vasculature of mice using ultrafast doppler ultrasound

Author Spotlight: Advanced Ultra-Fast Ultrasound for Precise Liver Vascular Imaging Without Contrast Agents

Ultrafast Doppler ultrasound (UFUS) with high spatial resolution (100 &#956;m) and sensitivity represents a suitable noninvasive imaging modality for obtaining a comprehensive qualitative overview of the hepatic vasculature. UFUS also facilitates quantitative measurements of the microvasculature, aiming to enhance our understanding of vascular disease...

Imaging and Quantification of the Hepatic Vasculature of Mice Using Ultrafast Doppler Ultrasound

functional transcranial doppler ultrasound for monitoring cerebral blood flow

Functional Transcranial Doppler Ultrasound for Monitoring Cerebral Blood Flow

Functional transcranial Doppler ultrasound complements other functional imaging modalities, with its high temporal resolution measurement of stimulus-induced changes in cerebral blood flow within the basal cerebral arteries. This Methods paper gives step-by-step instructions for using functional transcranial Doppler ultrasound to perform a functional imaging...

Rodent models of cardiac pathophysiology represent a valuable research tool to investigate mechanism of disease as well as test new therapeutics.1 Echocardiography provides a powerful, non-invasive tool to serially assess cardiac morphometry and function in a living animal.2 However, using this technique on mice poses unique challenges owing to the small size and rapid heart rate of these animals.3 Until recently, few ultrasound systems were capable of performing quality echocardiography on mice, and those generally lacked the image resolution and frame rate necessary to obtain truly quantitative measurements. Newly released systems such as the VisualSonics Vevo2100 provide new tools for researchers to carefully and non-invasively investigate cardiac function in mice. This system generates high resolution images and provides analysis capabilities similar to those used with human patients. Although color Doppler has been available for over 30 years in humans, this valuable technology has only recently been possible in rodent ultrasound.4,5 Color Doppler has broad applications for echocardiography, including the ability to quickly assess flow directionality in vessels and through valves, and to rapidly identify valve regurgitation. Strain analysis is a critical advance that is utilized to quantitatively measure regional myocardial function.6 This technique has the potential to detect changes in pathology, or resolution of pathology, earlier than conventional techniques. Coupled with the addition of three-dimensional image reconstruction, volumetric assessment of whole-organs is possible, including visualization and assessment of cardiac and vascular structures. Murine-compatible contrast imaging can also allow for volumetric measurements and tissue perfusion assessment.

This video demonstrates use of a rail-mounted high-frequency ultrasound probe to perform echocardiography on an anesthetized mouse. The methods describe both conventional two-dimensional and M-mode measurements of cardiac function in addition to newer, more powerful tools such as color Doppler, strain analysis, as well as general and targeted contrast imaging.

Rodent models of cardiac pathophysiology represent a valuable research tool to investigate mechanism of disease as well as test new therapeutics.1  ...

Fetal alcohol syndrome (FAS) is a severe manifestation of embryonic exposure to ethanol. It presents with characteristic defects to the face and organs, including mental retardation due to disordered and damaged brain development. Fetal alcohol spectrum disorder (FASD) is a term used to cover a continuum of birth defects that occur due to maternal alcohol consumption, and occurs in approximately 4% of children born in the United States. With 50% of child-bearing age women reporting consumption of alcohol, and half of all pregnancies being unplanned, unintentional exposure is a continuing issue2. In order to best understand the damage produced by ethanol, plus produce a model with which to test potential interventions, we developed a model of developmental ethanol exposure using the zebrafish embryo. Zebrafish are ideal for this kind of teratogen study3-8. Each pair lays hundreds of eggs, which can then be collected without harming the adult fish. The zebrafish embryo is transparent and can be readily imaged with any number of stains. Analysis of these embryos after exposure to ethanol at different doses and times of duration and application shows that the gross developmental defects produced by ethanol are consistent with the human birth defect. Described here are the basic techniques used to study and manipulate the zebrafish FAS model.

In order to understand the molecular mechanisms of the ethanol-induced developmental damage, we have developed a zebrafish model of ethanol exposure and are exploring the physical, cellular, and genetic alterations that occur after ethanol exposure1. We then seek to find potential interventions and rapidly test them in this animal model.

Fetal alcohol syndrome (FAS) is a severe manifestation of embryonic exposure to ethanol. It presents with characteristic defects to the face and organs, ...

A Novel Surgical Approach for Intratracheal Administration of Bioactive Agents in a Fetal Mouse Model

Prenatal pulmonary delivery of cells, genes or pharmacologic agents could provide the basis for new therapeutic strategies for a variety of genetic and acquired diseases. Apart from congenital or inherited abnormalities with the requirement for long-term expression of the delivered gene, several non-inherited perinatal conditions, where short-term gene expression or pharmacological intervention is sufficient to achieve therapeutic effects, are considered as potential future indications for this kind of approach. Candidate diseases for the application of short-term prenatal therapy could be the transient neonatal deficiency of surfactant protein B causing neonatal respiratory distress syndrome1,2 or hyperoxic injuries of the neonatal lung3. Candidate diseases for permanent therapeutic correction are Cystic Fibrosis (CF)4, genetic variants of surfactant deficiencies5 and &alpha;1-antitrypsin deficiency6.
Generally, an important advantage of prenatal gene therapy is the ability to start therapeutic intervention early in development, at or even prior to clinical manifestations in the patient, thus preventing irreparable damage to the individual. In addition, fetal organs have an increased cell proliferation rate as compared to adult organs, which could allow a more efficient gene or stem cell transfer into the fetus. Furthermore, in utero gene delivery is performed when the individual's immune system is not completely mature. Therefore, transplantation of heterologous cells or supplementation of a non-functional or absent protein with a correct version should not cause immune sensitization to the cell, vector or transgene product, which has recently been proven to be the case with both cellular and genetic therapies7.
In the present study, we investigated the potential to directly target the fetal trachea in a mouse model. This procedure is in use in larger animal models such as rabbits and sheep8, and even in a clinical setting9, but has to date not been performed before in a mouse model. When studying the potential of fetal gene therapy for genetic diseases such as CF, the mouse model is very useful as a first proof-of-concept because of the wide availability of different transgenic mouse strains, the well documented embryogenesis and fetal development, less stringent ethical regulations, short gestation and the large litter size.
Different access routes have been described to target the fetal rodent lung, including intra-amniotic injection10-12, (ultrasound-guided) intrapulmonary injection13,14 and intravenous administration into the yolk sac vessels15,16 or umbilical vein17. Our novel surgical procedure enables researchers to inject the agent of choice directly into the fetal mouse trachea which allows for a more efficient delivery to the airways than existing techniques18.

We developed a novel surgical approach for intratracheal administration of bioactive agents into the mouse fetus. The delivery route is more efficient in targeting the fetal mouse lungs than the commonly used intra-amniotic injection. This procedure has to date not been described in a mouse model.

Prenatal pulmonary delivery of cells, genes or pharmacologic agents could provide the basis for new therapeutic strategies for a variety of genetic and ...

Transgenic mice displaying abnormalities in cardiac development and function represent a powerful tool for the understanding the molecular mechanisms underlying both normal cardiovascular function and the pathophysiological basis of human cardiovascular disease. Fetal and perinatal death is a common feature when studying genetic alterations affecting cardiac development 1-3. In order to study the role of genetic or pharmacologic alterations in the early development of cardiac function, ultrasound imaging of the live fetus has become an important tool for early recognition of abnormalities and longitudinal follow-up. Noninvasive ultrasound imaging is an ideal method for detecting and studying congenital malformations and the impact on cardiac function prior to death 4. It allows early recognition of abnormalities in the living fetus and the progression of disease can be followed in utero with longitudinal studies 5,6. Until recently, imaging of fetal mouse hearts frequently involved invasive methods. The fetus had to be sacrificed to perform magnetic resonance microscopy and electron microscopy or surgically delivered for transillumination microscopy. An application of high-frequency probes with conventional 2-D and pulsed-wave Doppler imaging has been shown to provide measurements of cardiac contraction and heart rates during embryonic development with databases of normal developmental changes now available 6-10. M-mode imaging further provides important functional data, although, the proper imaging planes are often difficult to obtain. High-frequency ultrasound imaging of the fetus has improved 2-D resolution and can provide excellent information on the early development of cardiac structures 11.

Fetal and perinatal death is a common feature when studying genetic alterations affecting cardiac development. High-frequency ultrasound imaging has improved 2-D resolution and can provide excellent information on early cardiac development and is an ideal method to detect the impact on cardiac structure and function prior to death.

Transgenic mice displaying abnormalities in  cardiac development and function represent a powerful tool for the understanding  the molecular mechanisms ...

A Mouse Fetal Skin Model of Scarless Wound Repair

Early in utero, but not in postnatal life, cutaneous wounds undergo regeneration and heal without formation of a scar. Scarless fetal wound healing occurs across species but is age dependent. The transition from a scarless to scarring phenotype occurs in the third trimester of pregnancy in humans and around embryonic day 18 (E18) in mice. However, this varies with the size of the wound with larger defects generating a scar at an earlier gestational age. The emergence of lineage tracing and other genetic tools in the mouse has opened promising new avenues for investigation of fetal scarless wound healing. However, given the inherently high rates of morbidity and premature uterine contraction associated with fetal surgery, investigations of fetal scarless wound healing in vivo require a precise and reproducible surgical model. Here we detail a reliable model of fetal scarless wound healing in the dorsum of E16.5 (scarless) and E18.5 (scarring) mouse embryos.

During mammalian development, early gestational skin wounds heal without a scar. Here we detail a reliable and reproducible model of fetal scarless wound healing in the cutaneous dorsum of E16.5 (scarless) and E18.5 (scarring) mouse embryos.

Early in utero, but not in postnatal life, cutaneous wounds undergo regeneration and heal without formation of a scar. Scarless fetal wound healing occurs ...

Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo

A complex integration of molecular and electrical signals is needed to transform a quiescent uterus into a contractile organ at the end of pregnancy. Despite the discovery of key regulators of uterine contractility, this process is still not fully understood. Transgenic mice provide an ideal model in which to study parturition. Previously, the only method to study uterine contractility in the mouse was ex vivo isometric tension recordings, which are suboptimal for several reasons. The uterus must be removed from its physiological environment, a limited time course of investigation is possible, and the mice must be sacrificed. The recent development of radiometric telemetry has allowed for longitudinal, real-time measurements of in vivo intrauterine pressure in mice. Here, the implantation of an intrauterine telemeter to measure pressure changes in the mouse uterus from mid-pregnancy until delivery is described. By comparing differences in pressures between wild type and transgenic mice, the physiological impact of a gene of interest can be elucidated. This technique should expedite the development of therapeutics used to treat myometrial disorders during pregnancy, including preterm labor.

Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo

This manuscript provides a protocol for implanting telemeters in the mouse for the purpose of measuring intrauterine pressures during pregnancy.

A complex integration of molecular and electrical signals is needed to transform a quiescent uterus into a contractile organ at the end of pregnancy. ...

Cell and gene therapy are exciting and promising strategies for the purpose of cardiac regeneration in the setting of heart failure with reduced ejection fraction (HFrEF). Before they can be considered for use, and implemented in humans, extensive preclinical studies are required in large animal models to evaluate the safety, efficacy, and fate of the injectate (e.g., stem cells) once delivered into the myocardium. Small rodent models offer advantages (e.g., cost effectiveness, amenability for genetic manipulation); however, given inherent limitations of these models, the findings in these rarely translate into the clinic. Conversely, large animal models such as rabbits, have advantages (e.g., similar cardiac electrophysiology compared to humans and other large animals), whilst retaining a good cost-effective balance. Here, we demonstrate how to perform a percutaneous contrast echocardiography-guided intramyocardial injection (IMI) technique, which is minimally invasive, safe, well tolerated, and very effective in the targeted delivery of injectates, including cells, into several locations within the myocardium of a rabbit model. For the implementation of this technique, we also have taken advantage of a widely available clinical echocardiography system. After putting in practice the protocol described here, a researcher with basic ultrasound knowledge will become competent in the performance of this versatile and minimally invasive technique for routine use in experiments, aimed at hypothesis testing of the capabilities of cardiac regenerative therapeutics in the rabbit model. Once competency is achieved, the whole procedure can be performed within 25 min after anaesthetizing the rabbit.

Novel therapeutic strategies in cardiac regenerative medicine require extensive and detailed studies in large preclinical animal models before they can be considered for use in humans. Here, we demonstrate a percutaneous contrast echocardiography-guided intramyocardial injection technique in rabbits, which is valuable for hypothesis testing the efficacy of such novel therapies.

Cell and gene therapy are exciting and promising strategies for the purpose of cardiac regeneration in the setting of heart failure with reduced ejection ...

A Pleural Effusion Model in Rats by Intratracheal Instillation of Polyacrylate/Nanosilica

Pleural effusion is a prevalent clinical finding of many pulmonary diseases. Having a useful animal pleural effusion model is very important to study these pulmonary diseases. Previous pleural effusion models paid more attention to biological factors rather than nanoparticles in the environment. Here, we introduce a model to make pleural effusion in rats by intratracheal instillation of polyacrylate/nanosilica, and a method of nanoparticle isolation in the pleural effusion. By intratracheal instillation of polyacrylate/nanosilica with concentrations of 3.125, 6.25 and 12.5 mg/kg&#8729;mL, the pleural effusion in rats presented on day 3, peaked at days 7-10 in 6.25 and 12.5 mg/kg&#8729;mL groups, then slowly decreased and disappeared on day 14. When the concentration of polyacrylate/nanosilica increased, the pleural effusion is produced more and faster. This pleural fluid was detected by ultrasound examination or CT chest scanning and confirmed by dissection of rats. Silica nanoparticles were observed in the rats' pleural effusion by transmission electron microscope. These results showed that the exposure to polyacrylate/nanosilica leads to the induction of pleural effusion, which was consistent with our previous report in humans. Additionally, this model is beneficial for the further study of nanotoxicology and the pleural effusion diseases.

Here, we present a protocol to construct a pleural effusion model in rats by intratracheal instillation of polyacrylate/nanosilica.

Pleural effusion is a prevalent clinical finding of many pulmonary diseases. Having a useful animal pleural effusion model is very important to study ...

A preload challenge (PC) is a clinical maneuver that, first, increases the cardiac filling (i.e., preload) and, second, calculates the change in cardiac output. Fundamentally, a PC is a bedside approach for testing the Frank-Starling-Sarnoff (i.e., "cardiac function") curve. Normally, this curve has a steep slope such that a small change in the cardiac preload generates a large change in the stroke volume (SV) or cardiac output. However, in various disease states, the slope of this relationship flattens such that increasing the volume into the heart leads to little rise in the SV. In this pathological scenario, additional cardiac preload (e.g., intravenous fluid) is unlikely to be physiologically effective and could lead to harm if organ congestion evolves. Therefore, inferring both the cardiac preload and output is clinically useful as it may guide intravenous (IV) fluid resuscitation. Accordingly, the goal of this protocol is to describe a method for contemporaneously tracking the surrogates of cardiac preload and output using a novel, wireless, wearable ultrasound during a well-validated preload challenge.

The Frank-Starling-Sarnoff curve is clinically important and describes the relationship between cardiac preload and output. This report illustrates a novel method of simultaneous jugular venous and carotid arterial Doppler velocimetry as transient surrogates of cardiac preload and output, respectively; this approach is enabled by wireless, wearable Doppler ultrasound.

A preload challenge (PC) is a clinical maneuver that, first, increases the cardiac filling (i.e., preload) and, second, calculates the change in cardiac ...

A Modified Sonographic Algorithm for Image Acquisition in Life-Threatening Emergencies in the Critically Ill Newborn

The use of routine point-of-care ultrasound (POCUS) is increasing in neonatal intensive care units (NICUs), with several centers advocating for 24 h equipment availability. In 2018, the sonographic algorithm for life-threatening emergencies (SAFE) protocol was published, which allows the assessment of neonates with sudden decompensation to identify abnormal contractility, tamponade, pneumothorax, and pleural effusion. In the study unit (with a consulting neonatal hemodynamics and POCUS service), the algorithm was adapted by including consolidated core steps to support at-risk newborns, aiding clinicians in managing cardiac arrest, and adding views to verify correct intubation. This paper presents a protocol that can be applied in the NICU and the delivery room (DR) in relation to three scenarios: cardiac arrest, hemodynamic deterioration, or respiratory decompensation. 
This protocol can be performed with a state-of-the-art ultrasound machine or an affordable handheld device; the image acquisition protocol is carefully detailed. This method was designed to be learned as a general competence to obtain the timely diagnosis of life-threatening scenarios; the method aims to save time but does not represent a substitute for comprehensive and standardized hemodynamic and radiological analyses by a multidisciplinary team, which might not universally be on call but needs to be involved in the process. From January 2019 to July 2022, in our center, 1,045 hemodynamic consultation/POCUS consults were performed with 25 patients requiring the modified SAFE protocol (2.3%), and a total of 19 procedures were performed. In five cases, trained fellows on call resolved life-threatening situations. Clinical examples are provided that show the importance of including this technique in the care of critical newborns.

Here, we present a protocol that can be applied in the neonatal intensive care unit and the delivery room in relation to three scenarios: cardiac arrest, hemodynamic deterioration, or respiratory decompensation. This protocol can be performed with a state-of-the-art ultrasound machine or an affordable handheld device; an image acquisition protocol is carefully detailed.

The use of routine point-of-care ultrasound (POCUS) is increasing in neonatal intensive care units (NICUs), with several centers advocating for 24 h ...