Ultrasonography of the Adult Male Urinary Tract for Urinary Functional Testing
August 14th, 2019
We describe the use of high frequency ultrasound with contrast imaging as a method to measure bladder volume, bladder wall thickness, urine velocity, void volume, void duration, and urethral diameter. This strategy can be used to assess voiding dysfunction and treatment efficacy in various mouse models of lower urinary tract dysfunction (LUTD).
This protocol uses high-frequency ultrasonography to evaluate the changes in the bladder and lower urinary tract in mice. This technique allows a longitudinal examination of the urinary tract in vivo for anatomical and physiological measurements and is complementary to other techniques for measuring urinary flow. The ability to assess in vivo voiding changes in rodent models that recapitulate aspects of human urological diseases can provide insight into disease progression and treatment efficacy.
Begin by connecting a probe with a center frequency of 30 megahertz to the active port of the ultrasound system and presetting the application to abdominal imaging. For optimum contrast enhancement, shake an appropriate contrast agent in a vortex mixer for 45 seconds to encapsulate the microbubbles in solution. After confirming a lack of response to toe pinch, place an anesthetized 24-week-old C57 black 6 male mouse in the supine position on a heated platform and shave the hair from the abdomen.
Remove any remaining hair with depilatory cream and position the ultrasound probe parallel to the long axis of the bladder. Then using the XY manipulator to move the mouse as necessary, obtain long and short axis images of the bladder. Before imaging, make a midline incision through the skin and abdominal wall to expose the bladder and pre-fill a piece of flexible polyethylene tubing with saline.
Then connect a syringe equipped with a 27 gauge needle to the piece of tubing and insert the needle into the bladder. To confirm correct placement of the needle, instill 10 microliters of saline into the bladder. Replace the saline syringe with a syringe containing the vortexed contrast agent and collect another B-mode image of the catheterized bladder.
To facilitate measurement of the urethra diameter, instill a 0.5 milliliter bolus of microbubbles over a three-second period into the bladder until a urination event occurs. To facilitate measurement of the velocity, correct the Doppler sample window angle until it is parallel to the urine flow and instill a second bolus of microbubbles into the bladder. Then collect a 3D image of the full bladder.
When all of the images have been obtained, trace the outside to inside edges of the bladder wall and use the linear distance measurement tool to measure the bladder. Use the volumetric tool to trace the inside of the bladder walls to create a contour. Multiple contours can then be generated through the thickness of the bladder to calculate the volume of the bladder in the 3D acquisition mode.
For the first voiding event, use the linear distance tool to measure the urethral diameter edge to edge at the point of the Doppler velocity sample window. For the first voiding event, use the velocity time integral tool to measure the urethral velocity. Here, representative ultrasound images of a mouse bladder acquired without contrast agent are shown.
The bladder wall is echogenic and the bladder wall thickness can be measured using a software measurement package. A bladder size and shape can be rendered in 3D to determine bladder volume. Here, a representative image of a microbubble filled echogenic bladder is shown.
After low frequency ultrasonic burst, the bubbles are destroyed and the bladder becomes transiently echogenic before the bubbles reform confirming the structure as the bladder. During a urination event, the urethral lumen diameter can be acquired during urination along with the flow velocity of the urine passing through that region of the urethra. Using contrast imaging, diameter measurements can be made throughout the entire visible length of the urethra.
From these measurements, further calculations can be made to assess the urinary flow and bladder compliance. Activating the contrast agent correctly is essential to the success of this protocol. We can reconstruct rodent urinary tracts with MRI and microcomputed tomography to confirm the ultrasound flowmetry as well as introduce new complex models of urologic flow dynamics and 3D reconstruction.
We have begun using this technique to quantitatively examine models of lower urinary tract disease, the effects of aging, and the efficacy of therapeutic treatments on urinary function.
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Overview
This article describes the use of high-frequency ultrasound with contrast imaging to measure various parameters related to bladder function in mice. It provides a method for assessing voiding dysfunction and treatment efficacy in models of lower urinary tract dysfunction.
Key Study Components
Area of Science
- Neuroscience
- Urology
- Imaging Techniques
Background
- High-frequency ultrasonography allows for in vivo evaluation of the urinary tract.
- This technique provides anatomical and physiological measurements.
- It is complementary to other urinary flow measurement techniques.
- Rodent models can help understand human urological diseases.
Purpose of Study
- To assess bladder volume, wall thickness, and urine velocity.
- To evaluate void volume, duration, and urethral diameter.
- To investigate voiding dysfunction and treatment efficacy.
Methods Used
- Connect a 30 MHz probe to the ultrasound system.
- Prepare a contrast agent for optimal imaging.
- Use anesthetized mice positioned on a heated platform.
- Conduct longitudinal examinations of the urinary tract.
Main Results
- Successful measurement of bladder parameters in vivo.
- Demonstrated the ability to assess voiding changes over time.
- Provided insights into disease progression in LUTD models.
- Showed efficacy of treatments in mouse models.
Conclusions
- High-frequency ultrasound is effective for bladder assessment.
- This method enhances understanding of urological diseases.
- It can aid in evaluating treatment outcomes in research.
Frequently Asked Questions
Executive Industry Relevance
Quantitative ultrasonography of the adult male urinary tract in mouse models enables precise, longitudinal assessment of bladder and urethral function, directly supporting preclinical evaluation of lower urinary tract dysfunction (LUTD) and benign prostatic hyperplasia (BPH) therapies. This approach enhances predictive confidence in disease modeling and therapeutic efficacy, addressing a critical inflection point in translational urology research. The method's ability to deliver reproducible, in vivo functional measurements positions it as a strategic asset for portfolio triage and mechanistic de-risking in early discovery and preclinical pipelines.
Strategic Applications in Biopharma R&D
Early Discovery & Target Validation
- Enables quantitative interrogation of urinary tract function in genetically engineered and disease-relevant mouse models.
- Supports mechanistic de-risking by correlating anatomical and physiological changes with therapeutic interventions.
- Facilitates functional target validation through direct measurement of bladder and urethral parameters.
Screening & Assay Development
- Provides standardized, reproducible imaging outputs for downstream pharmacological screening.
- Delivers quantitative readouts such as bladder volume, wall thickness, and urine flow velocity for assay development.
- Enables high-throughput evaluation of candidate compounds in validated preclinical systems.
Translational & Preclinical Research
- Aligns preclinical urinary function measurements with clinical endpoints relevant to BPH/LUTS.
- Supports longitudinal tracking of disease progression and therapeutic response in vivo.
- Improves risk-adjusted advancement decisions by integrating functional biomarkers into preclinical workflows.
Pipeline & Workflow Integration
This ultrasonography protocol integrates into the discovery-to-preclinical continuum, bridging early mechanistic studies with translational efficacy assessments in mouse models of LUTD and BPH.
- Discovery Biology: Enables hypothesis testing and pathway clarification by quantifying urinary tract changes in response to genetic or pharmacological perturbations.
- Screening: Provides assay-ready, reproducible measurements for comparative evaluation of therapeutic candidates.
- Analytics: Generates quantitative outputs—bladder volume, wall thickness, urethral diameter, and flow velocity—for robust statistical analysis.
- Translational Research: Facilitates continuity from discovery through preclinical validation by modeling disease-relevant urinary dysfunction.
- Enterprise Reuse: Establishes a reusable imaging and analysis platform for diverse urological disease models and therapeutic modalities.
Operational & Enterprise Impact
- Scientific Value: Increases predictive confidence and reduces mechanistic ambiguity in urinary tract disease models.
- Operational Value: Standardizes functional testing, enabling reproducibility and scalability across studies.
- Strategic Value: Informs go/no-go decisions and enhances capital efficiency by providing early, quantitative efficacy data.
- Portfolio Impact: Supports risk-adjusted prioritization and advancement of urology-focused therapeutic programs.
Implementation Considerations
- Requires expertise in high-frequency ultrasound imaging and small animal handling.
- Demands access to advanced imaging instrumentation and analytical software for 3D reconstruction and flowmetry.
- Necessitates cross-team standardization of imaging protocols and measurement criteria.
- Adaptation may be needed for different mouse strains or disease models to ensure data comparability.
- Contrast agent preparation and administration are critical for optimal imaging and measurement accuracy.
