Pressure-Volume System

What does the Millar Pressure-Volume System (MPVS) and the PVAN software measure?

The MPVS measures high fidelity left ventricular pressure and relative volume continuously in the intact beating heart of small animals such as transgenic mice and rats. Millar's PVAN software uses the pressure and volume data in a variety of hemodynamic calculations to assess cardiac performance.

Why use MPVS?
With hundreds of P-V systems installed across the globe in pharmaceutical companies, biotech firms, research organizations, hospitals and universities, Millar has a proven track record in the rapidly expanding field of small animal hemodynamic research. The MPVS is the newest generation of Millar’s groundbreaking cardiovascular research systems.

Designed specifically for use with transgenic mice and rats, the MPVS allows beat-by-beat monitoring of ventricular performance. It enables pressure-volume studies using a single ultra-miniature catheter coupled with data acquisition and analysis tools. It is the least invasive, highest precision, most comprehensive and cost-effective solution available on the market today.

How does the mPVS link genetics & hemodynamic research?

By allowing continuous assessment of left ventricular pressure and volume from the intact beating hearts of transgenic mice, the MPVS provides the first link between molecular biology, biochemistry, and the physiological phenotype. Using the system, investigators have the power to study the physiological expression of a specific gene manipulation. By studying pressure and volume changes in intact circulation, researchers can evaluate the genetics of heart disease and the importance of individual genes in normal physiology.

Why Use Mice and rats?
Over the past few decades there has been a dramatic increase in the use of mice for cardiovascular research. Many investigators are scaling down from their large animal research models to mice and rats because of their small size, low maintenance, rapid gestation, highly characterized genome, and similarities to the human cardiovascular system.

Characterizing the cardiac phenotype through the left ventricular P-V analysis of transgenic mice and rats had been problematic because of the small size of their hearts and rapid heart rates. By developing the MPVS, Millar solved the problem and has provided the research community with the miniature tools necessary to comprehensively assess hemodynamic parameters and cardiac function in extremely small animal models.

How does the MPVS measure volume in the Left Ventricle of the heart?

The MPVS utilizes the principle that the volume of a uniform electrical conductor is directly proportional to its electrical conductance, or inversely proportional to its resistance. The MPVS hardware emits a low amplitude, constant current signal through the excitation electrodes on the catheter and into the left ventricle.

This output signal creates an electric field within the ventricle. The resistance of the ventricular blood pool changes as the heart expands and contracts during the cardiac cycle. The resulting voltage potentials are measured by the sensing electrodes on the catheter and are inversely proportional to the internal volumes of the heart chamber.

The sensed signal is amplified, rectified, filtered, and inverted by the signal conditioning electronics within the MPVS hardware to provide a digital and/or analog output that can be sent to a data acquisition system for recording and analysis.

Since the MPVS uses an indirect method to measure volume, the output is presented in Relative Volume Units (RVU's) until a blood calibration is performed to establish a relationship that converts RVU's into standard units of volume.

How can one ensure the reliable performance and accurate data from MPVS?

The user should read all of the associated instruction manuals before experimenting with the system. Care in the experimental preparation and care in handling of the catheters during use and during cleaning after each use are extremely important. Care in calibration will help ensure collection of accurate data. When using a volume calibration cuvette, make sure that:

• The cuvette is maintained at body temperature.
• Blood clotting in the cuvette is minimized.
• The cuvette is properly cleaned before performing the experiment.
• The catheter is centered within the cuvette.
• All four catheter electrodes are submerged in the blood used to fill the cuvette.

Can a researcher purchase just one component of the system?

Yes. Each component of the MPVS may be purchased separately with the exception of the catheter. However, the MPVS was developed to fulfill the need for a complete, "turn-key" small animal pressure-volume system; each component is designed to best complement the other, affording optimum results when used together. For optimal integration of your own equipment and components of the MPVS, please contact our Customer Service Department directly.

Does Millar provide on-site training for the MPVS?

Yes. However, the purchase price of the MPVS packages does NOT include on-site training. The MPVS is inherently user-friendly and comes with step-by step installation instructions and detailed operation manuals; therefore, instead of building this cost into the price of the system, Millar affords you the option to purchase on-site training and service at your discretion. The cost of on-site training may be obtained from our Customer Service Department.

What kind of warranty or repair service is available with the MPVS?

Millar Instruments, Inc. offers a six-month warranty on catheters covering defects in workmanship and/or materials. The MPVS pressure-volume instrumentation is warranted for one year against defects in workmanship and/or materials. Computers and associated data acquisition systems are covered under the original manufacturer's warranty. For more details, please contact Millar's Customer Service Department.

Conductance Method

What is the significance of conductance output being processed as the reciprocal of resistance?

The general equation for volume measurement is given to be:

Where alpha is the volume calibration factor, rho is the resistivity of the blood, L is the segment length, G is the conductance and Gp is the parallel conductance of surrounding structure.

Since this equation is written with volume directly proportional to conductance, a system that outputs conductance rather than resistance, such as the MPVS, allows real-time viewing of pressure-volume loops on an oscilloscope without the need for computer pre-processing.

How does the conductance method compare with other direct or indirect methods of volume measurement?

A direct method of measuring blood volume is to euthanize the animal and collect blood in a graduated cylinder. Similar direct measurements of blood volume can be made with a number of isolated heart preparations.

Where physiological parameters are to be studied, a combination of indirect methods is normally required. Such methods include sonomicrometry, thermodilution or saline dilution, and conductance. Each method has its advantages and disadvantages and, because they are indirect, the accuracy of each depends on several factors. For example, the thermodilution technique commonly used in human studies is simple, standardized, and relatively safe, but it provides only an average value of cardiac output with potential for significant error.

Sonomicrometry requires extensive, careful surgery, and accuracy depends on how well the three axes selected represent true volume throughout the cardiac cycle. The conductance method requires a relatively simple catheterization and provides continuous, high fidelity, beat-by-beat relative volumetry.

However, factors such as parallel conductance must be considered since they may introduce absolute errors even when a careful blood calibration is used to convert Relative Volume Units (RVU's) to standard units of volume.

Why not use another method such as echocardiography?

When comprehensive assessment of cardiac performance was first established through studies with dogs, ultrasonic imaging (a.k.a. echocardiography) was soon abandoned for the more quantitative data provided through the pressure-volume catheter method.

This method, as opposed to ultrasonic imaging, plotted pressure against volume, generating a pressure-volume loop. These P-V loops enabled real-time monitoring of pressure and volume in the whole heart, rather than sections, on a beat-to-beat basis.

Now, with the MPVS, researchers finally have the same comprehensive, accurate pressure-volume conductance methods available for use with mice and other small animals.

RVU's

How are Relative Volume Units (RVU) useful?

Until a blood calibration is performed to develop a relationship that converts RVUs to standard units of volume, RVUs may be used in many calculations where relative volume changes in the subject are important.

Tracking relative changes in left ventricular volume are important with progressive changes in preload or afterload, with changes in heart rate and/or blood pressure during exercise, or with changes in level of anesthesia.

Parallel Conductance

What is parallel conductance?

Parallel conductance refers to the conductivity of the heart muscle that surrounds the left ventricular blood pool. Ideally, the current applied to the excitation electrodes on the catheter should go through the blood only.

In reality, some of the applied current flows into the surrounding muscle, which is also a conductor, often causing an overestimation of the blood volume within the ventricle.

Because the heart muscle acts as a shunt to the applied current, this effect is referred to as parallel conductance or parallel resistance, or in volume calculations as parallel volume (Vp).

How does one obtain a value for the parallel volume (Vp)?

To obtain a value for Vp a researcher must perform a saline bolus calibration. It is recommended that the researcher inject a hypertonic saline bolus into the research animal near the end of the experiment. The volume of the bolus will depend on the size of the animal but must be low compared to the total blood volume (ex. 10 µl of 15% hypertonic saline is common for mice, where the approximate total blood volume is 5.5 ml per 100g of body weight).

The goal of injecting a saline bolus is to change the conductivity of the left ventricular blood pool without changing the volume or the pressure within the ventricle. If performed successfully, the saline calibration will cause an offset in the volume data visible as a shift in the P-V loops to the right.

The resulting data file can be entered into Millar's PVAN software to estimate a value for Vp.

How does PVAN use the saline calibration data file to calculate Vp?

Millar's PVAN software is configured to help researchers account for the parallel volume of the heart muscle surrounding the ventricle.

The parallel volume is found by solving a system of linear equations to locate the intersection of two lines, one represented by the saline calibration data, and the other by Ved = Ves.

The Ved = Ves line represents equal end-systolic and end-diastolic volumes; thus, a heart chamber empty of blood. As a result, the value at the intersection of the Ved = Ves and saline calibration lines represents the parallel volume of muscle tissue only (Vp).

The PVAN calibration screen contains a calibration conversion formula that allows the researcher to include a value for Vp. The parallel volume is subtracted from the calibrated volume in the volume conversion formula.

Catheter Design

What does "French size" mean?

"French size" is a scale used to identify the outer diameter of a catheter. French scale units are obtained by multiplying the outer diameter of the catheter in mm by 3. Likewise, multiplying the French size by .33 will give the outer diameter of the catheter in mm.

Another useful conversion relationship is the multiplication of inches by 25.4 to obtain mm. For example, 1.4F = 0.46 mm (O.D.) = .018" (O.D.).

What size catheter should be used?

The size catheter required depends on the preferences and specific requirements of the researcher as well as the size of the experimental subject. Millar catheter sizes generally refer to the largest diameter along the usable length of the catheter. For example, on model SPR-839 for mice, the outside diameter (O.D.) at the pressure sensor is .018" (.46 mm, 1.4F), at the electrodes the O.D. is .015" (.38 mm, 1.15F), and at the catheter body the O.D. is .011" (.28 mm, .84F). Millar calls the SPR-839 a 1.4F catheter because that represents the largest dimension along the working length of the catheter body. Diameters are reduced where possible to minimize obstruction to blood flow. The smallest available catheter should be used with mice, but larger diameter catheters are recommended, where practical, with rats and other larger experimental animals.

Customer Service will be happy to help you with the selection of the correct catheter for the particular animal you are using.

Why use four electrodes?

Conductance measurements to assess blood volume ideally should include conductance of blood alone. In reality, they also include conductivity of surrounding tissues, often called parallel conductance, and of the electrodes-to-blood interface, or series conductance. On a typical single segment catheter there are four electrodes and a pressure sensor. There is a pair of electrodes on each side of the pressure sensor. The electrode-to-blood interface factor can be eliminated with the four-electrode system; two excitation electrodes (outer electrodes) are used to apply a constant current to the subject and two signal electrodes (inner electrodes) are used to sense voltage changes in the electric field. Constant current assures that the excitation signal through the subject is independent of excitation electrode size or contact area. The use of a high impedance amplifier assures that there is negligible current drawn from the two signal electrodes and that the voltage detected is independent of signal electrode size or contact area.

What electrode spacing should I use?

Standard electrode spacing is useful for specific research subject sizes. A 4.5 mm signal electrode spacing is appropriate for mice. A 9 mm signal electrode spacing is appropriate for normal sized rats. A 6 mm signal electrode spacing is appropriate for small sized rats. Customer Service will be happy to help you evaluate your specific research requirements to establish the correct electrode spacing for the particular research animal you are using. Custom spacing is also available.

Why use different electrode spacing?

The electrode spacing helps insure the creation of an electric field that is adequate for measuring volume changes within the ventricle. The 4.5 mm segment is derived from the average length of a normal mouse left ventricle while the 9 mm segment is derived from the average length of a normal rat left ventricle. Again, custom electrode spacing is available. Please contact Customer Service with your specific requirements.

Why are these catheters so short?

The catheter length is minimized to help avoid electrical interference. This interference may come from lab equipment but also from "cross-talk" between the wires inside the catheter itself. To avoid signal disruption, keep signal cables as short as possible and neatly coiled up if they are too long. Keep appliances and power lines as far away from the subject and signal cables as possible.

Catheter Handling

What precautions should be taken to get the most use from a pressure-volume catheter?

Treat these small pressure-volume catheters as the delicate precision instruments they are. Do not kink, fold, pinch (as with forceps), or cut the catheter. The pressure sensor on the catheter tip is extremely sensitive in order to detect small changes in blood pressure. Treat both the electrodes and the pressure sensor with care. Immediately after withdrawal of the catheter from the animal, submerge the catheter (w/out immersing the connectors) in saline or distilled water until there is time to properly clean the catheter per the procedures outlined in the instruction manual. Following the cleaning instructions will avoid the formation of clots as well as protein build-up on the pressure sensor and electrodes and keep the catheter prepared for the next use.

Some flexing of the catheter is necessary to insert it into a rodent vessel. How much is too much?

Avoid right-angle bends! Pressure-Volume catheters are built to handle any flexing normal to good surgical practice. If the catheter has been flexed too much the electrodes could become separated from their connecting wires or solder joints, resulting in an "open electrode" and a noisy signal. If the catheter has been flexed too much, generally it will appear bent or kinked. Once a catheter has a weak point of flexure, it may not function properly. The pressure sensor and/or wires may be damaged in a similar manner resulting in a noisy pressure signal or a pressure signal that cannot be properly balanced.

How does one know if a catheter needs repairs or replacement?

Look at the catheter itself and also at the data acquired. If the catheter has kinks, folds, or cuts its reliability is suspect. If no data can be acquired or if the data acquired is noisy, different from what was expected, or different from what it was during the previous use, the catheter may be at fault.

Millar's technical support staff will be happy to discuss any problems encountered with the catheters, catheter signals, signal cables, or data acquisition components. Please call us at (832) 667-7000 or send an E-mail to info@millarmail.com.

The catheter is not kinked or cut and the signal is still noisy. What is wrong?

Look for electrical interference. The MPVS uses AC current and measures very small voltages. Possible sources of electrical interference include:

• Electrocautery devices
• Transformers
• Ungrounded operating tables
• Fans
• Ventilators/Respirators
• Warming or hot plates
• Fluorescent lamps

Often, the source and type of interference can be diagnosed by observing the frequency of the noise. A noise frequency of 100 Hz (Europe) or 120 Hz (USA) may be coming from room lights or a DC power supply. Power lines generate a 50 Hz (Europe) or 60 Hz (USA) noise signal. If possible, use DC current for lab equipment. Otherwise, shut off the interfering appliance briefly while recording data or disconnect it from the wall outlet.

Surgical Procedure

How does surgical procedure affect data acquisition?

Position of the catheter within the ventricle is critical to obtaining good pressure-volume data. The position of the catheter tip and electrodes within the ventricle depends on the surgical approach. In an open chest procedure with an apical approach, a stab must be made near the apex of the heart with a needle to create an insertion site for the catheter. The distal electrode (tip) should be placed near the aortic valve and the proximal electrode just inside the ventricle near the heart wall. Another option is a minimally invasive approach through the carotid artery and into the ventricle, with the proximal electrode at the aortic valve and the distal electrode at the apex.

Observation of the pressure-volume loops during insertion is the best way to judge whether or not the catheter is positioned properly. The researcher will learn by experience that when the catheter is positioned correctly, the loops will look clean and physiologic, with minimal signal artifact or discontinuities.

How many procedures can be expected from a pressure-volume catheter?

The catheter's life span depends on good surgical technique as well as proper handling, cleaning, and storage. Some catheters have been used in several hundred procedures, lasting many years without maintenance, while others have been damaged by traumatic procedures after only a few uses. With proper care, it is not unusual for a catheter to last for fifty or more procedures before requiring repair, refurbishing, or replacement. However, as a measure of protection, we suggest that our customers purchase a backup catheter so their experiments will not be put on hold in the event their primary catheter fails and needs to be sent back to Millar for evaluation.